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·The Symposium
on
THYRO-GONAD-ADRENAL-PITUITARY
RELATIONSHIPS
BOVENCESE
EASTITUTE CA
National Institute of Sciences of India
New Delhi
Price
Rupees Ten and Sixty-wo Naje Paise

·Council of the National Institute of Sciences of India : 1960 President ; Prof S K Mitri, DSC,TRS, Calaisia Vice Presidents : Prof K Banerjee, D SC, Calaitta Dr D M Bose, MA, PhD, Calcutta Dr B P Pal, MSc, PhD, FLS, FBS, Delhi. Prof P Parija, D Sc, Chitach. Prof NR Sen, D Sc, Calcutta Treasurer Prof Ram Bchars, MA, PhD, SCD, Delhi Foreign Secretary : Dr B Mukerji, D Sc. Lucknou Secretaries : Shri S Basu, MSc, Delle Prof P Miheshwari, D Sc, Delhi Editor of Publications : Prof R C Majumdar, Dr Phul Nat, Delhi Members of Council Dr K N Bagchu, BSc, MB, DTM,FRIC, Calcutta Dr UP Basu, DSC, Calcutta Dr KR Dixit, PhD, Bombay Prof C S Ghosh, M SC, SM (MIT), Men ATEE, Roorkee Prof S Ghosh, DSC, Allahabad Prof P S Gill, MS, PhD, FAPS, Aligarh Prof B C Guha, DSC, Calata Dr A C Joshi, D SC, Chandigarh Prof DS Kothari, MSc, PhD, Delhi Dr B C Kundu, PhD, FLS. Barrackpore Prof RP Mitra, D $c, Delhi Dr M A Moghe, MSc, PhD, Nagpur Prof B N Prasad, Dr es Sc, PhD, Allahabad Dr Atnia Ram, DSC, FSGT, Calata Dr M S Randhawa, D Sc,ICS, Delle Dr J C Ray, MD, Calcutta Denf Rhach.- Non ... D 1 + INVA IH, DLUI Past Prof P C Mahalanobis, MA, B SC, FRS, Calcutta Presidents Dr A C Ukıl, MB,FCCP, FSMF, F.AS. Calculta) (ex-offico)

·Bulletin
of the
National Institute of Sciences of India
No. 17, March, 1960
Proceedings
of
The Symposium
on
THYRO-GONAD-ADRENAL-PITUITARY RELATIONSHIPS
held at New Delhi
on October 2, 3, 4, 1959
SCIENCES
FISTITUTETES
National Institute of Sciences of India
New Delhi

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·PREFACE On the recommendation of the Committee for organisation of Symposia the Council of the National Institute of Sciences of India at their meeting iti May, 1959 deaded to organise 1 Symposiuım on Thyro-gonad adrenal-pituitary relationships at New Delhi on October 3, 3 and 4 1959 A Strcing Committee consisting of the following members was formed accordingly - Dr B Mukerji Luck # (Convenor) DE V R Khanolkar Boribay Dr S Mitra Calcutta Dr BB Dixit New Delhi Dr AB har Lucknon Dr P Flattacharya Izatnagar The Steering Conunittec secured the co-operation and help of those working in this field in different institutions both in this country and abroad As a result, abstracts of 27 papers were received three of which were from abroad It has however, not been possible to include all of them in the Bulletin as only 18 full papers were received from the authors for publication The Symposium was maugurated by Dr K N Bagch in the absence of Prof 1 and to those who helped the Institute to make this Symposium a success EDITOR

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·CONTENTS Prefirc Opening remarks , B Mukerji INI Pace V SECTION A Thyro-Gonad-litusary Afechamsus The nature of influence of the thyroid on genadaputuscary mechanisms by Amya 3 Kar Effect of mild hyperthyroidism on the reproducts performance is male and feniale rats under climatic stress (reat) by D N Mullick Sestabelary of 1131 (radio-todme) test in evaluation of thyroid function in thyroid-pituitary-gonad uiter-relationship by S R Mukherjec, Amita Basu, Parimal Kanta Chowdhury, Bijon Roz MK Chhetri Asus Mukherjee and C L Mukherjee Thyroid and Physiology of accessory sex organs by j S Rawat and A Roy Discussion Kar, S R Mukherjec, Shah, Prasad Ahuja Chairman Mukerja 3 YO 2 30 38 SECTION B Adrenocortical Gonad-Pecustary Afechanisms Infusence of castration wpon the adrenal responsiveness of hypophysectomized male rats gn en corticotrophm by SN Karken and I w Landgrebe Increased tolerance to corticoid overdosage during pregnancy by L Chedid Adrenocortical-gonad-pituitary mechanisnis by N Kakati Response of plasma coracosteroids to ACTH in various states of adrenocortical activity by Prabhakar N Shah Damodar K Mahujan aid Ernest J Borges Discussion Kar, Shah Chairman Mukerji 30 37 39 SECTION C Gonad-Pettory Alich misis The Efcut of restradiol monobenzoate on the oudative enzymes of the tat testis hy B Chowdhury, An Chowdhury and D P Sadhu Regulation of gametokanac function in intact and hypophysectomized male tond Berfume westictus Schneider by U K Banik $1 ·V1 CONTENTS Page $8 76 Studies on the effect of administration of gonadotrophins in augmenung fertility in farm animals by P Bhattacharya, F N I and S N Luktuke Clinical reports on cesacular hypoplasia in Govt breeding bulls of West Bengal by B K Basu Thyro-gonad-adrenocortical mechanisms as affected in scurvy hy Sachchidananda Banerjee Action of drugs on the release of oxytocin in unaniaesthetzed rabbits by Ranjit Roy Chowdhury Thyroid status and adrenal cortical activity by J S Rinat and A Roy Discussion Subrahmanın, Banerjee, Krishna Murthy, Prasad, Shah, Das Gupta, Kar, Ahuja 7) 03 101 SECTION D 107 113 Pituitary-Adrenocortical Mech 1MISIIS Studies on endocrinal inter-relationship using a histochemical niethod for the cvaluation of succnic dehydrogenase activity as an inder by E J de Souza and T H Rindani Influence of adrenals on the cytology and histology of gastric mucosi und the gastric secretion of pylorus obstructed rats by G B Singh and SH Zaidi Demonstration of the existence of Pre-corticotrophın precurser of cortacotrophın in the ory anterior pituitary gland by P R Das Gupca Histochemical study of adrenal cortical activity in Toad (Bufo melanoctcris) by Chandicharan Deb and Mrityunjoy Mukherzi Investigations on autotransplantation of the adrenal gland in the spleen with special reference to pituitary-adrenal relationships by A S Krishna Murthy and MB Sahasrabudhe Discussion Kar de Souza Chairman Mukerji, Singh Prasad Malaviya Krishna Murthy 117 127 130 LIST OF PARTICIPANTS 133 ·OPENING REMARKS by B MUKERJI the nervous and the endocrine systems The role of nervous system in homcostasis has been the theme of brisk study and research since the beginning of the present century when Sherington's magnun opus on this subject was published As a result neuronal interrelationships and the function of the various integrative portions of the central nervous system have been clarified A fundamnental concept of hormonal interrelationships, however, and their integrative relationships with the "milieu interieur" has been slower in evolving, most of the progress having becu made only in the past two or threc decades latter either through thus medium or through nervous pathways As a regulator of endocrme activity, the anterior lobe of the pituitary occupies an putstanding position le sunulates the adrenal cortex (ACTH and perhaps growth hormone, adrenal weight factor'), the pancreas (diabetogenic hormone), the thyroid (thyrotrophic hormone) and the gonads (FSH, LH and LTH) In this inanner it partiapates in almost all aspects of bodily functions Thus, under the influence of ACTH and possibly growth hormone, the adrenal cortex us somulated to produce glucocorticoids, perhaps mineralocorticoids and certain sex hormones particularly the androgens The coracoids influence carbohydrate metabolism, electrolyte metabolism, blood pressure, smooth muscle contraction, thymus and other lymphatic structures Theoretically some of the adrenocortical androgens should be able to maintain the male accessory genital organs in castrated animals but in practice thus possibility has not been satisfactorily realised This is but one of the many unsolved quite understood The thyrotrophic hormone (thyrotrophin or TSH) acts on the basal metabolic rate, growth, differentiation and metabolism via the thyroid hormone (thyroxine) whose production it sitmulates The thyroid and the adrenal cortex have a close partnership perhaps to maintain the cellular energetics at the optimum level The thyroid hormone also has profound influence on picuitary-gonad mechanisins several wayar directors (1) timed release of FSH and LH (2) peripheral passage of gonadotrophuns to the gonads ·1.ok.. namath.ntent of irsi Luinha haracturana he can estrogens to maintain this inhibition so that FSH and LH production again rises, and the Yanom tn widely different stimuli such as infections, intoxications, 111 B MUKFRJI OPFNING REMARKS 3) physiology of the gonads with especial reference to follicular maturation and estrogenecrction by the ovary, and sperinatogenesis and androgen sccrction by the testis (4) physiology of the accessory, sexual organs and particularly their sensitayity to homo The intricate and closely articulated nature of gonad-pituitary mechanisms can be best appreciated by a casual examination of the hormonal interrelations during the FSH and LH production In addition, they stimulate luteotrophin (prolactın) and 'lutcal phase changes lobe and inhubit FSH The regular recurrence of the female sexual cycle perhaps may be explained by the periodic production of FSH with a resultant increased Formation of estrogens, the Yatter preturn whubit FSH and LH formation to a point where the ovary fails to form suficient Some 20 years ago, r fatiguc or X-irradiation. Each of these stressor organ ped m . ummati active pi mammary gland 1 terone. process is reinitiated -Co ny cases . cally opposed The common te of biologic stress'. The dless of the nature of the rimposition of the specific y 'n n-specific stress'. prouocanye ageul actions stressor ·MUKERJT OPENING REMARKS ܬܢ - The prefactory nonspecific adaptave reaction the so-called 'alarm reaction' whuch characterizes body's response is in the nature of a call to arms of the organism's defences Detailed studies indicated that this was merely the initial phase of development of a more alarm reacm which ced adapta tion is lost Thus continued stress after adaptation leads to the stage of resistance' At this time inurement to the evocative stressor increases but resistance to other damaging agents diminishes When adaptation fails the stage of exhausaon' ensues with neutralzation of al defence mechanisms and eventual death The stressor such as trauma infection or bum acting on the targets evokes a stimulus which exates the anterior pituitary to produce ACTH under certain arcunstances If may also cause a discharge of TSH growth hormone or even gonadotrophins The nature of this first mediator between the directly injured organ and the anterior pituitary is as s, yet unknown Corticotrophın stimulates the adrenal cortex to produce pre- 1 1 reactions in the target organs such as anabolism and augmentation of granuloma formation and of allergic responses, primarily stimulaong the connective tissuc Part of this action is undoubtedly not mediated through the adrenal cortex but this direct effect sensitizes the connective tissue elements to the essentially sumular actions of the mineralo-coracoids In other words there is a peripheral synergism between growth hormone and mineralocorticoids However, by itself growth hormone cannot maintain the adrenal cortex in a functional state hence its 'corticotrophic effect is dependent on the simultancous availability of ACTH It may be mentioned in passing that growth hormone has becn considered by some to be associated with the so-called adrenal weight factor The mineralo-corticoids and the gluco-corticoids have many opposing effects Deficiencies and unbalances in either the production of the activity of these coraicords result in derangements of the normal adaptative mechanism and may result in the production of certain maladies considered to be essentially discases of adaptation 1 to stress Among these particular attention has been given to dietary protein which appears to increase the production and/or activity of growth horinone and to sodium whuch augments the effect of mineralocorticoids on certain target organs especially the kidney 1 bolic derangements which abnormally alter the sensitivity of the target organs to growth hormone ACTH or corticoids and (d) although the pituitary adrenal system plays prominent role in stress syndrome other organs which participate in the latter such as ·B MUKERJI OPENING REMARKS the nervous system, lover or Ldncy, may also respond abnormally and become the cause of disease during adaptation to stress I would like to cmphasize here that vigorous and concerted research on thyro-gonad adrenal-pituitary mechanisms might help to solve one of our grave nagonal problems Lilaca wiele role woulu OLI U Ise AVC existed, pregranicics llave veell saveu u would have terminated, diseases have been thwarted which would have proved fatal and finally the span of life has been significantly extendựd. In most respects the impact of endocrine rescarch upon man's life has been similar to that provided by other areas of biological study, all of which taken together bara in a relatively short time had a profound effect on world population If this trend 21 population increase is not checked, a serious situation resulang from food shortage 1 bound to arisc in not too distant future The only effective manner by wluch a balance been ecn food availability and population increase can be maintained us by evolving some method of controlling human fertility This does not appear to be an impossible task of the endocrine mterrelations are better understood and some method adopted through which the extremely delicate mechanism of fertilisation of the ovum by spermatozoa can be attucked at some vulnerable point A study of sex gland endocrinology alone as 11 1 ties of human fertility control and can adopt physiologic or pharmacologic means to control it at will when necessary Lastly I would like to elaborate a little more on the fact that the nervous and endocrine systems are integrated structurally and functionally in various ways and in various which are elaborated and secreted by the neuroendocrine system, are members of a untary constellation of metabolic acuvators and inhibitors that exhibit a phenomenal spectrum of individual effects and interrelated activities in the performance of its ubiquitous metabolic task, the neuroendocrine system, is interlocked functionally with the vascular system The vascular system supplies the neuroendocrine system with the raw materials required for synthesis of its internal secretions and it serves coincidentally as the body-wide distribution agency of the manifold hormones which it produces and secretes 1 1 ting the precipitate functional or organic pathology of clinical consequence However, very little

·B MUKERJI
OPENING REMARKS
has been discovered as yet about the psychobiologic mechanisins and pathways
through which environmental stress espçatállythatof a soaocultural nature, us perceived,
L.
11.
8
1) Hypogonadism in both sexes
Menstrual disorders of psychogenic origin
3) Eropthalmic goiter
4) Obesity
$) Cushung hyperadrenocorticalısm
6) Psychoscs of schizophrenic and epileptoid types
7) Hypertension
Diabetes of pancreatic pituitary and adrenocortical origin
9) Hypo and hyperpigmentation
Patuitary dwarfism and acromegaly
11) Cancer of various endocrine glands or their target organs like.
Ovarian and testicular tumors
Adrenal carcinoma with vırılısm in female
Patuitary and thyroid cancer
Breast and prostatic cancer
It will thus be realised that the field of neuroendocrinology is gaining in strength and
10)
SCEE
national welfare

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·SECTION A
THYRO-GONAD-PITUITARY
MECHANISMS

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·THE NATURE OF INFLUENCE OF THE THYROID ON GONAD
PITUITARY MECHANISMS
by AMIYA B KAR
Central Drug Research Institute, Lucknow
Hypo-or hyperthyroidism induced by pharmacologic means provoked various
types of alterations in gonadal physiology Thus, persistent thiourea administration to
TL
--
weight, the cholesterol concentration of the testis too registered a significant rise. The
latter tended to signify that the rate of genesis of androgen from cholesterol was reduced
treatment whercas, it may be recalled that 91 6 per cent of the animals not injected with
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L
3
1 1 1
L - 1
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f
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LI.1.

·4 A. B KAR INFLUENCE OF TITYROID ON GONAD PITUITARY MECILANISMS To retum to gonad-pituitary system, the cxistence of a reciprocal relationship Ăn 1 1 -.- L...hatent 11. L to sensitize the adrenal cortex to endogenous ACTH rcfcased through the agency of me thyroid hormone (S N Roy, and Kar, 1958) nharmacologic 11 IMIT 1..-t. 1 .1 The histological picture of the testis of the hormone treated rats was in no way, ditterent from that of the controls, spermatogenesis continued vigorously and the Leydig cells did not show any evident atrophic changes In contrast to the testis, 1-TIT varte interesting effects on the accessory genit reduced significantly after T gard to the absolute weight but the decrease in relative weight was not significantly ditterent from that of the controls although the trend of change was similar to that in the TIT group However, TIT exerted more drastic effects on the SV than T and this was evident from was L. conof the SV of TIT treated animals revealed that the villa-like folds of thus organ appeareu ·A. # KAR INFLUENCE OF TISYROID OV GONAD PITUITARY MECHANISMS S somewhat stunted and the secretary epithelium showed signs of atrophy at certan places tesembling that of the castrates Nevertheless such atrophy of the epuhelum was focal and not getarabized as in the castrates the organ appeared actne with innumcrable granules in the epithelmum and copious amounts of secretion in the lumen Repeated exams nation of sperms of the treated animals failed to show any particular abnormalites In view of such unusual changes in the accessory genital organs the effects of thyroid hormones on fertility was investigated However as the influence of the two on the genital organs as qualtaovely more or less semular the one (TIT) which produced more marked changes was used in these and che subsequent studies The results of thus test showed that TIT had no effect on ferality of the recipient males the latter size and Keight of the individual young remained uachanged (Kar et al 1959) The SV weight showed consistent reduction 25 before These studiсs therefore indicated that the nas tre of effect of hyperthyroidism on the genital organs of young male rats was different from that in old rats In the former an induced hyperthyroid condition interfered with testicular activity probably through 3 disturbince of pituitary gonadotrophın output and the changes in the accessory genital organs were evidently secondary to an mhubicon of androgen production by the atroplued Ley dig cells in older rats or the other hand a similar hyperthyroid condition did not seem to disturb testicular function cither gametokineuc or endocrinic although a direct anlubitory influence on the genital accessories as a consistent finding Cursot sly such effect on the accessories did not result in any commensurate reduction in ferality Nevertheless it was evident that the morphogencac nature of testicular changes whenever they occurred were fundamentally similar in hypo- or hyperthyroidism This fact suggested the normal testicular acuvey was largely dependent on an optimal ratio of the level of thyroid hormone to that of GTH any imbalance of this rado was detrimental to the functional status of the testis (S N Roy el el 1955) The reciprocal thyro-gonad pituitary mechanisms were adjusted suitably to maintain this optimal rats between these bormones Further studies were carried out to establisa that the thyroid hormone did in fact exert such a direct influence on accessory genital organs (Kar et al 1959) It was noticed that the admwistration of TIT to castrated male rats caused a significant declmc in absolute wergkt of the already infantle SV but the relative weight did not reveal a sign of cant change TP alone injected to castrates daily for 12 days caused the expected stimulation of the SV weight But when TP was given conjointly with the thyroid hormone begin ning from the seventh day of TIT administration the characterisac increase in SV weight was less in comparison to the group injected with TP alone However this difference was statistically s gnificant only with regard to the absoluce weight but when the relative weight of the orgas was considered the difference was not statistically significant in contrast prostaoc acid phosphatase activity tended to show noteworthy alternatons Thus castration caused a signif cans declinc (418 per cent) in enzynic activity from the normal controls but administration of TIT evoked as much as 72 per cent indubibon Phosphatase activity in the castrates given TTT was significantly ( sper cene) less than that in the untreated castrates TP therapy restored the enzyme activity to normal level but when TP was anjected along with TiT phosphatase activity tended to be less though not in a significant manner On percentage basis prostatic acid phosphatase activity in the group treated with TP alone was 67 ? per cent higher than in the untreated castrates but in the group simultaneously injected with TP and TIT the enzyme activity was only 21 7 per cent higher in this connection our unpublished observations indicated that the duration of action of TP too was anuenced by TIT ·6 A B KAR INFLUENCE OF TATYROID ON GOVAD PITUTTARY MECHANISMS 1 1 Studies on influence of TIT on androgen action led us to explore whether the thyroid hormones had any comparable effect on gonadotrophins at the periphery (Kar ét al 1954) Accordingly, prepubcral female rats scre injected with PMS gonadotrophin twice daily for 10 days Another group received thyroxine daily in addition to gonada trophic hornionc at the same dose and for the same period It was noticed that the ovarın weight was significantly mercase in combination with thyroune of the ovin was significantly k Simularly, the uterine weight was stimulated in gonadotrophin and gonadotrophin plus thyroxınc groups and there was virtually no difference between them in this respect Cholesterol concentration of the ovary declined significantly after gonadotrophın ad ministration but the difference between the two gonadotrophin groups was insignificant. Histological cxamination of the ovary of control animals presented a typical infantile picture with ovocytes and immature follicles in various stages of growth and atresia, The beginning of antrum formation was seen in only occasional follicles Thecal fibrocellular shin treated forniation 1 s were rare 11 CL 1 Apart from these there was hypertrophy of the interstral cpitheloid clements but the interstitium as a whole was considerably obliterated owing to the cnlargement of the follicles and bulky luteinization The entire organ was highly vascularized The gross histologic appearance of the ovary of GTH plus thyroxine treated rats was also indicative of 1 stimulatory response However certin details were noteworthy Thus healthy mature follicles and corpora luter were common as in gonadotrophin treated animals These findings suggested that thyroxine tended to reduce some of the untoward consequences of excessive and precocious excitation of the immature osary by gonado trophic hormone Thus the tendency of gonadotrophın to stimulate all of the available normal follicles to maturity irrespective of their extent of differentiation (and thereby leading to artresia hemmorhage and cyst formation) was curbed by thyroxine Howcver this effect of thyroxine was not directed against the normal physiologic action of gondo troplun on the ovary as follicular maturation ovulation luteinization and evokition of endocrine activity as shown by uterme changes were consistently observed Only the pathological effects of untimely and excessive ovarian stimulation by gonadotrophuc hormone were mmimized It was possible that thyroxine erther acted at the level of the ovary, and caused slower utilization of gonadotrophim or more likely thyroxine per mitted the sojoum of only physiologic amounts of gonadotrophın from circulation to gonads and the excess was inactivated maybe by stimulation of anti-hormone formation 171 ·A B KAR INFLUENCE OF THYROID ON GONAD PITUITARY MECHANISMS 7 tantiated by further studies it will be an important peripheral mechanism in the maintenance of gonad-pituitary bomeostasis Inadentally, we are also exploring the possibility of thyroid participation in the mechanism responsible for the formation of anagonadotrophin anti-bodies or antz-lormones Coming to the reaprocal influence of sex hormones on the thyroid mention may be made of one of our studies concerned with the effect of TP on thyroid is in doses known to evoke a differential response in the tests It is now a commonplace that low 1 C) -- 1 1 L3 trophin output 7 1 . Chris ܙ . ܬ ܬ 1 - 11 7 cularity was conspicuous and even some engorgement of the vessels was noticeable In order to determine a stimulation of pituitary che low dose of TP plus" Ath. - - A nan avaart An the shared..che 1.1- ܘܚܕܐ r ܙܘ i devoid of vacuoles The vascularity of the organ was poor in the T plys TP treated S. N Roy et al, 1956) We also examined the testis of these animals and obtamed interesting informations (Roy et al, 1955) Thus T alone caused a loss of tests weight and histological indications or real af SLA - ·8 AB. KAR INFLUENCE OF THYROID ON GONAD PITUTTARY MECHANISMS rarely beyond the primary spermatocyte stage and there was profound disorganization and sloughing of the senuiniferous eleinents The adverse effects, therefore, secmed 1 1. L. -11 T but not in a statisacally significant manner; the histological appearance too was normal. Cholesterol concentration was similar as in the controls These facts suggested that (1) the low dose of TP stimulated the thyroid through TEJ uch 2 noche noha-hand, T per se tended to siske che conde ased both TSH and gonadotrophin output of the pituitary as shown by the stimulation of both the thyroid and the testis However, depression of TSH production by T did not dampen the capacity of TP to excite pituitary gonadotropnun release. Even T with all its intrinsic ability to suppress pituitary gonadotroplun activity was evidently ineffective against such powerful seamulus provided by TP for outpouring of pituitary gonadotrophun. In other words, these studies indicated that at the primary level at least the differential action of the two doses of TP on gonadotrophın output was independent of their influence on TSH release. In the panoroma of endocrine inter-relationships which we are privileged to witness 111 this symposium the thyro-gonad-pituitary mechanisms occupy an outstanding post ton The true nature of relationship between these three endocrine glands is yet to be -..,.,.1 1 - 1,5-a - keo 501 REFERENCES Kar, A B. Karkun, j N and Datta, S, N (1959) Tch influence of thyroxine on thyrotrophic action Kar, A B, Roy, A C, Karkun, J N, and Roy, S N (1955) Responses of the adrenals of thyroide treated rats to adrenocorticotrophın hormone (ACTH) Arch Int Pharmacodyn , 107, 247 253 TT - A T T C Ter. A iani cerninal itent lity 10 lase activity and its prevention by testosterone propionate J. si induste Res, 15, 246-249 ·A 8 KAR INFLUENCE OF THYROID ON GONAD PITUITARY MECHANISMS 9 Roy A C Kar A B and Sur R N (1953) The effect of testosterone proponate on total cholesterol r 27, 276-224 Roy SN Kar A B and Dutta S N (1955) The i fluence of testosteronc propionate on the response of the restis of young rats to thyroxine Ad 11 Pl art.cosyn 102, 450-458 Roy S N Roy SK and De NN (1958) Effect of testostcronc prop orrate on the thyroid of thiou rea treated rats I dat ed Res 46 396-402 Roy S N Roy S K and Mukerj B (1956) Effect of testostero ic proponate on the thyro d of of you g male rats. I d 44 427 432 ·EFFECT OF MILD HYPERTHYROIDISM ON THE REPRODUCTIVE PERFORMANCE IN MALE AND FEMALE RATS UNDER CLIMATIC STRESS (HEAT) by DN MULLICK Anmal Nutrition Division, Indian Veterinary Research Institute, Izatnsgar, V P ABSTRACT The results obtained in the present experiment indicate that summer months depress sexual perfor mance or ferality in adult inale and female rats There was a significant difference in the weight of the tests and the diameter of the follicles of textes in sumnier and winter months Injections of different doses of thyroxine solutions have constderable positive effect in the activity of the testes as studied by the weight and the histological appearance In female rats at higher atmospheric temperature the decrease in the number of estrous cycles and their longer duration and increase in the gestation period were recorded These conditions can be improved by supplementing critical amount of thyroprotem u the dice. Considering the lower reproductive efficiency of the animals in the tropics, the present investigation was started first to study the effect of natural fluctuation of the atmospheric conditions with the activity of the reproductive capacity of the male and female rats and their rcactions with the thyroidal supplement during thermal stress groups were EXPERIMENTAL Eight niale normal adult rats of the Instituto stock of known performance were killed when they attained age of 180 days in winter months when the atmospheric temperature was 60°F In summer months (atınospheric temperature, 100°F) 48 male rats of the same strain and about 160-165, days old were divided into 6 groups One group was fed with Institute stock diet which was served control The other five supplemented by or per cent throuracil (BDH) and simultancously different doses of thyroxıne solution (BDH) ranging from 3 to 7 vom per 100 gm of live weight were injected daily for 15 days The dose of thyroxine solution to be injected was calculated from the daily body weight of the animals Food and water were provided ads The testcs of all the groups in summer and winter months were removed and weighed in a chemical balance corrected up to tenth of a nulligram They werc fixed in susa solution and paraffin cut sections were examaicd luscollogically after staining with hematoxylin and eosin The statusacal analysis of the data has been made according to the methods given by Snedecor (1946) Twelve adult female albino rats of the Institute strain were fed with stock diet in winter months when the room temperature was 60°F Similarly, 24 rats were fed on stock diet in summer months when the room temperature was 90°F Vaginal smears were taken daily in both these groups during a period of 4 weeks The selected rats have littered at least once In the summer experiment, after 4 weeks the aniinals were divided into two groups, one of which has continued on the stock diet and the other fed with the stock ·DN MOLLICK EFFECT OF HYPERTETYROIDISM ON MALE AND FEMALE RATS dict supplemented with a proprietary preparation ofo 02 per cent chyroprotein ("Protamone Durmg the period of observation the vaginal smears were obtained daily on a sude, dried m an incubator, stained with Giemsa stain and examined under the microse ape Evans and Long(1921) classification of the destrous was followed Atleast 6 oestrus cles were recorded before the animals were allowed to mato. Successful matings, the gestanon penod, and the number and weight of the liters were recorded The litters Here weighed in a group within 12 hr after birth RESULTS The average weights of the testes with the standard error were 1007186 and 7736226 mg per 100 gm of the body weight of the rat at the temperatures of 60°F nd 100°F (Table !) The weight decreased with the increase in the air temperature The ifference in the mean values between the seasons wasbughly significantaccording to Fisher's t'test in the summer month when the activity of the organ was minumum, the animals were injected daily with different doses of thyroxine ranging from 3 to 7 pgm per roogm body weight of the rats The weights of the testis gradually increased with the quantity of thyroxine injected and the values were examined statistically according to analysis of vanance to find out whether the differences in the weights of the organs due to treatments, were significant or not It was found that the F value beruv een the treatments and weights of the organs was highly significant at i per cent level TABLE 7 Show ng the average salues of ile weight or testes and the distrieter of the testolar foll cles at 60°F and 100°F and at offerru doses of thyroxule injected Weight of Testis (mg) Treatment with thyro uneugm 6 $ MUSE MUSE MUSE Asr temperature ºf MASE 300 MUSE 60 MUSE MUSE 773 +226 + 30075186 9756203 810419 8 850: 169 8925214 928+30+ Diameter of the testicular follicles (Macrons) 245469 30255 45 ** 344 61608 320 6:8 75 30606738 283 849 09 272-4+812 ** Hughly ugnificant at 1. level The histological examination of the testes of rats en winter months showed increased permatogenic acavay when compared with that in summer nionths (P11) The primary spermatocytes and spermatds were more prominent and showed marked mitouc acaviry Most of the lasına of the tubules contained nttnerous mature sperm, and the spermatogodia showed acave probferation and the cells were well organused Ar temperature 100°F testcs of rats showed decreased spermatogenic activity with atrophic and 'degenerative changes in some of the semniferous cubules and cellular disorganisaton The cells Mere very few With the injection of thyroxuse the picture gradually changed and the activity could be brought to one hke the winter stage especally in the two groups Icon friving 6 and 7 rgm of thyroxine which showed increased spermatogenic activity The ·12 DN MULLICK EFFECT OF MILD ITYPERTITYROIDISM ON THE REPRODUCTIVE 1 -fohater 1.-6.11 l. - kºtondino The effect of the different doses of thyroxine on the diameter of the follicles subjected for their significance test by analysis of variance It was found that size dif significantly at 1 per cent lose between the different doses of thyroxmnc The average number of oestrous scles and the durabon of each cyde in experimental group are presented in Table 2 The mean differences between the v for the summer and winter groups and beru een the thyroprotein treated and untr groups and untreated groups in summer are highly significant according to Fisher' test TAHLE 2 Siewing the reproduce performance of the rate Room Temperature 60°F 90°F 90' Treated with o oz Number of animals Average number of cycles Interval of cach cycle days Gestation period, days Successful mating, percent Average number of young/litter Average weight of young 12 87+0 34 4 SOLO 34 19 6+31 72 80-192 SILOS 12 611 08 496+OSI 23 OŁAI 66 76+1 04 4 5:50.4 1. 793 480 2009: 7 773 493 DISCUSSION The experiments, reported here, demonstrate rather clearly that higher atmos temperature has pronounced influence in the reproducave performances in male The level of injected thyroxine also influences the development and improvement sex organs according to the activity of the testes as judged by theu size and lustology Dempsey and Astwood (1943) and Mullick (1989) showed a decrease in the c xine secretion rate at higher temperature in rats Prestimbly, this is one of the ho thermic mechanisms that permits the animal to compensate for heat stress. The ani than good to improve the conditon of the animals, but the use of physiological may turn out to be bencficial rather than injurious to the health of the animals physiological dose should be worked out before actually any application in th therapy starts The mechanisms whereby the thyroid influences sexual function have not been gonadotropíc hormone content of the pituitary as a result of the thyroidectomy has ·DN MULLICK Istologkil Sections of testis of normal 1 luftrit uider different atmospheric temperature an i treatment with thyrowie A -Textus Scstron of rat it 100 F shou nie buted spermatoset esis and is geren truc cha ges in the tubulos 1 - Improvement of all the above conditions by suppliciting with 4 ign thyroxin per too ym bndy sucht (-Tistus section of rat it of showing numerous maturi spern and spermato BLOC cells showing activity with greater scular reg Bull Nat lust Sci Indu No 17, Place I

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·PERFORMANCE IN MALE AND FEMALE RATS UNDER CLIMATIC STRESS The 13 seported in rabbies (Van Dyke and Chew, 19+2) in rats (Stevi and Lisle, 1912) and in goats (Renske et al, 1941) Litle Information is available to indicate whether muld hyperthyroidism would increase the output of gonadotropins by the pituitary The admwistration of thyrord to rabbits resulted ma lowered content of pıtıntary É SH and mcrease in L H (Chu and You 1954) present results may also be explaned in part by a modifying effect of the thyroid hormone level on the response on ser organs to gonadotropins, since the response of the testes as increased by mild hyperthyroidism and reduced by hypothyroidism (Messes and Chandrashekar, 1948) Furtherinore the response of the accessary testasterone is increased by thyroxine (Masson 1947) The supplementacon of thyroprotein in summer months (Maqsood and Reneko 1950) to improve the general condition of the reproductive performances in males may be due to the well established role of this endocrine in controlling the general metabolusni in the body which may simultaneously help to increase the consumption of food Thus the nutrients and the higher metabolism may be chief factors in the improvement of sexual condition of the antmals in supplemented group The results of the present experiments support the view that hypothyroidism due to burgher environmental cemperature interferes with normal reproductive function in fernale albino rats also The decrease in the number of oestrous cycles and chesr longer duration in sunimer may be due to the effect of esther lack of stimulus of the general metabolic activity as a result of hypothyroidism or due to slowing down of the secretory activity of che pituitary or of both these factors on the function of the ovary The morease in the gestation period during summer observed in the present experiments is in conformity with the observations of Krohn and White (1950) on hypothyroid rats but as much less pronounced than in the case of Ukıra's (1919) bindings in rabbits The improvement in the reproductive performance of female rats during summer on supplementation of the diet with thyroprotein may be related to the well-established role of thus hormone in stimulating the general metabolism of the body and the increased consumption of food which may result therefrom Thus the mcreased intake of nutrieties and the higher metabolic rate may be the chief factors in the improvement of the reproductive performance of the animals in the group receiving the supplemented diet REFERENCES Chu J P and You S S (1945) J Endome 4, 115 Dempscy EW and Asa cod EB (1943) Endanin 32, 59 Evans HM and Long 1 A (1921) Anar lcc 21 1 Flurse V and Turner CW (1948) To Agose Exp Sta Res Bell 417 Krohn P L and White H (1950) Endocrinology 6, 375 Lac MO (1929) I 9, 410 Massort G (1947) Rev Canal De Bio 6, 78 Maqsood M and Reineke EP (1950) Amer J Physiol 162, 24 McKenzi F F and Chandrashekar B (1948)) Anon Se 7. 542 Mullick D N (1999) Indian med Res 47 393 Reineke EP Bergaman Ay and Turner CW (1941) Endarinoloer 29, 306 Richter CP (1933) Ibd 17. 153 Snedecor W (1946) Statessical methods Collegiate Press Jova Sem F and Lisle M (1942) Endowsclogy 30, 16 Vandyke }} B and Cher G (1945) Pros Soc Exp BNY 31, 327 Vera T (1919) fra Sd Ved Um Kiolo 3, 337 ·SUITABILITY OF [135 (RADIO-IODINE) IN EVALUATION OF THYR FUNCTION IN THYROID-PITUITARY-GONAD INTER-RELATION! by S R MUKHERJEE, ANITA BASU, PARINAL KANTI CHOX DIURY & Bijos R Department of Experimental Medical Sciences and MK CUTRI, ASWINI MUKITERJER & C. L MUKTIERJEE Departments of Aledicine and Aftdu tfery, Institute of Post-Gradu ite Aledical Education & Rescurch, Calaut ABSTRACT Dy po uprake in the Thyroid gun pescentage exctction of futi Ume and bound lodure (PBJ"*) in blood (indicating thyroxine or such compound) have all been quanto measured M Since the mtroduction of radio-sodine (1134) the functon of thyroid glan role of T S H (chyrotrophuc hormone of antenor pituttary), and influence of che interferring with the action of T S H on synthesis of thyroune, harc been sub re-investigation in various laboratories (Hertz et al, 1938, Hamiltos, 1938, Hertz, Already, a volununous literature (Freyers 1956, Ansell et al. 1953, Storaasls, Myant and Pochin 1949 Berson e al, 1952. Myant, 1952, McConahey and 1956, Pochin iner Howard 1988, Sheling of different sland These parameters are directed manly to assess (1) the rate of up-take of aodone by the gland (2 hours to 48 hours) (:) the rate of release of 111 labelled a hormone, and (3) rate of clearance of 1132 from thyroid gland But lodide in thyroid space" (uodine trapping by thyroid gland and subse yroidal i rord $P3 or suit on of 11 A In the last few years (Fraser et al, 1953, Goodwin et al, 1951, Rotblatt and M 1953 Paley es el 1955) importance has been attached to (1) rate and percentage ofert * Junior Research Fellow Nit10112) Institute of Saences of India ·IS SR MUKHERJLE c al . 1131 IN EVALUATION OF THYROID-FUNCTION of 1131 in che urinc and (2) sate and percentage of release of different fractions of thyronine and (3) capacity of thyroid to trap and/or bind odure Influence of exogenous' T SH, on these is being studied extensively in health and discase by German, Swedish and other European workers (Brown-Grant, 1957, Storm, 1959) and these data are being analysed along with experimental data obtained by hypophysectomy However, anterior pituitary gland releases many hormoncs, so it is rather difficult to believe that the results obtained from hypophysectomy will be comparable to thosc obtained by TSH administration or inhibition of TSH But a careful search in Iterature revealed that disorders in thyroid gland were envisaged, when patients reported for treatment of disturbances of growth or metabolism e g. cholesterol, or piginent metabolism, diabetes, cushing syndrone etc Available methods were not enough to study thyroid function In recent papers, monographs, and publications of colloquia or symposia, results of long terin clinical use of different tests for studying thyroid-function have been published and different methods have been compared The general impression from such papers is that there is little to choose between any of the similar tests as regards extreme precision (Fraser 1956. Oliver and Elis, 1957, Chaniman and Maloof, 1955) But if the tracer test result is compared with patient's clinical condition, in 80-90 per cent cases, interpretation is correct and in doubtful cases, further enquiry reveals valuable data which would have been ignored So in our unit for the last three years 1181 has been used in assessing the function of thyroid gland in cases reporting to out-patient's department or admitted in-doors with diagnosis of primary or secondary or doubtful chyroid disorders, but associated with some chnical manifestations of disturbances in growth or metabolism We have also attempted to assess the thyroid function during pregnancy (full term) before the expected date of onset of labour so that we can mcasure the radio-active rodine accuminulated in the chyroid of the new born child - which may throw some light elating to transgress of maternal hormone across the placenta in the foetus The purpose of this paper is to apprise you of these data which will reveal clearly that ne thyroid gland is intimately linked with alınost all the hormones of the anterior uztary METHIODS AND MATERIALS The subjects of the present series did not have any treatment with sodinc compounds thyroid cxtract, antithyroid compounds, inhubitung drugs beforc the investigations were carried out, nor any X-Ray exanunation was carried out previously with contrast medium Percentage uptake and Thyroid Thigh Ratio For the purpose of study of the absolute quantity of it (expressed as percentage of the administered dose) taken up by the thyroid glands, radioiodune in the form of odium iodide* having activity between 30-50 AC (11 adults) is administered orally, on mpty stomachs Counts over the thyroid glands are made at selected intervals with ze help of santillation counter and counting set No PW4032 At the time of cach yunt, care is taken that the geometrical relationship between the counting set and the Radio-10dine was obtained in the Carrier free state from M's NV Philips Roxane Pharma-Chem Industries Isotope Laboratory DUPHAR ·16 SR MUKJIERJEE et al SUITABILITY OF 1131 IN EVALUATION OF THYROID-TUNCTIU subject is kept constant As the radioactive decay occurs in radial directions, the ab: disintegration of the isotope can not be measured Therefore we measure a co 1 activity in the neck and are deducted together with the background from the values to come to the absolute thyroidal uptake expressed as counts per minute i over the area In order to convert the thyroid count as percentage of the dose administered: model of thyroid of approximately normal shape and size, is used in which : identical to that adınınıstered orally is instilled This model is known as phantom count over the phantom kept at the saine constant geometry as observed in the c individual subjects and using the same counting set is taken from these two percentage uptake of 1131 by the thyroid at any desired interval can be calculated second hour thyroid and thigh readings corrected as above the thyroid/thigh c: determuned Urary Excretion (Percent) Before administration of [131 thc subjcct is made to empty his bladder Aft dose of 1131 is administered urine during the next 24 hours is collected The volu urme is measured Sample pan contaming o 1 ml of urine is prepared and drie counted in a G M counter The total activity excreted in the 24 hours urine is esti as com het co inted dmini ring 1 expressed as the percentage of dose admmistered Protein Bound radio-Iodine (PB/131) 9 The precipitate is washed with trichloroacetic acid several times in order to elimunat inorganic iodine The precipitate is then dissolved in (2N)NaOH and the whole qu is transferred to a glass planchet and dried A standard of activity 002-005C 1 prepared at the same time and dried Counts of the PBI 131 sample and the standa taken in the same counting set under the same geometrical set up. The plasma bound activity is then calculated and expressed as percentage of the dose per plasma of contamination minimised In all estimations whether in VIVO over the neck and thigh or in the case of plan the backgrounds of scintillation and GM counters are recorded and considered.

·IN THYROID PITUITARY-CONAD INTER RELATIONSHIP RESULTS The results of investigations in normal conditions and in different disorders of thyroid gland - diagnosed cluncally are shown in Tables I & 17 TABLE 1 TEST I Hypo-Function (myroedema)etc <10% Normal range Four hours thyroid uptake Twenty four hours thyroid uptake Two hours thyroid thigh Twenty four hours urine excretion Hyper function 3 10-40% <20°. > 40° 20-50° >50% 5 Forty eight hours Plasma PB 11 2 10 > 50° >IO 40-70° <40% or 0.2% -4° per lit >0.4% per ut DISCUSSIONS Compared with data in normals the analysis of Figures in Table Il permit us to arrange disorders in Thyroid functions (diagnosed clmically) with relation to Radiolodine test as follows I Second hour tip-take (a) Very poor a Crecins (b) Poor in hypothyroidism (c) High in pregnancy (higher linut of normal or laetle above st) (d) Very high in hyperthyroidism (thyrotoxicosis) · Fourth hour up-take (a) Low or lower limit of normal range in myxocdema hypothyroidism with or without diabetes (b) Lower limit of normal in cretins (c) Very high in hyperthyroidism Twentyfourth hour up-tale (3) Very low in cretis hypothyroidism with or without dnbetes (b) High above maximum limit of normal range in hyperthyroidism Percentage excret on of 1131 in Urine up to twentyfourth hour (a) Very low in hyperthyroidismı (thyrotoxic) (b) Low in hyperthyroidism and mongolism (c) High in hypothyroidism (d) Very high in myxoedema 3 ·TABLE II REMARKS Age & Sex Case No Thyroid ThighRatio at the Second hour (2-10) Percentage uptake of [131 by Thyroid Gland at 4th hour 24th hour (10-40) (20-50) Percentage er Protein bound cretion of illodne (PBI**) in urine upto at the 48th 24th hour hour (40-70) (01-04) Cho cally referred Laboratory Diagnosis 22 м I 65 1885 29 08 (o 59 Normal 0218 Normal 2 3 F 07585 123 207 not collected Cretu 00434 Hypothyroidism 3 IZ F 6 77 19 63 33 09 27 22 029 Mongolism Normal 4 7 M 3 87 12 20 29 14 33 14 Goitre Hyperthyroidism 5 16 м 40 23 30 45 44 38 45 07308 Gigantisin Hyperthyroidism 6 41 F 8 47 23 76 45 S 38 00 032 Acromegaly Normal 18 $ R MUKJIERJEE ct al : SUITABILITY OF 131 IN EVALUATION OF THYROID-FUNCTION 7 25 M 224 833 4 65 47 37 110 activity Myxocdema Hypothyroidism 8 29 M 173 to 26 17 22 77 95 O 054 Anvicty state Treniblog Palpitation Hypothyroidism 47 M 20 84 20 23 36 82 20 76 00023 "? Cushing "My raederna 10 40 94 S 74 S 68 25 Thyroxine deficiency Hyperthyroidism Hyperthyroidism 944 D 547 ThytotoxicOSIS II 28 M 456 36 39 71 63 30 20 OSIS Thytotoxicosis IZ 46 M TO 91 16 86 36 02 4271 0 229 Normal(Cortisone-rcccded) Erophthalmos (Hyperthyroidisı) Under livestigation (?Eff Angi 1a) 13 M & 78 125 27 38 $2 23 o 2019 Normal ·TABLE II (Conced) Case No Thyroid/ Age & Sex Thigh Ratio at the Second hour (2-10) Percentage uptake of [131 by Thyroid Gland, at 4th hour 24th hour (10-40) (20-50) Percentage cxcretion of ſui in unne upto 24th hour (40-70) Proteu bowd REMARKS lodine (PBI131) at the 48th hour Clinically referred Laboratory Diagnosis (01-04) Congestive Cardiac 5619 failure with ThyrotoXICOSIS (Thyrotoxirosis?) 28 14 F TOS 46 79 07 91 81 8.45 IS 37 F 6 22 18 52 39 42 39 96 Normal 0 1617 Congestive Cardiac failure with (Myxocdema) 16 63 M 995 I8 29 44 30 35 97 0459 Diabetes Hyperthyroid Hypothyroid IN THYROID-PITUITARY-GONAD INTER-RELATIONSHIP 17 S2 M I 83 4 59 4 16 5603 no activity Diabetes 18 40 F 8 IS 14 15 39 35 39 70 0 232 Normal Normal 19 24 F 13 99 Normal Pregnancy (Full term) 20* 22 F 27 2 Pregnancy (Full term) Slightly above Normal 21. Child (Newbom) 219 New bom Norinal 22(A) 16 M 3717 Gigantism Hyperthyroidism 22(B) 18 M 683 Normal Normal 22(C) м 43 Normal Normal 19 • Indicates Intravenous administration ·THYROID AND PHYSIOLOGY OF ACCESSORY SEX ORGANS by J S RAWAT and A Roy UP College of Veterinary Science and Animal Husbandry, Mashura The hormones secreted by the various endocrine glands are directly or indirectly involved in the normal functioning of the reproductive processes in both the seres, and the thyroid hormone appears to be one of those which is intricately linked with reproduc tion in the present communication is reported the effects of hypothyroidism on the developnient of male reproductive organs and the composiaon of the semunal sesicles . 1 1 ted The seminal vesicles were dissected out alongwith their contents, weighed and macerated with sand in a glass mortar containing trichloracetic aad Fructose was esa mated in the extract The results are given in Tables I to 8 Progress of hypothyroidism as a result of thiourea administranon resulted in a decrease in weight of the seminal vesicles and statistically significant from 14th day With advancement of age, the seminal vesicles increased in size and weight inspite ·IS RAWAT & A ROY THYROID AND PHYSIOLOGY OF ACCESSORY SEX ORGANS vesicles was found to be very much higher and was nearly the same as those which were Teated with chyroxine and gonadotrophin alongwith thiourca (Table 3) The effect of the planc of nutrition is also shown by the changes in the weight of the Estis The weights of the testis of hypothyroid and pair fed control rats were markedly wer than the weight of the testis of animals fed ad Lib (Table 3) It was also noted that hough the simultaneous administration of gonadotrophın alongwith thyroxine and ourea increased the weight of testis, it was still significantly lower than those of unated animals fcd ad lib It was further observed that as hypothyroidism establishes itself, not only the weight the fructose concentration in seminal vesicles decreases markedly indicating thereby decreased testicular hormone claboration (Table 8) Adminustration of thyroxine along with throurca checks the derangement It is also evident that the seminal vesicles could not get back to normal size and function when thiourea fecding was continued for 42 days and then stopped The size of the seminal vesicles as well as the fructose content failed to return to normal during the observed post thiourea feeding period Chronic hypothyroidism, therefore, appears to produce a more permanent damage to the reproductive organs It appears that the alteration in the weight and function of the male reproductive organs during hypothyroidism is to a significant extent due to the lower caloric intake and the mam effect of undernouristuent is on the elaboration of the gonadotrophic hormones by pituitary. This is substantiated by the observation that when animals of samic weight are allowed to eat ad lib the size of the seminal vesicles as well as the various constituents it contains or produces are as high as those which received besides thtourea and thyroxine, gonadotrophen also (Table 3) I cight w me of sermal i esides and testis during the progress of hypothyroidism TABLE I D TU Fed 7 Y 14 S 21 28 95 42 minal Vesicles PFC. 76 + 20 89 + 21 ITO 214 + 21 20 TIS IU Fed 22 104 + 24 111 + 23 223 + 20 365 1620 27 Testas PFC 1340 1800 61 35 61 1450 + 30 1980 + 31 1480 1180 1040 + S5 41 1200 1820 40 34 ·24 J S RAWAT & A ROY. THYROID AND PILYSIOLOGY OR ACCESSORY SEX ORGAYS TABLE 3 Weight 111 mg of serunal vesicles and testis during the regress of hypotliyroidism following withdra thtourea after being fed for 28 and 42 days After 28 days DAYS After 42 days DAYS 7 21 28 7 14 172 417 TU Fed Seminal Vesicles 306 + 26 421 + 249 + 18 300 + 21 25 28 PFC 236 401 + 35 434 1 30 410 + 29 336 + 22 398 + 27 27 TU Fed 1660 1940 1830 + SO 2020 + 40 2000 + 31 2010 + 31 Testis 40 80 PFC 2110 2100 1560 + so 1980 + 30 2040 + 70 2150 + 32 40 34 TABLE 3 11 eight in wg of semimal vesicles and testis as aflected by simult mneous administration of Thourea and Hix continuously for 28 days Figures under the last column show weight of glands of animals fed ad lub (For ready reference weight of these glands from 28 day throurca fed animals and their pa controls have also been incorporated in the table) Pair Fed Thiourea Thiourca Thiourea Thiourca Fed at control thyroxine ACTH TX,G Semnal Vesicles 223 + 20 ΙΙΟ I. 21 250 + 32 121 Ł 31 850 st 125 Testos 1430 1760 2 1200 + 40 1350 + 44 1350 + 45 30 61 T28284 Fructose content of seinnal vesicles of anonals dining the progress of hypothyroidism DAYS Group Absolute content in mg Per 100 gm of S V m 7 TU Fed Trace Trace PFC Trace Trace 14 TU Fed Trace Trace PFC Trace Tracc ·S NWAT &A XOY THYROID AND PHYSIOLOCY OF ACCESSORY SEX ORCANS 25 TABLE 4 (Concld) DAYS Group Absolure content in mg Per 100 gm of S V an mg Tricc 2t TU Fed Trace PFC S7 + 814 70 28 TV Fed Trace Tracc PFC 47 104 + 035 130 TU Fed Trace Trace PFC 158 036 + 30 TABLES Frucose consent of seminal ves des of mals d ning regress of ypothyro d m ma netersed four 4 wecks DAYS Group Absolute contenit in my Per 100 gm of SV img 7 TU Fed 069 10 DII 60 PFC 094 40 053 60 14 TU Fed so 154 + 31 PFC 200 47 + $0 022 TV Fed 781 + 422 44 + $0 PFC 45 194 + 020 $o 23 TU Fed 188 45 OJI 7.0 PFC 190 45 + 60 023 ·26 J S RAWAT &A ROY. TITYROID AND PHYSIOLOGY OF ACCESSORY SEX ORGANS TABLE 6 Fructose content of seminal vestcies of animals during regress of hypothyroidism painta ned for 6 veces DAYS Group Absoluts content in mg Per 100 gm of SV in Tg 7 TU Fed Trace Trace PFC 43 145 + 042 13.0 14 TU Fed ooo 22 Ł 4D OTO PFC 144 + SO 020 21 TU Fed 41 149 + 053 ISO PFC .235 + 084 40 + 140 TABLE 7 Fructose content of seminal vesicles as affected by simultaneous administranon of Thiourea and Hormones.com tinuously for 28 days Figures under last column show fructose content of seminal vesicle of animals fed ad it (For ready reference the composition of these glands of 28 day thjourea fed animals and their part fed controls have also been incorporated in the table) PFC Content in Absolute mg Thiourca Thiourea Thiourea Thiourea Fed ad lib Thiourea Tx ACTH TX TXG ACTH Fructose Tracc Trace 1042 + 0350 II22 + OZgo SS10 £ 0450 6120 + 0612 ·IS RAWAT & A ROY THYROID AND PITYSIOLOGY OF ACCESSORY SEX ORGANS 27 TABLE 8 Suntrarise the results on the weight and fructose content of seminal vesicles during progress and regress of kyporthyroidism During progress of Hypothyroidism Weight of semnal vesicles Fructose 7th day 14th day 21st day 28th day 42nd day During regress of Hypothyroidism (A) Maintained for 4 weeks Х XXX XXX XXX XXX XXX (B) Maintained for 6 weeks XXX XXX 7 days 14 days 21 days 28 days 7 days 14 days 21 days XXX XXX XXX XXX XXX XXX REFERENCES Rawat JS and Roy A (1960) Thyroid status and adrenal cortical activity Bull Nat Inst Sa India Mann, T (1955) The Biochemistry of Semen Methuon and Co Ltd New York. No 17 PP 96-101 ·DISCUSSION Shah har Dr Kar do you think that the adverse effect of thyroid hormones on the testis of young puberal rats are direct or secondary to a primary effect on the pituitary? This point is engaging our attention We are also studying the FSH and LH content of tb pituitary of such tats injected with thyroxine and 3S 3 triiodothyronine I think the effect of these hormones on the testis are largely pitutary mediated although the possibility of some direct effect cannot be entirely ignored. You may recall Eartly and Leblonds work in this connection who noted some intensification of hypophysectomy effects on the rats testas by thyroxine Prasad A confusion dathyron nc on 318! 1 5 - 1. somewhat simular observations on guineapigs in Rhe Meyer s laboratory At www were able to show that LH had a direct effect on guincapig seminal vesicles Kar Have you published your results Dr Prasad? This is indeed an interesting finding because to my knowledge prolactin us the only gonadotrophic factor which has been implicated by some to have a direct trophic effect on a male accessory sexual organ and that is the prostate. Prasad These results are pending publication in Endors iology Kar Dr Mukherjee your use of 30-40 cof 1" as a tracer dose surprised me particularly when you arc using scintillation counters This is in my opinion a rather high dose for tracer purposes. Would you like to comment on this point? SR Mukherjee I agree with you Dr Kar that this is a rather hugh dose In collaboration with che Insutute of Nuclear Physics Calcutta we are trying to find out the minimum possible dose of 11 which could be used for tracer purposes Kar Could you tell me De Mukherjee whether your hyperthyroid patients with diabetes were insulin resistant or insulin responsive ? I have a feeling that diabetes in such patients were of the corticoid type due to attendent hyperadrenocorticalism and as such they were resistant to insulin SR Mukherjee Your diagnosis is probably correct Dr Kas The hyperthyroid diabets were tes stant to insulin Ahuja I fully agree with Dr Mukherjee about the absolute necessity of diagnosing the nature of thyroid disorders first before resorting to therapy Chairman Mukerji Dr Mukherjee s comprehensive studies have emphasised once again #bara va luable tool the radio-isotope technique has been mn medicine

·SECTION B
ADRENOCORTICAL - GONAD-PITUITARY
MECHANISMS

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·INFLUENCE OF CASTRATION UPON THE ADRENAL RESPONSIVENESS OF HYPOPHYSECTOMISED MALE RATS GIVEN CORTICOTROPHIN by J N KARKUN and F W LANDGREBE II elsh National School of Afedicine CardF UK ABSTRACT Adult male rats weighing between 120-160 g wete castrated and hypophysectomused 21 days L. The gonadal influence on the size and function of the adrenal gland is well documented In most mammalian species orchidectomy is followed by an increase in the size of the adrenal The process can be reversed by replacement therapy with androgen (Chester Jones 1957). Testosterone propionate is found to influence the adrenal ใน 1. 1 1 1 1 1 1 + 1 4 1 . I T 1 L 1. 3 AL 1 d L 1 1 1 1 1 1 - - with this end in view MATERIALS AND METHODS Adult rats having body weights ranging from 120 to 160 g were used for this experiment They were divided into two groups and kept under uniform husbandry conditons throughout the experimental period The first group was castrated through bilateral incasion into the scrotum while the second group was sham operated and served as the control After 21 days both the groups were hypophysectomised and the adrenal responsiveness was determined by measuring the depletion of adrenal ascorbic acid following the administration of os mg ACTH (Acthar Corticotrophin Armour) in 02 cc saline per sat into the animals The method of hypophysectomy che mode of administration of the hormone and the method of determination of adrenal ascorbic aad were the same as described by Karkun et al (1953) *Colombo Plan Trainee from Central Drug Research Institute Lucknow Ind a ·32 J N KARKUN & T W LANDCREDE INFLUENCE OF CASTRATION UPOV TILE ADA RESULTS Table 1 embodies the data regarding the change in adrenal ascorbic and foll ACTH treatment into the tivo groups of animals. The body weight of the ar the weight of their left adrenals at the time of sacrifice are also included therein TABLE I Cffect of Castration upon the Change in Adrenal Ascorbic And following Treatment with AC1 Depletion of Group of Treatment Body Weight of L. Adrenal Wt Ascorbic A Animals Animals GISE Mg SE 100G + I II Castrated Normal 133 S31643 129 0+1811 127+195 II 6+1 34 180-51 134 3+1 Depletion signuficantly greater than that in the control, t=23,05>p>02 DISCUSSION 1 7 hormone is supported by the finding of Gompertz (1958) who observed that treatm castrated rat with methyl testosterone reduced the sensitiveness of the adrenal to nous ACTH In this connection it is interesting to not the findings of Caster (1950) administered oestrogen to hypophysectomised rats and observed that the sex hor produces an increase in the sensitivity of the adrenal to cxogenous ACTH As known, oestrogen and androgen are often found to possess opposing physiological tions and this property is inade use of in their therapeutic applicanons Our conclusion, based on the present experimental results is opposed to advanced by Rennels et al (1953) In view, however, of the limited data at our du and also our different approach to the problem, it is difficult to account for discrepancy It may, however, be pointed out that Rennels et al (1953) used ac weight changes as parameter of the tissue function, while we have used a bioche change in the adrenal gland, viz adrenal-ascorbic acid change for the same purpose criterion used by us is considered to more sensitive than that employed by the fo group of investigators (Sayers et al 1948) The result of the present experiment thus indicates that testosterone 1 organ under the influence of a particular hormone is well understood REFERENCES Carter, S B (1956) J Endocrin, 13, 161 Chester Jones I (1997) The Suprarenal Cortex Cambridge University Press Gompertz D (1998) | Endocrin, 17, 107

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·38 BN KAKATI ADRENOCORTICAL-GONAD-PITUITARY MECHANISMS SCX (2) Syndromes in adults-When the adrenal cortex products severe derrangement in adults there is a tendency to "conversion to the secondary characters of the opposite " Thus adult women with cortical hyper function, gencrally develop male sccot dary scxual characters "adrenal vırılısmı " Commonest change noted in the adult male so afflicted is enlargement of brcast, sometines considerable glandular development occurs and may be possible to squeeze out milk from the gland Libido is diminished interest in women decreased, sexual potency and less frequent occurrence of nocturnal crections and seminal cnimissions is seen Gonad Pittary Relationships Gonad pituitary relationsłup has alrcady been established by various workers The chemistry of the Gonadal hormones have been established about 25 years ago Somic of then arc -(1) Oesterone (CG H, 0) (w) Ocstriole, (...) Pregnandiol, (w) Androgens (v) Androsterone and (1) Dehydroepiendrosterone (vi) Testosterone, and (you) Pro gesteronc The actions of the oestrogens is well known to all of us in normal life In state of oestrus, production of the oestrogen is due to stimulation of the anterior pituitary hormones the F'S H During the later stage of pregnancy the physiological activity of the c. 1. HAN in animals continues beyond the physiological limic, barring the uterus to seal its os and cervix and disallowing development of the foetus At times it is observed that ocstrus will rcappear on the next second heat ic after about 42 days of first service or insemination even if the anunal is pregnant This condition develops possibly due to the anterior piturtary excess as well as corpus luteal deficiencies Along with other calises oestrogen level of pregnancy has been observed to be so high at times that prolapse of the vagina repeatedly takes place, though abortion is not effected The above mentioned syndromes of ocstrogenic activities has been attempted to control with the corpus luteal hormone supplementation The condition of suppression of oestrus has been effectively achieved In certain cases of pregnancy, anti-partum prolapse of the vagina which could not be retained permanently by replacement Test har fare L1 11 ·RESPONSE OF PLASMA CORTICOSTEROIDS TO ACTH IN VARIOUS STATES OF ADRENOCORTICAL ACTIVITY. by PRABHAKER N SHAN, DAMODAR K MAHAJAN and ERNEST ) BORGES Indigot Cancer Research Centre & Tata Afemorial Hospital. Bombay 12 ABSTRACT 1 The usefulness of the plasma 17. 21-dihydroxy--20-ketosteroid response to atravenously administered ACTH in the assessment of adrenocortical function has been demonstrated II The adrenocortical response to a standard dose of ACTH in Indian subjects is comparable to that observed in American subjects III After a standard ACTH test changes m plasma 17-hydroxycorticosteroid levels were helpful in differentiating various ecological categories of Cushings syndrome and in detecting partial adreno cortical deficiency in apparently normal subjects It is now well established that hydrocortisone is the predominant adrenocoracal homone in the peripheral blood of primates including Homo Sapiens (Bush). A direct measurement of alterations in circulating concentration of hydrocortisone and some of its metabolites therefore appears to be the most practical approach for a study of the dynanucs of adrenocornical activation in man, both in health and disease We are here 1 - 1 1 1 1. 1 1 The first type of adrenal hyperactivity represents thc Cushing's syndrome, definable in simplest terinis as a sort of symptom-comples due to increased secretion of hydrocortisone The second type of hyperactivity will be described under the adrenogenital syndrome characterised by excessive androgen production without increased hydrocortisone secretion On the other hand, the clinical state of adrenocortical hypoactivity is characterised biochemically by durunution in hydrocortisone and electrolyte regulating factors of the adrenal cortex 1 1 L. syndromes MATERIAL AND METHODS In all 30 normal subjects of both sexes were selected for the study of these 27 were females and 3 were males Our criteria of normalcy were (1) Age range of 16-39 years (1) 4 *This investigation was supported in part by a grant from Me A V Mody of Unichem Laboratones ·40 P N SHAII, D X MAUAJAN &E J BORGES RESPONSE OP PLASMA CORTICOSTEROIDS (tt) Quandtatively normal menstrual flow and evidence of ovulatory menstrual cycles in females Froin each subject 20 ml of blood was drawn in a heparinised syringe These speamens were collected in the morning, between 8-9 am since there is a rhythmuc variation in the levels of plasma corticosteroids - they being at their highest carly al Thertor 17 21-dihydroxy-20-ketosteroids Of the 27 normal females, a standard ACTH test was performed in 6 cases to record adrenocortical responsiveness by infusing intravenously 2 units of ACTH in 500 cc of physiological saline solution over a period of four hours The blood samples for determination of plasma corticosteroids were drawn prior to starting and at the end of this test In addinon, there were 7 patients in whom along with the measurements of urinary . one case OBSERVATION AND COMMENTS Fig i represetits the range of values of corticosteroids in our subjects lt varted of plasma corticosteroids employed by us is closely similar to that described by these NORMAL VALUES OF 17– HYDROXYCORTICOSTEROIDS 15 NUMBER OF SAMPLES 34) 12 16 20 24 MICROGRAM PER 100 ML PLASMA FIG

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·42 P N SILAII, D K MAILAJAN & E. J BORGES RESPONSE OF PLASMA CORTICOSTEROIDS from the anterior pituitary stimulates the production of adrenal steroids Evidence exists for a 'feed back" system, which means that steroids produced by the ACTH action on the adrenal cortex, act back on the pituitary to control ACTH synthesis Thus there is a reciprocal relation between the antenor pituitary and adrenal cortex and bence rather POSSIBLE CAUSES OF CUSHING'S SYNDROME OLM us RUTAR ACTN I-OH CS MA OOTED NORMAL BILATERAL ADRENAL HYPERPLASIA TUMOUR FIG 3 ACTH nor hydrocortisone arc produced in excessive amounts in a normal person In Cushing's syndrome, on the other hand, there is always an excessive secretion of hy dro- 1. cortisone, a'2 about For philism or 'com 3 certain adrenal enzyme systems or is due to excessive secretion of another factor TABLET PLASMA 17-OH-CS VALUES IN ADRENOCORTICAL SYNDROMES NO OF PATIENTS 17-OH-OS /rogol CUSHWNG S SYNDROME 3 316 7) ADOISONS DISEASE 2 ADRENOVERTA 2 FIGURES THE WALUES SOME OTHER

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·TO ACTII IN VARIOUS STATES OF ADRENOCORTICAL ACTIVITY 43 from pituitary which potentares the steroidogenic action of ACTH is still not settled (Grumbach et al, 1953, Jaller et sl, 1956) A third possibility exists where the hugh production of hydrocortisone may be by the tumour tissue which essentially 35 independent of ACTH stimulanon (Jarlet et al, 1993) In thus particular situation, hugh arculatng level of hydrocortisone on the contrary depresses ACTH production and therefore the other gland gets atrophied Table 1 summarises the findings of plasma corucosteroids in the three chical syndromes It is evident that all the 3 cases of Cushang's syndrome have persistently elevated levels of plasma corricosteroids, while an Addison's disease, plasma corticosteroids arc dininished in Onc case and in the other, it is in the lower range of normal variation In the adrenogenital syndrome, these valucs fall in the normal range Fig 4 demonstrates the close resemblance in the clinical appearance of our 3 patients having Cushing's syndrome though the mechanism involved in cach may be quite different. One of the most difficult of clinical problems is the differentiation of the Custung's syndrome arising from hyperplasia of the adrenal cortex from an identical condition caused by a benign or malignant tumour It cannot be overemphasised that a knowledge of the underlying adrenocortical pathology is of practical importance in the therapeucic management of Cushing's syndrome We feel that, of che various tests evolved so far to delineate the various etiological categories en Cusłung's syndrome the measurement of hormones from blood and urine is probably the most dependable Table of presents the alterationis in plastna and urinary corticosteroids and urinary 17-ketosteroids in these chrce patients of Cushing's syndrome It is evident front this table that in cach patient che plasma levels of corticosteroids are sigmficantly and persistenely clevated The untary values of 17-ketogenic steroids however do not reflect the true brochemical status in one of these 3 cases The findings of 17-ketosteroids in all the chrec cases are in the normal range In general, very high 17-kecosteroids in Cushing's syndrome is strongly indicative of adrenal carcinoma but not pathognomonic of an adrenocortical carcinoma since a few such cases having normal levels of 17-ketosceroids ase reported Thus direct measurement of plasnia corticosteroids dors help in quantitative evaluation of adrenocortical function yes at fails to solve the difficult problem of differentiation PLASMA AND URINARY STEROIDAL VALUES N CUSHING S SYNDROME TABLE 3 PATIENT Rx PLASMA 11 -OH - C$ Lups/130 ml TN -kos (ngira mp3 XK 17 - * 39 5,040 01 31 6. (30 O) SO 208 99 300, (275) 6 015 B) 3 8 14 € 20 0 NORIAL EXCRETORY VALUES OF 17-KETOGENIC STEROIDS 17 KGS) OF WOMEX DIR LABORATORY ARC 40 mg / HOUR NORMAL EXCRETORT VALUES OF 19 - *ZTOSTEROIDS OF WOMEX * OUR LABORATORY ARE 13 m/24 HOUR ·P N SHAI, D K MASAJAN & E J BORGES RESPONSE OF PLASMA CORTICOSTEROIDS of various cuiological categories of Cushing's syndrome In recent years two approaches, the one anatomical and the other chenucal are carried out simultancously and perhaps would give higher accuracy in differentiating hyperplasia from tumour We, however, believe that the chemical approach is a dynamic one and therefore always preferable co start with The anatomical approach should be employed to locate the pathological lesion or to remove doubts if any still remained after chemical studies have been done Thc chemical approach appcars promising m the light of accumulating knowledge that characteristic responses are recorded following administration of a standard dose of ACTH in different types of adrenal and pituitary discases Fig s illustrates the individual variations of adrenocortical response as measured by changes in plasma corticosteroids to 25 units of ACTH given intravenously over a pcnod of 4 hours in healthy adult women This response in Indian subjects is comparable to that observed in a comparatively larger group studied by Christy et al in USAL appears that no correlation between the resting level on the onc hand and the height to wluch the steroid concentration rose following infusion, on the other The ACTH test was then perfornied on our patients having Cushing's syndromcor Addison's discase. 1 r L.A. m ITN levels remain almost unaffected in the remaining to patients It is interesting to meetion here that this cxcessivc response seen in K K is after removal of one of the hyper plastic adrenal gland, five years ago 1 1 It by husca cnocortical d evidence of presence of pituitary tumour RESPONSE TO ACTH IN NORMAL SUBJECTS WOMAN AMERICAN 71 70 PLASMA. 17-OH CORTCOSTEROIDS (MICRODA PER CET) JOS 29- 20 13 fD TNE I HOURS TIME IN HOURS FICS ·TO ACTII IN VARIOUS STATES OF ADRENOCORTICAL ACTIVITY 45 L r 275 to 2 s microgram per cent on the eleventh post-operative day, and in the patient TN the level fell from 400 to 11 t microgram per cent during the deep X-ray therap: for pituitary tumour The second part of Fig presents the results of the ACTH test donc 30 791 TO PLASMA 17-OM CORTICOSTENDOS CROUANS PER CENT) RESPONSE TO ACTH IN ADRENOCORTICAL SYNDROMES 10 DO TS 75 70- 10-11 KR 65 65 60 BO 55 os 50 301 TN 0431 3040 40 35 09- 30 30 29 25- 20 20 19- 15 10 10 5- - 0 CUSHING S SYNORONE ADDISON D(SCASE CUSHINGS SYNDROME (TN) FIC 6 during treatment in the patient TN Curve i represents the response before treatment In this state, the adrenocortical cells are working at the maximum capacity because of high or maximalendogenous secretion of ACTH by pituitary tumour Additional exogmous ACTH, therefore, fails to stinulate further production of plasma corticosteroids r.- also present as thus stage Curvę 3 depicts the response, 3 weeks after the full tumour dose 1 and conversely normal adrenal may be present with lower range of normal variation A single low initial plasma steroidal value may therefore be consistent with adequate adrenal funcuon and 10 ngid interpretation be given unless assessment of the response of adrenal cortex to ACTH is made Early recognition of partial adrenocortical deficiency is of practical importance because under conditon of severe stress acute adrerial insuffi- ·46 P N SHAII, D K MAIIAJAN & E J BORGES RESPONSE OF CORTICOSTEROIDS TO ACTH 11 1 . ACTH The ACTH test, therefore, is the final arbiter in all doubtful cases of Addison's disease It is very pertinent to mention here that salıne solution and not 5 per cent glucose in water should be used as a vehicle for ACTH since a full Addisonian crisis has been known to be provoked by the inadvertent use of the latter vehicle ACKNOWLEDGEMENTS The authors are indebted to Dr V R Khanolkar, FNT Director, Indian Cancer Rescarch Centrc, for his interest and support of this work These sincere thanks are due to Dr IC Paymaster, Superintendent, Tata Memorial Hospital for giving them the hospital facilities without which this type of work is not possible Grateful acknowledgement is made to Dr W J Tindall of Orginon Laboratories England, for Irberal supplies of ACTH used in this study REFERENCES Bush I E (1953) Ciba Foundation Collog Endocrinol 7 210 Christy N P Wallace E Z and Jailer) W (1955) jan I tvest 34, 899 Eik Nes K Sandberg A A Migeon Ć J Tyler FH and Sainucls L T (1955) J on Endocrin 19, 13 VaR KIT and W1k me T (194) Jolms Α Α Cholen Samuels LT ibid 13, 1445 ·DISCUSSION Dr Shah froni the p cture of Cushings disease whuch you have shown it seems to ine that the case is of the impure type because there are frank indications of hirsutism You may recall that according to Dorfman pure Cushings should not have any symptoms ascribable to excessive androgens In fact most of the Cushings belong to the impure type DShah what method did you employ for estimation of plasma 17-OH corticoids We used Porter Silber technique which w our hands is giving consistently good results Dr Shah I am very much interested in your finding about increase in plasma 17-OH corticoid

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·SECTION C
GONAD PITUITARY
MECHANISMS

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·THE EFFECT OF OESTRADIOL MONOBENZOATE ON THE OXIDATIVE ENZYMES OF RAT TESTIS by B CHAUDHURIA K CHOUDHURY and D P SADHU Department of Physiology & Nutrition and the Department of Med aine Bengal Veterinary College, Calcutta 37 ABSTRACT The effect of oestradiol monobenzoate (OMB) has been studied on the oxidative enzymes of rat testis There is an indubitory effect in the earlier phase which is followed by an incrcased activity of the succinoxidase system after prolonged injection of OMB The mechanism of this differential action has been discussed Histopathological changes in the testis such as atrophyand the loss of weight (Hermann and Stein 1916 Spencer et al 1932) are observed after administration of oestrogens L11 IN administration of oestrogens EXPERIMENTAL Forty male albino rats from a local strain weighing from 109 to 135 gm were used All the rats were put on the laboratory basal diet for 3 weeks and then divided into two groups Ten rats served as the control wlule the other thirty were subjected to daily intramuscular injection of 300 rgm of Oestradiol Monobenzocte (OMB) in oil suspension In Ware en fred of intervals of twelfth thirteenth were sacrificed simultaneously and determinations of tissue respiration were carried out under identical conditions Tissue suspension was made such that 2 3 ml. of the suspension when added to os ml of the substrate final concentration of the enzyme preparation in the reaction vessel would be ten per cent This was incubated at 38°C Sodrum succinate los M) and sodium ascorbate (o 114 M) were used as substrate for the determination of che succinc dehydrogenase and the cytocrome oxidase respectively The enzymaac activities were studied by determination of oxygen consumption in a Warburg's apparatus Present Address Junior Saentific Oficer Defence Science Organisation Delhi ·52 B. CHLAUDHURI, A. K CHIQUDITURY & D. B. SADITU. THE EFFECT OF OESTRADIOL RESULTS TABLE 1 Oxygen consumpuon in (lambda) per mg of wet testis ttssue of rat on different days of treatment with 3000 vgm of oestradiol monobenzoate oil suspension No of animals Days of Treatment Without substrate With Succinate With Ascorbate 0492 10 6 6 0 188 Or88 O 12 13 IS 18 24 0968 0332 0 377 0-460 0704 I 328 I 212 392 O 520 o 588 1.696 2390 o 184 392 0433 injection of OMB DISCUSSION It will be evident from the results of enzyme activity that injection of OMB sets up two different processes in the body, one causing a decrease and the other an increase of the sucanoxidase system of the teses Such a phenomenon has not been recorded so far. The amount of depression of enzyme activity at any moment will depend on the 11. nini Mc Shan, 1950) ht cu L.L,1, Nomally oestrogen would be great enough to destroy its activity completely. Besides the increased enzyme activity on the twentyfourth day suggests some definite stunulating factor and causes increased secretion stimulate the enzyme actvit are in progress to demonstrate a possible increase in adrenocoracal and anterior pituitary eriments

·MONOBENZOATE ON THE OXIDATIYE ENZYMES OF BAT TESTIS
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ACKNOWLEDGEMENTS
.' M Lahiri, Director of Veterinary Services &
Principal K. C Mukherjee of the Bengal
REFERENCES
Hermann, E and Stein, M (1916) IVier Klon II od nschr , 29, 177
Meyer, R K and McShan, WH (1950) Recent Prog Hormone Res S. 464
Morrione, TG (1944) Arch Path 37, 39
Roberts, S and Szego, C. M. (1953) J biol Chem 201, 21
Spencer, J. D'Amour, F E. and Gustavson, R G (1932) Amer ) Anat 30, 129
Tepperman J, Tepperman H M and Dick J (1949) Erdorrmology 45, 491

·REGULATION OF GAMETOKINETIC FUNCTION IN INTACT AND HYPOPHYSECTOMIZED MALE TOAD, BUFO MELANOSTICTUS SCHNEIDER by U. K BANIK Indian Institute for Biochemistry and Experimental Mediane, Calais INTRODUCTION Gametokinetic function in male includes both spermatogenesis and spermination The regulation of gametokinetic function in South American toad Bufo arenarum Hensel has been studied by Houssay and Lascano-Gonzalez (1929), Burgos (1949), and Burgos and Rufino (1952) Recently, Burgos and Ladman (1957) have studied spermatogenic mination-test in B melanostictus However, precise information is lacking regarding the regulation of spermatogenesis and spermuition in intact and hypophysectomized toad. The present paper relates to the study of gametokinetic function in male toad B melanos- Ictus EXPERIMENTAL PROCEDURE The common Indian male toad, weighing 4-40 g. were used All anunals everc maintained in most wooden box under uniform conditions throughout the period of investigation. Expo Hypophysectomy cavity of only adul d... Por extract 1. CI Spermuation test was performed according to the method of Banık and Chakravarti (1957) All injections were given subcuraneously by sungle dose OBSERVATIONS It will be evident from Fig 1 that testicular weight is reduced after hypophysectomy, which becomes marked only after four weeks Spermatogenesis is also alccred progressively In the semınıferous tubule of adult intact toad all types of germ cells (spermatogoria Annracen. In the period of --- . . naratonal wraca 1 ministration of pituitary extracts (2 mg. wet) twice a week, however, mantainca wie ·V K BANK REGULATION OF GAMETOKINETIC FUNCTIOY IN MALE TOAD TESTICULAR WEIGHT OF TOAD BUFO MELANOTICIAS 70+ SS CONTO WE 687 or TESTEI KYPPWKTORIO DAYS ANTER XVORWYSET TOWY FIG spermatogenesis and intersanal tissue like normal road up to 30-40 days But the injection of HCG 20 LU twice a week for four wecks was unable to maintain normal function of spermatogenesis The hormons anduced the formation of spermatozoa without any effect on primodel germ cells and as such the lumen of seminiferous tubule was packed with large number of spermatozoa only Other types of geran cells were virtually absent The intersacial elements too remained unconspicuous The same phenomenon was also observed in the lumen of semimferous tubule in ancact animal af 1001 U of hormone nas Tven for five days All germ cells starting from spermatogonia were transformed into spermatozoa through usual successive spermatogenuc phases It will be evident from Table i that spermuation in coad anduced by HCG remaxes Practically unchanged for 20 days after hypophysectomy and apprecable response is observed even after four weeks The influence of HCG on sperm stron of Bufo melanostictus before and after hypophysectomy Values 380 expressed as per cent of poyave response TAALI HCG Before hypophysectory 15 20 2$ Days after hypophyscomy 10 *Numbert in parcatbeses indicate the number of arunais usai in each experiment, 80 roj 100 ofu) 100 017) 20 7717) 100 (2) 100 (6) 30 66 6(0) 75 014) 33 3(6) 5004) 63 366) $0 0(0) so of4) 60 OS) ·$6 U K BANIK REGULATION OF GAMETOKINETIC FUNCTION IN 1 P. are re of hc toad was almost devoid of spermatıds and spermatozoa However, 20-25 IV of HCG accelerated the transformation of spermatogonia through chronological phases to sper matozoa and eventually the spcrmation It is also evident from Table 2 that small sustained doses of the hormone (2 s IU/day for 4 days) do not cvoke spermuations although the tempo of spermatogenesis starting from sperinatogonia was found to be accelerated TABLE 2 Influence of HCG on the spermiation of immature intact toad Bufo melanostictus Toads weighing 4 10 8 were used HCG was injected at random and sex was determined by sacrificing the animals after completion of the experiments HCG Percentage of positive response 25 ool 7) 50 10 (10) 10 0 40 0(10) ISO 60 (10) 200 100 (10) 350 100 (7) 2 s/day for 4 days 0 0(10) *Numbers in parentheses indicate the number of an mals used in each group The effect of HCG in gametokinenc function in toad scems interesting and different froin mammalian male Simpson et al (1951) believed that HCG and interstitial cells comparable to that obtained by administration of pituitary gonadotrophin However 17 1 L ا- 1 This study shows that whule spermiation could be induced by HCG the completo spermatogenesis in B melanostictus is not regulated by HCG alone ACKNOWLEDGMENT The author is grateful to Dr JC Ray, F N I for his keen interest and constant encouragement in the work Thanks are due to Mr B Basak and Mr N C Mitra for technical assistance and to Miss Anita Barua for her help in many ways ·INTACT AND TYPOPHYSECTOMIZED MALE TOAD, Bufo Melanositotis SCHNEIDER $7 25, 206 REFERENCES Batuk, U K (1959) Substances modifying spermiation in the common Indsan male toad Bufo melanos- Ictus Schnieder Ant Biochent expel Afet, 19, 219 Banik U K and Chakravartı HS (1957) Some factors influencing the common Indian malc-toad test for pregnancy Ibid, 17, 8s Bhadur: J L (1951) The role of salienta in human and mammalan pregnancy test Proc 38th Indian Sc Congr Pt 11, 171 Bhaduri,J L and Bardhan, N R (1949) Male frogs and toads as test arumal for early pregnancy and certain related conditions Saence, 109, 517 Burgos M H (1949) Estudio histologico del testiculo del sapo tupofisopny Rev Soc argent Biol. Burgos M H and Ladman, A T (1957) The Effect of purified gonadotrophins on the morphology of the testes and thumb pad of the normal and hypophysectomized autumn frogs (Rana pipiens) Endocrinology, 61, 20 Burgos, M H and Rufino, MA (1952) Accion del las gonadotrophinas suenca y corionica sobre el testiculo de sapo Rev Soc argent Biol 28, 159 Houssay B A (1954) Hormonal regulation of the sexual function of the male toad Ada physiol Latinoamer, 4.1 Houssay, B A and Lascano-Gonzalez, JM (1929) Relaciones entre la hypofisis y el testiculo en el sapo Rev Soc argent col, S, 77 Kartzman P A, Godfrd, M, Cain, C K and Doisy, E A (1943) The preparation of chononic gonadotrophen by chromatographic absorption buol Chen, 148, SOI Simpson, M E L C H and Evans H M (1951) Synergism between pituitary follicle stimulating hormone (FSH) and human chorionic gonadotrophin (HCG) Endocrinology. 48, 370 ·Experiments and clinical observations on the use of gonadotrophins STUDIES ON THE EFFECT OF ADMINISTRATION OF GONADOTROPHINS IN AUGMENTING FERTILITY IN FARM ANIMALS by P BHATTACHARYA, F N I and S N LUKTUKE Indian Veterinary Research Institute Izatnagar, UP ABSTRACT Coss and heifers with mactive ovaries were in ccted with varying doses of PMIS Oestrus Wa: induced in most of the treated cows within 3-4 days after treatmus Pregnances were obtained in 22 out of 28 animals as a result of administratio : f PMS PUIS was admuistered to 102 buffalo females un ano strum various stages of oestrous cycle and carly pregnancy Multiovulations occurred when the hormone was injected during the follicular phase of the destrous cycle and during the mid-oestrous cycle in the presence of corpus luteum which was subsequently enucleated at various intervals after treatment Singk ovulation occurred in a large number of the anoestrous animals The ovaries of the pregnant a umals treated with PMS were considerably enlarged The ovarian weight varied from 17--82 sms Rams were trcated with PMS with a view to determining the effect of administration of hormone during summer when the atmospheric temperature is high and humid and when the quali of semen is comparatively poor Semen obtained from the experimental animals was of a superio quality than that obtained from the treated oncs Buffalo bulls simularly treated with PMIS did na give satisfactory response The discovery of a potent gonadotrophic hormone by Cole and Hart (1930) in the blood serum of pregnant mares led to a considerable amount of rescarch on the possibility of augmenting fertility in livestock by hormone administration Cole and Miller (1933) were first to report the effect of administration of gonadotropluc hormones on anoestrous sheep They could induce ovulation in the quiescent ovary of the anoestrous ctve by a single or repeated injections of equme gonadotrophın but although ovuladon was obtamed it was generally unaccompanied by oestrus With this work was initiate a series of investigations in various species These studies were reviewed by Luktuke (1954) Hammond and Bhattacharya (1944) conducted an claborate investigation on the control of ovulation in the cow by hormonal methods They demonstrated that superovulation could be obtained by injections of horse pituitary extracts or PMS if the corpus luteum was regressing or was manually removed Folley and Malpress (1944) injected cows with PMS in the luteal phase of the cycle Though the development of follicles was stimulated, no ovulation occurred of functional infertility in cattle have also been reported by vanouska et al 1940. Bhattacharya et al 1941, Asd, 42 Cameron, 10 Luktuke and Bhattacharya, 1948 Abmaf. Effect of administration of gonad ovaries and in Folley and M by (1934 sons hat estic anir 1949) 典 It fecundity in cattle was studied by C Hamnond and Bhattack (1944) TT lation in cattle has (1949) Umbaugh (1 suitable condicons of as fifty follicles ovul ial u acharya 1 RT ons cart int ·P BILATTACHARYA & S N LUKTURE FERTILITY IN FARM ANIMALS 59 malaovulation has become a problem of great importance m researches peruaug to the transplantation of fertilszed ova in cattle Casidi et al (1943) stimul red calf ovaries by antenor pituitary gonadotrophins Marden (1953) and Black et al (1953) attempted to explost the juvenile calf ovary for the production of viable ova la india the intercalving period on an average is iery long and a large proportion of cows take a long time to come to heat after parturition An attempt was therefore mnade to reduce the mtercalving period by inducing oestrus and os ulation with PMS These mvestigations are reported in Experiment No 1 As in cow's 1 condition known as anoestrus characterised by marked gonadal subacaviry possibly due to the lack of stimulation of the hypophysis is commonly encountered in buffaloes as well Delay in getting these anınıals in calf and the resultanc loss an milk yield necessarily construtos a serious problem During a countrywide survey on bovine infertility conducted from the Division of Animal Genetics Indian Vetermary Research Institute it was found that 66% of buffalo heifers over 33 years age and s3 per cent of buffalo cows which bad calıcd more than 4 months back had subactive ovaries (Bharracharya 1954) The investigations reported in Experiment No If in this paper were aimed at inducng oestrus and ovulation in noestrous buffalo with cquine gonadotroplun and for studying the effect of administration of the hormone during various phases of the destrous cycle and carly pregnancy an this species. The study was thus also concerned wich the potential fertility of eggs chat have been ovulated by arufidal mcans The quality of seinen produced by bulls buffalo-bulls rains and goats deteriorates during the part of the year when air ternperature is hugh coupled with high humidity (Mukherji and Bhattacharya 1952 Shukla and Bhattacharya 19 b Kushusha e al 1955) Studies concerning the attempts made to improve the semen quality w rams (Luktuke 1954) and in buffaloes (Goswami 1957) by activating the testes with the ad munistration of PMS in unfavourable season have also been included 1 chus paper (Experiments III & IV) MATERIAL AND METHODS eper mest do I Experimental animals for this study consisted of 34 cows and 3 heifers The cows were anoestrous for over four months after parturition The heifers though long nature werç anioestrous The animals were divided in tvo groups in the first group 9 cous were injected with 1000-2000 IU PMS These animals ncre slaughtered 4 to days after injection in the second group comprising of 25 cows and 3 heifers the dosage of PMS administered varied front 1000 to 3000 1 U Eight cows from this sct were slaugh tered after a period of 2 months or over all the cxperimental animals were either arti baally mseminated or were given a natural sense at the induced or subscquent heats Exper a ent No II Experimental anuals for this study consisted of 103 buffalocs including to buffalo peifers and three buffalo-cer cakes Twelc buffalo cows were purchased from the local cattle markets Sex buffalo cows tere treated at the Military Dairy Farm Bareilly and fifteen animals were offered by private cattle owners for treatment The remainmg anunals used m this invest gation acre obruined through an arrangement made with the local abattoir agents ulicrcby buffalocs desained for slaughter were te nporarily kept at the Institute for experimental purposes. These animals were almost of umform agegroup and were in good condinon of health ·60 P BHATTACHARYA & S N LUKTUKE STUDIES ON THE EFFECT OF Out of the 12 purchased animals, eleven were slaughtered in the Institute's post mortem rooms and one was later auctioned on her becoming pregnant The animais obtained on contract were maintained at the Institute for obscrvations and treatment and were slaughtered at the local abattoir Six animals at the Military Dairy Farnı, Barcilly and is animals offered by private cattle owners, were all anocstrous These animals were 1.food-S. naturally at the induced heat and were left to calve The private cattle owners were re quested to present their animals for insemination as soon as ocstrum was noticed after treatment All injections of PMS were given subcutaneously in the neck region, with a light riiassage for a few seconds after the injection The dosage varied from 750 IU to 6000 IU After injections, the animals were frequently cxamined for the onset of oestrum ard the examinations were carried out at still lesser intervals till the animals went off heat All anımals except those at the Military Dairy Farm, Barcilly were inseminaced during the declınıng phase of the oestrus The inseminations were repeated after 18 hours of tha animals continued to remain in oestrus for more than one day Semen from two buffalo bulls was used throughout for ensenunating animals in this mvcstigation The quality of semen was good and contained at least 80 per cent moule spermatozoa Ea amination of matenal Animals were slaughtered at appropriate times at the local inunicipal slaughter housc (except the eleven purchased animals mentioned above, which were destroyed in the postmortem rooins of the Institute) and after slaughter, the reproductive organs were removed as quickly as possible and ucre brought to the laboratory wrapped in wet towels for investigation In the laboratory, the organs were carefully dissected and the following particulars were recorded 1 Weights of ovaries and uterus Ovulation points 3 Number of unovulated follicles 4 Number of old corpora lutes and luteal scars Condition of uterus, cervix and vagina The diameters of follicles and corpora lutea were measured with vernier callipers The follicles were classified as 1 Very large (VL) - Over 15 mm in diameter 2 Large (L) - Between 10-15 mm 3 Moderate (M)- 5-10 (S) - Ś Very Small (vs) - less than 2 mm in diameter Estimate of the approximate age of all induced corporalutea were inade from their size and colour 2 11 1 ?? 17 4 Small Recivery of ova In case of animals in which ovulation had taken place, the fallopian tube on the side ·4 ADMINISTRATION OF GONADOTROPINS I AUGMEVTING FERTILITY N PARM ANIMALS 61 of ovulation was carefully dissected to straighten ut out Warm physiological salınc soluton was rinsed through the fallopian tube carefully with the help of a glass syringe mto medium sized flat bottomed watch glasses in every watch glass about 20 drops were dramed out As a routine washings were obtamed to 6-8 watch glasses These washings were soon cranncd andet microscope for the recovery of oyr and she evidence of cleavage Thc ovum, if present in the oviduct usually canic out with the first few drops of Aud The diameter of the ovum was measured with the help of a micro-meter Classfication of groups The animals were grouped as shown below Grcup S: No 3 Buffalo heefers 4 Buffalo Cows Total S Injection of PMS an anoestrous animals Ljection of PMS during med destrous cycle in the presence of a well developed corpus luteum Injection of PMS during mud-oestrous cycle and corpus luteun removed at various intervals thereafter I yjection of PMS at various intervals after the enucles non of mid-ocstrous cycle cospus luteum Injection of PMS simultancously wth the < aclca tson of corpus luscum Injection of PMS during follcular phase of Destroux cycle Injection of PMS durung carly pregna cy Anoestrous ansals treated with PMS and left to calve 28 6 37 6 8 ΤΟ 20 4 4 6 6 TO al 4 11 6 18 11 13 89 102 experiment Mo III Ten razts were divided into two equal groups on the basis of their semen production Onc group received during summer thrce subcı tancous injection of 400 IU of PMS at intervals of 2 weeks Tuo ejaculates of semen yere obtamed ın quick succession at ucckly intervals froni cach animal The semen samples wicte examined for different attributes Experiment Mo IV Eighteen buffalo bulls were divided into three equal groups Animals of one group Nere administered PMS in doses of 1000 IU per week those in the second group were guyen pec os 100 mg of sodsuni thyroxine per day The treatment period lasted for 14 wecks The semen from all the balls was cxamined at regular weekly intervals for Various attributes RISULTS Data collected on the ovarian response to administration of PMS of nine cousslaugh. cred m the first group of Experiment i ase presented in Table No 1 Single Osulation cusred in six cons and ovulations in one with PMS treatnient. Pregnancies seetc obtamedin - out of Sanımıls of the second group in Experiment a result of treatincnt with PMS Data are presented sa Tabic No 2 The results of experiment No II are summarised v Table Nos ; $ ands ·64 P BILATTACHIARYA & SN LUKTURE STUDIES ON THE EFFECT OF Administration of PMS in subactive condition of ovaries caused single ovulation in a majority of animals. In the present investigation in ovulation was obs-rved in only two anoestrous buffaloes It has been obseried that when the anoestrous condition is very deep and the ovaries are atrophied, even large doses of PMS have failed to develop follicles There was no incidence of multiovulation in this group of animals The first follicle which attained maturity probably ovulated spontaneously and the formation of new corpus luteum inhibited ovulation of other follicles According to Hammond (19 7) ovulation occurs in bovines in follicles which are 10-15 mm in diameter In an investi gation on the development of follicle and corpus luteum in the buffalo Luktuke and L 1 ILover 10 days and no ovulation occurred Examination of ovarics by rectal palpatio? Folley and Malpress (1944) as Hormone Hypertrophy Pituitary activity in calves and in very young heifers is at a much lower level com pared to the adult females and hence the mode of response is cypected to be somewhat different Casıda et al (1943) experimented with 9 calves of unknown age and obtained 129 follicles but only 9 corpora lutea Marden (1953) has shown that ovarian stiniulation can be induced in the juvenile bovine thro igh the administration of gonadotrophic extracts Both these workers have found that relatively few ova are recoverable from calves in relation to the number of corporr lutea present The percentage of fertilized Ova recovered is also low In the present study only 3 herfer calves and 6 heifers were treated with PMS Ovulation could be induced only in one heifer calf As the heat symptoms were not evinced the animal was not artificially inseminated The ovum recovered from this animal was therefore unfertilized Out of the hçrfers 3 ovulated and in two of them the ovum was fertilized Casıda et al (1943) made attempts to induce superfecundity in the cows by treating them in the luteal and follicular phases of the oestrous cycle No pregnancies were obtained in the cows when treated during the luteal phase but a normal percentage of pregnancies occurred in the animals in which treatment was given in the follicular r (Isth day) of the oestrous cycle a single subcutaneous injection of 2000 IU PMS. W given and followed by an intravenous injection of sheep pituitary extract five days later All the sows were mited or artificially inseminated at the time of intravenous injectior and slaughtered 48-56 hours later ·ADMINISTRATION OF GONADOTROPEOINS IN AUGMENTING FERTILITY IN FARM ANIMAIS 65 Feralized ova were not found in any of the 'luteal sows' although an average of 106 ova was recovered per sow In the follicular group, 19 6 ova per sow were re covered and of these si serc feralized in the investigadon conducted by Tanabe et al (1949) trich 22 sows and 45 giles, there was also a very low incadence of feruilizacion of the ova produced, during the luteal phase of the oestrous cycle Hammond and Bhattacharya (1944) created 17 cows with varymg doses of PMS, durng the follicular phase of the oestrous cycle They obruned multiple ovulations in myo ways, ccher by injection of PMS towards the end of normal cycle or by its administranon cather m the cycle and subsequent expression of the curpus luteum Ovulation normally occurred only after rexioval or regression of corpus luteum present at the time of injection Mulnovulations were produced in 7 of these cows Gonadotrophıs have been used to produce superovulation in rats and mice. There was relatively a high rate of mortality in the new borts but larger than normal size of Later was produced (Engle 1927, Cole 1927. Evans and Simpson 1940) This problem of superovulation was also investigated in both cattle and sheep (Casıda et al, 1943, 1944) Response obranned varied with the gonadotrophins used and the stage of oestrous y de at a huch the treatment was given (Murphres es al 1944) Hammond and Bhattacharya (1944) and Hanumond (1939) induced twin and mulaple ovulazon in cattle by the use of PMS given as a single subcutancous injection Various methods to increase the number of viable ova shed by the cow have been tried (Dowling, 1949, Hammond, 1950, Rowson 1991 Brock and Rowson 1952) Dowling (1949) achucved considerable success by using three daily injections of 100 mg of horse pritustary extract during the follicular phase of the acle Injection of 3000 1. U of PMS subculaneously at any time between 4- och day of oestrous cycle was found by hum to result m marked folhcular stuulapon and high ovulatoa rate (81 per cent) after I H was 1967 tetan enously Hon ever, none of the ova were fertilized Casida et al (1943) ad previously shown chat ova shed during the lutcal phase of oestrous cycle were not pable of fertilizagon The cochnique of induction of multiovulations is of great practical importance in studios pertamıng to the transplantation of fertilized ova (Chang. 1949. Umbaugh, 1919. Hammond, 1950, Lamning and Rowson 1952) In the present mvestigation, mulgovulations were induced in buffaloes by injecting PMS during mud-oestrous cycle and the corpus luteum was removed at various intervals after injections. Twentyseven Wulzaons were obtained from nmc animals Out of the 17 ova that were recosered only ere found to be fertilized (1765 per cent) Muloovulations have also been induced in 9 other buffaloes by trcaong them with PMS during the follicular phase of the oestrous de 24 Ovulaaons were obtained from o anumals and out of the 16 ova u bich were recovered, 14 were alvar sous stages of development (87 50 per ont) Casıda (1943) and Rowson (1956) had found while induang superovulation in mature animals during luzeal phase. that the resulting ferulity is exceedmgly low Wale et al (1952) have tried to deposit the semen in the uterus of cow dursng the luteal phase of the cycle and found hat fertility continued to remain low Robuson (1950, 1951) has obrane promumg experimental results 2n raising the vel of ferulicy in ewes in the breeding season, so that a high perceritage of twins are born * This has been obtamed by mjecting the ewes with 450 of PMS on the 12th day of oestrous cycle (follicular pbase) A number of follicles mature during oestrous. The follicles which are woyilated m ·TABLE I Ovar an response of cows to administration of PMS Animal No Approx. time of Weight of ovulation Days after Uterus RO LO injection (mg) $1 No Remarks Interval in days between Follicles Ing and Inj and VL L M S VS H heat slaughter Dose i u 3 I 2 3 8 II 23 1000 I 500 1000 8 8 8 1 14 8 3 1 1 121.4 114 S 1165 309 23 7 IZS 20 4 24 1000 6 9 - 3 2 2 93.0 1 500 4 6 6 62 63 30 Ovulation LO IS7 LO 125 No ovulation Old CL. present in LO 19 Ovulation L O Ovum recovered 135 Ovulation L.O 38 No ovulation Very smalt old CL present in LO 55 Ovulation RO 31 1 Two ova recovered. Onc ovulation in 8 7 10 9 S 3 11 11 1316 980 14 8 83 1 500 3 BHATTACHARYA & S N LUKTUKE STUDIES ON THE EFTECT OF 1 65 66 1200 1200 IO 8 Z 14 S 21 7 1130 1166 40 22 1 13 DO 8 cach ova 9 72 2000 3 4 1 6 s 7 13 10 4 Ovulation Lo + 103 S SS VL. F265 1111 Right Ovary Left Ovary Corpus luteuin Haemorrhagic VS Very Large Large Moderate Small - Very Small ·TE Ffed of PAIS - restoring feuyi cows & hef rs Interval in days between Animal No Mode of Admnistration Breed SI No Dose iu Result Remarks Inj and Ist heat Ist and and heats COWS 2 non descript Subcutaneous 1 200 1 46 Pregnant 24 Slaughtered N foetus recovered 3 28 3 + 2 6 7 2 4 2 45 Calved at term + S 1 8 TO TO 25 I ISA 21A 15 120 185 344 12 16 19 4 3 Hanana 1200 1000 1600 2000 2000 1200 1000 1 200 I 200 1600 I 900 I 500 1 500 I 500 T000 1200 1000 ISCO 1000 1200 3000 3000 3000 3000 ADMINISTRATION OF GONADOTROPHINS IN AUGMENTING FERTILITY IN FARM ANIMALS 69 9 10 1 12 13 14 IS 16 17 18 19 20 21 22 23 24 as 4 4 IL 9 2 22 169 Repeated Pregnant on-descript Intra muscular Ded Calved at term Aborted (two months) Calved at term 190 Subcutancous Sahiwal Hartana Carrying 3 2 4 22 68 56 22 39 18 18 25 9 non-descript 06D 182D 39 14 48 to HEIFERS 13 PX Pira Repeating Pregnant Repeating Pregnant Repeating Rcreating Pregnant Carrying 3 Carrying 8 26 27 28 Gur non-desce pt 25 1900 1600 1000 1 Not Pregnant Pregnant Calved at term *Conceived after repeating a number of times N-Normal ·TABLE 3 SI No Arumal No Age Yrs Body weight lb Dose of Interval (in days) PMS between injection and We ghts (in 8) Ovulations Ova IU Heat Slaughter Uterus R.O LORO LO recovered State of Development of ovum Injection of PMS in heifer calves 1 2 3 1} It 2 308 394 392 1000 1000 3000 5 S 8 120 TOO 89 os ro 84 07 07 14 8 1 Unfertilized 52 Injection of PMS in heifers BHATTACHARYA & SN LUKTUKE STUDIES ON TJIE EFTECT of 16A 23 27 6 6 7 7 4 1 1 ammo 4 cell stage 16 cell stage 390 $08 SO2 $73 332 $88 1200 3000 3000 3500 3000 3000 Iliww 12 7 8 7 10 7 196 203 179 155 195 240 17 32 13 1 13 72 140 18 126 33 08 92 90 I Unfernuized 1 SS 17 ·TABLE4 I jest on of PAS 11 de ce crous b falo Cous ADMINISTRATION Si No Arnal No Body weight Ib Dose of Interval (in days) PMS between inject on and We ghts (10 g) Ovulations No of ova IU Heat Slaughter Uterus RO LORO LO recovered State of Development of ovum to 7 1 1 2 cell stage 1 2 9A TIA 12A 4 4 7 12 7 9 26 Unfertilized Unfert lized Unfertilized 1 760 830 740 690 500 650 450 472 500 $50 SIS 13A 18A T9A 20A 21A 23A 25A 3 6 8 10 7 64 20 4 4 19 IS 6 24 47 14 29 11 2 cell stage 11 26 11 20 7 2$ 17 8 & IO 19 6 6 2 I 31 IS 63 I 8 113 42 18 2 ZZ 50 3 2 19 142 11 I1 ---|-!|-!|-||---- 813 1 1 163 U fertil zed 2500 1900 1500 I 00 1 200 1200 1200 1200 5 500 2000 1500 1500 1500 1300 ISCO 1900 1500 3000 4500 3000 4500 4500 4500 4500 3000 4500 3000 4500 7 +7 135 21 94 135 268 199 22 145 282 182 455 460 299 146 329 718 196 337 237 405 273 340 715 474 391 356 390 13 14 13 16 17 IX 19 O 21 22 23 24 8 cel stage 10 GONADOTROPHINS IN AUGMENTING FERTILITY IN FARM ANIMALS ID 23 22 25 36 43 4 cell stage Unf talized 700 640 sho 768 702 585 619 512 784 (88 616 360 868 674 804 772 4 9 8 134 75 6 119 50 09 45 63 S2 21 O 137 13 9 90 130 70 98 Unfort lszed 2 cell stage Unfort I zed 3 15 so 16 cell scage 7 6 7 11 12 7 7 162 141 II 8 77 10 57 129 27 28 1 178 3 3 Z 1 4 cell stage 3 4 cell stage 1 71 ·72 TABLES Treatment of buffalo-cows with Pregnant Mfare Scrum Gonadotropham (P415) and the recovery of ova No of animals treated SI No Group No of animals ovulated No of ovulations L T No of ova recovered L Percentage of fertilized Ova recovered No of fertilized Ova R L T Percentage recovery of ova R T 9 23 7 3 6 9 1 18 ? 3 IZ 2 2 28 3 6 6 10 9 Z 2 16 1 3 5625 00 00 76 19 100 00 100 00 3 1 3 I 1 I 2 IA IB IC II III IV V VI 66 67 S 1 I 9 3 4 9 10 4 Z 13 17 I 2 II 27 5 4 24 8 4 2 10 17 5 9 1 2 6 63 00 100 00 100 00 66 67 2 4 2 9 S 4 7 8 9 17 65 100 00 TOO 00 87 50 2 16 11 6 3 Vil P BIIATTACIARYA & SN LUKTUKE STUDIES ON THE EFFECT OF Total 80 47 44 41 85 36 26 62 72 94 21 16 37 59 68 R Right ovary L Left ovary IA Injection of PMS in anoestrous buffalo cows IB Injection of PMS in anoes. trous heifer calves IC Injection of PMS anoestrous heifers 10 T. Total 11 Injection of PMS during midocs. trous cycle in the presence of a well developed corpus luteum. III Injection of PMS during midoestrous cycle and corpus lutcum removed at various intervals thereafter IV Injection of PMS at various intervals, after the cnucleation of muud ocstrous cycle corpus luteum V Injection of PMS, simultaneously with the enucleation of corpus luteum V Injection of PMS during follicular phase of oestrous cycle VII Injection of PMS during early pregnancy ·ADMINISTRATION OF GOVADOTROPINS IN AUGMEVTING FERTILITY IV FARM ANIMALS 73 TABLE 6 Average of sen en qual t es of treated and u treated rams during tle expenmer lal per od - Ist Ejaculate Control Experimental group Difference between two means group t value 0 60 0 69 Significance level oog 0 Volume of semen (ml) 1 Sperm concentration (millions/ml) Total No of spermatozoa (millons/ejaculate) Percentage of abnormal spermatozoa nitial fructose (mg/100 ml semen) uctolysis (%) 2841 I 90 Not signs 3814 ficant. 18.45 973 2792 3 10 II 09 947 1. 6 26 504 583 4 83 SII 300 2º. S4 72 72 189 Not sign- 18 2.46 ficant 5% TABLS 7 Averages of semen qual es of treated and untreated ra ns dur gile expet neral per od -- d Ejaculate Difference Control Erperimental between group group to incans t value 047 59 Significaacc Icvel Volume of semen (ml) (Sperm concentration (mi) ons/m!) ( ) Total No of spermatozoa millons/ejaculate) (*) Percentage of abnormal spermatozoa (u) Intial fructose (mg/100 ml of semen) (2) Fructolysis () 012 2326 3531 231 1231 1.OS sº. 2296 r/ 1005 1307 689 618 647 39 46 399 s33 6515 114 25 69 1% 272 3 99 2 1 10 ·Range of Vanation and the Afean of Reactson-Time and Semen Characteristics of Buffalo Bulls during the Pretreatment, Treatment and the Post-Ireatment Periods 74 Pretreatment period Treatment period Post-treatment period Reaction time, and Semen Characteristics Group Range Mean Range Mean Range Mc3n Reaction Time in Seconds с TH PMS 18 O-59 8 14 6-22 1 16 8-453 38 0(34) 19 85(35) 28 83(36) IO 3-343 II 0-286 IT 3-443 18 6(83) 17 6(81) 24 7(83) 14 8-435 93-540 91-426 23 0(76) 22 9(78) 22 2(76) Volume с TH PMS I 24-209 1 21-2 32 I 17-2 IS I 81(35) I 60(3) I 69(36) 1 62-2 54 1 08-2 12 I 15-2 42 I 99(83) 1 SS(81) I 81(83) I 20-2 16 087-2 16 I 31-2 50 1 76(76) 13878) I 82(76) Initzal Motility of Spermatozoa с TH PMS 19-27 20-25 I 4-23 207(35) 2 20(35) 1 95(0) 20-32 2 2-3 4 19-34 270(81) 2 92(80) 273(13) 20-34 25-37 22-36 275(72) 3 28(75) 3 00(74) Sperm concentration in millions/ml с TH 536 6-796 0 573 8-879 1 Sos 0-909 1 721 0(3) 713 6(35) 677 (36) 418 0-870 8 355 8-835 8 340 0-8030 605 0(81) $87 8(80) 538 1(83) 231.6-13330 481 6-1394 1 283 3-972 5 726 $(72) 571 0(75) 630 3(74) PMS Total sperm in millions/ ejaculate с TH PMS 710 9-1861 S S53 3-21930 775 7-1887 8 607 2-1638 5 496 8-15589 $09 S-14807 IIS4 7(81) 1029 2(80) 1020 9(83) 346 3-2623 6 856 7-19507 432 8-2077 8 1418 6(72) 1252 5(75) 1201 0(74) 1321 2(3) 1275 (35) 1257 (36) 543 4(25) 578 3(25) S56 (30) P BILATTACHARYA & S N. LUKTURE. STUDIES ON TILL EFFECT OF Initial fructose content in mgm/roo ml of semen C TH PMS 472 2-3926 551 3-5928 394 3-5919 664 3-9502 767 9-9439 OIS 3-896 5 821 9(82) 839 8(80) 777 5(83) 793 G-IOS4 4 755 7-11029 730 6-8726 902 6(71) 872 5(76) 820 11(74) Fructolysis Indes C TH PMS 099-3 52 074-276 070-3 49 I 90(17) I 93(24) 1 81(27) I 12-3 30 I 52-3 49 I 06-235 198(64) 2 31(76) 2 11(61) 0 55-4 56 1 04-320 1 1-3 49 198(83) 2 03(67) 2 07(62) %of abnormal spermatozoa C TR PMS was not cstimated 9 89-25 86 642-22 41 II 05-29 25 15 82(81) II 83(80) 18 89(83) 12 01-20 ST 742-10 07 11 06-28 29 17 09(67) 8 60(71) 1798(70) ·ADMINISTRATION OF COVADOTROPITINS IN AUGMENTING FERTILITY N FARM ANIMALS REFERENCES 75 Abmayer H. (1950) Terar il Us 1 59 Ctat on from Arm Beed Abst 19 IISI Asdell S A., Facher M G Smith S E and Ellot F I (1942) Comell Un v ag Cep S: 17 243 Bhattacharya P Harumond J Jnr and Day F T (1941) Tc Rec 33 450 Black, W C., Ulberg L C Christian R É and Casıda LE (1953) J Dary S 36 274 Bottomley A C Folley S J and Watson H ME (1940) Jagr Sa 30 235 Bock, H and Rowson LE (1952) ibd 47 479 Cameron H. S (1942) J AI et Afed Asso 100 60 Cas da EE Warwick E J and Meyer R I (1944) J An Sa 32 Cas da L E. Meyer R K McShan WH and Wacky W (1943) Anset J vet Res 4 76 Chance MRA Rowlands I W a d Young F G (1939) I dont 1 239 Chang MC (1949) Proc Nat Egg Tran fa Bed & Conf f 39 Cole HH (1937) Amer J Plysol 119 704 Cole H H Hart GH and Miler R F (1945) E doerin 36 370 Cole H H and Miller R F (1933) Ame J Ply of 104 165 Howling D F (1949) J ag Sa 39 374 Jutt RH (1953) J An Sa 12 SIS Engle E T (1927) Soc exp Bol N Y 25 84 Evans H M and S mpson M E. (1940) Endocr clogy 27 jos Folley S J and Malpress F H (1944) Proc roy So B 132 164 Gosa am S B (1957) Effect of administration of Thyroun a d PVS Horst ones on semen qual ty of buffalo bulis Thesis for Ph D Nagpur Un vers ty Hunmond, ) (197) The Physiology of Reproduction n th co v Cambr dgc (1948) 111 lemnat Congr Phyol & Palol A - Rep od a dil cm M lan (1949) Jag Sd 39 222 Hammond J Jar and Bhattacharya P (1944) Jag S 34 Kushwaha N S Mukherjee DP and Bhattacharjee P (1955 1JAJ SC 25 317 Lamming GE2d Rowso1 LE A (1952) Pro 2.d1 Do I Cory ol and P lol Anm R pod ad At Inc Copenhagen P 144 Lubin E (1946) J Amer ve med As 49 352 Luktuke S N (1954) Thess for Assoc IV R I Izatoagar UP Luktuke S N and Bhattacharya P (1948) Ind an J Diy Sa$9 -(1981) Induct on of multiovuls ons Hariana co vs Unp blh jd la (1954) Unpubl sled data al 954b) Oestrous cycle n murah buffalo he fers tpb hdd Marden, W GR. (1953) agt Sa 43 381 Mukherjee D P and Bhattacharya P (1952) Ind in ve Sa 22 73 Mu phree RL Warw ck, EJ Cas da LC and Mcshan W H (1941) J A Sa 3 12 Noble R. L Rowlands i'w Warwick M H and Wiliams P C (193) ] Edoorn 1 22 Robinson T J (1950) ) agn Sa 40 275 (1951) ibd 411 Rowlands I w and Williams P C (1943) J Endoc a 3 310 Rowson L. L. (1951) Ib 2 7 260 Shukla D P and Bhattacharya, P (1952) Ind J vet Sa 22 109 I d 23 179 Umbaugh R E (1949) Amet J vet Res 10 295 ·CLINICAL REPORTS ON TESTICULAR HYPOPLASIA IN GOVERNMENT BREEDING BULLS OF WEST BENGAL by B K BASU Department of Obstetrics & Gynaecology including Artificial Insemtination, Bengal Veterinary College, Calaitta 37 ABSTRACT Observations on some clinical cases of hypoplaste testes in Hariana bulls have been reported Two of these animals showed bilateral hypoplasia while the other two showed unilateral hypoplastic testes. It was observed that the secondary sex characters and organs were apparently normal i unulateral hypoplasia but in cases of bilateral hypoplasia there was definitc underdevelopment of secondary sex characters and organs and feminisation of the bulls The nroblem of crarinnnd fin. -1 11 1 ficial Insemination work, has not so far been appreciated Extensive rescarch has been made in Europe and America on the subject of sterility and infertility due to Hypoplasia in bulls by eminent workers like Williams, (1947) Lagerl of (1938) and many others Here in the Eastern Hemisphere we consider the question of infertility and sterility L power or become responsible for the production of defective offsprings s as a cow" So far as infertility be one of the commonest causes It may be unilateral or bilateral lys The present work describes the results of some clinical observations on some Govt bulls distributed for breeding purposes Investigation ܘܬ to follow up the case for detailed Morphological, Histo-pathological and other exam. nations Occasional cases of hypoplastic testes encountered in the animals of other breeds have not been considered here The study consisted of clinical examinations of the primary and secondary ser organs and histopathological examinations of some of these organs after death Out of such 20 Hariana bulls examined, 4 cases of Hypoplastic testes were detected Of these 4 cases, 2 were of bilateral in nature and the other two of unilateral type-the right testes being the affected one ·BK BASU TESTICULAR HYPOPLASIA N BREEDING BULLS OF WEST BENGAL 77 Observations - 1 Bilateral hypoplastic testes - (w) Clinical symptoms-The testes were much smaller than the usual size They were unduly hard with a roundish contour It was extremely difficult to locate the tail of epıdıdymis on palpation The spermatic cords were very short and distorted (6) Morphology-The testes were highly underdeveloped in consideration of breed, size and weight of the animals (about 700 lbs) and age (s years) and weighed for 60-80 Gms (4) Histo-Pathology–Majority of the seminiferous tubules were vaculated Different formative stages of the spermatozoa could not be seen Most of the tubules were devoid of spermatozenic epithelium and paratubular fibrosis though present was not very prominent (4) Secondary Sex organs and characters-- Animals presented a feminine appearance with slender neck and small head and showed a feminine temperament The secondary sex glands e & seminal vesicle prostratc, etc were comparatively smaller, though the penis showed almost nornal development, II Unilateral Hypoplastic testes (a) 1 2 Hyperplasia of the epididymis of the other side arcumscribed by an unusually clear limiting linc (6) Morphology-The normal testes with apparent hypertrophy of the epididymis was suggestive of its tendency to carry on some compensatory function 'But the size of the normal testes did not show any metaplastic changes, spermatic chord was shorter on the affected side (9) Histopathology-The hypoplastic testes was found to have reduced cell layers and in hypofunctioning state (4) Secondary sex organs and cluaracters-Development of the secondary scx organs was not affected Serving ability, secondary sex characters and fertility appeared to remain mostly unimpaired DISCUSSIONS According to Enkson (1943) tesacular hypoplasia is aused by a recessive autosomal gene with incomplete penetrance His earlier finding has been confitmed by wide genicic nvestigations (1950) ·80 SACHCHIDANANDA BANERJEE: TIIYRO-CONAD-ADRENOCORTICAL less glycogen The scorbutic guinea pigs thus showed lowered glucose tolerance and they deposited in the liver. These observations suggested diminished insulin secreton in the scorbutic animals. Insuln content of the pancreas of scorbutic guinea pigs as well as thc paired fed normal guinea pigs were extracted and assayed by the rabbit method of insulin assay. The results are given in Table III. TABLE III Insulin content of pancreas of guinea pigs Animals Insulin content per gm Weight of pooled pancreas (gm ) pancreas (International Units) Normal (10) II 91 045 Scorbutic (10) 10 38 OII Figures in parenthesis indicate the number of animals, The insulin content of the pancreas was markedly diminished when the anmals developed scurvy. from the tail end of each pancreas, stained with iron hematoxylın, the number of aslees was counted and the size of the individual islets was measured (Banerjec, 1944) The results are given in Table IV. TABLE IV Number and size of islets of Langerhans of pancreas of guinea pig's Animals Number (mcan) of Total (mncan) size of the the islets in each setion islets in each section (sq mm) Normal (10) Scorbutic (10) 76 II 2 3 og 273.52 1168 25 21 Figures in parenthesis indicate the number of animals The sections which were stained with azan stain were examined for the different types of cells present in the islets of Langerhans. In the scorbutic guinca pigs, alpha cells were found to be increased in number in proportion to the beta cells, and the beta cells were found to be mostly degranulated. Lain the sections studied The increase in the size and alsa attempt to react against the fall in insulu any degenerative changes seems to be i place when the scorbutic guinea pigs are given supplements of vitamin C. ...1 held it was 1C ·RELATIONSHIPS AS AFFECTED DY SCURVY 81 Guinea pigs were separated into several groups, cach group consisting of one normal, one scorbutic and one insulin treated scorbutic gumca pig The gunea pig in each group was fed equal amounts of the scorbutic diet Regular insulin was injected subcutaneously into the animal intended for insulin treatment with a dose increasing from or to o 3 Inut per 100 gm of body weight per day from the beginning of the second week On he day of the experiment insulin injection was stopped. An oral glucose tolerance was rade in the 4th week of the regime with the scorbutic diet on eleven groups of animals ter they were fasted for 15 to 18 hours Animals were killed after the glucose tolerance t and liver and muscle were removed for the glycogen estimation (Banerjee, Biswas 1 Singh, 1958) The results are given in Tables V and VI TABLE V Glucose tolerance test in guomea pigs after different treatments Animals Fasting blood Blood sugar (mg) after glucose feeding sugar (mg) 45 min Jo mun ISO min Normal (11) Scorbutic (11) Insulin treated scorbutic (11) III E46 124+53 1544116 1975170 180 min. 128 +44 181+131 244+184 Figures un parenthesis indicate the number of animals 226-510) 1445 10.4 248+18 8 242 5189 123ITO 227+176 1984 79 161 98 Glycogen content of hver and muscle of guinea pigs (gm 1100 gm fresh tissue) TABLE VI 9 norinal animals 9 scorbutic anuals Liver Muscle 11 insulin treated scorbutic animals Liver --- 2053 +0295 Muscle 0 697 0133 Laver 0 391 +0083 Muscle 0137 +0039 I 349 +0 181 350 toos7 Insulin treatment of the scorbutic guinea pigs did not siguficantly alter the fasting blood sugar and the blood sugar values of samples of blood uken 45 and 90 minutes after the fecding of glucose However, after 150 and 180 minutes there was a marked lowering of the elevated blood sugar in the insulin treated animals although the sugar level was stall significantly higher chan that of the normal animal The peak of the blood sugar value of the scorbutic guinea pig was also shifted from the 1soth mmute period to the goth minute period after glucose was fed under the action of insulin In this respect it imulated the glucose tolerance pattern of normal controls Glycogen content of the Incr and skeletal muscle dumnushed in scorbutic gunca pigs in comparison with that of the normal controls and the decrease was bughly significant staastically Treatment with insulin strikingly improved the glycogen content of the liver and the skeletal muscle of the scorbutic guinea pigs, though the values did not reach the normal level It was also observed that the dosage of insulor used in these experiments did not affect the glucose tolerance of normal guinca pigs These results 11 ·82 SACHCHIDANANDA BANERJEE TITYRO-GONAD-ADRENOCORTICAL indicated that treatment with insulin for a prolonged perod could correct to a great extent the disturbed carbohydrate metabolism in scurvy At present there are increasing evidences that insulin is involved in the intermediate nietabolism of carbohydrates at the level of the Krebs cycle It is possible that in scurvy there may be a disturbance in carbohydrate metabolism at the level of the Krebs cycle due either to diminished insulin production or to the direct effect of lack of vitamin C on the enzyme systems concerned with the ondation of intermediates through the Krebs cycle The tissue contents of atric, malic and lactic acids werc, therefore, determined in normal, scorbutic and insulin treated scorbutic guinea pigs (Banerjce, Biswas and Singh, 1958) The results are given in Table VII, VIūt and ix TABLE VII Citric dad content of tissues of guinea pigs (mg/100 gm wet tissue) Animal Blood Liver Kadacy Brain Normal (12) Scorbutic (12) Insulin treated scorbutic (8) S08E0 30 6 47+0 26 11 27+0 59 701+0 61 787+017 1065 ŁO S7 17 96085 12 04+075 6 88+070 6 18+0 32 13 970 92 760-035 TABLE VIIT Malic and contents of tissues of guinea pigs (mg (100 gm u et tissue) Animal Normal (8) Scorbutic (8) Insulin treated scorbutic (6) Blood Liver Kidncy Brain Cardiac muscle 3 42£0 34 20 3240 88 12 985095 13-43+1 63 16 93 +1 91 8 12+0 80 26 97+2 09 26 60+2 11 25 74+1 67 24 03:1 73 4 SI:£0 37 17 55+:2 20 18 24+1 24 TABLE EX Lactic and contents of tissues of gunea pigs (mg/100 gm wet tissue) Animal Blood Liver Kidney Brain Normal(10) Scorbutic (10) Insalın treated scorbutic (8) 94 – 45 204+159 142169 223584 228160 366–179 173+: 86 251-119 55+ SO 96+17 1703177 109+ 46 Figures in parenthese indicate the number of animals Table VII shows an increased content of atrıc aad in the tissues of the scorbuto animals in comparison with that of the normal controls Treatment with insulin brought the level of atric acid to normal This defect in citric aad metabolism may possibly be due to the presence of a metabolic block below the level of citric acid in the Krebs tricar boxylic acid cycle Insulis corrected the defect m citric acid metabolism, possibly by activating the aconitase enzyme and thereby facilitating metabolism of citrate ·RELATIONSHIPS AS AFTTCTED IN SCURVY 83 Malic and contents of tissues are increased in scorbutic guinea pigs in comparison with those of the normal controls Insulin treatment lowered the level of malic acid to normal in blood, liver and cardiac muscle The increased malic acid content may be related to the increased accumulation of citric aad which is reversibly converted to malic and through the condensing enzyme-malıc dehydrogenase system Malic and may also be formed by the reaction of carbon dioxide, pyruvic aad and TPNH Pyruvic acid excretion is increased in scurvy (Banerjee and Biswas, 1959) The observation that treatmeut with insulin lowers the lescls of both atric acid and malic aad to normal levels supports the conclusion that lack of insulin is an important factor in the carbohydrate metabolic derangement in scurvy Table IX shows a greatly increased lactic and content of the tissues in scorbutic gunca pigs in comparison with that of the normal guinea pig Treatment with insulin lowered Lactic acid value even below normal The accumulation of lactic aad may be duc to decreased glycogen formation from lactic acid in scurvy To study further the operation of the tricarboxylic aad cycle in scurvy the urinary excretons of ketone bodies, citric acid and malic and were determined in normal, scorbutic, insulin treated scorbutic and Fe**and cysteine treated scorbutic gunca pigs after the feeding of butyrate After the collecaon of a 24 hour sample of urme under tolucnc cach animal was fed daily in three divided doses (Banerjee and Kawishwar, 1959) a neutral solution containing 25 per cent of butyric acid (i ml/1oogm body weight) for 3 consecutive days Urine samples were collected over 24 hours After the above experiments, the animals were rendered scorbutic by the withdrawal of the ascorbic acid supplement. They were divided into three groups Animals of one of the groups were injected with protamine zinc insulin one injection per day, in doses increasing from o I to o 3 units/100 gm of body weight from the second wcck of the scorbutic regime To a second group of animals, mtraperitoneal injections of a muture of o.s ml. of oos M Fetas ferrous ammonium sulphate and o s ml of o M cysteine hydrochloride were given from the third week of the scorbutic regime The animals of he third group served as untreated scorbutic controls When the animals showed gross signs of scurvy, they were fed butyrate for three days Twenty four hour urige samples, collected before and after the administration of butyrate, were analysed for ketone bodies, citric aad and malic acid The results are given in Tables X and XI Twenty-four hour urinary excretion of kerone bodies atre and and malic aad by guinea pigs TABLE X Anuals Ketonc bodics Catric aad molcs Malc aad ft-moles Normal (6) Insulin-treated normal (4) Scorbutic (6) Insulin-treated scorbutic (6) Fe** + cysteine-treated scorbutic (6) noles 487+0.436 27500014 573ŁO S41 493 +0 208 14 805 231 13 20+ 210 388714-45 12 175 315 3 02+0.26 35602 43300903 3 080 204 6 100.245 Figures in parenthess indicate the number of ammals Values are mean + tandard error IS 91 145 1 84+0 342 ·84 SACHICHIDANANDA BANERJEE: THYRO-GONAD-ADRENOCORTICAL TABLE XI Twenty-four-hour urinary excretion of ketone bodies, atre and, and music acid by guinea pigs fed sodium butyrale u-moles pornoles f-moles Normal (6) Insulin-treated normal (4) Scorbutic (6) Insulin-treated scorbutic (6) Fe** + cystcnc-treated scorbutic (6) 62 6454-473 10 13+0973 79 39+8 OSI 28 863 095 55 87–13 17 46 99+ SSS 45-46+11983 438 43.1:85 03 68 36+11 33 282 10+ $0 65 8 770 904 6.44+1 143 20 9241.475 404ŁO SH4 663 10 198 The urinary excretion of ketone bodies increased enormously in animals of all che groups after they were fed butyrate The increased urnary excretion of acrate may be due to decreased oxidanon of citrate in scurvy. The mcrcased urnary excretion of malic T 11,- genous insulin possibly promotes increased oxidation of ketone bodies by periphera? assues, stimulates the production of morc oxaloacetate through restoration of the normal operation of the Krebs cycle or stimulates fatty acid synthesis It seems that insulin released in normal guinea pigs was not sufficient to deal with the excessive amount of ketone bodies formed as a result of the fecding of butyrate , Common of tjet, and mre to normal after 1 T seem to indicate . In order to see if ascorbic aad has any effect in the treatment of diabetic patients, the effect of administration of vitamin C on the glucose tolerance test and urmary and nonnchen lsr du hatır monents sense studied Diabetic papets ad hus the oral glucose tolerance test was performed. The patients were then fed ascorbic and (10 mg/kg body weight) for three weeks and the glucose tolerance test was repeated The results are given in Table XII (Banerjee and Ghosh, 1950). Twenty-four-hour urine samples were collected in these patients, before and durag the administration of ascorbic aad for three weeks. The urinary excretions of sugar were determined In 10 out of 14 cases urine became sugar free thuee weeks after the administration of ascorbic aad In order to determine if the diabetic patients were suffenng from Vitamin C deficiency the 24-hour unnary excretion of ascorbic aad was determined after administration of ascorbic aad (icmg/kg body weight) for 2 to 4 days in 20 diabetic and s normal subjects The results are given in Table XIH ·RELATIONSŁUPS AS AFFECTED IN SCURVY 85 TABLE XII Glucose tolerance test and abetic patients (10 patients) Treatment -hour Mg glucose per 100 ml blood -hour 3 hour 1-bour after the feeding of so gm glucose 2-hours 143 +10 206+13 256+13 271:14 243316 None After feeding ascorbic agd (10mg/kg) for three weeks 113ĖS 177+ 8 -303:12 14 231+11 22 204+11 26 20 TABLE XII Urmury excretion of ascorb c acid by odabere and rormal subjects Twenty four hour urmary excretion of ascorbic acid u mg Before ascorbic Days after the administration of ascorbic acid aad I-day 2-days 3-days 4-days Subjects Diabenc Normal 132+27 1833 16+4 06 59.21 34992 178 +48 33+ 7 288 +67 38 The fasting blood sugar level of diabetic subjects was significantly lowered after the administration of ascorbic and The blood sugar values it and 2 hours after the feed up of al fonet -11 La L. . . 1 th the inc. results indicate that diabetic patients suffered from ascorbic acid defiaency The absencc of sugar in the urine and the improved sugar tolerance after the diabetic patients were fed ascorbic acid for three weeks indicated that ascorbic aad led to the increased produce tion of insulin in these patients 1 r ·84 SACHCINDANANDA BANERJEE THYRO-GONAD-ADRENOCORTICAL TABLE XI Twenty-fow-tour urmary excretion of letone bodies, atre and, and malic acid hy guinea pigs fed sod um butyrate μ πιoles f-moles fr-moles Normal (6) Lasubin-treated normal (4) Scorbutic (6) Insulin treated scorbutic (6) Fett + cysteine-treated scorbutic (6) 62 64+4-473 10 13+0975 79 39+8 Osi 28 86£3.05 SS 87+13 17 46 99 553 45 46+II 983 438.43+8503 68 36+II 33 282 10+5065 877 +0904 6.44+1 143 20 92 +1.475 404+0 $44 6 63+0 198 The urinary excretion of ketone bodies increased enormously in animals of all the groups after they were fed butyrate The increased urinary excretion of citrate may be due to decreased ovdation of citrate in scurvy The increased urinary exerction of make L.L bodies was distinctly lowered by the insulin treatment of the scorbutic animals Exo genous insulin, possibly promotes increased oxidation of ketone bodies by peripheral c 1 206 JCC ind lactic dehydrogenases in scurvy The activity of the enzymes was restored to normal after prolonged treatment of the animals with insulin All these observations seem to indicate chat vitamin C-deficiency leads to hypoinsulinısın In order to see if ascorbic aad has any effect in the treatment of diabeac patients the effect of administration of vitamin C on the glucose tolerance test and urinary 1 ments diet during their stay in the hospital After the patients received this diet for one week the oral glucose tolerance test was performed the patients were then fed ascorbic aad (10 mg/kg body weight) for three weeks and the glucose tolerance test was repeated The results are given in Table XII (Banerjee and Ghosh, 1950) Twenty-four-hour urme samples were collected in these patients, before and during the administration of ascorbic acid for three weeks The urinary excretions of sugar were determined In 10 out of 14 cases urine became sugar free three weeks after the admmestration of ascorbic acid In order to determine if the diabetic patients were suffering from vitamin C deficiency the 24-hour urinary excretion of ascorbic aad was determuned after admunistration of ascorbic acd (somg/kg body weight) for 2 to 4 days in 20 diabetic and s normal subjects The results are given in Table XIII ·RELATIONSHIPS AS AFFECTED IN SCURVY 85 TABLE XII Glucose tolerance test mn diabetic patients (16 patients) Treatment -hour Mg glucose per 100 ml blood +-hour I-hour Il-hour after the feeding of so gm glucose 3-hours 14310 206+13 256+13 271 4.14 213 +16 None After feeding ascorbic acid (tomg/kg) for three weeks 113+3 26 177+ 8 18 -30+12 14 231 +11 22 204 +11 20* TABLE X Unnary excretion of ascorbic acid by 20 diabetic and s normal subjects Twenty four-hour urmary excretion of ascorbic aad in mg Before ascorbic Days after the administration of ascorbic and acid I-day 2-days 3-days 4-days Subjects Dubenc Normal 18:53 132+27 178+48 16+4 06 33+ 7 288 +67 38 59.321 349+92 31 The fasting blood sugar level of diabetic subjects was significantly lowered after the administration of ascorbic and The blood sugar values if and 2 hours after the feeding of glucose were significantly lower when the patients received ascorbic and for three weeks Thus indicated that the utilization of glucose was slightly improved when the diabetic patients were given ascorbic acid The normal subjects excreted siguficantly increased amounts of ascorbic aad than diabetic patients, when fed ascorbic and The results indicate that diabetic patients suffered from ascorbic and deficiency The absence of sugar in the urine and the improved sugar tolerance after the diabetic patients were fed ascorbic acid for three weeks indicated that ascorbic aad led to the increased producton of insulin in these patients It was of interest so find out how insulin production might be influenced by vitamin C-nutriqon of the body Insulin contains a large amount of cystine and it is probable that under the influence of enzymes cystine is taken up by beta cells to form the insulin molecule In the body cystne may be derived either from methionine or cysteine, and glutathione, the naturally occurring tripepade, cortasrus cystcane If the glutathione Content of the body is durunished, it may interfere with the normal synthesis of insulin Certan enzymes of the body depend on-SH groups for theu activity, and if the -SH naria The activity, however, may be Glutathione thus is supposed to glutathione content of tissues is , we bota cell sulfhydryl enzymes, necessary for the synthesis of insulin, mas not be well protected from the touc action of unknown substances leading to the hypofunction and diminished secretion of insulin intravenous injection of dehydroascorbic acid in fats leads to chronic hyperglycemia and glycosuna Glutathione if injected ground 74 +.1.2 2 ·88 SACHCHIDANANDA BANERJEE THYRO-GONAD-ADRENOCORTICAL significantly Cholesterol content of the different tissues varied differently in the scor butic animals, it diminished in the adrenals spleen, and lungs considerably during scurvy This may be due erther to diminished synthests by these tissues or to reduced transport The increase in the cholesterol content of the testes during scurvy might be associated L 1 TL 1 production Cholesterol content of intestine increased and that of liver or kidney did not show any change during scurvy From thus study it could not be ascertained if the lower adrenal cholesterol in scurvy was duc to diminished synthesis of cholesterol or increased production of adrenial cortical hormone pigs (Banerjee and Singh 1958) The results are given in Table XIX TABLE XX Total body cholesterol content of normal scorbutis, and insul n treated scorbune gu ned pix (mg per 100 gm wet issue) No of animals - 6 Total cholesterol content I values Insulin treated scorbutic Between normal and scorbut Between normal and insulin treated scorbutic Between scorbutic and insulin treated scorbutic Nornal Scorbutic 180+14 277+27 173+15 20 07 30 The total body cholesterol content of scorbutic guinca, pigs increased significandy in comparison to that of normal controls This possibly indicates increased synthesis of cholesterol in scurvy The decreased cholesterol of adrenals in scorbutic guinea pa therefore might be due to increased formation of adrenocortical steroids Insulin treat ment lowered the body cholesterol of scorbutic guinea pigs to normal level The incrcased cholesterologenesis in scurvy therefore might be due to the utilization of an increased pool of acetate which is not burned through tricarboxylic cycle in scurvy Vitamin C-defidency results in enlargement of adrenal glands (Banerjee and Ghosh 1946) A decrease in liver and muscle hexokinase activity in scorbutic guinea pigs, also becn reported (Banerjee and Ghosh J958) Adrenal cholesterol is diminished in scurvy All these investigations indicated deranged activity of the adrenal cortex in scurty, The urinary excretions of 17-ketosteroids and corticosteroids were therefore determined during the progress of survy to determine the adrenocortical activity Gumea pigs and monkeys were used as the experimental animals After determinng the normales cretions ascorbic acid supplement was withdrawn and these excretions werc estimated during the different weeks after withdrawal of ascorbic acd The results are given Tables XX XXI and XXII (Banerjee and Singh 1957) ·RELATIONSHIPS AS AFECTED IV SCURVY TABLE XX Twenty-four 4037 m ary excret on of 17tis teto ds by femaeg Neagu (ges reeds al day) 89 An Before ul withdrawal No of ascorbic zed Wecks after wihdrawal of ascorbic acid 1 3 2 3 3 4 8 36 3 64 31 135 89 96 ios 168 83 164 76 88 129 7 26 60 36 83 S02 57 [$4 94 so 138 9 12 13 Ded Dod Dud 4 ro 117 202 245 95 71 5 70 133 303 176 Dieu 123 ed 87 43 Ded 85 270 125 Died 27 TABLP XX] Twmniy-four hout story exret ost of 17 ie i rods dy fc nae yon Y (de erst tel un al day) Disd Before Mon- withdrawal kry of ascorbc No raid Months at er withdsanal of ascorbica 1 2 3 809 644 732 685 $17 555 682 673 873 3 600 Jos 42 hos TABLE XXIL Twenty-four-bout ur try escono af 19 keto zero ds (17 KS) and enh o toro ds (CS) by femalt * nespus ex retson an o tay OJO 34 54 787 OZ $08 959 2712 789 3003 Ded 1042 Wecks after wil drana) of ascorbs and Par No 7 17-KS 2 CS 17 KS CS 2 17 AS CS 17 AS Normal Scorbutic Normal Scorbutic 1 Normal Scorbutic 4 Normat Scorbuess Normal Scorbutic 225 22 266 160 571 .22 Cs 2;8 175 GO LAO 202 too 162 37 207 137 [1 $ 223 416 258 365 217 350 239 209 23 379 250 32 169 324 107 257 Ped 277 118 20% 293 370 192 200 375 ws 237 357 280 417 691 392 362 435 ing 331 297 321 ·90 SACHCIODANANDA BANERJEE THYRO-GOVAD-ADRENOCORTICAL It was observed that the excretion of 17-ketosteroids diminished in the first sicek the animals were on the scorbutic diet, then the cxcretion was irregular and when the animals became acutely scorbutic, 7 out of 10 guinea pigs and 3 monkeys excreted larger amounts of 17-ketosteroids The remaining three severely scorbutic gunca pigs however showed a diminished urinary excretion of 17-ketosteroids When scurvy is developed in the animals, food consumption is diminished and inanition itself mught alter adrenocortical function Excrcuon patterns of both 17 ketosteroids and corticosteroids were, therefore, studied m guinea pigs during the process of development of vitamin C-deficiency and in paired fed control animals While the excretion of these steroids did not vary considerably in the control animals during the experimental period, four out of five severely scorbutic guinea pigs excreted increased amounts of both the steroids which indicated hyperfunction of the adrenal cortex those anımals during the later part of the experimental period One of the severuly scorbutic animals, however excreted diminished amounts of both these steroids The results obtained prompted to conclude that the levels of steroids of adrenocorti cal origin in uriner body It is possible be responsible for Thyroid activity in normal and scorbutic guinea pigs 1 of the Guinea pig f. ja...1 dret ..oth ranır diet only (b) the scorbutc with o os% desi ascorbic acid pc supplement of C The oral glucose tolerance test was performed on the 22nd day in all we a meals The results are given in Table XXIII ady mun TABLE XXIII Glucose tolerance fest in guinea pigs on different diets Fasting blood sugar Blood sugar after feeding glucose (mg) Animal (mg%) 180 min 45 min 90 min 135 min 3064311 281371 333:24 4 266 +372 251317.2 1941149 210+16 4 136+ 41 Scorbutic (8) 150+ 22 326+239 333 +224 Scorbutic with methylthiouraal (8) IS1107 298 +235 316+31.2 Scorbutic with thyroid (8) 151 S3 331 : 34 317+170 Normal (8) 132 5.4 279-117 251+172 Normal with methylthiouracil (7) 121 57 261 +10 8 256 82 Normal with thyroid (8) 147+ 35 271 +IOS 262173 Figures in parenthesis indicate the number of animals 196+126 146+149 "10202 170+339 ·RELATIONSHIPS AS AFFECTED IN SCURVY 91 There was not much differences in the glucose tolerance test in the animals of the diferent groups The altered carbohydrate metabolism observed in scorbutic guinca pigs, therefore, may not be due to either hypo or hyperthyroidism. These observations indicate that thyroid function may not be disturbed in scurvy Structural changes in testes in scorbutic guinea pigs Chronic scurvy was produced in guinea pigs by feeding them a scorbutogenic diet long with a daily oral supplement of o 25 mg of ascorbic acid Histological studies ere carried out in normal, scorbutic and chronically produced scorbutic guinea pigs Mukherjee and Banerjec, 1954) Complete degeneration of Leydig's cells with increased Ábrosis and coagulative necrosis of the seminiferous tubules were observed in chronic seurvy. In acute scurvy the Leydig's cells only showed early degenerative changes but semunsferous tubules were normal The degencrative changes of Leydig's cells in the testes of scorbutic guinea pigs indicate hypofunction of testes The observation of increased cholesterol content of the testes of scorbutic guinea pigs (Belavady and Banerjec, 1954) indicated that possibly cholesterol could not be utilised by the degenerated Leydig's cells for the synthesis of the testicular hormone The absence of degenerative changes in the ovaries of scorbutic gunca pigs indicated that the gonadotrophic hormone of the anterior utustary was not involved in the degenerative changes observed in the testes of scorbutic umca pigs It was not possible to study the function of the pituitary gland in scorbutic inea pigs as it had not been possible to remove the pituitary gland from guinea pigs REFERENCES Banerjee, S (1944) Relation of scurvy to histological changes in thc pancreas Vature, Lont, 153 344 (1945) Relation of scurvy to the adrenalin content of the adrenal glands of guinca pigs j biol Chem, 159 327-331 Banerjee, S., and Biswas, D K (1959) Unnary excretion of pyruvic acid x-ketoglutanc acid, and oxaloacetic and in scurvy Ibid, 234, December issue Banerjee S. Biswas, D K, and ſingh HD (1958) Studies on carbohydrate metabolism un scorbutic gwnca pigs Ibid, 230, 261-270 1959) Dehydrogenase activity of the tissues in scurvy ibid 234, 405-408 Banerjee, S. and Deb, C (1951) Lđect of scurvy on the cholesterol and ascorbic aad contests of adrenal glands of guinca p!Ss Ibid. 194, 177-179 Baacrjes, $, Deb, C. and Belavady, B (1952) Effect of scurvy on glutathione, and dehydroascorbic acid in gunea pis tissues Ibed 195, 271-276 Bezerjec, S, and Ghosh. N C. (1946) Adrenalin in scurvy. Ibid 166, 25-30 (1947) Relation of scurvy to glucose tolerance tess glycogen of liver and sisulin content of pancreas of gwnca pugs ibid, 268, 207-211 ---(1949) Studies on the effect of thyroid and methylthioureal on the glucose tolerance test in pormal and scorbutic guinea pigs Ibud 180, 189-191 (1950) Effects of ascorbic aad on the glucose tolerance test and on the unnary excretion of sugar and ascorbic aad by diabetic patients Indian J Physiol & All SC 4. 129-536 Banerjee, S, and Ghosh, P K (195) Effect of seurvy on hexokinase activity of tissues of guinea pigs Proc Soc cxp Biol N Y 88, 415-416 anerjee, S, and Kawishwar, W K (1989) Studies on the operation of the trarboxylic zad cycle In scurvy bol Chem 234, 1347-1349 nerec, S, and Singh, H D (1957) Adrenal corucal activity in sorbutic monkeys and gunca pass Amer J Physiol., 190, 265 267 (1959) Effect of exogenous insulin on the dehydroascorbic acid and glutathione content of tissues in survey sa & Ends Res, 17C, 1-4 -1958.1) Cholesterol metabolism in scorbunc gunca pigs J brol Chem, 333, 336-339 rady, D, and Banerjee, S (1954) Metabolism of cholesterol in korbuoc gunea pigs Ibid., 203, Mukherjee, A K., and Banerjee, S (1954) Studies on histological changes ia experimental curry Anet Ret 120, 907-911 ·ACTION OF DRUGS ON THE RELEASE OF OXYTOCIN IN UNANAESTHETIZED RABBITS by RANJIT ROY CHAUDITURY Department of Pharmacology, All India Institute of Medical Sciences, New D The 16 ABSTRACT Alcohol and water which cause a block in the release of ADH also cause a block in the release of oxytocin This pharmacological tool can be made usc of to investigate if sperm migration at co tu is dependent on the release of orytocin The results also confirm earlier work that simultaneous release or block of the neurohypophysial hormones always occurs Chlorpromazıne caused a block the release of orytocin Oestrogen in doses which blocked the anterior pituitary and reserpine in the doses used caused no block in the release of orytocin It is generally believed that uterine contractions presumably caused by orytoan tigated EXPERIMENTAL PROCEDURE Lactating rabbits 2 5 to 3 kgm in weight were taken 10-12 days after birth of the 1 L.LT L. An the 10$ Weight aller SucKIN & Oltre pups alict sepulauiou ovlug il ds Luwu . (1956) When the pups guined the same weight after feeding on three consecuave mornings the drug was administered to the mother prior to feeding on the next day L the pups did not gun weight after suckling for fifteen minutes then an intravenous injection of oxytocin (20-100 milli units) was made into the marginal car vein of the rabbit and the pups allowed to suckle agun When the pups increased in weight after this second feeding chen only was it concluded that the release of oxytoan had spec. fically been blocked by the administered drug The percentage block has been calculated Acht an Arevious 4 Oxytocin was administered in the marginal ear van Chlorpromazıne was adminis tered immediately before feeding by intravenous injection in the ear vern of the animal. Oestrogen and reserpine were administered intraperitoneally at various intervals before feeding of the pups Alcohol and water were administered by means of a soft rubber catheter 30-40 minutes before feeding ·RANJIT ROY CHAUDHURY ACTION OF DRUGS OV TIIE RELEASE OF OXYTOCIN 93 Synococinon (synthetic oxytocil, Sandoz Products) was used in all experiments Ovoc din Ploestradiol dipropionate Coba Led) serpasıl (Reserpine Ciba Led ) and Largacal (chlorpromazine May and Baker Ltd) were the substances used RESULTS T! 1 f.1 TLIT 41 1 linn-ren 10 per as before he block was reversed by intravenous injection of oxytocin (20-100 inilliunits) TABILE Blockage of A1 IL Eject: * 11 Rabbt Drug Litter B Litter C Litter D Litter E Litter G Litter H Litter 1 Blocked Blocked Blocked 100°. Blocked 1000 Blocked 850 100°. Blocked 100% 100° Blocked Blocked No Block No Block Blocked 85 Blocked 74 Blocked 100°. 93º. 100°. Alcohol 175. 10% orally) Chlorpromazine Img/kgmP) Reserp ne (25 mgm-6 mgm kgm I P) Oestrogen (100-200 Microgram 1.P) No Block No Block No Block No Block No Block No Block No Block No Block No Block No Block No Block Blocked Blocked Blocked Blocked Water 5 ml/100 gm Body weight) 100% 800. 100. 100% Chlorpromazine administered in the dose of smem kem body weight intra venously immediately befor Seven arumals In those anum the dose of chlorpromazine Reserpine was administered to six anumals intraperitoneall; (1 5-40 mg/kgm body weight) at intervals of 1 hour 2 hours and 31 hours before allowing the pups to suckle In none of these animals was the suckling reflex abolished When the dose of reserpine was further increased it proved to be touc to the animals which died the next day In these cases the milk ejection reflex was abolished but the blockırg action of reserpore on the suckling reflex was not reversed by intravenous injection of oxytoan Oestrogen was administered in doscs 150-200 micrograms intraperitoncally, 5-31 hours before suckling In none of the five ammals to whom I was administered did it cause a block of the mulk ejection reflex Water--In four experiments the action of hydration on the milk cjection reflex as invesagated It was found that when tap water (5 ml/100 gms body weight) tvas ·9.4 RANJIT ROY CHAUDHURY ACTION OF DRUGS OV TALE admirustered to lactating rabbits 30-40 minutes prior so suckling the milk ejection reflex was abolished and this block could be reversed by intravenous administration of oxytocin 1 DISCUSSION It has been shown by Abrahams and Pickford (1954) in bitches and Harris (1955) in rabbits that whenever one of the neurohypophys:al hormones is released the other is also released These observations based on the measurement of uterine motility and antidiuresis have been confirmed by direct measurement of the hormones m blood after stimulation of the neurohypophysis by drugs (Chaudhury and Walker, 1958). Since the hormones are always released simaltaneously it is not surprising that alcohol and water which are known to block the release of ADH also block the release of oxytocin as has been demonstrated in these experiments. This may be used as a pharmacological tool now be incker (1959) days after puerperium the uterine contractions were not in any way affected by hydration of the subject He argues that a differential release of the two hormones as therefore possible. It is contended that uterine contractions as observed by Dicker are not a specific criterion for assessing the release of oxytocin as the exact role of oxytocin in parturition is not yet clear Milk ejection is a more specific action of orytocin and in the above experiments on the rabbit it has very definitely been blocked by alcohol and water and then reversed by administraton of intravenous oxytoan The above results go against the possibility caused no block at all in two of the animals be added that Moon and Turner (1959) have recently shown that Reserpine inhibits the release of oxytocin as judged by milk-let down yield per time nursing in rats * 1 1 1: mammary glands and even Oxytocin injections could not get out any mulk. The oestrogen had it appeared completely blocked the secretion of mulk. ACKNOWLEDGEMENTS He would like to acknowledge the willing and efficient technical assistance of Mr TC Joseph Added by the Convener

·RELEASE OF OXYTOCN IN UNAESTHETIZED RABBITS
93
REFERENCES

·THYROID STATUS AND ADRENAL CORTICAL ACTIVITY by J S Rawat and A Rox UP College of Veterinary Science and Ammil Hisórndry, Mathura 11 corte cortiu On gross pathogenesis of exophthalmıc goiter partly on the basis of under functioning of the adrenal cortex Certain manifestations are common to over function of the thyroid and under function of the adrenal, Zondek (1935) has stated that thyroidectomuzed animals survive total adrenalectomy longer than those possessing thyroids Koelsche and Kendall (1935) have found that adrenocortical hormones exert a sparing action against the negative nitrogen balance induced by thyrovne A systematic study was undertaken to elucdate the changes that take place in the adrenal cortex physiology in hypothyrc +10 gms , and age 90 0-7 days were Is mg thiourea mixed in their food perods and thus different stages of of animals was maintained as control and thts animal was given same amount ol food as consunied by the respective thiourca fed animal Animals in the various other groups received alongwith chioarca, chyroxine administered orally and/or ACTH subcutaneously, "The dosage of the hormone was sy of 1-thyroxine sodium and o 2 IU ACTH daily --- killed following anaesthetization ministered intraperitoncally The -fore slaughter when completely anaesthetized the adrenals were dissected out and weighed in a torsion balance and mousture and cholesterol was estimated in the left adrenal gland and ascorbic acid in the right Cholesterol and ascorbic acid were estimated as described by King (1947) The results obtained from animals of various groups are given in Tables 1 to 8 and che observations are discussed below - -1 WEIGHT OF ADRENAL Weight of adrenal glands at weekly intervals during the progress of hypothyroidism as a result of thiourea feeding and also during its regress by stopping thiourea administration after feeding it for 28 and 42 days was noted at moet hy thyroid hypertrophy, In weight on the third 1- During the post thiourea teeding period te dulendi gulus were ganing in weight but the degree of improvement in those where thourea was administered for 6 wecks, was less marked, it appears that prolonged hypothyroidism results in more lasting injury to the adrenals (Table 2) ·JS RAWAT & A ROY THYROID STATUS AND ADRENI CORTIN The atranh durng this harice ACTI maucrice oft รพ ศ Lapussive to ACTH admastrabort MEHUn of thyroxine and ACTH alongwith thiourea on the othes hand cause slight hypertrophy of the adrenals The implication of these observation are discussed in detail later dj 1 MON CHOLESTEROL Cholesterol content of the } a ndition for so long a per Weight of tissue matter or 3 fontcho tragon of thyrox 2ed content of: unorcared qasr fc may, therefore, 1 ACT cortical horr and Rosen 3 was adianist more than th Here capable o ACTH durms] Icroll carcy duct teret Size 2 nalcc It is por ciearly known that fort of cholesterol We offe ether factors is depen content of pituitary 15 12 our experiments i decreased in or with throarea noto laden with choleste cholesterol which tom also account for the 13 553!1 ·100 J S RAWAT & A ROY THYROID STATUS AND ADRENAL CORTICAL ACTIVITY TABLES Composition of adrenals of animals during regress of hypothyroidisul mamtained for 4 weeks Absolute content (ang) per 100 mg of adrenalin mg Moisture Ascorbic Choles Moisture Ascorbic Choles- Days aad terol acid terol 7 TU Fed 68 + 242 70 I 200 020 +0003 039 +0004 PFC 48 + 022 80 I 054 71 11.25 72 320 221 #003 396 SO + 23 Xo + $3 +004 XXX XXX XXX XXX 14 TU Fed 72 20 73 + 204 80 + 220 PFC 036 £ 0004 041 0004 75 E 048 88 + 054 377 004 394 75 + 48 78 + 48 #I +004 21 TU Fed PFC 76 + 263 79 Ł 229 039 +0003 044 t: 0004 83 037 91 +044 72 +2S 72 +21 354 +003 402 +004 78 35 83 + 40 TU Fed 67 88 + 362 040 + 0005 047 tooos 80 - 072 80 74 +30 73 +28 359 E 004 407 PFC + 60 60 94 £ 369 £ 044 004 TABLE 6 Composition of adrenals of animals during regress of hypothyroidisin maintained for 6 u erks Absolute content (nig) Per 100 mg of adrenalin mg Moisture Ascorbic Choles Moisture Ascorbic Choles- Days acid terol aad terol 7 TU Fed 68 + 043 76 + 283 81 + 31 O22 +0002 039 +0003 72 #31 72 241 + 002 356 + m2 76 48 74 sz PFC 81 063 +28 XXX > х 14 TU Fed 68 + 312 77 + 189 033 £ 0003 047 +0003 68 +040 68 034 71 +30 69 +22 383 +004 443 #003 47 = 42 56 + 30 PFC XXX XXX 21 TU Fed 69 + 248 023 + 0002 038 +0003 os + 031 68 * 042 69 +2s 70 +30 254 # 002 332 + CO2 66 Est SO + 35 PFC 84 E 252 XXX XXX ·} S RAWAT & A ROY TITYROID STATUS AND ADRENAL CORTICAL ACTIVITY TAELE 7 Composition of Adrenals es offered by simulimepus adminstration of thicures anal Harinones, ronimously for 3 days (For ready reference the compositos of adimals of 28 day tourca fed animals and their pusr fed controls have also bere incorporated in the tabic) Absolute cuntent in mg Par fed Thou.cz Thourez Throutca, Thiourea, control TX MCTH TX, ACTH 690 09 Morsture 77 + 220 70 240 Ascorbc acid 0 270 +0003 Dj& 0003 026 0001 020 + 2002 Ozo +0002 0914 300 SI5 Cholesterol +61 0524 5038 200 3.071 -oor TALE 8 Glycogere content on ms of whole fresh lover of animale as affected by amull.mpom administration of thronted ond hormones continuously for 18 days (For ready reference the composition of layer of 28 day tourea fed animals asid theur par fed controls have also been incorporated in the table) Pair fed Thiourea Tissourca Tuourea Thiourea, costsol TX ACTB TX, ACTH 40 40 Glycogen as glucose iza + 28 In 135 - JO ·DISCUSSION Subrahmanian Dr Banerjee, what is the effect of vitamin C-deficiency on protein metabolism Different fractions of plasma proteins were determined by paper electroPOLESIS !! - Land genesis is not clear Prasad Dr Banerjee, what is the relationslup between decrease in hexokinase activity and ascorbic aad deficiency? ----- there we *% decrease in liver hexokinase activity and 8% decreasc very interesting Could ovaries > I any observation on the Banerjee No histological changes were observed in the ovaries of scorbutic guinea pigs lt 15, therefore, unlikely that the degenerative changes in the testes was of pitutary origin due to defect in the 1. ... that sernthic and wluch plays a role in the testicular 1 resulting in its degeneration Das Gupta De Banerjee, what was the level of ascorbic and in the adrenals of scorbutic guinea taal > - *00 gm rmione s and as cretion .bserved se these

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·SECTION D
PITUITARY-ADRENOCORTICAL
MECHAMSMS

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·STUDIES IN ENDOCRINAL INTERRELATIONSHIP USING A HISTO- CHEMICAL METHOD FOR THE EVALUATION OF SUCCINIC DEHYDROGENASE ACTIVITY AS AN INDEX by E) DE SOUZA and T H RINDANT Department of Anatomy and Physiology Topiwala Vional Mcd cal College, Bombay 8 ABSTRACT Interrelated activity between the thyroid and testis, the thyroid and the adrenal corter, and becuceri the adrenal cortex and the testus has been histochemically demonstrated in experimentally treated this Alterations in thyroid status affect both the levels and activity patterns in the testes and supra renal cortex Compensatory atrophy of the adrenal cortex while also showing changes in lesel and pattern of SD activity in itself also brings about general lowered activity in the testis as well as interferug with the activity level in spermatogesis Both thyroidectory and compensatory adrenocortical atrophy lowered the activity of S. Dan the liver as well as altered the zonal pattem The functional interrelattonship among the various endocrines is well known That the activery of the gonads is anfluenced by the adenohy pophysis the adrenal corter and the thyroid kas been shown both in experimental animals as well as in clinical syndromes associated with the anomalies of these glands The methods used so far for demonstration of these interrelationships have been mostly clinical evidences of altered function or morphological and biochenucal changes In experimental animals (Rından 1957 1958) The application of histocherucal methods to the study of the biological activity of Importanc cellular elements of various organs in the body has howcier the disana advantage of locating sites of altered activity under various conditions Thus the localization of sucanic dehydrogenase is of great value in this respect as the speafiary of the enzyme -substrate reaction and the demonstratwn of its activity by means of a tetrazobum (Nitro 3.T) reduction technique renders it as one of the deal indices of organ metabolism at histochemical and even cytochemical lesel (De Souza and kochare 1959) Succinc dchydrogenase activity has been shown in be altered in the liver tissues under conditsons of change in gonadal function (Rındamı 1958) Even the highly specialized structure like the sperinatozoon lends itself to such a method of cs alusdon of the physiological normality or otherwise (Kothare and de Souza 1958) This study was therefore undertaken to investigate the thyrordgonadal relatonship by roang the variations froni normal in the sucanic dehydrogenase activity of the testis under alccred conditions of thyroid activity Simularly the samic enzyme was used to note the relationship between thyroid status and adrenocortical activity and the influence of the adrenal corter on the gonads MATERIALS AND METHODS The studies were carried out on male albino ro waghing about 100 g cach The animals were divided into four groups and treated as under ·ITO E J DE SOUZA & T II RINDANI STUDIES IN ENDOCRINAL INTERRELATIONSHIP USING The basis of this techniquc lies in the fact that whenever the enzyme sucanic dehydrogenase is active, electrons are released during the breakdown of sucaric aad to fumanc and The electrons are normally utilised in the reduction of the cytochromes and further made available for the synthesis of ATP and protein or for energy requirements of cells in their burst of metabolic, motor or secretomotor activities By the use of tetrazolium salts in general and p Nitrophenyl substituted ditetrazole (Nitro BT) in particular these electrons reduce the normally soluble protein binding colourless tetrazolium mto a blue insoluble diformazan at the exact site of enzyme activity (Nachlas et al 1957) Nitro BT has spcaal advantages in a study such as this The first is the extremely finely particulate deposition of diformazan rendering it cytocheniucally an excellent index of the degree of activity in the various layers of the seminiferous tubules and hence in the various stages of spermatogenesis Another advantage is the fact that the reaction resulang in an alcohol-xylol fast diformazan has great speed and is a high electron compctitor, minimizing falsification by a possible cytochcome alteration and the staining result is thus more truly that of succinic dehydrogenase than is possible with other tetrazolium salts Inadentally thus makes possible permanent sections mounted in balsam (one of us E J S has secaons four years old without appreciablc loss of staining results) Further, the fact that a fat soluble monoformazan is at a mimimum is important in organs with hugh fat content like the adrenal (especially during adrenal hypoplasia) as diffusion artefacts and false cytological localization are prevented This is best exemplified in the increased fatty deposition that takes place in a broad belt in the peripheral portion of the zona fasiculata and the glomerulosa in induced hypoplasia of the adrenal This is seen as a belt of almost total inactivity Thuis is quite different from normal fat cells where even though the fat is negatively stained the cytoplasın of normal adipose tissue is quite heavily loaded with formazan The method besides classifying zonal patterns of enzyme activity and intracytoplasmic localization can also be used for a rough quantitative estimation of activity as differences un colour intensity denoted differences in activity levels when scctions of fautly identical thickness were incubated under unform conditions for cqual times As the adrenal hypoplasia had to be effected by the administration of hydrocortison acetate the animals were sacrificed after the second day to ensure best effect with no res dual cortison action This was checked histologically by the complete band of far change in the distal fasculate zone the smaller adrenal weight and by the fact that i lowered SD activity in the check liver sections is in consonance with the findings of Mal: and Bourne in adrenalectomised animals (Malaty and Bourne, 1953) The studies reported here indicate that the hormones of the thyroid and adrenal cortex influence the activity of the testicular function at a metabolic level where the adrenocortical hormones seem to be concerned with a regulation of the general metabolic activity of all the cells of the semınıferous tubules, the thyroid hormone appears to have an effect on the processes involved in the differentiation of these highly specialised cells Similarly the adrenocortical activity also seems to be influenced by the thyroid hormone acting at a metabolic level, beng probably concerned in the intermediate chemical changes concerned in the synthesis of the cortical hormones So far as the general metabolic effects of the thyroid and adrenal cortex are concerned it seems quite certain that as judged by the influence of the deficiency of their endocrines on the SD activity --

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·Bull Nar Inst Sa Indii, No 17, Plate IV DE SOUZA & RINDANI Fic 6 Photumia roztapla stwisng SD activity the normal rit adrenal (* 100) TH 7 Photomicrocryph slow sity SP activity 11 the adrenal after thyroidecfunt Theri 15 a Acirally lower activity The high activity 51 duced to a few sills du po m thi pusti medullwynu (X 100) FK & Ph xanncrograph sh - ing Stuty in the rat adrenal afres confistory atraphy There is formazed acn vity with almost coral absence ut the fasisulit but fruly h ch activity 22 the glomerului 100)

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·A INSTOCHEMICAL METHOD FOR EVALUATION Or SL CCIVIC DUYDROCLS ASL ACTIVITY m the laer the chenucal reactions occurring in lycr round the Krebs atric acid cycle arc substantally influenced by these hormones 10 ACKNOWLEDGEMENTS The Authors are thankful to Di L Monteiro Dcan, TN Medical College four the interest Laken in the studies and to MNM Chavan for the assistince rendered in the work, RETERENCES 1. Souza E J and Kothare N (1939) A method t r the cytocl cinn a demonstration of suluu dehydrogruase on Human leucocytes | Hiflcm and perhom 2, 77. Kothare S N and de Souza, L) (1958) Cytox hemn al dem istration of sucaruc de hydrogenase patterns in human spermatozoa | Post Jus te vind 4. Mality, II A and Bourne C II ( 13) The Effect of adrenalecta ny and a zject of steroid horntonics on the sucanic dehydrogenase reaction of tissue ] That Land 87.449 Nachlas M M., Tsou k C de Souza L. (heugs and Seligman A M (1957) Cytochemical demonstration of succinx dehydrocnaxe by the use of a new p-Nazophenyl substituted ditetrazole J Hi dachem and (y hom . 420 Rundani, T 11 (1957) Relationship of mak kth monc iu stress mduced activation of adrenal cortet un rat Inds) nied Res 45, 1.1 Rundam T H (1958) Is fluctc ftest sterone propie nate on the succinic dehydrogenase activity of male rat laver And J Plyst / 193 (1) 73 ·INFLUENCE OF ADRENALS ON THE CYTOLOGY AND HISTOLOGY OF GASTRIC MUCOSA AND THE GASTRIC SECRETION OF PYLORUS OBSTRUCTED RATS by G B SINGIL and S H ZAIDI Central Drig Research Institute, Lucknow ABSTRACT Studies on the influence of adrenals on gastric mucosa and tlic gastric secretion mı normal and pylotus obstructed fats have been made There was no ulceration of the gastric mucosa m normal or admalectonuzed rats without pyloric obstruction Pyloric obstruction in adrenal intact animals caused various grades of ulceranon in th. forestomach Adrenalectomy completely protected the gastric mucosa from ulceration in pylorus obstructed rats and also significantly reduced the free acidity peptic activity and total volume of gastric Pepsinogen contents of the glandular mucosa was diminished after adrenalectomy in normal raus and could not be detected when adrenalectomy was associated with obstruction The influence of adrenals on the gastric mucosa of pylorus obstructed rats has beca investigated and various views regarding its significance expressed Adrenals played juce on gastric mucosa and the gastric secretion in pylorus obstructed rats MATERIAL AND METHOD 1 Histopathological technique-Autopsy was performed on all the animals sacrificed After opening the abdominal cavity the stomach was inspected in situ for distension, congestion and perforation It was then taken out, a small slit made in the duodenum along the greater curvature and the contents drained in a beaker The stomach was then opened, inspected for any ulceration and graded by the method of Barrett et al (1983) The tissue blocks from different portions of stomach were fixed in 10 per cent formal salıne and Rrgaud's Aud After complete fixation the blocks were embedded in paraffin and section cut at si Sections were stained with haematoxylin and cosin, Pas for muan (Pcarse, 1949) and for pepsinogen by Bensley's neutral gentian stain (Cowdry, 1948) Brochemical technique-- The gastric contents from each animal were centrifuged to remove food particles Peptic activity of gastric juice was estimated as described by Glick (1958) ums phenol reagent and casein substrate The results have been expressed as 'pepsin unit Frec and total aadity was titrated with o oi N-NaOH using Topfer's reagent and phenolphthalein as indicators and expressed as meq/L.

·CB SINGH & S 11 ZAIDI INTLLENCE OF ADRES ALS 113 - PLAN OF EXPERIMENT The follon ing groups were consatuted -- Group I Six normal rats Group II Six adrenalectomized rats Group III Ten non-adrenalectomized pylorus obstructed rats Group IV Ten adrenalectomized and pylorus obstructed rats EXPERIMENTAL PROCEDURE All the animals were fasted for forty-eight hours Pylonic obstruction was done in groups 1 and IV by the procedure previously stated and animals sacrificed sitteen bours after the obstruction Rats of groups I and II were further fasted for sixteen hours and sacrificed along with the pylorus obstructed rats This was done because the fasting period of fortyright hours before the pyloric obstruction followed by sixtcen hours when the animals were sacrificed brought about maximum structural changes in the gastric mucosa RESULTS Gross appearance--The naked-eye examination of the stomach in situ in groups I (Normal control) and I (adrenalectomized) showed smooth shay serous layer On opening the stomach was seen to have normal mucous layer Group (pylorus obstructed without adrenalcctomy) showed marked distension congestion of the serous laser On opening the stomach the ulcers were seen to be confined mostly in the frestomach Small ulcers appeared as any rounded lesions while the larger ones were irregulara sacacular in share Onc animal showed ulceration measuring ? inm ene hadi lcers 4-6 mm three showed large number of ulcers 1-3 mm and in the remaining three animals few small ulcers measuring 1-3 mm in onc diameter were present Tuc animals died in the process of investigation, the cause of death could not be ascertained Group IV (adrenalectomized and pylorus obstructed) showed slight congesaon and distensial of the scruus layet No ulceration was present in this group of animals Three as inuals died in the course of experiments The gastric mucosa showed greyish brown appearance The gastric contents were dirty brown in colour (Table 1) TABLE ! Summary of results of brod omical invest gat ons of gastne ju e and degree fulcreto adrenal inf.K adrenalectomized pyl rus obstru ted rais Volume Aad ty l'opia Degter of Group and animal numbet Focs Total activity ulcerabo сс PU.X10* Group III (Adrenal satact) mcg L 1 3 3 33 56 SS SO So SI S 6 7 30 10 48 54 74 22 100 100 95 Tos 8 69 56 3 PU --Peptic Unit. 039 0.40 040 0.44 0.43 040 0 1 037 SS 0 45 50 ·114 COSINGR & S 7ASBE INFLUENCE OF ADRÉNALS ON CYTOLOGY AND HISTOLOCY TABLE (Concld) Volumc Group and anumal number Aodity Free Total Peppic activity PU*X10 Degree of ulceration CC meq/L Mean Median Range 442 830 52 410 880 47-S 220740 S6 D--1080 042 041 o 37-46 26 30 ID 4.0 Group IV (Adrenalecton mized) 0 1 2 3 30 1.0 24 a 0 20 O 10 017 023 0.20 0.10 013 08 OS 18 10 O 0 0 7 16 24 Mean Medinm Range 13 88 27.2 IO 240 OS-30 06-22 0 20 D460 60 018 0 20 010-023 00 00 00 Groups I and II -Sufficient quantity of gastric juice was not available for biochemical investigations The mean values of gastric juice, free acidity and peptic activity for group 111 are significantly higher than for group IV at 95 per cent level. granules in all the animals in group IV, ulceration was not present. Mucosa showed norinal appearance, except congestion in the submucosal layer Peptic cells in all the animals were mostly devoid of pepsinogen granules (Fig. 4) Mucous cells were not affected in any of the groups Parietal cells were slightly diminished in size in groups III and IV Biochemical group IV at 95 per cent level ·GB SINGIT & SH ZAIDI. Hull Not Inst Sc India, No 1, Mate V 1. FIC I Pupsiogin granules sa normal stouah (Bensley's neutral gental xam to 2 rinola 10 noul adre. Ich Banknow- 3 FIG. 3 Larly croston shuns mg de struction of the supertorial lantat the summer the hun B and I 4U 4 Ayia of the ATAX ITI hells the Tal 1981

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·OF GASTRIC MUCOSA AND THE CASTRIC SECRETIOV OF PYLORLS OBSTRUCTED RATS 115 Fig.5 MEAN VALUE OF FREE ACID. PEPTIC ACTIVITY & DEGREE OF ULCERATION 064 607 os 50- 2.6 44 401 FREE ACID PEPSIN UNIT X10 THE PEPTIC ACTIVITY FREE ACID,m eq/ E DEGREE OF ULCERATION 204 10 DEGREE OF ULCERATION O 18 ADRENALINTACT RATS ADRENALECTOMIZED RATS FIG. 5 Mean value of free acid. peptic activity and degree of ulceration mi adrenal ustact and adrenalectomized pylorus obstructed rats Anduy--The mean values and the range of free and total aadity are summarised in Table 1 and Fig $ Enough gastric juice was not available for brochcnucal estimation of free and total acdity in groups I and II The mean value of tree acdity in group I was !!2m cq/L and in group IV only & 8 m cq/L The mean value of free acidity in group III is significantly higher than group IV at 95 per cent level Peptie echivity-- The results of peptic activity csamations of the gastric juice are shown in Table I and Fig Enough gastric juice was not available for estimation of portic activity in groups I and II The mean value of peptic activity was 0 42 X 10 and 0 18 X 80P. U. in groups III and IV respectively The difference in peptic actiun is significantly higher in group III chan group IV al 95 per cent level DISCUSSION Gray and Ramsey (1956), poured out that under normal arcumstances the stomach has under parual control of the adrenal cortex and sequired normal adrene cortical function for peptic activity The diminuzun of gastric pepsinogen a normal adrenalcctomuzed rats in the present experiments indicates that there may be a direct effect of adrenocorecords in the formation of gastric pepsinogen or on the control on the ability of the pepac cells to synthesize pepsinogen Pyloric obstruction in adrenal intact rats brought about deplcaon of mucosal popuhogen and shrinkage of the pepac cells. Pepuc acuvity in the gastric juice was also norcased svluch showed a posiave relationship with the increased degree of gastric ulceraon This confirmed the findings of Singh, Zaidi and Balkrishna (1958) sho showed that psingoen in pylorus ligated rats was significantly lower than in the normal, and or creased pras activity was directly proportional to the degree of pepoc ukeraton Gray stol 1957) suggested that under conditions of stress the stomach was more directly under the sumulatory control of the adrenocorticoids through hypothalamic-pituitary-drenal 2024. Pyloric obstruction in adrenal intact rats, by process of stress, possibly brought the somach under the direct stimulatory control of adrenals and thus depicted more the Dengan than normally forned ·116 G. SINGH & SH. ZAIDI : INFLUENCE OF ADRENALS In the present experiments adrenalectomy significantly lowered the volume of gastrac juice as well as free acid and further marked inhibition of peptic activity. The cytological 1 ACKNOWLEDGEMENTS Thanks are due to Dr. B Mukerji, F. N. I for his interest in this work The authors are grateful to Mr M M Soni and Mr. Mulkraj for their technical assistance. REFERENCES Barrett, W E. Rutledge, R, Plummer, A J and Yonkman, F F (1953) Inhibition of ulcer formation in the Shay rat and reduction of gastric acidity in dogs by antrenyi (oxyphenonium) (BA-5473) Diethyl (2-hydroxyethy!) methylamonium bromide a-phenyl-cyclohexane-glycolate, an anticholinergic agent) Pharmacol, 108, 305 Cowdry, E V (1948) Laboratory Technique, 2nd ed p 167 Williams and Wilkins Co, Baltimore ." shers, Inc, New York >mach peptic ulcer in 73 cent Progr Hormone Res Vol 13 583, Academuc Press Inc Publishers, New York Harouturuan, L M and Segal, H L (1952) The effect of adrenalectomy and nephrectomy on pylorus legated rats Gastroenterology, 27, 356, ·DEMONSTRATION OF THE EXISTENCE or PRECORTICOTROPIN-A PRECURSOR OF CORTICOTROPIN (ACTH) IN THE OXY ANTERIOR PITUITARY GLAND by P R Diena Central Drug Research Institure Lucknu In addigon to ACTH (adrennan called AW-factor causmg only adrenals of hypophysectomized . 1193 and Dixon et al (1951, 1953) to us į It sent in the ox pick utan tissue Stack-Dunn. " Young (1954) and Cater and Surk-on-1 acha 2 uds or complete absence Ludus ul activity detectable in Sayers festas intriguing particularls since frak or anterior pleustary issue contained much crac te spin cxtractable by acid acetonte The onginal aim of the peren AW-factor presc to be des oid of sha 1 € c Urucot rin It is intended to describe in chus paper only threr son trate the existence in the or antcriot pitstary & of corticotropus The postaon of AW-facto subject matter of a separate communication MATERZALS & ALETI1ODS (0) Rets. Male albino fats, 8-12 ucсks old kere in a constant mitronment sett used. Adult female sats were excasional choice (1) Crude sterk alkalas auto Jorbo minutes after killar extracted according to will be referred to here igm glind - 4 mls FGS 00-80 mg profcun بل ·RESPONS & md RDRECORSK ACID 118 PR DASGUPTA DEMONSTRATION OF TILE EXISTENCE OF PRECORTICOTROPINA (3) Crude ACTH from FGS-A certain volume of FGS was dried wlule frozen The solid obtained was treated with hydrochloric acid and acetone according to the method of Lyons (1937) and the extracted material corresponds to the L-fraction of Dixon and Stack-Dunne (1955) (4) "Precorticotropi" - Extract- Fresh or anterior pituitary lobes were dissected, minced, dried by filter paper, promptly weighed and then homogenized tvith glass homogenizer in chilled (40) 09 per cent sodium chloride soluton The homogenate (20 mgm dry tissue/ml solvent was spun (18 000 $) for 30 minutes The clear reddish supernatant flud (pH 65 to 67) which contained the "Precorticotropin" was stored in the cool chamber (0°-4°C) (s) Bioassay-The ascorbic aad-depletion method of Sayers et al (1948) was used with minor modifications The positive response is indicated by a fall of adrenals ascorbic acd-level (in vg/100 mg tissue) caused by an intravenous injection of the test material (025 ml/100 gm body weight) RESULTS Sayers test of crude corticotropin extracted from FGS Table r shows the adrenal ascorbic acid depleting activity of crude corticotropin extracted from FGS as compared with the activities of a standard ACTH preparation (Armour 84-85 H - kindly supplied by Armour Laboratories, Ltd, U SA), of the ongual FGS and of FGS at pH 3 0 it us found that although the original untreated FGS possesses virtually no adrenal ascorbic acid-depleting activity yet the crude cortin cotropin extracted from it is active Further, activity appears in the FGS when its pH is adjusted to pH 3 o by adding Hcl (2N) The reason for changing the pH of the original FGS to pH 3 is that in the Sayers assay the test material is administered after dissolving it in a trace-acetic aad-containing og per cent sodium chloride solution whose pH is 3 Effect on the Sayers activity of FGS of lowering the pH Irrs 100! bu oar/bar 70 60 50 40 10 to PH F61 FEFECT ON THE ACTIVIV OF KGS IN THE SAYERS TESTOF TOWER NG THE PH FOR APPROXIMATELY 10 MIN ·Tapu ! Sayers actis sty of the crude corticotropent extracted from IGS {Tigures sielun parenthesis indicate rotal sumber of animals minus vign indicates positive response) Change of adrenal ascorbic and levels (in ug/100mg adrenal weight) produced by Armour 84-85 H Original TGS (pll Crude ACTH TGS atpil 3 (20 mg om tu per ml) 85) (2 mg per mi) protein per ml) Mcan response standard error of the incan -46+5 830) 10+124(4) 101 + 198(3) -7219 (3) TABLE 3 Original TGS (p!! 8 s) Incubated Mican rffect on the Sayers actaily of TGS of lou cring the pll md then maubosting at 37° for lor (rigures unthin parentheses indicate total number of animal Mnus su indicates posstue responses) Change of adrenal ascorbic acid levels lug/100 mg adrenal weight ) byponse Not incubated 7 SEM FRECURSOR OF CORTICOTROPIN (ACTH) IN TIE OXY AYTIRIOR PITUTTARY CLAND 119 5 TGS mculated at 37* for 1 hr after adjusting the pil to 1 (2) 3 3 + IM 91(4) +10.13 50 -18 01 (3) --44936) --74461(3) SCM - Standard error of the incan 65. TOO(3) $6L13 2(3) ·120 PR DASGUPTA DEMONSTRATION OF THE EXISTENCE OF PRECORTICOTROPINA Effect on the Sayers activity of FGS of addition of urea Table 3 shows the effect of addition of urea on the activity of FGS as compared with the shots of Lwas free pink soli amount of crystalline urea (6 6 M) when a clear transparent solution was obtained It is clear that Sayers activity appears in the FGS after addition of urca to it (Table 3). TABLE 3 Sayers activity of FGS treated with stea (66%) (Almus sign indicates positive responses figures withm parentheses indicate Total number of animals) Change of adrenal ascorbic acid content (ugm /100 mgm adrenal tissue weight) by FGS after various treatments - TREATMENT Mean response ASEM (*) Nonci e original FGS Adjustment to pH 3 Urea (66 m) Blank 6 om urca solution only -1983 (3) -35 +60(3) -66 + 36 (4) +11 42 (3) SEM Standard ctror of the mean extracts Sayers activity of "Precorticotropin" extract Table 4 shows the ascorbic acid-depleting activity of the "Precorticotropin' (see page 3 for preparation) before and after treatments with various activating agents Effect of Sodum chloride on "Precorticotropin" Supernatants were obtained in the identical way as in the previous experiment out of a fresh pituitary-homogenates prepared in os per cent, 09 per cent and in so per cent sodium chloride solution without adjustment of pH The adrenal ascorbic aciddepleting activity of such supernatants was determined Further, the activity of the supernatants in o 9 per cent and in os per cent sodium chloride solutions was determined separately after adding fresh quantity of sodium chloride to it such that its final concentration was so per cent The results have been summarised in Table s It appears that sodium chloride also is capable of modifying the native precorticotropin into the material active in the Sayers test Fffect on "Precorticotropın" content of pituitary of acetone treatment and of repeated freezing and thawing A similar experiment (Table s) was conducted using supematants of homogenates prepared not from fresh untreated glands but from glands which were initially subjected ·Pol Sru Sayers add ty of ile supernatant flu d of the hon yone e offehera! 'p #syi suc (F gures x chin parentheses nd cate totalnun ber of an als min ssgn ni cares posit ve respo x) Fresh or anterior p tu tary tssuc hon ogenate (o ing per mi) d v ded 1 to 2 port of $ Sold diy ded tel omog 1 sed otwor ons Sugenda dv det 3 portos Poron 11 d luted 10-fld (Solro A) MR SEM 110$ 7 (s) Port w d lu ed (& pr ctn ml) P u" adju ed to p}} (Sal on dle (s mg tissue nl) MR SEMA 13 014) (Sol on C) MR SEM 138 P MR Jlued (Sol D) SEM 1 KOC 7 PRECURSOR OF CORTICOTROPIN (ACTII) IN THE OXY ANTERIOR PITUITARY GLAND 121 u MR !! adjut di pll di (15 8 prot (Sid SE! -4 No 9) ini) Pill urca added (6 6 M) d lutet (!smg prote (Solon n MR SIM 61+ 15 0(1) NB) D lut s were of Id cach cac mal wihoy par ce sau chir le sol co 2) MR Mean response 3) S EM --Standard come of the na ·TTI TABLE 5 Effect of Sodium chonde en Precosiscotropin Change of adrenal ascorbic aad (ug/too mg adrenal weight) produced by precorticotropin ---Extract made in sodium chloride solution of various concentrations 50% NaCl (1 ml extract=10 09% NaCl (1 ml extract 07 mg N.) os% NaCl (1 ml extracto s mg N.) mg Nh) Untreated & Adjusted to pH Untreated & Adjusted to PH Treated with Untreated & Adufsted to TH Treated with diluted 3 & diluted diluted 3 & diluted NaCl & diluted diluted 3 & diluted NaCl & diluted -48+21 (3) -567 (3) -454 0(2) -82+26 (2) -5072(4) -8£77(3) -694:9 4(3) 6258 3(4) MR. + SEM P R DASGUPTA DEMONSTRATION ON THE EXISTENCE OF PRLCORTICOTROPINGA NB Requisite amount of sodium chloride was added to the extract such that the final concentration was s per cent the extract was then incubated at 37°c for hr before testing 1) In each case the dilution was 20-fold and was made with distilled water 2) Figures within parentheses indicate the total number of animals taken 3) Minus sign indicates positive response 4) SEM --Standard error of the mean s) MR --Mcan response ·123 PRECURSOR OF CORTICOTROPIN (ACTH) IN TILE OXY ANTERIOR PITUTTARY GLAND either to (1) drying with acetone, or to (2) repeated freezing and thawing Table 6 DISCUSSION 1 L 15 justed to 30 (lable uy the reason lor dujastu tu yli juu OLU! the test inaterials (Table 1) is that in Sayers assays done rouunely by us the test inaterials are dissolved in 09 per cent sodium chloride solution containing accuc aad (o 25 per cerit) pH of hich is 30 This latter finding should naturally prompt one to study the influence of pH on the unknown process associated with the production of Sayers active material in the FGS However, there appears to be no such effect of pH change on the production of Sayers activity in TGS (Table 1, Fig 1) w kuch could be regarded as optimal indicating the unvolvement of enzymes in the process It is of interest to Sayers activity appears in FGS when its pH is brought just to the acid side (pH 6 ) and the virtual lack of response at pH's 60 and so is due to the fact as observed by Geschwand Here it can be argued that the absence of activity in the Sayers test of the crude alkaline extract us due to the presence of proteolytic enzymes in such extracts which affect material active in Sayers test much more adversely in the alkaline side than in the acid side That this, however, is not the case would appear fronı what follows Suce some biological substances of neutral nature such as trca have got profound actions upon the native structure of proteins the effect of urca on the abosc prinaple of FGS was studied Interestingly enough, the prinaple macerse u Sayers test has been changed into the active form (Table 3) in the presence of urca also fycatment required that the change brought about in His is not ol arasc nature, and fertains rather to the Internal condition of the above fanarle this to tos ceremaal The question now arises whether this new prmarle detected in the FGS is a native tubstance of the pituitary gland or an artifact of preparation One way of ansu TINS ·124 PR DASGUPTA DEMONSTRATION OF THE EXISTENCE OF PRECOTRICOTROPINA this question would be to demonstrate the presence of the principle in an extract prepared from very fresh pituitary glands under such muld conditions which preclude the chance om sinan C. above conditions Moreover, since this principle is present also in FGS it appcars to be stable indefinitely at pH 7-9 It should be recalled that the activity of corticotropin is lose in presence of alkalı Does the pituitary gland synthesize the release solely this proaple into blood where it changes rapidly into corticotropin under the conditions of stress Is then the corticotropin obtained from pituitary an artifact of preparation : However, this does not appear to be the case In the fresh homogenate of anterior pituitary tissue only a portion of the total corticotropin content is present as precorticotropin and is ineffective in the Sayers test (Table 1) It is possible that the conditions of killing the animals from which the pituitary glands are obtamcd, or the tinic which elapses between the death of the animal and the preparation of the homogenate, may affect the content of precorticotropin of the glands it is also possible that the pituitary elaborates and releases a mixture of corticotropin and precorticotropin in a fixed ratio which is mamtained by breakdown (or otherwise) of the latter inco corticotropin (homocostats): Is needs further investigation to answer these questions What is clear at present is that the ox anterior pituitary gland contains a principle which though inactive by itself in the Sayers test is convertible into a material active in this respect It is, therefore, proposed that this pituitary principle acting like a precursor of corticotropin is as designated as **Precorticotropin It was now decided to try extraction of fresh pituitary tissue for precorticotropin f c 11.1 -11. that drying of tissues with acetone or subjecting it to repeated freezing and thawing is times) prior to extraction with o9 per cent sodum chloride produced similar cffects TABLE 6 Adivities in Sayers test of the Supernatants obtamed from homogenates of (1) acetone dried (2) frozent or anterior pituitary tissue in o 9% NaCl solution (Figures within parentheses indicate total amber of animals, minus sign indicates positive response) Change of adresyal ascorbic acid level (ogma (taq mgm adteaal weight) by- Supernatant from acetone dried Supernatant from frozen gland gland homogenate homogenate Untreated & Treated to pH3 Untreated & Treated to PH3 diluted 10-fold and diluted 10 diluted 20-fold and diluted 20- fold fold Mean Response + SEM --127-59 0(6) -139+16 3(7) -70+10 8(4) -98+1 4(4) SEM= Standard error of the mean ·PRECURSOR OF CORTICOTROPIN (ACTH) IN THE OTY ANTERIOR PITUTTARY CLAND 135 The observation made in presence of acctone is in agreement with that by Carlisle (1958) 17 respect of a crustacean chrom-activator Whatever may be the nature of actions of these trcaonents upon precoracotropm It is unbkely that the change brought about by PH-lowering urca and sodium chloride would go beyond breaking of hydrogen bonds of the native materul Hall (1944) Mates and Tume (1948) and Carlisle (1957) obtawed highly lactogenic material trom pituitary extracts which by itself was inactac in this respect when tested in rabbies In fact, Carlisle (1957) believes that the conventional prolactin (luccutropin) is really an artifact of preparation le secins quite probable that the presence of precorticotrupin in a crude alkaline extract of or pituitary tissue accounts for the striking discrepencies between Ad-activity and AW-activity of such extracts Furthermore this demonstration of the presence of prou corticotropin in the pituitary tissue can pertinenchs be expected to shed some new lighe on the probable relationship among (1) the factor of pregnant marc serum described by Golla and Reiss (1941-42), (11) the activable corticotra pm of Addisonidan blow reported by Moruzzı et al (1951) and (w) the adrenal cght mcreasing factor of plasma of patients with Cushing's Syndrome due to bilateral adrenal hyperplasia and of pregnane women farcly described by Jarler et al (1997), lo is tcmpung.co speculate that our precorticotropin is identifiable with the actrvable cutticotropin of Moruzzi et al (1954) It is probable that our procorticotropin is present in blood and that the high level of sodum and chloride sons in the blood of patients with Cushings Syndrome users precortcotropin into corticotropin as soon as the former enters the circulatie n and that the high leel of urca (another of our activating agents) > rendered operate in the blocul of Addisonizans due to the high level of potassium u hich haver as low in the blood of patients with Cushing's Syndrome ACKNOWLEDGEMENTS A note on some of the earlier tindings made in the mastigat un described in this paper was published 12 Nature (182, 32 1958) jomth with Professor i G Young TRS who suggested the problem and to whom the aucher is greatly mdebred The author as grateful also to Dr B Mukerji, Director Central Drug Research Institute Lucknow for allowing him to continue the study at CDR I Thanks are due "Aus M B Thous of the Biochemistry Department University of Cambridge and co Spax Narain of the Endocrinology Section CDRi Luckus fr their skilled assistants REFERENCES Carlsle D B (1957) Physiol Comp 1959) Hurt Lond 182, 32 Catst D B and Stack Duane MP (1958) Cuba Foussanon Call quia on Joctuiel y 8.31 Den H T F Stack-Dunne M P Young FG and Cute D 13 (1951) Wfure Lens 108. toks Thron, BF Stack-Dunne N and Young F G (1993) The Supremuleren Prffs Syair Colston Ra Soc Bristol (April 1952) Odt) M Yoffey 1 (Butterw hi Hi Puh London 1953) Deon B P F. and Suck Danne M P (19şs) Bachom 61, *k! Geschwind IL and LCR (1932) Ind cromley 50, 2.0 Golla Y M L and Reiss, MA (1941-42) J Phyu / 100, ! HNL, S R (1944) End minor 34.14 Juk ) w. Longson D and Christy N P (1957) J du laveu 36, tros lrom, U R. (1937) Cold Spring Higher Sympa Qrant Bal 5,197

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·HISTOCHEMICAL STUDY OF ADRENAL CORTICAL ACTIVITY IN TOAD (BUFO MELANOSTICTUS) by CHANDICAJARAN Des and MUTYUNJOY MUXILERJI Department of Physiology, University College of Saence, Calcutta The study of comparative endocrinology have gained importance in recent years The adrenal gland in toad is such that it can not be separated from the kidney, making brochemical estimations impracticable So adrenal cortical activity in this spears onder inous conditions was studied by histochenucal methods Seasonal Variation Mukherji and Deb(1959) have studied seasonal valudos m neutral Ispads, wasaturated lipids, plasmalogen and cholesterol en toads adrenal during hisbeya nacion (February), post luberation Aptu), breeding season (August) and pre hibernation (November) Maximum amount of sudan positive lipids and cholesterol was observed ditriang February and minimum during April In August and Novcmber there was a gradual rise from April The unsaturated lipids and plasttulogen also reached thew miximen durung February and the minimum hiving been attained during April in most of the species the adrenal cortex was found contam a hugh amount of cholesterol (Cowme and flagce 1934) Injection of A CTH in rats and guinea pigs lowered both accurbic sad and cholesterol (Sayers o al 1946) in toad, howeser we failed to detect any hister chemally demonstrablc ascorbic and in adrenal gland Recendv Eltın Larrou and Zarrow (1959) have obser cd chenucally a very low value of ascorbic acid is oud's adrenal compared to other species so estimauon of ascorbic acid was not included in the presene investigation. Sayers and his ci-workers (1990) have observed an mamate relationshup between the density of sud unophilic material and the concentracion of cholesterol in adrenals He has also corrclared distribution of ficutral lipids in adrenals with its secretory activity and have concluded that in general lipud depiction denoted hyperactivity and lipid accumulation hypo rctivity of the gland Our findings of wstreased uudenophulua and cholesterol during November and February thus suggested hypostaty of the gland, an increased activity as observed by reduced sodan porsitase substance have been observed durmig April-Auguse Marimuni amount of plasms has beer, herved durang February and minimum during Apret Albert and Leblond (1910) and alw fcmp, Ky and Wisłocka (1944) have suggested that plasmal reaction like phenylhydrazine could be used for detection of ketostetards Accordingly maginum accumulan nf ketosteroids In adrenals during February indiczted a non functional accumulation of 1 Our obserzbor of hypofunction of adrenals during hisbernation was in confirmatory sath the in golden hamsters deemg hubernation Experiments are in progress to obserse of tres Hypofunction is due to lovered A CT production Effect of acure dehydration The effectof stress produced by acuie in 2!17 deprivason fix haurs on touf's adrenals were studied by Deb and Mukhery(1959) Tarbsense the effect of kaxonal vartanon on adrenal response to stress the experiment was carried out durung and cholesterol were detected huseochemically in the nonhibranting towad's adrenal there was a depletion of all the four hustach mcally demonstrable substance compared to normal ones Toads were also subjected to similar stress dening bubertudom u bien chase ·128 C DER AND M MUKHERJI. HISTOCIIEMICAL STUDY OF was no alteration m the lipids Sayers and his co-workers (1944) have observed that when there was an increased demand for adrenal cortical hormonc such as stress, there was a decrease in the cholesterol concent adrenals Our findings of the fall in lipids including cholesterol might be due to similar reasons It has also been observed that in the absence of anterior pituitary the adrenal cholesterol did not change to stress (Patt et al 1948, Sayers et al 1945) Duc possibly to blockade of pituitary adrenal axis in hibernating toad. no alteration in the lipids were observed by us .. the cytoplismuc inclusions which were further depleted after 48 hours. Long (1947) longed exposure has not yet been studied Elcon, Zarrow and Zarrow (1959) have failed to observe any alteration in the cholesterol content of toad's adrenal on exposure to cold fonsoC) for 1-4 hours The authors, however, have not maintained if they have used hibernating or non hbernaung road le can be concluded that tord's adrenal also respond to different stress condtions, but anly during non hubcrnatng period. ACKNOWLEDGEMENT Thanks arc due to Prof P Sen, Professor of Play siology, for his dund suggestsons and encouragement REFERENCES Albert S, and Leblond C P (1940) Distribution of Fulgen and 2,4-dunutrophenylhydrazınc react in normal castrated, adrenalectomized and hormonally traded sats Endocrinology, 39, 386-460 Cowx, D M and Mage, M C. (1934) Lipoids and lipid descases Ill Schuller Christian's disease (Xanthomatosss) and review of Isterature on lipod content of human tissues. Atch unter and 53. 391-399 Deb, c, and Mukherji, M (1989) Unpublished Explomation of Plate I'II FIG 1 Adrenal, cholesterol, after 48 hours' dehydration, during August, Compare with Fig S x 91 ric 2 Adrenal, neutral lipid duttig February Compare with Fig 3 x 96 Fic 3 Adrenal, neutral lipid, during April, Compare with Fig 2 x Dis FK 4 Adrenal cholesterol durmg February Compate with Fig s ro FC Compare with Fig 4 x 95 Fic Compare with rig 706 FIC mpare with Fig 6 x 96 Fic Adrenas, ncuitas uplu WILCI 48 wouls dehydration during April Compare with Fig 1 x 90 Fic 9 Adrenal, cholesterol, after "Cold shock for 3 hours", during August Compare * Figs X 90 FIG 10 Adrenal, cholesterol, after "Coid shock for 48 hours", during August Compare with Fig 5 * 96. . 5

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·129 ADRENAL CORTICAL ACTIVITY IN TOAD (BUFO MELANOSTICTUS) Dempsey, E. W and Wislockı, G B (1944) Observation on some bastochemical scactions in the lumi placenta, with some special reference to the significance of the lipoids, glycogen and in Endocrinology 35, 401-429 Elton, R L Zarrow. I G and Zarrow, M X (1959) Depletion of adrenal ascorbic and ad choler cerol a comparative study Ibid., 65, 152-157 Long CNH (1947) Recent progress in hormone research Vol I Lyman C. P. and Chatfield, PO (1955) Physiology of hubemacion in inammals Physiol Rev., JS, 403-425 rel rat Saence, 108, 475-476 Sayers G. Sayers MA, Fry, E G, White, A, and Long CNH (1914) Effect of adrenocortico tropluc hormone of anterior pituitary on cholesterol content of adrenals-adrenal cholesterol Yale J Biol Mied. 16 361-392 Sayers, G, Sayers, M. A, Liang T. Y., and Long CN (1945) Cholesterol and ascorbic acid content of adrenal, liver, brain and plasma following hemontage Endominology. 37.9-110. (1946) Effect of pituitary adrenatropluc hormone on cholesterol and ascorbic and content of adrenal of it and guinea pig Ibid, 38, 1-2 - (1950) Thu adrenal cortex Physiol Rev. 30. 244-320 ·5 INVESTIGATIONS ON AUTOTRANSPLANTATION OF THE ADRENAL GLAND IN THE SPLEEN WITH SPECIAL REFERENCE TO PITUITARY- ADRENAL-RELATIONSHIPS by AS, KRISTINA MURTILY and M B. SAHASRABUDIIE Indian Cancer Rescarch Centre and Biology Division, Atomic Energy Establishment, Trombay In connection with a research project under the auspices of the I. C. M. R. for the "Development of a Histochemical Method for the Localisation of Adrenocorticotropic IAAT 11 + 1 cuinaine 1 probably get detoxicated and made physiologically inactive. This technique of autotransplantation of the adrenal into spleen has been employed in the present investigation to study its response on pituitary histology at different intervals of time. In the present investigation Wistar rats of both the sexes and about eight to ten weeks of age were used Both the adrenals were removed under aether anaesthesia, and piece of adrenal assue introduced in the spleen with a trocar The animals were maintained on isotonic salınc for about eight to ten days after operation Saline was later withdrawn and the annals were reverted on water Mortality rate was high due to operative trauma The operated animals were divided into four groups. The first, second, third and a 7 The changes in the graft and pituitary gland were as follows. control animals -- fones the foregoing, that autotransplantation of adrenal into spleen pituitary gland Basophils are supposed to tropic hormones (TSH), gonadotropic hormones (GH) and ACTH In the present investigation, however, it appears that cells asso Mated with the production of ACTH were increased, as evidenced by the probferation ·CRISTINA MONTITY & SAHASRADUDIE TYYESTIGATIONS ON AUTOTRANSPLANTATION 131 of adrenal graft on the spleen TSH OF GH however were not appreasbly altered as the thyroid gland and gonads did not show any proliferaton ·DISCUSSION Kar If I remember correctly Seligman and Rutenburg found that there was a difference in starnung intensity with acrobic is anaerobic incubation De De Souza, what type of incubation did you use in your studies > De Souza It is correct that a difference in stammg intensity under aerobic and anacrobic conditions was found in carly studies where tetrazolium salts were mostly used We used acrobic 11- cubation with nitro B T - a fast electron acceptor To my mund the difference is not very marked as the cytochrome system cannot compete as efficiently with nitro B T for the electron acceptance 1. L LE 11.' L. action at the cellular level The recent attempts by workers like Glick to make these techniques more refined and quantitative are provung to be extremely valuable Kar Singh Prasad Singh Malaviya Dr Sagh, your experiments are well planned and your results are interesting But, I feel that it is a little too much to draw any resemblance with gastric ulcer as found in human 1 ·LIST OF THOSE PARTICIPATING IN OR ATTENDING THE SYMPOSIUVI ON "THIYRO-GONAD-ADRENAL-PITUITARY RELATIONSHIPS-NEU DELHI OCTOBER 2 3 & 4 1959 A Ahuja X N Bagchi S Banerjee UK Danik gital Medicure Calcutta B Chowdhury Defence Science Organisation Dcl? RR Chowdhury All India Institute of Medical Sciences, New Delhi D Das Gupta 19 Panditya Road Calcutta PR Das Gupta Central Drug Research Institute Lucknow DB Duxit All India Institute of Medical Sacnces New Delhi E De Souza Tofiwala Nanonal Medical College Bomhay A B Kar AS Krishna Murthy BK Malaviya B Mukerji SR Mukherjee MRN Prasad University of LCL Leu L'al Indian Veterinary Research Institute Izaugar MB Sahasrabudhe Acomic Energy Research Establishment Bombas P N Shah Indian Cancer Research Centre Bombay GB Singh Central Drug Research Institute Lucknow V Subrahmanan Central Food Technological Research Institute Myxom VN Talawar All India Institute of Moulcal Saences, Nen Dchhi

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