Effects of Superior Cervical Ganglionectomy and Melatonin Replacement on Intra-hypothalamic LHRH Content and Pulsatile Luteinizing Hormone Release in the Mink

Long-term effects of subcutaneous melatonin implants on intrahypothalamic LHRH content and on pulsatile luteinizing hormone release have been investigated in ganglionectomized male mink. Animals were submitted to bilateral removal of the superior cervical ganglion in mid-April. A preliminary study revealed that plasma LH concentrations remain at a basal level throughout the year following ganglionectomy. In a second experiment, one month after ganglionectomy and transfer from the natural photoperiod environment to short daylengths (LD 4:20), melatonin pellets were subcutaneously implanted to overcome deafferentation of the pineal. Progressive effects of treatment were studied 7 days, 15 days, and one, two and three months after insertion of the melatonin implants. The intra-hypothalamic LHRH content in ganglionectomized mink was at a basal level similar to that observed during seasonally sexual quiescence, or after exposure to inhibitory long days (LD 20:4). A significant and transient elevation in LHRH content was observed already after fifteen days, and also one month after insertion of melatonin implants. This resulted in mean values similar to those observed during the breeding season, or after exposure to stimulatory short days (LD 4:20). A decrease in hypothalamic LHRH content started after two months. No pattern of pulsatile LH secretion was recorded in ganglionectomized untreated mink. A significant increase in all parameters of pulsatile LH secretion was observed fifteen days after the elevation of LHRH content induced by melatonin treatment, and maximum values were reached after two months. Pituitary activity tended to decrease after three months, characterized in particular by a significant decrease in the mean frequency of LH pulses. In addition, the increase in pulsatile characteristics of LH release occurred two months before the peripheral renewal of testicular activity. Apparently, the reproductive endocrine function in ganglionectomized mink treated with melatonin implants is restored more rapidly at the hypothalamic level than at the pituitary or testicular levels.     

Alterations in hypothalamic content of luteinizing hormone-releasing hormone associated with pineal-mediated testicular regression in the golden hamster

The adult male golden hamster will undergo testicular regression when exposed to a short photoperiod, blinding, or late afternoon injections of melatonin. The present study was conducted to compare the effects of all three treatments on serum gonadotropin levels and testicular weights, and to evaluate the effects of these treatments on hypothalamic content of both immunoreactive and bioactive luteinizing hormone-releasing hormone (LHRH) levels. Hamsters were blinded (BL), exposed to a short photoperiod (SP), or received daily injections of melatonin (MEL) for 15 wk. Each treatment (BL, SP, MEL) induced a temporally similar decline in serum luteinizing hormone (LH), serum follicle-stimulating hormone (FSH), and testicular weight. Spontaneous recrudescence occurred earliest in the MEL group, with serum gonadotropins and testicular weight returning to normal by 15 wk. The SP group exhibited recovery of serum gonadotropins but not testicular weight by 15 wk. The BL group demonstrated partial recovery of serum FSH levels by 15 wk, with no recovery in either serum LH or testicular weight. Each treatment group demonstrated increased hypothalamic content of immunoreactive LHRH which was temporally correlated with the decreases of serum gonadotropins. Additionally, the MEL and SP groups demonstrated decreased immunoreactive LHRH levels during spontaneous recrudescence. Extracts of hypothalami from all treatment groups were bioactive on control hamster pituitary cells. These results indicate that there are temporal differences among the three common treatments and that these differences are manifested in serum gonadotropins, testicular weight and hypothalamic LHRH. Hypothalamic LHRH levels determined by radioimmunoassay and bioassay show periods of increase and decrease which coincide with periods of altered serum gonadotropin levels in all groups.     

Pituitary response to LHRH, LH pulsatility and plasma melatonin and prolactin changes in ewe lambs treated with melatonin implants to delay puberty

Prepubertal ewe lambs were treated with empty or filled melatonin implants. The implants were placed s.c. at birth and pituitary responsiveness to various doses of LHRH, LH/FSH pulsatility and prolactin and melatonin secretion were examined at 10, 19, 28, 36 and 45 weeks of age. Control animals (N = 10) showed no consistent alteration in pituitary responsiveness to LHRH during development. Ewes treated with melatonin (N = 10) had puberty onset delayed by 4 weeks (P less than 0.03) but no effect of melatonin on LH or FSH response to LHRH injection was observed at any stage of development. In the control and melatonin-treated ewe lambs the responses to LHRH injection were lower during darkness than during the day at all stages of development. No consistent differences in LH or FSH pulsatility were observed between treatment groups or during development. Prolactin concentrations, however, failed to decrease at the time of puberty (autumn) in the melatonin-treated group. Melatonin-treated ewe lambs maintained normal rhythmic melatonin production which was superimposed on a higher basal concentration and showed the same increase in melatonin output with age as the control ewes. These results indicate that the delayed puberty caused by melatonin implants is not due to decreased pituitary responsiveness to LHRH or to dramatic changes in basal LH or FSH secretion.     

Effect of melatonin implants on secretion of luteinizing hormone in intact and castrated rams

Rams were treated with melatonin implants in 2 experiments designed to examine the control of reproductive seasonality. In Exp. 1, rams (n = 12) were allocated to 3 treatment groups: 2 groups were treated with 2 melatonin implants per ram for 4 months from 11 November (N) and 9 December (D) and the remaining group was untreated (C). The seasonal increase in luteinizing hormone (LH) pulse frequency and testes size was advanced in Groups N and D. A second seasonal cycle in LH secretion and testes size occurred in Groups N and D after melatonin implants became exhausted. In Exp. 2, rams (n = 20) were allocated to 4 treatment groups: 10 rams were castrated on 6 October and 1 group of entire rams (EM) and one group of castrated rams (CM) were treated with 2 melatonin implants per ram each month from 3 November until 8 January. The other group of entire rams (EC) and castrated rams (CC) was untreated. An increase in LH pulse frequency occurred after castration. Melatonin treatment increased LH pulse frequency in entire rams and reduced LH pulse frequency in castrated rams. The results demonstrated that the advanced reproductive development as a result of treatment with melatonin implants was due to an effect of melatonin on the hypothalamic pulse generator to increase LH pulse frequency. The ability of melatonin to influence LH pulse frequency in entire and castrated rams indicated that an effect of melatonin on the hypothalamic pulse generator is independent of testicular steroids.     

Neuroendocrine regulation of pulsatile luteinizing hormone secretion in elderly men

Leydig cell function is driven by LH, secreted in a pulsatile manner by the anterior pituitary in response to episodic discharge of hypothalamic LHRH into the pituitary portal circulation, under control of a yet to be defined neural mechanism, the "hypothalamic LHRH pulse generator". The normal aging process in elderly men is accompanied by a decline in Leydig cell function. Whereas primary testicular factors undoubtedly play an important role in the decrease of circulating (free) testosterone levels with age, recent studies demonstrated that aging also affects the central compartment of the neuroendocrine cascade. Hypothalamic alterations comprise changes in the regulation of the frequency of the LHRH pulse generator with an inappropriately low frequency relative to the prevailing androgen impregnation and opioid tone, and with an increased sensitivity to retardation of the LHRH pulse generator by androgens. As observed by some authors in basal conditions and by others after endocrine manipulations. LH pulse amplitude seems also to be reduced in elderly men as compared to young subjects. This is most probably the consequence of a reduction in the amount of LHRH released by the hypothalamus. Indeed, challenge of the gonadotropes with low, close to physiological doses of LHRH in young and elderly men reveals no alterations in pituitary responsiveness when looking at either the response for immunoreactive LH or bioactive LH. Deconvolution analysis on data obtained after low-dose LHRH suggests a markedly prolonged plasma half-life of LH in elderly men, a finding which may explain the paradoxical increase of mean LH levels in face of the reduced or unchanged frequency and amplitude of LH pulses.     

Stress induced changes in testis function

The mechanism through which chronic stress inhibits the hypothalamic-pituitary-testicular axis has been investigated. Chronic restraint stress decreases testosterone secretion, an effect that is associated with a decrease in plasma gonadotropin levels. In chronically stressed rats there was a decrease in hypothalamic luteinizing hormone-releasing hormone (LHRH) content and the response on plasma gonadotropins to LHRH administration was enhanced. Thus the inhibitory effect of chronic stress on plasma LH and FSH levels seems not to be due to a reduction in pituitary responsiveness to LHRH, but rather to a modification in LHRH secretion. It has been suggested that beta-endorphin might interfere with hypothalamic LHRH secretion during stress. Chronic immobilization did not modify hypothalamic beta-endorphin, while an increase in pituitary beta-endorphin secretion was observed. Since we cannot exclude that changes in beta-endorphin secreted by the pituitary or other opioids may play some role in the stress-induced decrease in LHRH secretion, the effect of naltrexone administration on plasma gonadotropin was studied in chronically stressed rats. Naltrexone treatment did not modify the decrease in plasma concentrations of LH or FSH. These findings suggest that the inhibitory effect of restraint on the testicular axis is exerted at hypothalamic level by some mechanism other than opioids.     

Studies on the development of growth hormone and prolactin cells in the rat pituitary gland by in situ hybridization

Summary An antiserum raised against N-amino-3-propyl melatonin bound to a protein carrier was used to visualize melatonin by immunohistochemistry and to measure melatonin concentration by radioimmunoassay in the pineal gland of intact mink females killed throughout the 24 h cycle and females killed after a bilateral ablation of the cervical superior ganglion. Melatonin immunoreactivity revealed by immunofluorescence or by the peroxidase-antiperoxidase complex was observed in the cytoplasm of presumed pinealocytes of all the females. Circadian changes in pineal melatonin content were not visualized by immunohistochemistry; furthermore, immunoreactivity was also present in the pineal gland of the ganglionectomized females. However, the melatonin content measured by radioimmunoassay was significantly higher in the pineal gland from intact females killed during the night compared with that of intact females killed during the day or of ganglionectomized females. The discrepancy between the results obtained using the two methods may arise because immunohistochemistry can detect very small amounts of melatonin.     

Effects of intermittent pulsatile infusion of luteinizing hormone-releasing hormone on dihydrotestosterone-suppressed gonadotropin secretion in castrate rams

The feedback effects of dihydrotestosterone (DHT) on gonadotropin secretion in rams were investigated using DHT-implanted castrate rams (wethers) infused with intermittent pulsatile luteinizing hormone-releasing hormone (LHRH) for 14 days. Castration, as anticipated, reduced both serum testosterone and DHT but elevated serum LH and follicle-stimulating hormone (FSH). Dihydrotestosterone implants raised serum DHT in wethers to intact ram levels and blocked the LH and FSH response to castration. The secretory profile of these individuals failed to show an endogenous LH pulse during any of the scheduled blood sampling periods, but a small LH pulse was observed following a 5-ng/kg LHRH challenge injection. Dihydrotestosterone-implanted wethers given repeated LHRH injections beginning at the time of castration increased serum FSH and yielded LH pulses that were temporally coupled to exogenous LHRH administration. While the frequency of these secretory episodes was comparable to that observed for castrates, amplitudes of the induced LH pulses were blunted relative to those observed for similarly infused, testosterone-implanted castrates. Dihydrotestosterone was also shown to inhibit LH and FSH secretion and serum testosterone concentrations in intact rams. In summary, it appears that DHT may normally participate in feedback regulation of LH and FSH secretion in rams. These data suggest androgen feedback is regulated by deceleration of the hypothalamic LHRH pulse generator and direct actions at the level of the adenohypophysis.     

Changes in the testicular binding of luteinizing hormone and plasma testosterone concentrations in the Djungarian hamster subjected to different photoperiods and temperatures and effects of long-term testosterone treatment on the binding

The effects of artificial photoperiod, temperature, and long-term testosterone treatment on testicular luteinizing hormone (LH) binding were studied in adult male Djungarian hamsters. In hamsters transferred to long-day (LD; 16 hr light, 8 hr dark) photoperiod 8 weeks after adaptation in short-day (SD; 8 hr light, 16 hr dark) photoperiod of 25 degrees C, testicular growth was associated with an increase in the total LH binding per two testes and a decrease in LH binding per unit testicular weight. Plasma testosterone levels reached a peak 47 days after transfer to LD and tended to decrease thereafter, while the testes continued growing. In contrast, when hamsters reared under LD conditions at 25 degrees C for 12 weeks were transferred to SD, testicular regression was associated with a decrease in plasma testosterone and the total LH binding per two testes and an increase in LH binding per unit testicular weight. A significant decrease in LH binding per unit weight compared to SD controls was observed in those hamsters exposed to SD with continuous testosterone treatment. The testosterone treatment tended to induce decrease in the total LH binding. Scatchard plot analyses of the binding suggested that changes in LH binding were due to changes in the number of binding sites. When sexually mature male hamsters were subjected for 8 weeks to two different ambient temperatures (7 degrees C and 25 degrees C) and photoperiods (LD and SD), the difference between the two temperature groups was statistically not significant regarding the weights of testes, epididymides, and prostates; plasma testosterone levels; and LH binding in either LD or SD group. These results suggest that photoperiod is a more important environmental factor than temperature for the regulation of testicular activity and LH receptors and that testosterone reduces the number of LH receptors per unit testicular weight in adult male Djungarian hamsters.     

Continuous light after 2 months of long days stimulates ram testis volume and increases fertility in spring

Seasonal reproduction is one of the major biotechnical and economic constraints of sheep production in temperate latitudes. Treatments using extra light followed by melatonin implants have been used satisfactorily in open barns, farms and artificial insemination centres to produce out-of-season sexual activity in rams. The aim of the present study is to explore the possibility of replacing melatonin implants with continuous light (LL), which was recently shown to increase LH secretion similar to melatonin and/or pinealectomy. Four experiments during 4 consecutive years were conducted in ‘Ile-de-France’ rams. In each study, one group was systematically exposed to permanent light after a first photoperiodic treatment of 60 long days (LD-LL) during the winter and compared with various other control groups subjected either to a natural photoperiod or the classical LD-melatonin treatment. As expected, blood nocturnal melatonin secretion was suppressed by LL. In all four experiments, LL treatment produced a highly significant and robust increase in ram testicular volume in the spring compared with the testicular volume of control rams or of that of treated rams at the end of the LD. For the two experiments in which fertility was tested, fertility after hand-mating was significantly higher in LD-LL rams than in control rams (76% v. 64%). Therefore, permanent light after an LD treatment may be an interesting alternative to LD-melatonin treatment to induce out-of-season sexual activity in rams.     

Comparative effects of LHRH agonist and ovine LH administration on testicular steroidogenesis in intact adult rat

In order to better understand the effects of LHRH administration on testicular function in adult rat, we compared the inhibitory effects of LH and the LHRH analogue [D-Ser-(TBU)⁶, des-Gly-NH₂ ¹⁰]LHRH ethylamide upon testicular steroidogenesis and LH, FSH and prolactin receptor contents. Administration of LH as well as LHRH analogue resulted in a marked decrease of LH receptor levels, accompanied by a blockage at the level of 17-hydroxylase activity. We have been able to demonstrate that multiple LH administration can achieve a testicular desensitization comparable to that observed after LHRH agonist treatment.     

Effect of time after castration on secretion of LHRH and LH in the ram

Hypophysial portal blood and peripheral blood were obtained from conscious, unrestrained rams to measure simultaneously the secretion of LHRH and LH in entire rams and rams which had been castrated for 2-15 days (short-term castration) and for 1-6 months (long-term castration). The apparatus for portal blood collection was surgically implanted using a transnasal trans-sphenoidal approach and, 4-5 days later, portal blood and peripheral blood were collected simultaneously at 10-min intervals for 8-9 h from 15 sheep. LHRH was clearly secreted in pulses in all three physiological conditions, but there were marked differences in pulse frequencies, which averaged 1 pulse/2-4 h in entire rams, 1 pulse/70 min in short-term castrated rams and 1 pulse/36 min in long-term castrated rams. In entire and short-term castrated animals, LH profiles were also clearly pulsatile and each LHRH pulse in hypophysial portal blood was associated with an LH pulse in the peripheral blood. In long-term castrated animals, LH pulses were not as well defined, because of the high basal levels and small pulse amplitudes, and the temporal relationship between LHRH and LH pulses was not always clear. These results demonstrate the pulsatile nature of LHRH secretion under the three physiological conditions and suggest that the irregular LH profiles characteristic of long-term castrates are due to an inability of the pituitary gland to transduce accurately the hypothalamic signal. The very high frequency of the LHRH pulses may be one of the major reasons for this, and is probably also responsible for the high rate of LH secretion in the long-term castrated animal.     

The different mechanisms for suppression of pituitary and testicular function

The differential mechanisms reducing androgen secretion by LHRH agonists are discussed with relevance to clinical therapy. LH secretion can be desensitised by exposure to agonists using high doses, frequent injections or sustained release/constant infusion. The desensitized pituitary is refractory to hypothalamic stimulation. Pituitary receptor suppression is associated with depletion of pituitary gonadotrophin content, and a decline of LH and FSH secretion to a basal rate. Recovery of LH responsiveness to endogenous LHRH stimulation requires restitution of gonadotrophin content (about 7 days in rats). After long-term infusions in normal men, testosterone secretion recovers within 7-10 days. The binding capacity of testicular LH/hCG receptors is reduced in rats after supraphysiological gonadotrophin stimulation, by agonists or directly by hCG, concomitantly the steroidogenic capacity of the testis in vitro is impaired. Qualitative changes in androgen biosynthesis are a marked fall in testosterone production and dose-dependent enhancement of progesterone production. After 12 months of buserelin injections, the changes in hCG-stimulated rat testes are an increased ratio of progesterone/17-OH-progesterone (inhibition of 17-hydroxylase), a reduced capacity for secretion of androstenedione and testosterone (block of 17,20-desmolase), and increased 5 alpha-pregnane-3,20-dione (this steroid inhibits the 17,20-desmolase, similarly to progesterone). After treatment, Leydig cell function recovers completely. Leydig cell hyperplasia is observed as a result of the steroidogenic changes. These findings in rats have not been observed in dogs, monkeys or in humans.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pituitary and gonadal function in hypogonadotrophic hypogonadal (hpg) mice bearing hypothalamic implants

GnRH receptor values are 30-50% of normal in pituitaries of hpg male mice, and testicular LH receptors only 8% of normal (160.4 +/- 17.6 and 2013 +/- 208.1 fmol/testis respectively). In male hpg mice bearing fetal preoptic area (POA) hypothalamic implants for 10 days there was no change in pituitary GnRH receptors, pituitary gonadotrophin content, or seminal vesicle weight. However, testicular weights and LH receptors were doubled in 4/10 mice and 2 had increased serum FSH levels. Between 26 and 40 days after implantation pituitary GnRH receptors and pituitary LH increased to normal male levels, although at 40 days serum and pituitary FSH concentrations had reached only 50% of normal values. Testicular and seminal vesicle weights increased more than 10-fold by 40 days after implantation and LH receptors to 70% of normal. In hpg female mice bearing hypothalamic implants for 30-256 days pituitary gonadotrophin concentrations were normal, even though GnRH receptors reached only 60% of normal values (6.18 +/- 0.4 and 9.8 +/- 0.4 fmol/pituitary respectively). Serum FSH was substantially increased from values of less than 30 ng/ml in hpg mice to within the normal female range in hypothalamic implant recipients. Ovarian and uterine weights increased after hypothalamic grafting from only 4-5% to over 74% of normal values. LH receptors increased from 6.5 +/- 1.3 fmol/ovary for hpg mice to 566.9 +/- 39.2 fmol/ovary for implant recipients. Vaginal opening occurred about 23 days after implantation and these animals displayed prolonged periods of oestrus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Correlation of hypothalamic LHRH, pituitary LH and plasma LH in developing rats.

Hypothalamic LHRH, pituitary LH and plasma LH levels were measured in rats of both sexes from day 5-60 after birth. The content of hypothalamic LHRH was very high in one-week-old male and female rats. It declined gradually till day 17 in the female rat and sharply on day 10 in the male rat. Subsequently the content of hypothalamic LHRH increased and showed peak values on day 25 in the female rat and on day 45 in the male rat. It decreased markedly at respective times of puberty in both sexes (day 37 in the female rat and day 52-60 in the male rat). Results of the study suggest that maturation of hypothalamo-hypophyseal-axis proceeds in three distinct stages. Observations on days 17, 25 and 37 in the female rat and on days 5, 7, 10 and 22 in the male rat clearly show an inverse relationship between hypothalamic LHRH and plasma LH and a parallel relationship between pituitary and plasma LH. Marked decline in the content of hypothalamic LHRH at respective times of puberty in both sexes indicates that the release of threshold levels of LHRH from the hypothalamus may apparently be the event initiating the pubertal changes in rat.     

Blockade of pulsatile LH, FSH and testosterone secretion in rams by constant infusion of an LHRH agonist

Adult Soay rams were infused for 21 days with 50 micrograms buserelin/day, using s.c. implanted osmotic mini-pumps. The continuous treatment with this LHRH agonist induced a supraphysiological increase in the blood concentrations of LH (15-fold) and testosterone (5-fold) followed by a decrease below pre-treatment values after 10 days. The blood concentrations of FSH showed only a minimal initial increase but the subsequent decrease was dramatic, occurring within 1 day. By Day 10 of treatment, the blood concentrations of all 3 hormones were low or declining, LH pulses were absent in the serial profiles based on 20-min blood samples and the administration of LHRH antiserum failed to affect the secretion of LH or testosterone. By Day 21, the secretion of FSH, LH and testosterone was maximally suppressed. The i.v. injection of 400 ng LHRH was totally ineffective at stimulating an increase in the blood concentrations of LH while the i.v. injection of 50 micrograms ovine LH induced a normal increase in the concentrations of testosterone; this confirmed that the chronic treatment with the LHRH agonist had desensitized the pituitary gonadotrophs without markedly affecting the responsiveness of the testicular Leydig cells. The ratio of bioactive: radioimmunoactive LH did not change during the treatment. The long-term effect of the infusion was fully reversible as shown by the increase in the blood concentrations of FSH, LH and testosterone and the return of normal pulsatile fluctuations in LH and testosterone within 7 days of the end of treatment.     

Induction of early rutting in male red deer (Cervus elaphus) by melatonin and its dependence on LHRH

Eight red deer stags, 2 control, 3 control-immunized (i.e. a low titre of LHRH antibodies after active immunization 2 years earlier) and 3 superior cervical ganglionectomized, were given a s.c. implant of melatonin in May at the nadir of the sexual cycle; 5 other stags remained untreated. All the melatonin-treated animals shed the velvet-like skin from the antlers in June or July at least 1 month before the untreated controls, and had an early increase in blood plasma testosterone concentrations. The treated stags were also precocious in the development of rutting behaviour, although this inductive effect was blocked dramatically in the control-immunized stags after a booster immunization against LHRH; these animals failed to show any further reproductive development and cast their antlers. It is concluded that continuous exposure to melatonin in early summer will induce premature seasonal testicular development, an effect dependent on the secretion of LHRH, and similar to that produced by exposure to short daylengths.     

Effect of protein deficiency on some hypothalamic and pituitary hormones in growing male lambs. An immunohistochemical study

Growing male lambs were fed with diets containing 14.0, 10.8 and 7.6% protein for 3 months to determine their effects on the content of hypothalamic LHRH and SRIH and pituitary LH and GH, using immunohistochemical methods. Lowering the concentration of dietary proteins caused marked changes in the immunoreactivity of these hormones. The immunoreactive (IR) content of LHRH stored in the median eminence was enhanced, and the proportion of LH cells and their IR content increased. Opposite effects were observed in the SRIH/GH system; IR SRIH content stored in the median eminence markedly diminished, and, although hyperplasia of GH cells was observed, their IR content was diminished. This study indicates that prolonged restrictions of protein in the diet of growing male sheep affects the immunoreactive content of the investigated hormones. It seems that they suppress LHRH/LH release and augment GH release, possibly via suppression of hypothalamic somatostatin.     

Effects of intrahypothalamic testosterone implants on LHRH levels in the preoptic area and the medial basal hypothalamus.

Following castration LHRH levels in the MBH but not in the POA decreased. Testosterone implants in the medial POA following castration failed to alter the LHRH activity either locally in the POA or in remote sites in the MBH. On the contrary, similar T implants in the MBH blocked castration-induced depletion of MBH LHRH stores without affecting either the POA LHRH content or the post-castration hypersecretion of pituitary LH. These findings identify the MBH as the focal site of T action in the regulation of hypothalamic LHRH activity.     

Effect of neonatal castration on the content of hypothalamic LHRH, pituitary LH and plasma LH in developing male rats.

The content of hypothalamic LHRH and concentration of LH in pituitary and plasma were measured on day 5, 7, 10, 14, 17, 22, 25, 30, 45, 52 and 60 in male rats which were bilaterally castrated on day 2. The levels of plasma LH were significantly higher in all the groups of castrated rats than in normal male rats of corresponding ages. The concentration of plasma LH did not rise progressively but showed day to day fluctuation apparently due to alteration of sexual differentiation of the hypothalamus. The concentration of pituitary LH was significantly lower in neonatally castrated rats compared to normal male rats except on days 17, 25 and 30. The content of hypothalamic LHRH declined initially following castration, but from day 17 onwards significantly higher levels of hypothalamic LHRH were maintained in neonatally castrated rats than in intact control. Initial decline in the content of hypothalamic LHRH may be because of stimulation of release of LHRH which exceeds maximal rate of synthesis and subsequent increase in the content of hypothalamic LHRH may be due to enhanced LHRH synthesis as a result of castration.