Sertoli cell differentiation in rhesus monkey (Macaca mulatta) is an early event in puberty and precedes attainment of the adult complement of undifferentiated spermatogonia.

In primates, the time course of Sertoli cell proliferation and differentiation during puberty and its relationship with the expansion of undifferentiated type A spermatogonia that occurs at this critical stage of development are poorly defined. Mid and late juvenile and early and late pubertal male rhesus monkeys were studied. Testes were immersion fixed, embedded in paraffin, and sectioned at 5 µm. Sertoli cell number per testis, S-phase labeling (BrdU), and growth fraction (Ki67 labeling) were determined and correlated with corresponding parameters for undifferentiated type A spermatogonia (A dark and A pale). Dual fluorescence labeling was used in addition to histochemistry to monitor spermatogonial differentiation during the peripubertal period using GFRα-1 and cKIT as markers. While the adult complement of Sertoli cells/testis was attained in early pubertal monkeys after only a few weeks of exposure to the elevated gonadotropin secretion characteristic of this developmental stage, the number of undifferentiated type A spermatogonia several months later in mid pubertal monkeys was only 50% of that in adult testes. Both A dark and A pale spermatogonia exhibited high S-phase BrdU labeling at all stages of juvenile and pubertal development. Spermatogonial differentiation, as reflected histochemically and by relative changes in GFRα-1 and cKIT expression, was not observed until after the initiation of puberty. In the rhesus monkey and maybe in other higher primates including human, the pubertal proliferation of undifferentiated spermatogonia is insidious and proceeds in the wake of a surge in Sertoli cell proliferation following termination of the juvenile stage of development.

Adrenal androgen concentrations increase during infancy in male rhesus macaques (Macaca mulatta).

This study investigated adrenal androgens (AA), gonadotropins, and cortisol in castrated and gonad-intact male rhesus macaques from birth through infancy. Blood samples were collected longitudinally from castrated (n = 6; weekly, 1-40 wk) and intact (n = 4; every other week, 1-17 wk) males. Plasma concentrations of AA were determined by liquid chromatography-tandem mass spectrometry, and plasma concentrations of cortisol and gonadotropins were determined by RIA. Dehydroepiandrosterone sulfate (DHEAS) concentrations increased almost threefold (to 8 wk), dehydroepiandrosterone (DHEA) increased more than eightfold (to 11 wk), and androstenedione doubled (to 15 wk) in five castrated infant males and declined continuously thereafter. A sixth castrated male had markedly different temporal patterns and concentrations (many times more than 2 SDs from the cohort mean) of AA and gonadotropins from first sampling (3 wk) and was excluded from analysis. Cortisol increased over 16 wk but correlated poorly with DHEAS. Luteinizing and follicle-stimulating hormones increased to peaks at 3 and 7 wk, respectively. Testis-intact males exhibited similar profiles, but with earlier peaks of DHEAS (5 wk) and DHEA and androstenedione (7 wk). Peak concentrations of DHEAS were lower and those of DHEA and androstenedione were higher in intact than castrated infants. Testosterone was undetectable in castrated males and >0.5 ng/ml in intact males but was not correlated with DHEA or DHEAS. These are the first data documenting a transient increase in AA secretion during infancy in an Old World primate and are consistent with the previously documented time course of zona reticularis development that accompanies increases in androgen synthetic capacity of the adrenal. The rhesus is a promising model for androgen secretion from the human adrenal cortex.

A re-examination of proliferation and differentiation of type A spermatogonia in the adult rhesus monkey (Macaca mulatta).

BACKGROUND: Companion studies using an experimental non-human primate paradigm known as a testicular clamp indicated that the behavior of undifferentiated type A spermatogonia did not conform fully to earlier classical models. This issue was therefore re-examined in normal monkeys. METHODS: Adult male rhesus monkeys (n = 4) received an i.v. bolus of 5-bromo-2'-deoxyuridine (BrdU): one testis (first) was removed 3 h later and the remaining testis (second) was removed after 11 days and 3 h. Tissue was fixed in Bouin's solution, and numbers of A dark (Ad), small A pale (Aps) and large A pale spermatogonia, differentiating B spermatogonia, S-phase-labeled and degenerating cells were enumerated. Data are given as mean +/- SEM. RESULTS: During the early stages of the seminiferous epithelial cycle in the first testis, Ap spermatogonia (1.3 cells/cross section) were predominantly Aps (nuclear dia., 7.1 +/- 0.1 microm). Aps were never S-phase labeled. Apl (nuclear dia., 8.8 +/- 0.5 microm) appeared in Stages IV-VI and were maximal in Stages VII-X when S-phase labeling of this phenotype at 3 h was greatest. The first generation of B spermatogonia appeared in Stages XI-XII (0.84 cells/cross section). Using cells/cross section, the ratio of Ap (Stages I-V):B1:B2:B3:B4:preleptotene spermatocyte was 1:0.7:1.4:2.8:5.6:11.2. In the second testis, labeled Aps (and Apl) were observed. Ad were not BrdU labeled, and degenerating cells were rarely observed. CONCLUSIONS: The results are not entirely consistent with earlier models of spermatogonial proliferation and differentiation in the monkey. Most notably, our findings suggest that in any one cycle of the seminiferous epithelium only a fraction of Ap spermatogonia is mitotically active.

A selective monotropic elevation of FSH, but not that of LH, amplifies the proliferation and differentiation of spermatogonia in the adult rhesus monkey (Macaca mulatta).

BACKGROUND: Unilateral orchidectomy in monkeys increases spermatogenesis in the remaining testis in association with elevated follicle-stimulating hormone (FSH) secretion and testicular testosterone. The present study examined the relative importance of FSH and testosterone in driving the primate testis toward its spermatogenic ceiling. METHODS: Adult male rhesus monkeys were treated with a gonadotropin-releasing hormone receptor antagonist to inhibit endogenous FSH and luteinizing hormone (LH) secretion. The gonadotrophin drive to the testis was replaced with a pulsatile recombinant human FSH and LH infusion to maintain testicular volume and circulating testosterone and inhibin B at physiological levels. A selective monotropic elevation of FSH or LH that doubled the concentrations of inhibin B or testosterone, respectively, was then imposed for 4 weeks, each in a group of four monkeys. In a third group (n = 4), the gonadotrophin drive remained clamped at physiological levels. Bromo-deoxyuridine was administered 3 h prior to castration, and the effects of the monotropic hormone increments on germ cell number, S-phase labeling and degeneration were determined. RESULTS: Increased FSH, but not LH, produced increases in testicular volume (P < 0.05), the proportion of A pale spermatogonia entering the cell cycle and the numbers of differentiated spermatogonia and more advanced germ cells (P < 0.05). Indexes for spermatogonia labeling and germ cell degeneration were not affected. CONCLUSIONS: Under physiological conditions, circulating concentrations of FSH directly dictate sperm output of the primate testis by regulating the proportion of Ap spermatogonia in the growth fraction. An effect of FSH on survival of the first generation of differentiated B spermatogonia is not excluded.

Control of the rhesus monkey menstrual cycle: permissive role of hypothalamic gonadotropin-releasing hormone.

In rhesus monkeys with hypothalamic lesions (which appear to abolish the endogenous production of gonadotropin-releasing hormone), normal ovulatory mestrual cycles were reestablished by an unvarying, long-term replacement regimen consisting of one intravenous pulse of synthetic gonadotropic-releasing hormone per hour. This finding is in accord with the hypothesis that the pattern of pituitary gonadotropin secretion throughout the menstrual cycle (basal secretion interrupted, once every 28 days on the average, by a preovulatory surge) is not directed by alterations in hypothalamic gonadotropin-releasing hormone secretion but by the ebb and flow of ovarian estrogens acting directly on the pituitary gland.

Hypophysial responses to continuous and intermittent delivery of hypopthalamic gonadotropin-releasing hormone.

In rhesus monkeys with hypothalamic lesions that abolish gonadotropic hormone release by the pituitary gland, the constant infusion of exogenous gonadotropin-releasing hormone (GnRH) fails to restore sustained gonadotropin secretion. In marked contrast, intermittent administration of the synthetic decapeptide once per hour, the physiological frequency of gonadotropin release in the monkeys, reestablishes pituitary gonadotropin secretion. This phenomenon is attributable to the pattern of GnRH delivery rather than to the amounts of this hormone to which the cells of the pituitary are exposed. Moreover, the initiation of continuous GnRH administration in animals with lesions and in which gonadotropin secretion is reestablished by intermittent GnRH replacement can result in a "desensitization" or "down regulation" of the processes responsible for gonadotropin release.

Gonadal regulation of hypothalamic gonadotropin-releasing hormone release in primates.

This review has examined, in primates, the action of sex steroids on the neural timing mechanism that governs the intermittent release of GnRH by the hypothalamus, the so-called GnRH pulse generator. Determinants of hypothalamic GnRH pulse generator frequency have generally been examined indirectly by monitoring moment to moment fluctuations in circulating LH concentrations. Studies using this approach have led to the hypothesis that negative feedback control of LH secretion by the testes is mediated by the action of T, or of one of its metabolites, to retard the frequency of the GnRH pulse generator. P also appears capable of decelerating GnRH pulse frequency during the luteal phase of the menstrual cycle, but the physiological significance of this phenomenon remains to be clarified. To date, a cognate action of E2 on the GnRH pulse generator has not been described. Because of limitations in contemporary technology, the factors underlying amplitude modulation of the GnRH pulse generator are less well understood. In addition to the ability of certain sex steroids to decelerate the frequency of pulsatile GnRH discharge, E2 and P appear capable of facilitating the secretion of this hypothalamic releasing factor. However, the increase in GnRH pulse frequency and/or GnRH pulse amplitude that must underlie these stimulatory actions remains to be fully defined. An inhibitory action of high levels of circulating cortisol on the hypothalamic GnRH pulse generator has also been noted.

The functional significance of FSH in spermatogenesis and the control of its secretion in male primates.

The aim of this review is to provide an integrative analysis of the role of FSH in the control of testicular function in higher primates, including man. Attention is focused on the action of FSH during neonatal development, puberty, and adulthood. Whether FSH is the major determinant of the adult complement of Sertoli cells and whether FSH is obligatory for the initiation, maintenance, and restoration of spermatogenesis is evaluated. The mechanism whereby the circulating concentration of FSH regulates spermatogonial proliferation to dictate the sperm production rate under physiological conditions in the adult is discussed in detail. Inhibin B is the major component of the testicular negative feedback signal governing FSH beta gene expression and FSH secretion, and the evidence for this view is presented. The review concludes with the presentation of a model for the operation of the FSH-inhibin B feedback control system regulating sperm production postpubertally in monkey and man, and with speculation on issues of clinical interest.

Evidence that down regulation of hypothalamic KiSS-1 expression is involved in the negative feedback action of testosterone to regulate luteinising hormone secretion in the adult male rhesus monkey (Macaca mulatta).

In the male monkey, luteinising hormone (LH) secretion is regulated by a negative feedback action of testicular testosterone that is exerted indirectly at the hypothalamic level to decelerate pulsatile gonadotrophin-releasing hormone release (GnRH). The purpose of the present experiment was to investigate whether the kisspeptin-G protein-coupled receptor 54 (GPR54) signalling pathway is involved in mediating the action of testosterone to suppress GnRH release in the monkey, as has been indicated by studies of nonprimates. To this end, 12 castrated adult male rhesus monkeys were implanted with either testosterone containing or empty Silastic capsules. Testosterone treatment produced a square wave increment in circulating testosterone levels within the physiologic range. After suppression of LH and follicle-stimulating hormone secretion was established at 5-6 weeks of testosterone exposure, the animals were killed and expression of the genes encoding for kisspeptin, GPR54 and GnRH determined in the mediobasal hypothalamus and preoptic area of both treated and control animals using RNase protection assays. The suppression in pituitary gonadotrophin secretion was associated with a reduction in kisspeptin mRNA levels in the mediobasal hypothalamus, but not the preoptic area. GPR54 mRNA levels, on the other hand, were not influenced by testosterone treatment. These results are consistent with those previously reported for the rodent, and suggest that the neurobiology of the negative feedback action of testicular testosterone on LH secretion in the monkey, a representative higher primate, may be mediated by kisspeptinergic neurones upstream to the GnRH network.

Induction of a hypothyroid state during juvenile development delays pubertal reactivation of the neuroendocrine system governing luteinising hormone secretion in the male rhesus monkey (Macaca mulatta).

The present study aimed to determine the influence of thyroid status on the timing of the pubertal resurgence in gonadotrophin-releasing hormone pulse generator activity [tracked by circulating luteinising hormone (LH) levels] in male rhesus monkeys. Six juvenile monkeys were orchidectomised and then treated with the antithyroid drug, methimazole, from 15-19 months until 36 months of age, at which time thyroxine (T(4)) replacement was initiated. Four additional agonadal monkeys served as controls. Blood samples were drawn weekly for hormonal assessments. Body weight, crown-rump length and bone age were monitored at regular intervals. By 8 weeks of methimazole treatment, plasma T(4) had fallen sharply, and the decline was associated with a plasma thyroid-stimulating hormone increase. In controls, plasma LH levels remained undetectable until the pubertal rise occurred at 29.3 +/- 0.2 months of age. This developmental event occurred in only half of the methimazole-treated animals before 36 months of age when T(4) replacement was initiated. The hypothyroid state was associated with a profound arrest of growth and bone maturation, but increased body mass indices and plasma leptin levels. T(4) replacement in methimazole-treated monkeys was associated with the pubertal rise in LH in the remaining three animals and accelerated somatic development in all six animals. Although pubertal resurgence in LH secretion occurred at a later chronological age in methimazole-treated animals compared to controls, bone age, crown-rump length and body weight at that time did not differ between groups. There were no long-term differences in plasma prolactin between groups. We conclude that juvenile hypothyroidism in male primates causes a marked delay in the pubertal resurgence of LH secretion, probably occasioned at the hypothalamic level. Whether this effect is meditated by an action of thyroid hormone directly on the hypothalamus or indirectly as a result of the concomitant deficit in somatic development remains to be determined.

A Reevaluation of the Question: Is the Pubertal Resurgence in Pulsatile GnRH Release in the Male Rhesus Monkey (Macaca mulatta) Associated With a Gonad-Independent Augmentation of GH Secretion?

A somatic signal has been posited to trigger the pubertal resurgence in pulsatile GnRH secretion that initiates puberty in highly evolved primates. That GH might provide such a signal emerged in 2000 as a result of a study reporting that circulating nocturnal GH concentrations in castrated juvenile male monkeys increased in a 3-week period immediately preceding the pubertal resurgence of LH secretion. The present study was conducted to reexamine this intriguing relationship, again in an agonadal model. Four castrated juvenile male monkeys were implanted with indwelling jugular catheters, housed in remote sampling cages, and subjected to 24 hours of sequential blood sampling (every 30 min) every 2 weeks from 19.5 to 22 months of age. Twenty-four-hour profiles of circulating GH concentrations were analyzed using the pulse detection algorithm, PULSAR, and developmental changes in pulsatile GH release with respect to the initiation of the pubertal rise of LH secretion (week 0; observed between 22.5 and 32 mo of age) were examined for significance by a repeated-measures ANOVA. Changes in the parameters of pulsatile GH secretion, including mean 24-hour GH concentration and GH pulse frequency and pulse amplitude for 3 (n = 4) and 6 (n = 3) months before week 0 were unremarkable and nonsignificant. These findings fail to confirm those of the earlier study and lead us to conclude that the timing of the pubertal resurgence of GnRH release in the male monkey is not dictated by GH. Reasons for the discrepancy between the two studies are unclear.

A striking sex difference in the gonadotropin response to gonadectomy during infantile development in the rhesus monkey (Macaca mulatta).

The present study was conducted to examine further the view that the early postnatal ontogeny of the neuroendocrine control system that regulates ovarian function in the rhesus monkey differs from that which governs testicular activity in this species. To this end, time courses of gonadotropin secretion were determined from birth until 40 weeks of age in neonatally ovariectomized (n = 6) and gonad-intact (n = 4) female rhesus monkeys by monitoring circulating LH and FSH concentrations in blood samples collected at weekly intervals. The developmental patterns of FSH and LH secretion in agonadal infantile females were then compared to those in a group of neonatally orchidectomized animals (n = 4). In addition, moment to moment changes in circulating LH concentrations were determined in agonadal male (n = 2) and female infants (n = 6) at approximately 1 month of age by collecting sequential blood samples every 10-20 min for 5-8 h. Ovariectomy at 1 week of age elicited a truncated and abbreviated rise in circulating LH concentrations, which contrasted to a postcastration LH hypersecretion of substantial magnitude and duration in males. The initial FSH response to neonatal ovariectomy and orchidectomy were similar, with both males and females exhibiting a striking and progressive rise in circulating levels of this gonadotropin during the immediate weeks following castration. Thereafter, however, a dramatic sex difference in the time course of open loop FSH secretion emerged; in males, plasma FSH concentrations declined to undetectable levels by 24 weeks of age, while in females, plasma concentrations of this gonadotropin were sustained at elevated levels for the duration of the 40-week period of observation. As a result of the foregoing sex difference in the early postnatal ontogeny of open loop gonadotropin secretion, circulating FSH to LH ratios in ovariectomized infantile female monkeys (2.3:8.1) were consistently greater than those in agonadal males (0.5:3.8). Interestingly, the elevated FSH to LH ratios in castrated females were associated with a relatively slow LH pulse frequency of approximately one pulse every 3-4 h. The expected circhoral frequency of pulsatile LH release was observed in agonadal males of comparable age. The persistence of sex differences in gonadotropin secretion in infantile rhesus monkeys several weeks after removal of the gonads suggests that the origin of these differences must reside at the hypothalamic-pituitary level rather than at the level of the gonad.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of orchidectomy and testosterone replacement treatment on pulsatile luteinizing hormone secretion in the adult rhesus monkey (Macaca mulatta).

The effect of bilateral orchidectomy and subsequent replacement treatment with testosterone (T) on the intermittent pattern of LH release were studied in six adult male rhesus monkeys. Frequent blood samples were withdrawn for extended periods via remote sampling devices that permitted continuous access to the venous circulation with minimal restraint of the animals. Plasma concentrations of LH were determined by RIA, and episodes of LH secretion (pulses) were identified using a computer-executed algorithm. In intact animals, the frequency of episodic LH secretion averaged one pulse every 4.5 h, and this increased within 2 days of castration to approximately one pulse every hour. The acceleration in LH pulse frequency occasioned by orchidectomy was associated with increases in mean plasma LH concentration and an enhancement of LH pulse amplitude. Implantation of acutely orchidectomized animals with T-containing Silastic capsules that resulted in sustained increments in plasma T concentrations in the mid to upper physiological ranges generally resulted, within 10 days, in a partial to complete reversal of the high LH pulse frequency characteristic of the castrated animal. The onset of the T-induced reduction in LH pulse frequency was associated with a marked increase in pulse amplitude. Since it is generally held that a LH pulse is occasioned by, and therefore reflects, a discharge of hypothalamic gonadotropin-releasing hormone, it may be proposed that in the rhesus monkey, the testes impose a decisive retardation on the neural mechanism that governs the timing of intermittent gonadotropin-releasing hormone secretion, and that this modulation of the hypophysiotropic oscillator by the gonad is mediated by testicular T secretion.

The effects of neonatal orchidectomy on the developmental pattern of gonadotropin secretion in the male rhesus monkey (Macaca mulatta).

Plasma concentrations of LH and FSH in immature male rhesus monkeys were determined at weekly intervals from birth until 300 days of age, and the effect of bilateral orchidectomy at 1 week of age on the neonatal pattern of gonadotropin secretion was studied. Plasma FSH and LH concentrations in intact animals were elevated during the first few months of neonatal life but then declined to undetectable values characteristic of the prepubertal state in this species. Neonatal castration, but not sham castration, resulted in a progressive and dramatic rise in circulating gonadotropin concentrations, which plateaued approximately 3 weeks later at values an order of magnitude greater than those observed in intact animals. The hypersecretion of LH and FSH after removal of the gonads was not sustained, however, and by 32 weeks of age plasma gonadotropin concentrations in the agonadal animals were indistinguishable from those in age-matched intact controls. These findings suggest that, in the rhesus monkey, the testes do not play a major role in determining the pattern of gonadotropin secretion throughout neonatal and prepubertal development. They further suggest that a profound inhibitory influence of nongonadal origin impinges upon the neuroendocrine control system governing gonadotropin secretion in the neonate, thereby initiating the diminished secretion of LH and FSH which is characteristic of the prepubertal stage of development in primates.

A study of the role of the postnatal testes in determining the ontogeny of gonadotropin secretion in the male rhesus monkey (Macaca mulatta).

To further examine the role of the testes in determining the ontogeny of gonadotropin secretion in the male rhesus monkey, the time courses of circulating LH and FSH concentrations were determined using established RIAs in daytime and nighttime blood samples collected at weekly intervals from birth until 4 yr of age in intact animals (n = 7) and in males orchidectomized at approximately 1 week of age (n = 6). Estimates of plasma androgen (A) concentrations were obtained on nonchromatographed samples with a RIA that reacts with testosterone and other androgens. Plasma PRL concentrations were also determined by RIA, and body weight was monitored at weekly intervals. Testicular volumes were measured at weekly intervals after 1.8 yr of age in five animals. In intact animals, mean daytime plasma A concentrations during the first 3 months of infantile development ranged from 1-6 ng/ml. Daytime plasma A concentrations then declined to reach 0.5-1 ng/ml by 9 months of age, where they were maintained usually until 3 yr of age when circulating levels progressively increased to reach, by approximately 3.5 yr of age, mean levels (6 ng/ml) characteristic of fully mature males. During the transition from infantile to perpubertal development, circulating LH and FSH concentrations showed a pattern similar to that of A, with elevations during the first 2 months of life followed by a decline to undetectable concentrations. In contrast to A, however, distinct pubertal increments in circulating LH and FSH concentrations were not observed in daytime samples from intact animals. The first indication of the pubertal reactivation of the hypothalamic-pituitary-testicular axis was provided by a reemergence of nocturnal elevations in plasma A concentrations between 2-3 yr of age. These were followed shortly thereafter by detectable plasma LH concentrations in nighttime samples. Orchidectomy at 1 week of age resulted in a progressive and dramatic rise in circulating gonadotropin concentrations, which plateaued approximately 3 weeks later at values 1 order of magnitude greater than those observed in intact animals. This hypersecretion of LH and FSH was not sustained, however, and by 10 months of age, plasma gonadotropin concentrations in agonadal animals were indistinguishable from those in age-matched intact controls. These low levels of the gonadotropic hormones were maintained, in the absence of the testes, for approximately 2 yr until a second or "pubertal" postcastration hypersecretion of LH and FSH was observed.(ABSTRACT TRUNCATED AT 400 WORDS)

Bilateral orchidectomy and concomitant testosterone replacement in the juvenile male rhesus monkey (Macaca mulatta) receiving an invariant intravenous gonadotropin-releasing hormone (GnRH) infusion results, as in the hypothalamus lesioned GnRH-driven adult male, in a selective hypersecretion of follicle-stimulating hormone.

The purpose of this experiment was to determine whether the testes of the juvenile male rhesus monkey, receiving an invariant intermittent iv infusion of GnRH, produce a specific FSH secretion-inhibiting hormone that exerts its action directly at the level of the pituitary gland. To this end, five male rhesus monkeys between 13-18 months of age were treated with a chronic intermittent iv infusion of GnRH (0.1 microgram/min for 3 min every 3 h) for 10 weeks to elicit an adult-like pattern in the episodic activity of the pituitary-Leydig cell axis. Animals were then bilaterally orchidectomized, and on the day of castration testosterone replacement with testosterone-containing Silastic capsules that maintained circulating levels of the steroid at approximately 6 ng/ml was initiated. Sequential blood samples were collected before castration and at 4-5, 11-13, and 18-19 days thereafter. Removal of the testes resulted in a marked and selective hypersecretion of FSH, a response very similar to that observed previously in hypothalamus-lesioned GnRH-treated adult males. This finding indicates that it will be possible to substitute the juvenile male for the much larger hypothalamus-lesioned adult in future studies requiring a hypophysiotropic clamp preparation. Such a modification of this experimental model will facilitate an examination of the effects on FSH secretion of passive immunization with inhibin antisera and of administering pure inhibin peptides.

Institution of combined treatment with testosterone and charcoal-extracted porcine follicular fluid immediately after orchidectomy prevents the postcastration hypersecretion of follicle-stimulating hormone in the hypothalamus-lesioned rhesus monkey (Macaca mulatta) receiving an invariant intravenous gonadotropin-releasing hormone infusion.

In the male rhesus monkey, the negative feedback regulation of gonadotropin secretion by the gonad involves a specific inhibitory action of a testicular hormone on FSH release at the level of the anterior pituitary gland. Neither circulating testosterone (T) nor estradiol appears to be able to account for the testicular inhibition of FSH in this species. The purpose of the present study was to begin to examine the role of gonadal peptides in this regard. To this end, an episodic pattern of activity in the pituitary-Leydig cell axis was restored in seven hypothalamus-lesioned male rhesus monkeys with a chronic and unchanging intermittent iv infusion of GnRH (0.1 microgram/min for 3 min every 3 h). This preparation, known as the hypophysiotropic clamp, has been described in detail previously. Charcoal-extracted porcine follicular fluid (pFF) was used as the source of gonadal peptides. In five animals, initiation of combined T replacement and pFF treatment (10-15 ml, sc, every 12 h for 8 days) maintained circulating FSH at concentrations similar to those observed before gonadectomy. Withdrawal of pFF treatment for 8 days while maintaining T replacement resulted in a progressive and dramatic rise in plasma FSH concentrations. Reinitiation of pFF treatment resulted in a return of circulating FSH concentrations toward precastration control values. Changes in LH secretion throughout the experiment were unremarkable. In an attempt to assess any nonspecific effects of porcine protein on gonadotropin secretion, the remaining two animals received charcoal-extracted pig serum instead of pFF. In these animals circulating FSH concentrations rose 7- to 8-fold during the 8 days of combined T replacement and pig serum treatment. These findings provide evidence to support the view that a testicular peptide, most probably inhibin, plays a major role in the negative feedback regulation of gonadotropin secretion in the monkey by exerting an inhibitory action on FSH secretion directly at the level of the anterior pituitary gland.

N-methyl-D,L-aspartate elicits hypothalamic gonadotropin-releasing hormone release in prepubertal male rhesus monkeys (Macaca mulatta).

In higher primates, the protracted delay from infancy to puberty results from an interruption in hypothalamic GnRH release. To determine whether the quiescent hypothalamic GnRH neurons of the prepubertal macaque are capable of discharging the decapeptide in response to a generalized neural depolarization, an excitatory amino acid analog, N-methyl-D,L-aspartate (NMA), was administered systemically to orchidectomized rhesus monkeys between 13 and 20 months of age. GnRH secretion was estimated indirectly by monitoring changes in circulating LH concentrations after the responsivity of pituitary gonadotropes to GnRH had been greatly facilitated by the chronic intermittent iv infusion of GnRH (0.1 microgram/min for 3 min every hour). The iv bolus administration of increasing doses of NMA (1.5, 4.8, and 15.0 mg/kg BW), 10-14 h after termination of the priming infusion of GnRH, elicited distinct discharges of LH, with magnitudes directly related to the amount of the excitant injected. Administration of a higher dose of NMA (48 mg/kg BW), however, failed to induce further LH release. The finding that pretreatment with a long-acting and potent GnRH receptor antagonist [( AcD2Nal1,4ClPhe2,DTrp3,DArg6,DAla10] GnRH-HOAc) abolished the LH-releasing activity of NMA provides compelling evidence for the view that the action of the neural excitant to induce gonadotropin release was exerted at a suprapituitary level. The additional observation that an N-methyl-D-aspartate receptor antagonist (D,L-2-amino-5-phosphono-valeric acid) blocked the NMA-induced release of GnRH suggests that the amino acid analog interacted with the N-methyl-D-aspartate receptor on neurons that synthesize and/or control the release of the hypothalamic hormone. Most interestingly, three sequential GnRH discharges, with a period and an amplitude apparently similar to those generated by the hypothalamus of the adult, were elicited from the brain of prepubertal monkeys by the sequential administration of three injections of NMA at hourly intervals. Taken together these findings demonstrate that the apparent dormancy of hypothalamic GnRH neurons, which is characteristic of prepubertal development in higher primates and underlies the protracted delay in the onset of puberty in these species, may be readily terminated by application of a generalized neural excitation. Plasma FSH, PRL, GH, and cortisol concentrations were also monitored during the course of some of these experiments, and release of each of these four hormones was observed after the iv injection of NMA (15 mg/kg BW).

Circulating leptin does not appear to provide a signal for triggering the initiation of puberty in the male rhesus monkey (Macaca mulatta).

The notion that circulating leptin may provide a somatic signal for timing the onset of puberty was examined in the male rhesus monkey. Circulating leptin levels were determined at weekly or biweekly intervals by RIA in intact (N=6) and prepubertally castrated monkeys (N=5) from approximately 18 to 30 months of age. Circulating testosterone (T) and gonadotropin levels were used as indices to identify the onset of the pubertal reaugmentation of pulsatile GnRH release (week 0) in intact and castrated animals, respectively. Time courses of the peripubertal changes in leptin concentrations in individual monkeys were normalized to week 0. In the intact group, mean leptin concentrations at this critical stage of development (week -26 to week +9) were unremarkable, ranging from 1.6+/-0.3 (SEM) to 2.4+/-0.6 ng/ml (P>0.05). In agonadal males, the pubertal onset of GnRH pulse generator activity, as reflected by sustained increments in plasma gonadotropin concentrations, also occurred in the absence of changes in circulating leptin levels (P>0.05). These findings indicate that the timing of the onset of puberty in male monkeys is not triggered by rising circulating leptin concentrations.

A study of the role of the adrenal glands in the initiation of the hiatus in gonadotropin secretion during prepubertal development in the male rhesus monkey (Macaca mulatta).

An earlier finding that the decline in LH and FSH secretion which typically occurs in neonatally orchidectomized rhesus monkeys between 8-32 weeks of age was temporally associated with a broad peak in circulating concentrations of immunoreactive testosterone has prompted us to evaluate the role of adrenal androgens in determining the ontogeny of gonadotropin secretion in this species. To this end, six orchidectomized (1 week) infantile rhesus monkeys were bilaterally adrenalectomized between 2 and 5 weeks of age and subsequently maintained on daily glucocorticoid (cortisone acetate or hydrocortisone acetate) and mineralocorticoid (desoxycorticosterone acetate) replacement. Plasma cortisol concentrations and electrolytes (Na+ and K+) were monitored at approximately weekly intervals, and the steroid replacements were titrated in an attempt to maintain the foregoing parameters within physiological limits. Plasma was also obtained at weekly intervals for 50 weeks in order to determine the time courses of LH and FSH secretion during this phase of postnatal development. Bilateral adrenalectomy resulted in an immediate and permanent decline in plasma testosterone to undetectable concentrations. This procedure together with appropriate steroid replacement, however, did not appear to influence the developmental pattern of gonadotropin secretion in neonatally orchidectomized monkeys; most notably, the reduction in the open loop secretion of LH and FSH between 8-32 weeks of age was neither interrupted nor masked by adrenalectomy. The growth rate of the adrenalectomized monkeys, as reflected by weekly body weight determinations, was less than that of adrenally intact animals. These findings suggest that adrenal androgens are not involved in the initiation of the prepubertal restraint of gonadotropin secretion in the rhesus monkey and are consistent with the notion that the adrenal gland does not play a major role in determining the timing of puberty onset in higher primates.