Experimental induction of puberty in the infantile female rhesus monkey.

Normal ovulatory menstrual cycles were initiated in prepubertal female rhesus monkeys by the infusion of gonadotropin-releasing hormone for 6 minutes once every hou;. When this regimen was discontinued, the animals promptly reverted to an immature state. These findings permit the conclusion that neither adenohypophysial nor ovarian competence is limiting in the initiation of puberty and suggest that this process depends on the maturation of the neuroendocrine control system that directs the pulsatile secretion of gonadotropin-releasing hormone from the hypothalamus.

Sexual differentiation of pituitary function: apparent difference bewteen primates and rodents.

Surges in luteinizing hormone secretion resembling those which occur spontaneously during the menstrual cycle were induced by acute elevations in circulating estrogen concentrations in both male and female rhesus monkeys gonadectomized in adulthood. These experiments demonstrate that in primates, in contrast to rodents, exposure of the hypothalamohypophyseal unit to androgens throughout fetal and postnatal development does not prevent the differentiation of the control system that governs cyclic gonadotropin secretion.

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.

The effects of ovariectomy, estrogen replacement, and neuraminidase treatment on the properties of the adenohypophysial glycoprotein hormones of the Rhesus monkey.

A comparison of pituitary extracts from intact and ovariectomized monkeys revealed that, like FSH, LH from ovariectomized animals is larger in average apparent molecular size and disappears from the circulation of test rats less rapidly than does LH from intact females. In contrast, ovariectomy had no effect on the apparent molecular size of TSH nor on its disappearance rate following injection into test rats. Reversal of the effect of ovariectomy on the molecular size of both LH and FSH was achieved by the chronic administration of estradiol. The differences in apparent molecular size of gonadotropins from intact and ovariectomized monkeys were either reduced or abolished by digestion with the enzyme neuraminidase. These findings suggest that the qualitative changes in the gonadotropins consequent to ovariectomy represent a specific response to the removal of estrogen and are consonant with the view that this phenomenon involves changes in the sialic acid content of the gonadotropins.

Qualitative changes in the pituitary gonadotropins of the male Rhesus monkey following castration.

Characterization of pituitary LH and FSH from intact and castrated male monkeys showed that gonadotropins from intact males are smaller in average apparent molecular size, and disappear from the circulation of test rats more rapidly than hormones from castrated males. Furthermore, intact male monkey LH and FSH are smaller and are cleared from the rat circulation faster than the gonadotropins from intact female monkeys. We conclude that 1) in the rhesus monkey changes in the properties of pituitary LH and FSH resulting from orchidectomy are similar to those which follow ovariectomy, 2) the response to gonadectomy of the male monkey with regard to the molecular properties of FSH is opposite to that of the male rat, and 3) the relationship between the influence of male and female sex hormones on the apparent molecular size of FSH in the monkey is the reverse of that in the rat.

A new substance resembling luteinizing hormone in the blood of rhesus monkeys.

Sera from a number of rhesus monkeys showed low or undetectable levels of LH according to radioimmunoassays which employ radioiodinated rhesus LH and antisera against rhesus LH or hCG. These same sera, when assayed by a system utilizing radioiodinated ovine LH and a unique anti-ovine LH serum which cross-reacts with LH from a variety of species, appeared to contain large and variable quantities of LH. The chromatographic behavior on Sephadex G-100 of the LH-like material in these sera was indistinguishable from that of authentic rhesus LH. Chromatographic fractions containing this LH-like material, as well as the sera from which they were derived, generated dose-response curves in the ovine:anti-ovine radioimmunoassay with steeper slopes than those produced by rhesus LH. These same chromatographic fractions had negligible activity in an alpha subunit radioimmunoassay which detects not only free rhesus alpha subunit but also the alpha component of undissociated rhesus glycoprotein hormones including LH. Treatment of these fractions with 4M guanidine-HCl produced a substance of smaller molecular size which, like rhLH beta, was active in the ovine:anti-ovine assay. A substance closely resembling the serum LH-like material but having a somewhat greater molecular size is also present in the rhesus adenohypophysis, but its relationship to the serum substance remains uncertain.

Surgical disconnection of the medial basal hypothalamus and pituitary function in the rhesus monkey. I. Gonadotropin secretion.

In an intial attempt to localize the central components of the neuroendocrine control systems whereby estrogens regulate gonadotropin secretion, the medial basal hypothalamus (MBH) was surgically isolated from the remainder of the brain in 18 female rhesus monkeys using a modified "Halasz knife". The completeness and localization of the resulting "cuts" were verified by examinations of serial 50 mugM coronal sections through each hypothalamus. The cuts extended from the optic chiasm to the mamillary bodies and from midline to the medial aspects of the optic tracts. The MBH "ISLANDS" ISOLATED BY THESE CUTS INCLUDED THE MEDIAN EMINENCE AND ARCUATE NUCLEI AS WELL AS PORTIONS OF THE VENTROMEDIAL NUCLEI, PREMAMILLARY AREAS AND MAMILLARY BODIES. Following complete MBH isolation, 6 of 7 otherwise intact monkeys responded to an estradiol benzoate (EB) injection with large LH and FSH surges. Three of these animals also displayed spontaneous gonadotropin surges which eventuated in ovulation as evidenced by sustained elevations in circulating progesterone levels. Similar patterns of LH and FSH secretion in response to EB administration were also observed in 6 of 7 intact animals in which only the superofrontal imputs to the MBH WERE INTERRUPTED. Four of these monkeys ovulated spontaneously. Circulating gonadotropin levels fell slightly in ovariectomized monkeys following complete MBH disconnection, but the circhoral, pulsatile rhythms of gonadotropin release persisted. These animals also responded to the negative and positive feedback actions of estrogen as evidenced by initial declines in circulating LH and FSH concentrations followed by surges of these hormones at the appropriate times following the injection of EB. It would appear from these observations that the sites of the negative and positive feedback actions of estrogen on LH and FSH secretion and, therefore, the sites of the central components of the neuroendocrine systems which control tonic and surge secretion of the gonadotropic hormones in the rhesus monkey may be resident within the MBH-hypophysial unit.

Functional luteolysis in the rhesus monkey: ovarian estrogen and progesterone during the luteal phase of the menstrual cycle.

In a continuing effort to elucidate the processes underlying in primates, the concentrations of estradiol, estrone and progesterone were measured in ovarian tissues and in ovarian vein plasma through the luteal phase of the menstrual cycle of the rhesus monkey. The concentration of progesterone in corpora lutea collected 4-6 days after the preovulatory LH peak (early luteal phase) was more than twice that found in those collected 8-13 days after the LH surge (late luteal phase) while estradiol and estrone concentrations in the latter had increased 4-fold. These changes in luteal steroid concentrations were paralleled by a striking increase in the concentration of estrone in the ipsilateral ovarian vein. Estrone predominated in the venous effluent of the ovary beaing the corpus luteum while estradiol concentrations were similar in both ovarian veins suggesting that estrone is the principal estrogen released by the corpus luteum. The results of this study are consonant with the hypothesis that estrogen produced by the corpus luteum is the physiologic luteolsin in the rhesus monkey.

Electrophysiological manifestation of luteinizing hormone-releasing hormone pulse generator activity in the rhesus monkey: influence of alpha-adrenergic and dopaminergic blocking agents.

Characteristic increases in neuronal electrical activity associated with the initiation of each LH pulse were recorded from ovariectomized rhesus monkeys bearing multiple chronic electrodes in the medial basal hypothalamus. These electrophysiological manifestations of hypothalamic LHRH pulse generator activity were inhibited by the alpha-adrenergic blocker phentolamine or the alpha 1-adrenoceptor blockers phenoxybenzamine and prazosin. At the dosages used, the effects of single injections of these drugs ranged from a reduction in the frequency of LHRH pulse generator activity to its complete arrest. This was faithfully reflected in the pattern of pulsatile LH discharges. The dopaminergic blocking agent metaclopramide similarly reduced the frequency of the pulse generator or arrested its activity altogether. The alpha 2-adrenoceptor blocker yohimbine had no demonstrable effect on hypothalamic electrical activity at the doses studied. These findings support the view of a central action of alpha 1-adrenergic and dopaminergic blockade on LHRH pulse generator activity and the concept that central adrenergic and dopaminergic inputs can modulate the frequency of the LHRH pulse generator.

Estradiol as a gonadotropin releasing hormone in the rhesus monkey.

The temporal relationship between cessation of GnRH delivery to the pituitary gland and the loss of responsiveness to the stimulatory action of estradiol (E2) was examined in 4 ovariectomized rhesus monkeys whose endogenous GnRH production had been abolished by hypothalamic lesions. Gonadotropin secretion was re-established by the intermittent administration of GnRH. The GnRH-replacement regimen was then discontinued and estradiol benzoate (EB) injected 24, 48, 72 and 96 hours later. Unambiguous gonadotropin discharges were induced when EB was administered 24 or 48 hours after discontinuation of GnRH replacement. We conclude that E2 can initiate gonadotropin discharges in the absence of circulating GnRH. E2 may, therefore, be viewed as a gonadotropin releasing hormone.

The sites of action of estradiol and phentolamine in the inhibition of the pulsatile, circhoral discharges of LH in the rhesus monkey (Macaca mulatta).

Pulsatile LH secretion was re-established in ovariectomized monkeys bearing hypothalamic lesions by an intermittent infusion of LHRH. The administration of estradiol to such animals resulted in a prompt cessation of these pulsatile discharges of LH and a resultant decline in the mean plasma concentration of the gonadotropin. The time course of this inhibition of LH secretion was indistinguishable from that observed after estrogen administration to ovariectomized animals with intact nervous systems. In contrast, phentolamine did not interrupt the pulsatile LH discharges occasioned by the hourly administration of exogenous LHRH to the lesioned animals. These results are consistent with the conclusion that the acute negative feedback action of estradiol on circhoral LH release in the monkey is at the level of the pituitary gland, whereas the inhibitory action of phentolamine on this mode of LH secretion is at a neural site.

Mode of action of progesterone in the blockade of gonadotropin surges in the rhesus monkey.

The action of progesterone (P) to block estradiol-induced gonadotropin surges in the rhesus monkey requires an intact hypothalamus. The present study was performed to determine whether the blocking action of P is the result of an inhibition in hypothalamic GnRH secretion or of some other mechanism. Ovulatory menstrual cycles were initiated in acyclic rhesus monkeys with intact nervous systems by a pulsatile GnRH replacement regimen previously shown to be effective in sustaining normal ovarian cycles in monkeys with hypothalamic lesions. Whereas unambiguous surges of LH and FSH were elicited by estradiol benzoate during the follicular phase of these cycles, no such gonadotropin discharges were obtained in the presence of luteal phase levels of P generated by sc P-containing Silastic capsules. Because exogenous GnRH replacement was provided throughout these experiments, it is concluded that P blockade of estradiol-induced gonadotropin surges is not the consequence of an interruption in GnRH release. The results suggest, rather, that P may cause the production of an inhibitory factor from the central nervous system which blocks the positive feedback action of estradiol on the pituitary gland.

Surgical disconnection of the medial basal hypothalamus and pituitary function in the rhesus monkey. III. Thyroxine secretion.

The effects of complete and anterior disconnection of the medial basal hypothalamus (MBH( on TSH secretion in the rhesus monkey were assessed by measuring serum thyroxine concentrations. Following complete MBH disconnection, serum thyroxine concentrations declined to 50% of preoperative control levels within 7 days. In 6 of 10 animals, thyroxine levels remained depressed throughout the postoperative observation period of 60 to 150 days. In the remainder, serum thyroxine concentrations returned to control levels after 3 weeks. Anterior disconnection of the MBH had variable effects on thyroid function. In 4 of 7 animals, serum thyroxine concentrations declined for 5 days following the operation but returned to preoperative levels within the next 3 weeks. In 2 others, serum thyroxine levels did not change while in the remaining animal thyroxine concentrations increased transiently to twice preoperative levels before returning to normal values. Thus, in the monkey, as in the rat, anterior MBH disconnection has little consistent effect on thyroid function, while complete isolation of the MBH causes a moderate decline which is most readily attributable to a decrease in thyrotropin secretion. The variations in the effects of anterior and complete MBH disconnection on thyroid function could not be confidently correlated with variations in the placement of the cuts.

The effect of luteinizing hormone releasing hormone (LHRH) antiserum administration on gonadotropin secretion in the rhesus monkey.

Single iv injections of a rabbit antiserum to synthetic LHRH promptly suppressed serum LH and FSH concentrations in ovariectomized rhesus monkeys. Gonadotropin levels remained depressed for 10-21 days, the approximate duration of enhanced LHRH binding activity in the circulation. Doses of LHRH antiserum sufficient to reduce tonic gonadotropin secretion did not modify the time course or magnitude of estrogen-induced gonadotropin surges. These negative findings, however, cannot be interpreted to signify that such surges are not caused by a release of LHRH.

On the site of action of progesterone in the blockade of the estradiol-induced gonadotropin discharge in the rhesus monkey.

The site of action of progesterone (P) in the blockade of estradiol-induced gonadotropin discharges was examined in rhesus monkeys bearing hypothalamic lesions which abolish endogenous GnRH production. Normal ovulatory menstrual cycles were re-established in these animals by the pulsatile, hourly administration of GnRH. In the follicular phase of these induced menstrual cycles, P-containing Silastic capsules were implanted sc yielding luteal phase plasma P concentrations which normally block estradiol-induced gonadotropin surges. P failed to block estradiol-induced surges in lesioned animals on GnRH replacement. In such animals, however, P advanced the initiation of these surges. While estradiol acts on the pituitary to cause gonadotropin discharges, P appears to block this effect by acting on the central nervous system. On the other hand, the results also suggest that the site of facilitatory action of P on gonadotropin release is at the level of the pituitary gland.

The arcuate nucleus and the control of gonadotropin and prolactin secretion in the female rhesus monkey (Macaca mulatta).

Attempts were made to destroy selectively the arcuate nucleus with radiofrequency current in adult female rhesus monkeys as a first step in identifying the areas of the mediobasal hypothalamus (MBH) that are responsible for the neural control of gonadotropin secretion in this species. Extensive or complete destruction of the arcuate region was produced in three animals and in two of these the lesion was confined primarily to the arcuate region and the dorsal aspect of the posterior median eminence. These lesions resulted in the cessation of LH and FSH secretion and blocked the positive feedback action of estradiol on gonadotropin release but did not appear to influence grossly basal thyroid and adrenocortical function, or to abolish GH discharge in response to insulin hypoglycemia. Adenohypophysial infarcts were not observed and exogenous LHRH and TRH induced marked discharges of the appropriate anterior pituitary hormones. In two additional animals with large hypothalamic lesions, destruction of the arcuate region was incomplete. In this group only partial inhibition of gonadotropin secretion was observed. LH and FSH secretion did not appear to be influenced in one animal bearing a large MBH lesion that entirely spared the arcuate region. Although serum prolactin remained at pre-lesion control levels after placement of the two relatively discrete lesions confined to the arcuate region, unambiguous increases in the secretion of this hormone were observed when the area of destruction encompassed tissue anterior and/or dorsal to the arcuate region. These observations suggest that the arcuate region is the primary structure mediating the hypothalamic control of gonadotropin secretion in the rhesus monkey. They also suggest that, in this species, the regions of the MBH involved with the regulation of gonadotropin release and those which control prolactin secretion are anatomically distinct.

On the sites of the negative and positive feedback actions of estradiol in the control of gonadotropin secretion in the rhesus monkey.

Ovariectomized rhesus monkeys bearing hypothalamic lesions which had abolished endogenous LHRH production, as evidenced by a profound reduction in gonadotropin secretion, but in which LH and FSH secretion was reestablished by a chronic intermittent iv infusion of synthetic LHRH (1 microgram/min for 6 min every hour) were used to investigate the sites of the negative and positive feedback actions of estradiol in the control of gonadotropin secretion. The administration of estradiol to such animals, while continuing the LHRH replacement regimen, resulted in a decline in circulating LH and FSH levels, followed by an unambiguous discharge of these hormones. The time course of this biphasic pattern of gonadotropin secretion was remarkably similar to that observed in response to estradiol administration in otherwise intact ovariectomized animals. These results suggest that, in the rhesus monkey, estradiol can exert both its negative and positive feedback actions on gonadotropin secretion at the level of the pituitary gland.

Surgical disconnection of the medial basal hypothalamus and pituitary function in the rhesus monkey. II. GH and cortisol secretion.

Eighteen female rhesus monkeys subjected to complete or anterior disconnection of the medial basal hypothalamus (MBH) were studied to assess the effects of these deafferentation procedures on GH and cortisol secretion. Basal serum levels of GH were not disturbed or were slightly elevated following complete or anterior MBH disconnection. GH secretion in response to vasopressin administration or insulin hypoglycemia, however, was abolished by complete isolation of the MBH. In contrast, the elevations in serum cortisol concentrations observed in response to these noxious stimuli were not noticeably affected. The normal diurnal rhythm in cortisol secretion remained fully evident following anterior deafferentation, but was severely attenuated or abolished when all neuronal inputs to the MBH were transected. Such observations suggest that the central components of the neuroendocrine systems which regulate basal GH secretion and which subserve stress-induced elevations in cortisol secretion are resident within the MBH-hypophysial unit. In addition, these data indicate that the mechanisms underlying the diurnal rhythm in cortisol secretion, as well as those mediating the discharges of GH in response to vasopressin administration and insulin hypoglycemia, are dependent on the integrity of neuronal connections between the MBH and other hypothalamic and/or extrahypothalamic areas.

Further studies on the effects of lesions in the rostral hypothalamus on gonadotropin secretion in the female rhesus monkey (Macaca mulatta).

Bilateral radiofrequency lesions were stereotaxically placed in the rostral hypothalamus of four adult female rhesus monkeys. These lesions resulted in extensive destruction of the ventromedial preoptic-anterior hypothalamic area (POA-AHA) and included the suprachiasmatic nucleus as well as, with the exception of one animal, the organum vasculosum of the lamina terminalis. In three of these four animals, gonadotropin surges similar to those observed before surgery were elicited in response to either a spontaneous increment in serum estrogen concentration or an estradiol benzoate injection. This stimulatory action of estradiol on LH and FSH release was not demonstratable in the remaining lesioned animal, but estradiol benzoate injections also failed to elicit a gonadotropin discharge in one of a series of five normal control animals. These findings fail to support the view that destruction of the ventromedial POA-AHA in this species compromises the ability of the hypothalamicohypophysial apparatus to respond to the positive feedback action of estradiol. The diurnal variation in serum cortisol concentration was not interrupted by placement of the lesions in the ventromedial POA-AHA.