Embryonic epsilon and gamma globin genes of a prosimian primate (Galago crassicaudatus). Nucleotide and amino acid sequences, developmental regulation and phylogenetic footprints.

Sequence analysis of epsilon and gamma genes and encoded globins and high-pressure liquid chromatography analysis of globin compositions in blood hemolysates obtained from embryos, fetuses and adults show that the prosimian primate Galago crassicaudatus expresses its epsilon and gamma genes only embryonically. Since rabbit, mouse and galago all have embryonic gamma genes but simian primates have fetal gamma genes, we conclude that gamma E evolved into gamma F in stem-simians. An elevated non-synonymous substitution rate characterizes this transition. The alignment of epsilon and gamma nucleotide sequences and the parsimoniously reconstructed evolutionary history of these sequences identify several anciently conserved cis-regulatory elements (phylogenetic footprints) important for gamma expression in primates and also cis-mutations which may have been involved in the recruitment of the gamma gene to a fetal program in simian primates.

The androgen status of aging male rhesus macaques.

Whether the rhesus male experiences changes in the dynamics of testosterone (T) and LH secretion with age was investigated in six young (6-8 yr old; 7-12 kg) and six aged (22+ yr old; 9-11 kg) intact rhesus males. Each male was fitted with an indwelling jugular catheter, which was attached to a cranial platform and stainless steel cable tether. Catheters passed from a swivel device at the top of each cage through a wall to an adjoining room. On four occasions, 1.0-ml blood samples were obtained from each male every 20 min for 24 h for plasma T RIA and plasma LH bioassay. Plasma T and LH data were analyzed by the PULSAR program to detect hormone peaks. Mean 24-h plasma T levels were similar in young and aged males as were the pulse amplitudes of T; however, the pulsatile pattern of T concentrations was different between young and aged males. Young males displayed a marked nocturnal increase in both T concentration and number of T pulses. Aged males demonstrated a significant nocturnal elevation of plasma T concentration, but displayed a significant nocturnal diminution in both concentration and numbers of T pulses compared with young males at night. Although LH concentrations and the total number of LH pulses per 24 h were similar in young and aged males, marked age-related alterations were evident in the pulsatile pattern of LH levels. Unlike young males, aged males failed to display a daytime reduction in the number of LH pulses. Our data point to coexistent changes in hypothalamic sensitivity to the negative feedback effects of T and testicular responsiveness to LH as a function of age in the rhesus macaque.

Sexual dimorphism in secretion of hypothalamic gonadotropin-releasing hormone and norepinephrine after coitus in rabbits.

Coital activation of the hypothalamo-hypophyseal ovarian axis (HOA) is well documented in rabbits, but coital excitation of the hypothalamo-hypophyseal testicular axis (HTA) is less well described. We and others have postulated that the response of the HOA to coitus, as characterized by a dramatic release of hypothalamic GnRH, may be mediated by an increase in norepinephrine (NE) neuronal activity. Herein, we studied selective HOA and HTA responses in New Zealand White rabbits before, during, and after coitus. Firstly, we determined the effects of microdialysis (mu D) and blood-sampling methods on plasma LH and testosterone (T) patterns in male rabbits during sexual performance. Subsequently, we compared the patterns of release in GnRH and norepinephrine (NE) from the arcuate nucleus-median eminence (AME) at 10-min intervals with changes in plasma LH levels in copulating male and female rabbits. Lastly, in 2.5-min AME mu D samples from females immediately after coitus, we measured NE and GnRH concentrations to determine whether NE release precedes that of GnRH. Tethered, freely moving rabbits were exposed to their partners for 10 min at the end of the third (10-min sampling for 5-7 h) or second (2.5-min sampling for 4 h) hour. Data from individuals that did not mate during the 10-min of pairing in the 3- to 7-h sampling trials were included as a control group (sham-mated). The results showed no changes (P > 0.05) in plasma LH and T in either mated (LH: pre, 0.13 +/- 0.08 ng/ml; post, 0.15 +/- 0.03 ng/ml; T: pre, 2.39 +/- 1.20 ng/ml; post, 0.85 +/- 0.26 ng/ml) or sham-mated males (LH: pre, 0.21 +/- 0.08; post, 0.25 +/- 0.10 ng/ml; T: pre, 1.46 +/- 0.51 ng/ml; post, 1.40 +/- 0.38 ng/ml). Likewise, coitus did not alter patterns of AME-NE (pre, 0.47 +/- 0.25; post, 0.56 +/- 0.25 ng/ml) and GnRH (pre, 0.61 +/- 0.45; post, 0.74 +/- 0.32 pg/ml) in mated or sham-mated males. The constant HTA activity during coitus in males appears to be independent of experimental manipulation per se because LH and T levels between mu D (0.18 +/- 0.05 and 1.72 +/- 0.85 ng/ml, respectively) and non-mu D (0.16 +/- 0.05 and 1.52 +/- 0.36 ng/ml, respectively) rabbits were not different (P > 0.05). In contrast to males, females displayed unambiguous and simultaneous increases in NE (P < 0.05) and GnRH (P < 0.01) release from the AME within 10-20 min after coitus; these elevated concentrations in mu D samples lasted for 3-4 h. Microdialysis NE levels averaged 0.02 +/- 0.01 ng/ml before mating, whereas postcoital values averaged 0.09 +/- 0.01 ng/ml. GnRH levels were 1.04 +/- 0.56 and 11.78 +/- 5.06 pg/ml before and after coitus, respectively. Concomitant increases in plasma LH levels were also observed after coitus in these female rabbits. Moreover, measurements of NE and GnRH in 2.5-min mu D samples revealed that the postcoital increase in NE preceded that in GnRH by 2.5-7.5 min (P < 0.05). The results suggest that neuroendocrine circuits in the two sexes of the New Zealand White rabbit respond differently to genital stimulation. In male rabbits, coitus does not alter patterns of AME NE or GnRH secretion, nor does it change the circulating levels of plasma LH or T. Conversely, in females, coitus induces the rapid release of hypothalamic NE, GnRH, and pituitary LH. The increase in coitally induced NE occurs before the rise in GnRH, which supports the hypothesis that NE is a critical neurochemical in coital activation of GnRH neurons.

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.

Preovulatory gonadotropin-releasing hormone surge in ovarian-intact rhesus macaques.

The occurrence and profile of the preovulatory hypothalamic GnRH surge in relation to plasma profiles of LH and ovarian steroids, i.e. 17 beta-estradiol (E2) and progesterone (P4), were examined in ovarian intact, freely moving rhesus macaques. Nine monkeys with active ovarian cycles were each fitted with a jugular venous catheter and two push-pull cannulae directed to separate sites within the median eminence (ME). Each female was connected continuously to a tether/swivel device through which daily blood samples or frequent blood samples and ME perfusates (simultaneously at 10- to 20-min intervals for 18-24 h) were obtained without disturbing the animals. An increment in the plasma E2 level (> 150 pg/ml) during the follicular phase (FP) was selected as the preovulatory ovarian signal and served as the index for initiating the ME push-pull perfusion (PPP). Daily increased P4 concentrations of more than 1 ng P4/ml plasma for several consecutive days were consistent with the assumption of ovulation and subsequent formation of a corpus luteum after PPP. A total of 18 PPP trials were completed; each in a fresh ME site. Five of these PPPs were performed during the mid- and late FP (3 were between 6-8 days before and 2 were 4 days before the E2 peak). The remaining 13 PPPs, each of 18- to 24-h duration, were performed between 24 h before and 48 h after the highest daily plasma E2 level, i.e. time zero. Of these 13 PPPs, 2 started within 12 h before (-12 to 0 h), 8 began within 12 h after (0-12 h), and 3 started between 12-24 h after this peak E2 value. During the FP, mean levels of GnRH and LH were less than 2 pg/ml and 20 ng/ml, respectively. During the periovulatory interval (-24 to 48 h around time zero), the release of hypothalamic GnRH (expressed in picograms per ml) increased to 6.63 +/- 2.35 between -12 to 0 h (n = 2), peaked at 20.70 +/- 6.09 between 0-12 h (n = 10), declined to 3.25 +/- 1.39 between 12-24 h (n = 11), and further declined to 0.89 +/- 0.18 between 24-36 h (n = 3). The mean GnRH value from 0-12 h was higher (P < 0.05) than other means (including those during the FP), except for the value between -12 to 0 h. Changes in mean plasma LH values during the same periods paralleled those in GnRH.(ABSTRACT TRUNCATED AT 400 WORDS)

Third-ventricular infusion of neuropeptide Y suppresses luteinizing hormone secretion in ovariectomized rhesus macaques.

Neuropeptide Y (NPY) has been shown to modulate gonadotropin secretion in an estrogen-dependent manner in the rat and rabbit, and to act centrally in these species to alter GnRH release. The present study examined the ability of centrally administered NPY to affect LH secretion in the primate. Human NPY (hNPY) was administered into the third cerebroventricle of unanesthetized, freely moving, ovariectomized (OVX), or estradiol (E2)-treated OVX rhesus monkeys. LH was measured in blood samples collected remotely at 10-min intervals throughout the experiments. An extensive range of NPY doses was tested in a preliminary study in which OVX monkeys received a 3-h control infusion of Krebs Ringer phosphate buffer (KRP) followed immediately by a 3-h infusion of hNPY (0.1-50 micrograms/h). Only doses greater than or equal to 5 micrograms/h produced a consistent and marked suppression of LH (5 micrograms/h; 35.1 +/- 7.2% reduction, P less than 0.05, n = 4). A longer duration study was performed to better characterize changes in LH pulse frequency and amplitude produced by hNPY treatment. We administered 5 micrograms/h and 15 micrograms/h to OVX (n = 5) and E2-treated OVX (n = 4) monkeys according to the following protocol: 8 h control KRP/8-h hNPY/6-h recovery KRP. In OVX monkeys, LH was suppressed after 2 to 3 h of peptide infusion (P less than 0.01); LH secretion returned to normal after treatment. Both doses of hNPY suppressed mean LH by approximately 55% (P less than 0.05) and LH pulse frequency by approximately 69% (P less than 0.025). LH pulse amplitude was unaffected. In E2-treated OVX monkeys, neither dose of hNPY affected mean LH or LH pulse amplitude. LH pulse frequency was suppressed by approximately 65% (P less than 0.05) during 15-micrograms/h treatment. Because centrally administered hNPY reduced LH pulse frequency and thereby mean LH levels, our results support a central, neural action of NPY to affect the GnRH/LH secretory system. The ability of estrogen feedback to alter the response to NPY treatment supports a physiological role for NPY in controlling reproduction in the primate.

Gonadotropin-releasing hormone and norepinephrine release from the rabbit mediobasal and anterior hypothalamus during the mating-induced luteinizing hormone surge.

The release of hypothalamic GnRH in association with the mating-induced LH surge was studied in the rabbit. Push-pull perfusion (PPP) of the mediobasal (MBH) or anterior (AH) regions of the hypothalamus was performed on conscious, unrestrained does for 3 h before and 5 h after exposure to a vasectomized buck. In experiment 1, GnRH concentrations were measured by RIA in 20-min fractions of MBH-PPP. An approximately 100-fold increase in GnRH release was observed within 1 h of coitus (pre, 1.15 +/- 0.29 pg/ml; peak, 106.67 +/- 37.42 pg/ml; n = 6; P less than 0.05). Concomitant surges of LH and PRL in the peripheral circulation were observed. In experiment 2, GnRH and norepinephrine (NE) were measured (the latter by radioenzymatic assay) in 10-min fractions of MBH-PPP. A 218% postcoital rise in NE levels (n = 5; P less than 0.05) in MBH-PPP accompanied an approximately 50-fold peak rise in GnRH in the same samples (pre, 1.57 +/- 0.23 pg/ml; peak, 76.52 +/- 50.14 pg/ml; P less than 0.05). MBH-NE, MBH-GnRH, LH, and PRL release began rising within 10 min of coitus. In experiment 3, GnRH was measured in 20 min fractions of AH-PPP. Coitus induced a marked rise in AH-GnRH release (precoitus, 0.31 +/- 0.03 pg/ml; peak, 2.25 +/- 0.80 pg/ml; n = 4; P less than 0.05) which differed from coitus-induced MBH-GnRH release both quantitatively (i.e. approximately 7-fold increase for AH vs. approximately 50-100-fold increase for MBH; 50-min lag time for AH vs. less than 20 min for MBH) and qualitatively (i.e. AH-GnRH release was discontinuous, while MBH-GnRH release rose sharply, plateaued, and then declined slowly over the 2-5 h following coitus). No changes in MBH-NE, MBH-GnRH, AH-GnRH, LH, FSH, or PRL were observed in sham-mated does (Exp 1, n = 7; Exp 2, n = 3; Exp 3, n = 4). These data support the hypotheses that: 1) hypothalamic GnRH release is a component of reflexive ovulation in the rabbit; 2) increased hypothalamic noradrenergic tone is related to the surge-release of GnRH; and 3) AH-GnRH release is enhanced following coitus.

Absence of regular pulsatile gonadotropin secretion during implantation in the rhesus macaque.

The secretory response of the primate corpus luteum (CL) to CG after implantation suggests that gonadotropin receptors are not depleted despite increasing CG production and continuous elevated tropic stimulation. Such continuous stimulation is known to cause down-regulation of receptors in other tissues. To determine if CG secretion is intermittent during the initial stages of CL rescue, we assessed the secretory pattern of CG during the periimplantation period by collecting frequent (4/h) blood samples in two studies (for 4 h on 3 separate days between days 8-13, or for 2 separate 13-h sequences between days 10-15 postovulation) in 13 chair-adapted females. Day 0 of gestation was defined as the day of ovulation, as estimated by peak urinary estrone conjugate excretion in females mated on days 9, 11, and 13 of the menstrual cycle. Hormone concentrations were measured by either RIA [irFSH; estradiol and progesterone (P)] or Leydig cell bioassay (bioLH or bioCG). In the first study, 4 of 6 females conceived, and the mean for bioLH was not significantly elevated until days 12-13. In the second study, 5 of 7 females conceived, and the episodic secretory pattern of circulating pituitary bioLH typically observed in cycling females (2.7 +/- 0.3 peaks/13 h) was replaced by a relatively nonpulsatile, but steadily increasing profile during days 12-15 of gestation (1.5 +/- 0.4 peaks/13 h). Although occasional large fluctuations in bioLH/CG and P were noted, the bioLH/P peaks were less congruent than those in nonfertile cycles, and there was no diurnal pattern in the secretion of either hormone. In contrast, irFSH concentrations did not fluctuate and were similar in the two groups of females [2.93 +/- 0.28 vs. 2.34 +/- 1.7 ng/ml (mean +/- SEM)]. These data demonstrate that 1) a steady, gradually increasing secretory pattern of CG is associated with rescue of the CL; 2) the circulating profile of CG during the periimplantation interval is not consistently episodic and does not support the hypothesis that intermittent CG release prevents LH/CG receptor down-regulation in the CL during early pregnancy; 3) increased bioLH/CG levels during conceptive cycles in rhesus monkeys are not detected until days 12-13 after the midcycle bioLH peak; 4) irFSH patterns on pooled aliquots appear to be uninformative with regard to gonadotropin dynamics in early pregnancy; and 5) urinary estrone conjugate measurements provide a practical method for the precise timing of infrequent events, such as implantation, in the laboratory macaque.

Unexpected virilization in male mice lacking steroid 5 alpha-reductase enzymes.

Mice lacking steroid 5 alpha-reductase 1 and 2 were produced by gene targeting and breeding. Male mice without 5 alpha-reductase 2 or without both enzymes had fully formed internal and external genitalia and were fertile, but had smaller prostates and seminal vesicles than controls. T accumulated to high levels in the reproductive tissues of the mutant mice. DHT administration increased seminal vesicle and coagulating gland weights in mice deficient in 5 alpha-reductase 2 and increased the weights of the prostate, seminal vesicle, and coagulating gland in animals deficient in both enzymes. An inhibitor of both 5 alpha-reductases (GI 208335X) decreased prostate and coagulating gland weights of control mice, but had no effect in those lacking 5 alpha-reductase 1 and 2. Castration reduced the sizes of these tissues in animals of all genotypes. Androgen-dependent gene expression was decreased in the seminal vesicles of mice lacking one or more 5 alpha-reductases and was restored by administration of T or DHT. Female mice missing both enzymes exhibited parturition and fecundity defects similar to those of animals without 5 alpha-reductase 1. We conclude that T is the only androgen required for differentiation of the male urogenital tract in mice and that the synthesis of DHT serves largely as a signal amplification mechanism.

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.

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.

Suppression of mediobasal hypothalamic gonadotropin-releasing hormone and plasma luteinizing hormone pulsatile patterns by phentolamine in ovariectomized rhesus macaques.

In gonadectomized animals, pulses of LH are secreted concurrently with pulsatile hypothalamic GnRH and it is hypothesized that pulses of GnRH are either driven or modulated by episodes of catecholamine release. The objective of this study was to determine if the alpha-adrenergic antagonist phentolamine (PHEN) can simultaneously block the release of GnRH and LH in ovariectomized (OVX) rhesus macaques. In Exp 1, simultaneous peripheral blood and mediobasal hypothalamic push-pull perfusion (PPP) samples were collected remotely at 10-min intervals for 24 h via a swivel/tether device in eight conscious, freely moving OVX rhesus monkeys. Phentolamine was continuously infused iv for 6 h at the rate of 4 mg/kg BW.h in five animals and 20 mg/kg BW.h in three animals. Infusion started at 6 h after the commencement of PPP. Sampling of PPP and blood continued for 12 h after the cessation of PHEN infusion. Exp 2 was carried out to determine if PHEN affects pituitary responsiveness to exogenous GnRH under conditions similar to those in Exp 1. Exogenous GnRH (5 micrograms, iv) was injected as a single bolus at 10-h intervals before, during, and after either a saline (4 ml/h for 6 h) infusion or, 3 weeks later, a PHEN infusion (4 mg/kgBW.h for 6 h) in three OVX females. The results of Exp 1 show that pulsatile patterns of hypothalamic GnRH and LH were either dampened or abolished by PHEN infusion. During the recovery period after PHEN infusion, pulse amplitudes of LH were enhanced, but pulse amplitudes of endogenous GnRH did not differ, as compared to those of corresponding LH and GnRH before infusion of PHEN. Data from Exp 2 suggested that the alpha-adrenergic blocking agent had no effect on the pituitary LH response to exogenous GnRH administration. These results directly support the hypothesis that adrenergic neuronal activities are critical for the pulsatile release of hypothalamic GnRH which governs the pulsatile release of LH in gonadectomized animals.

Estrogen-induced gonadotropin surges in decerebrated female rhesus monkeys with medial basal hypothalamic peninsulae.

Hypothalamic peninsulae, posteriorly continuous with the brain stem, were produced in ovariectomized rhesus monkeys by aspiration of the cerebral hemispheres and all brain structures anterior and dorsal to the optic chiasm. These preparations were sustained for 52 to 60 h postoperatively by maintaining blood gases and pH within physiological limits and by supporting arterial blood pressure, when necessary, with infusions of rhesus monkey blood and norepinephrine. The sc injection of estradiol benzoate (EB) to such animals upon completion of the ablation procedure was followed 18-24 h later by unambiguous surges of serum LH and FSH with durations in the excess of 24 h. These findings are consonant with the view that the central components of the control system that initiates the preovulatory gonadotropin surge in the rhesus monkey are resident within the medial basal hypothalamic-hypophysial apparatus.

Persistence of fetal zone function in the infant rhesus monkey adrenal gland.

Plasma dehydroepiandrosterone sulfate (DHAS) concentrations increase markedly in the rhesus monkey fetus at the end of gestation. A further increase occurs in the infant. To determine whether the changes in plasma concentration between the fetus and infant represent maintenance of DHAS production by the infant adrenal gland, we measured the t1/2, distribution volume (VD), MCR, and production rate of DHAS in the late gestation rhesus monkey fetus (129-155 days gestation; term is 165 days) and infant (14-42 days of age). A single bolus dose of [3H]DHAS was injected into five fetuses and four infants, and blood samples were collected serially from 5 min to 24 h after the injection. The amount of [3H]DHAS in the circulation was measured after solvolysis, extraction, and Celite chromatography. The concentration of DHAS in each sample was measured by RIA. DHAS was cleared significantly more rapidly in the fetus than in the infant [MCR in fetus, 2.4 +/- 0.4 (+/- SE); MCR in infant, 0.6 +/- 0.2 liters day-1 kg-1]. The t1/2 of DHAS was shorter in the fetus than in the infant (1.0 +/- 0.1 vs. 3.3 +/- 0.7 h). Absolute VD values were larger in the fetus than in the infant (231 +/- 29 and 143.8 +/- 11.6 ml kg-1); however, they were similar when the fetal VD was calculated including placental weight as a component of fetal weight. The production rate of DHAS, calculated as the product of MCR and integrated plasma DHAS concentration for the duration of the experiment, was not significantly different between the fetus and the infant (1.0 +/- 0.2 and 3.3 +/- 1.2 mg kg-1 day-1) in spite of the marked differences in plasma DHAS concentrations (445.8 +/- 103.8 ng ml-1 in the fetus and 5165 +/- 1296 ng ml-1 in the infant). These results indicate that the adrenal of the infant rhesus monkey continues to secrete DHAS at a rate at least as high as that in the late gestation fetus. Since the infant maintains DHAS production similar to that of the fetus in the absence of the placenta, a corollary of these studies is that the elevated DHAS secretion in the rhesus infant is independent of the placenta or the hormonal milieu of pregnancy. The maintenance of a functional fetal zone in the adrenal gland makes the rhesus infant a suitable model to use in studying the regulation of DHAS secretion and fetal zone morphology.

Acute administration of a 3 beta-hydroxysteroid dehydrogenase inhibitor to rhesus monkeys at the midluteal phase of the menstrual cycle: evidence for possible autocrine regulation of the primate corpus luteum by progesterone.

Colocalization of progesterone receptors and 3 beta-hydroxysteroid dehydrogenase (3 beta HSD), a key enzyme in progesterone biosynthesis, in macaque luteal cells suggest that progesterone has an autocrine role in the regulation of primate luteal function. To test this hypothesis, we administered trilostane, a 3 beta HSD inhibitor, to rhesus macaques at the midluteal phase of spontaneous menstrual cycles to rapidly and reversibly reduce progesterone production. Animals received trilostane (600 mg/dose; treated group; n = 5) or vehicle (control group; n = 4) orally on days 6-7 of the luteal phase. Trilostane significantly (P < 0.05) elevated pregnenolone levels within 1 h of treatment compared to those in vehicle-treated animals; after 1 day of treatment, the mean pregnenolone level (173 nmol/L) was 86-fold greater than the control value. Pregnenolone levels dropped after cessation of drug administration and became indistinguishable from control levels by day 13. Trilostane significantly reduced serum progesterone levels within 3 h of initial administration (P < 0.01), and levels remained near baseline (1.0 nmol/L) throughout the 2 days of treatment. Progesterone levels also remained low after cessation of trilostane treatment in four of five monkeys, and trilostane-treated animals experienced a shorter luteal phase than vehicle-treated animals (7.8 +/- 0.2 vs. 16 +/- 1 days; P < 0.01). Histological analysis (n = 3/group) revealed indexes of premature structural luteolysis by 4 days after the onset of trilostane administration. Exposure to trilostane had no effect on the percentage of luteal cells expressing progesterone receptors, as determined by immunocytochemistry. Serum LH levels were not different between treatment and control groups throughout the experimental period. As trilostane dramatically reduced serum progesterone and induced premature menses without major concurrent alteration in serum cortisol, we conclude that trilostane ios an effective, rapidly acting inhibitor of 3 beta HSD in the macaque corpus luteum during the midluteal phase of the menstrual cycle. Progesterone production did not typically resume after cessation of trilostane treatment despite continuing gonadotropin support luteolysis. Thus, progesterone or a related metabolite may be required to maintain the function and structural integrity of the primate corpus luteum during the normal menstrual cycle.

Endocrine and morphological maturation of the fetal and neonatal adrenal cortex in baboons.

Developmental changes in basal and circadian fluctuations in plasma dehydroepiandrosterone sulfate (DHEAS) and cortisol (F) from birth until 3 months of life were studied in conjunction with morphological characteristics of the fetal and neonatal adrenal cortex in baboons (Papio anubis). These studies were complimented by measurements of in vitro production of DHEAS and F (basal and ACTH stimulated) by adrenal tissue slices. Cortisol, DHEAS, estrone, estradiol, and progesterone were determined in plasma and incubation medium by specific RIA. After delivery, an initial rise in plasma DHEAS was sustained for 4 days, followed by a precipitous decline, which reached a nadir between days 10-12 postpartum. Thereafter, plasma DHEAS and F concentrations stabilized with minor fluctuations. A significant (P less than 0.05) and persistent diurnal rhythm in DHEAS and F secretion was evident by the end of the first week after birth. Administration of estrone acetate and progesterone in oil maintained neonatal plasma concentrations of estrone, estradiol, and progesterone at levels comparable to those in utero, but had no quantitative or qualitative effect on the pattern of plasma DHEAS or F in the neonate. The relative production of F to DHEAS by tissue slices in vitro (the calculated F/DHEAS ratios) indicates that DHEAS secretion by the fetal adrenal is 10-fold higher than that of F near parturition; ACTH stimulation does not alter this relationship. By postnatal day 10, the basal production rates of F and DHEAS are equivalent, and the response to ACTH stimulation favors production of F. With advancing neonatal age (at 30 and 100 days), there is an increase in the F/DHEAS secretion ratio both during the basal state and in response to ACTH. The baboon adrenal glands increased in weight during the last month of gestation and then stabilized during the early postnatal period; a gradual increase in weight was observed after 30 days postpartum. Within 2 weeks after parturition, the relative width of the fetal zone decreased dramatically to occupy less than one third of the total cortex. During involution, we observed a decrease in cell size and a reduction in cytoplasmic vacuolation. A zone of closely packed cells with numerous areas of cell death (the dense band) separated the zona fasciculata from the fetal zone. Cell proliferation was observed in the upper regions of the definitive cortex. We conclude the following. 1) The hypothalamic-hypophyseal mechanisms that regulate the diurnal adrenal secretory rhythm are established in the early neonatal period in the baboon.(ABSTRACT TRUNCATED AT 400 WORDS)

Comparison of plasma oxytocin and catecholamine concentrations with uterine activity in pregnant rhesus monkeys.

Pregnant rhesus monkeys exhibit diurnal changes in uterine activity (UA), with episodes of increased UA during the early hours of darkness. The estrogenic environment during late pregnancy serves a permissive role in the maintenance of nocturnal UA episodes and may involve myometrial interactions with oxytocin (OT) and/or alpha-adrenergic stimuli. In the present study we have used chronically catheterized pregnant rhesus monkeys to measure diurnal changes in maternal plasma OT, epinephrine, norepinephrine, and dopamine. We also determined the effects of infusing an OT antagonist (ORF 22164) and the alpha-adrenergic antagonist phentolamine on nocturnal UA episodes. Animals were exposed to a 16-h light, 8-h dark photo-period, with the hours of darkness between 2300-0700 h. Maternal plasma samples were collected at 3-h intervals for 36 h and analyzed by RIA for OT and by high performance liquid chromatography for catecholamines. Plasma OT was correlated with UA in animals that displayed nocturnal UA episodes (r = 0.76; P less than 0.01). Maximal OT concentrations occurred at 2400 h in these animals; plasma OT was higher during the hours of darkness compared to levels during the light phase (10.4 +/- 1.9 and 3.0 +/- 0.3 pmol/L, respectively; n = 4). Some animals did not display nocturnal episodes of increased UA and showed no increase in OT concentrations during the hours of darkness. Maternal plasma catecholamine concentrations were not correlated with nocturnal UA and were maximal during the light phase. Nocturnal UA was abolished within 30 min of infusion of the OT antagonist, but phentolamine infusions had no effect on nocturnal UA. We conclude that 1) changes in maternal plasma catecholamine concentrations are not involved in the generation of nocturnal UA; 2) the presence of episodes of increased UA at night results from increased maternal plasma OT concentrations; and 3) the absence of nocturnal UA in some animals can be explained by a reduced level of OT secretion.

Recombinant human inhibin-A administered early in the menstrual cycle alters concurrent pituitary and follicular, plus subsequent luteal, function in rhesus monkeys.

Inhibin, a suppressor of pituitary FSH secretion in nonprimate species, may also act in the ovary to regulate follicular development. To examine whether inhibin has similar actions in primates, female rhesus monkeys (n = 3/treatment), exhibiting regular menstrual cycles, received sc injections of either vehicle or 60 micrograms/kg recombinant human inhibin-A at 0800 and 1600 h for 5 days beginning at menses. The vehicle-treated monkeys displayed menstrual cycles of normal length, with the follicular (11.3 +/- 2.5 days, mean +/- SE) and luteal (16.3 +/- 2.5 days) phases demarcated by midcycle peaks in serum estradiol (E) and bioactive LH. After the first inhibin injection, levels of immunoreactive inhibin A peaked at 10 ng/mL within 1 h and returned to baseline (< 0.1 ng/mL) before the second injection 8 h later. Although serum E and LH did not change, bioactive FSH decreased (to 66% of pretreatment levels, P < 0.05) within 8 h. Within 1 day, circulating bioactive FSH was less (P < 0.05) in inhibin-treated monkeys, compared with controls. By 2-3 days, serum E levels were also markedly (P < 0.05) reduced in inhibin-treated animals, whereas bioactive LH rose 3-fold (P < 0.05). After inhibin treatment, the midcycle rises in serum E and LH were delayed; hence, the follicular phase was prolonged (15.0 +/- 2.6 days, P < 0.05), compared with controls. Although the patterns and levels of serum LH circulating during the subsequent luteal phase seemed comparable in both groups, mean progesterone levels were suppressed to 2-3 ng/mL (P < 0.05) during the midluteal phase in inhibin-treated monkeys. However, the length of the luteal phase in inhibin-treated cycles (13.0 +/- 2.6 days) was not significantly altered. We conclude that exogenous inhibin rapidly diminishes pituitary FSH secretion in female monkeys during the early follicular phase of the menstrual cycle. This action, and/or other actions directly on the ovary, leads to subsequent effects on follicular steroidogenesis and pituitary LH secretion that culminate in an aberrant ovarian cycle characterized by an insufficient luteal phase. The study identifies, for the first time, possible activities and roles of inhibin during the ovarian cycle in primates.

Inhibition of aromatization stimulates luteinizing hormone and testosterone secretion in adult male rhesus monkeys.

Experiments were conducted to examine the role of aromatization in the control of LH and testosterone secretion in adult male rhesus monkeys. Treatment of male monkeys (n = 7) with sc Silastic packets containing the aromatase inhibitor 1,4,6-androstatriene-3,17-dione (ATD) resulted in 1.5- to 3-fold elevations in serum LH and testosterone concentrations in six of seven animals. Concurrent treatment of ATD-treated monkeys with small quantities of estradiol-17 beta (n = 4) abolished the stimulatory effect of ATD. During ATD treatment, peripheral estradiol levels were reduced by 30% and hypothalamic aromatase activity, as determined in vitro, was reduced 80-90%. The lack of androgenic or antiandrogenic activity of ATD was demonstrated by its inactivity in either a mouse seminal vesicle bioassay or a highly sensitive penile spine bioassay. Furthermore, ATD did not react with rat prostatic or hypothalamic cytosol androgen receptors. 1,4,6-Androstatriene-17-ol-3-one, a possible metabolite of ATD in vivo, did react with prostatic and hypothalamic androgen receptors, but possessed no antiandrogenic activity in either bioassay. Thus, treatment of adult males with an aromatase inhibitor that inhibits both peripheral and central aromatization, and which has no apparent antiandrogenic activity, results in stimulation of LH and testosterone secretion. These data demonstrate that aromatization of androgens to estrogens plays an important role in negative feedback regulation of LH secretion and maintenance of normal testosterone levels in adult male primates.