The LHRH neuronal system in female rats: relation to the medial preoptic nucleus

Previously we have found that small lesions confined to the medial preoptic nucleus (MPN) or the suprachiasmatic nucleus (SCN) blocked the cyclic release of gonadotropins in the female rat, inducing a persistent estrous state. Since the MPN is located just caudal to the organum vasculosum of the lamina terminalis (OVLT) where LHRH cell bodies are most concentrated, we applied an immunocytochemical technique to examine the possibility that the lesions had simply disrupted LHRH neurons or fibers. Using a new anti-LHRH provided by Dr. V. D. Ramirez, we found that the distribution pattern of immunoreactive LHRH cell bodies and fibers was similar to that previously reported, although the staining was more intense and extensive with low background. There was no concentration of LHRH cell bodies and fibers in the MPN or SCN and, in fact, these nuclei generally showed a lower density of stained elements than did surrounding tissue. In persistent estrous animals with lesions confined to the MPN there was no detectable reduction of stained fibers in the median eminence. These results, along with the results of other workers, suggest that persistent estrus following lesions of the MPN or SCN is not due to reduction of LHRH neurons or fibers. Rather, they support the hypothesis that these nuclei are critical for triggering the ovulatory release of LHRH.     

Dietary fish oil delays puberty in female rats.

Marine oils contain eicosapentaenoic acid, a fatty acid that competes for cyclooxygenase and reduces the synthesis of dienoic prostanoids including prostaglandin E2 (PGE2). Since PGE2 plays an important role in the estrogen-stimulated release of hypothalamic GnRH on proestrus, it was postulated that a diet containing fish oil would delay first ovulation through inhibitory effects on GnRH release. Thirty, 22-day-old female Sprague-Dawley rats were fed a diet containing fish oil ad libitum. Controls were pair-fed an identical diet with the substitution of safflower oil as the dietary fat. All rats were killed on the morning of first metestrus after vaginal opening and the display of an estrous smear(s). Fish oil feeding did not affect growth as indicated by the lack of an observed effect on body weights or femur lengths. On the other hand, pituitary, ovarian, and uterine weights were significantly lower in the rats fed fish oil (p < 0.001). The age at first estrus of the rats fed fish oil was significantly increased compared with the controls (42.9 +/- 1.0 vs. 36.1 +/- 0.3 days; p < 0.001), whereas the number of rats with corpora lutea (CL), as well as the number of CL per ovary (2.3 +/- 0.4 vs 4.8 +/- 0.6 for controls; p < 0.001) was significantly reduced by fish oil feeding. GnRH concentration in the preoptic area/hypothalamus was significantly increased in the fish oil-fed rats (21.4 +/- 4.0 pg/mg vs. 7.6 +/- 2.2 pg/mg for controls; p < 0.01); radioimmunoassable hypothalamic PGE2 was concomitantly reduced (p < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Decoupling through synchrony in neuronal circuits with propagation delays

The level of synchronization in distributed systems is often controlled by the strength of the interactions between individual elements. In brain circuits the connection strengths between neurons are modified under the influence of spike-timing-dependent plasticity (STDP) rules. Here we show that when recurrent networks with conduction delays exhibit population bursts, STDP rules exert a strong decoupling force that desynchronizes activity. Conversely, when activity in the network is random, the same rules can have a coupling and synchronizing influence. The presence of these opposing forces promotes the self-organization of spontaneously active neuronal networks to a state at the border between randomness and synchrony. The decoupling force of STDP may be engaged by the synchronous bursts occurring in the hippocampus during slow-wave sleep, leading to the selective erasure of information from hippocampal circuits as memories are established in neocortical areas.     

Neonatal corticoid administration: retardation of adrenal rhythmicity and desynchronization of puberty.

The development of the adrenal corticosterone and progesterone diurnal rhythms was retarded by a single injection of cortisol acetate, dexamethasone acetate or progesterone into 3-day-old female rats. This effect was transient with the establishment of the corticosterone rhythm by day 70. The retardation of adrenal hormone rhythmicity may alter the normal process of sexual maturation, because cortisol and dexamethasone-treated animals but not progesterone treated rats exhibited a desynchronization of the events associated with puberty. Vaginal opening was delayed, and dissociated from first ovulation in these rats. Further, neonatal cortisol administration lengthened the estrous cycle in adulthood. It can be concluded from our study that the maturation of the neural centers controlling adrenal steroid secretion may be essential for the proper timing of pubertal events in the female rat.     

Neonatal androgens influence the social play of prepubescent rats

The results of six experiments designed to investigate the hormonal basis of the sex differences in the occurrence of social play in the rat are reported. From the time of weaning animals were housed in mixed-sex, peer groups of six, composed of some treated and some untreated animals. Observations were made of the animals in these groups each day between Days 26 and 40 of life in Experiments 1, 3–6 and between Days 31 and 40 in Experiment 2. In Experiment 1 it was found that males castrated on Day 1 of life engaged in less social play than did intact males, and did not differ from normal females. In Experiment 2, castration carried out at 23 days of age had no effects on the frequency with which males engaged in social play. In Experiment 3, it was found that neonatal ovariectomy had no effect on the frequency with which female pups engaged in social play. In Experiment 4, females treated on Days 1 and 2 of life with either 250 μg of testosterone propionate or 250 μg of dihydrotestosterone engaged in social play at rates comparable to those of normal males, whereas treatment with 5 μg of estradiol benzoate had no such effect. In Experiments 5 and 6 it was found that neither the reduction of testosterone-derived estradiol (by implants of the aromatization blocker, androst-1,4,6-triene-3,17-dione) nor that of testosterone-derived dihydrotestosterone (by implants of the 5α-reductase blocker, testosterone 17β-carboxylic acid) during the early neonatal period (Days 1 to 10 of life) changed the frequency of social play in intact males. The results of these experiments indicate that the sex difference in the social play of prepubescent rats is dependent on the neonatal exposure to testosterone or to its 5α-reduced metabolite, dihydrotestosterone. The reduction of testosterone to dihydrotestosterone, however, would not appear to be a necessary step.     

Activation of AMPA receptors inhibits prolactin and estradiol secretion and delays the onset of puberty in female rats

Previous experiments have evidenced the neuroendocrine role of AMPA receptors. Present studies were carried out to obtain information on the role of these receptors in the control of the onset of puberty. To this end, female rats were i.c.v. injected with vehicle or AMPA (agonist of AMPA receptors: 0.1 or 0.5 nmol/day) between 26 and 30 days (Experiment 1), or 30 and 34 days (Experiment 2) of age. Serum concentrations of PRL, LH and estradiol were measured before drug administration, 10 min after the last injection, at vaginal opening (VO) and at first estrus (FE) presentation. In both experiments, AMPA administration inhibited PRL and estradiol secretion without affecting LH release. When AMPA was administered between 26 and 30 days a significant delay in the day of vaginal opening was observed. These results confirmed the inhibitory effect of AMPA on PRL secretion and suggests a role of AMPA receptors in the control of puberty onset.     

Single administration of di(n-butyl) phthalate delays spermatogenesis in prepubertal rats

Morphological alterations in seminiferous tubules caused by single administration of di(n-butyl) phthalate (DBP) in 3-week-old rats were investigated throughout the first wave of spermatogenesis. Single administration of DBP (500 mg/kg) showed progressive detachment and displacement of spermatogenic cells and disappearance of tubular lumen at 3 h after treatment, and then showed thin seminiferous epithelia and wide tubular lumen at day 1 (D1). At D1, quite significant numbers of apoptotic spermatogenic cells were detected, and then they gradually decreased in accordance with the passage of time. In contrast, the testes revealed lower weight gain, even after completion of first wave of spermatogenesis in the DBP-treated group, compared to the control. In order to clarify whether spermatogenic cells differentiate into mature spermatids in the DBP-treated rats, immunohistochemical staining for Hsc 70t, a specific marker for elongate spermatids, was carried out. As a result, the decrease in mature spermatids in the DBP-treated testes, compared to the control, was demonstrated. For example, at D20 (41-day-old) after treatment, the most advanced spermatids in the tubules from rats in the DBP-treated groups were steps 2-4, while those of the control were steps 12-13. Moreover, in some tubules, pachytene spermatocytes were the most advanced spermatogenic cell. At D30 (51-day-old) after treatment, maturation of spermatogenic cells in the DBP-treated rats proceeded further, and the most advanced spermatids in tubules were steps 8-9, while those of the control were steps 15-19. These results lead us to the postulation that a single administration of DBP to prepubertal rats delays maturation of spermatogenic cells, even after completion of first wave of spermatogenesis.     

Experience with the use of an intranasal LHRH analogue for the induction of puberty

7 patients aged 16-25 years with hypogonadotrophic hypogonadism have been treated with 100 micrograms of a synthetic gonadotrophin-releasing hormone intranasal spray (HOE 766) on alternate days. In 1 patient, after an encouraging initial response there was a rapid decline in gonadotrophin responsiveness. 3 patients failed to show biochemical or clinical effects of treatment. 3 cases showed continuing responsiveness of gonadotrophin sex steroid levels to treatment for periods between 8 and 11 months.     

A miniature infuser for the pulsatile administration of LHRH

This paper describes a compact, battery-powered infuser that is light, unobtrusive and simple to operate. A novel direct-drive method is used to deliver intermittent boluses of hormone at predetermined time intervals via a special prefilled syringe. Normal pregnancy rates were achieved in anovulatory women when infused with luteinising hormone releasing hormone (LHRH).     

Dynamic alterations in luteinizing hormone-releasing hormone (LHRH) neuronal cell bodies and terminals of adult rats

Summary 1. The decapeptide lueteinizing hormone-releasing hormone (LHRH) is synthesized in neuronal cell bodies diffusely distributed across the basal forebrain and is secreted from neuronal terminals in the median eminence. Once secreted, LHRH enters the portal vessels and is then transported to the anterior pituitary, where it modulates the synthesis and secretion of gonadotropins, which are essential to gonadal function and reproduction.2. Because of the difficulties encountered in studying these diffusely distributed neurons, we have developed strategies which combine immunocytochemistry and computer-assisted techniques to examine individual LHRH neuronal cell bodies, as well as the entire population of LHRH neurons from the diagonal band of Broca to the mammillary bodies. In addition, we have examined LHRH neuronal terminals in the median eminence using computer-assisted imaging techniques to examine individual terminals by electron microscopy or across all rostral-caudal regions of the median eminence by light microscopy. In our most recent studies using confocal microscopy, we have examined the relationships of LHRH terminals to glial processes.3. These studies reveal a very dynamic system of LHRH neuronal cell bodies and terminals. The population of neurons in which LHRH can be detected varies as a function of time after gonadectomy, during the estrous cycle, and during the preovulatory surge of LH during the afternoon of proestrus. Dynamic changes are also observed in LHRH terminals in the median eminence as a function of time after gonadectomy and in specific rostral-caudal regions of the median eminence during the preovulatory surge of LH. Finally, confocal microscopy reveals that LHRH terminals are prevented from contacting the basal lamina of the brain by glial end-feet.4. We are currently examining the hypothesis that these relationships change as a function of endocrine milieu and, therefore, participate in the modulation of LHRH secretion. Ongoing studies focus on defining the sites of action and synergy of multiple sources of regulation of LHRH secretion and their relative importance to ensuring reproductive success.     

PACAP decides neuronal laminar fate via PKA signaling in the developing cerebral cortex

Laminar formation in the developing cerebral cortex requires the precisely regulated generation of phenotype-specified neurons. To test the possible involvement of pituitary adenylate cyclase-activating polypeptide (PACAP) in this formation, we investigated the effects of PACAP administered into the telencephalic ventricular space of 13.5-day-old mouse embryos. PACAP partially inhibited the proliferation of cortical progenitors and altered the position and gene-expression profiles of newly generated neurons otherwise expected for layer IV to those of neurons for the deeper layers, V and VI, of the cerebral cortex. The former and latter effects were seen only when the parent progenitor cells were exposed to PACAP in the later and in earlier G1 phase, respectively; and these effects were suppressed by co-treatment with a protein kinase A (PKA) inhibitor. These observations suggest that PACAP participates in the processes forming the neuronal laminas in the developing cortex via the intracellular PKA pathway.     

Effect of chronic administration of progesterone on the naloxone-induced LHRH release from hypothalami of ovariectomized, estradiol-primed prepubertal rats

To test the hypothesis that progesterone (P) administered in a high dose and for a long duration would influence in vitro LHRH release by naloxone, silastic capsules containing P (50 mg/ml) or vehicle were implanted sc in ovariectomized, estradiol-primed (OVX+E) prepubertal rats. Following a 48 hr exposure to P, rats were sacrificed and mediobasal hypothalamic (MBH) fragments were obtained. In addition, in other OVX+E primed rats, a single injection of P (1 mg) was given sc 6 hr prior to decapitation. Naloxone or control medium were infused into superfusion chambers containing MBH fragments derived from these animals for 2 hr following a 1 hr control period. Infusion of naloxone (1 x 10(-4) M) markedly stimulated in vitro LHRH release from MBH from rats pre-exposed to P for 48 hr, whereas it was unable to do so in other groups examined. These data clearly indicate that the ability of naloxone (1 x 10(-4) M) to stimulate LHRH release is dependent on the duration of P administration.     

The influence of hollyhock extract administration on testicular function in rats

It has been reported, recently that an aqueous extract from hollyhock flowers (Althaea rosea Cav. varietas nigra) induces weak metabolic changes in rat testes. In the present study, the in vivoinfluence of a methanolic extract was investigated on the metabolism and morphology of the rat testis. To this end, histochemical, morphometric and radioimmunological methods were used. The rats drank the extract at a dose of 100mg/day for 7weeks. The histochemical activities of glucose-6-phosphate dehydrogenase (G6PDH) and ⁵-hydroxysteroid dehydrogenase (⁵HSD) increased significantly statistically in the Leydig cells of the experimental rats in comparison with controls. There were no significant changes in either the diameter of seminiferous tubules or the height of seminiferous epithelium after hollyhock administration. Further, only a small amount of hyperplasia of the interstitial tissue was observed. The morphological and histoenzymatic changes in the Leydig cells indicate that the methanolic hollyhock extract has a direct but small influence on rat testes. The insignificant changes in testicular testosterone and estradiol content suggest that the extract does not disturb steroidogenesis.     

Portal GLP-1 administration in rats augments the insulin response to glucose via neuronal mechanisms

The incretin glucagon-like peptide-1 (GLP-1)-(7---36) amide is an important factor in prandial glucose homeostasis. Findings that GLP-1 is rapidly inactivated led to the hypothesis that the target of GLP-1 is close to the site of release. To investigate whether the target tissue is located in the hepatoportal system, we administered GLP-1 with glucose into the portal vein of rats and compared this with peripheral GLP-1 administration (jugular vein) and studied the effects of blockers of the nervous system. Portal GLP-1 augmented the insulin response to a portal glucose bolus by 81% (P < 0.01) and markedly improved the glucose disposal rate (P < 0.05). Peripheral administration of GLP-1 produced a similar augmentation of the insulin response (88%) and of the glucose disposal rate. However, only the effect of portal GLP-1 on insulin secretion was blocked by the ganglionic blocker chlorisondamine. The data suggest that prandial beta-cell stimulation by GLP-1 is evoked via a neural reflex triggered in the hepatoportal system. Because absorbed nutrients and GLP-1 first appear in the portal system, this mechanism may constitute a major pathway of GLP-1 action during meals.     

Neonatal androgenization of female rats during sex differentiation of the brain modifies neuronal organization of the amygdala complex

Sex differences in neuron dendroarchitectonics of the amygdala posterior cortical nucleus of adult rats were described for the first time using the Golgi method. Long-axon sparse-branched neurons in male rats possessed a larger number of primary dendrites, while female rats had long-axon dense-branched neurons with longer dendrites. Injection of testosterone propionate at 1250 g to females on day 5 after birth resulted in a greater number of primary dendrites of long-axon sparse branched neurons in adults, as compared to that in the control. Dendrites of long-axon sparse-branched neurons became much longer, thus enlarging the dendrite area.