Effects of gonadal steroids during pubertal development on androgen and estrogen receptor-alpha immunoreactivity in the hypothalamus and amygdala.

Perinatal development is often viewed as the major window of time for organization of steroid-sensitive neural circuits by steroid hormones. Behavioral and neuroendocrine responses to steroids are dramatically different before and after puberty, suggesting that puberty is another window of time during which gonadal steroids affect neural development. In the present study, we investigated whether the presence of gonadal hormones during pubertal development affects the number of androgen receptor and estrogen receptor alpha-immunoreactive (AR-ir and ER alpha-ir, respectively) cells in limbic regions. Male Syrian hamsters were castrated either before or after pubertal development, and 4 weeks later they received a single injection of testosterone or oil vehicle 4 h prior to tissue collection. Immunocytochemistry for AR and ER alpha was performed on brain sections from testosterone-treated and oil-treated males, respectively. Adult males that had been castrated before puberty had a greater number of AR-ir cells in the medial preoptic nucleus than adult males that had been castrated after puberty. There were no significant differences in ER alpha-ir cell number in any of the brain regions examined. The demonstration that exposure to gonadal hormones during pubertal development is associated with reduced AR-ir in the medial preoptic nucleus indicates that puberty is a period of neural development during which hormones shape steroid-sensitive neural circuits.

GnRH mRNA increases with puberty in the male Syrian hamster brain.

Puberty in the male Syrian hamster (Mesocricetus auratus) is characterized by decreased responsiveness to testosterone mediated negative feedback, but the neural mechanism for this change remains elusive. We hypothesized that decreased inhibition of the gonadotropin-releasing hormone (GnRH) system results in increased neurosecretory activity, which includes an increase in GnRH gene expression. This study examined GnRH mRNA in male hamsters before and after puberty, and sought to determine if any increase in mRNA was specific to particular subpopulations of GnRH neurones. Brains were collected from 21-day-old prepubertal males (n = 5) and 56-day-old postpubertal males (n = 5). Alternate 10 microm coronal sections from fresh-frozen brains were collected throughout the septo-hypothalamic region, and 25% of those sections were processed for in-situ hybridization histochemistry using an 35S-riboprobe complementary to hamster GnRH. No differences were observed in the number of GnRH mRNA expressing cells in any region, but in the diagonal band of Broca (DBB)/organum vasculosum of the lamina terminalis (OVLT) there was a significant increase in labelling intensity (defined as area of the cell occupied by silver grains) in postpubertal males. A second analysis compared the frequency distributions of cells based on labelling intensity between prepubertal and postpubertal males. This analysis revealed significant differences between the two frequency distributions in all areas analysed (DBB/OVLT, medial septum (MS), and preoptic area (POA)). Furthermore, examining the distribution of cells in these regions revealed a shift to the right in the postpubertal population of cells, which indicated an increased number of GnRH neurones with greater labelling intensity. These data clearly demonstrate increased GnRH mRNA during puberty. Furthermore, they suggest that the previous observation of brain region specific pubertal decreases in GnRH-immunoreactivity only within the DBB/OVLT and MS but not the POA are not due to differential levels of GnRH gene expression, but could indicate increased release from these neurones during puberty.

Androgenic regulation of hypothalamic aromatase activity in prepubertal and postpubertal male golden hamsters.

Doses of testosterone that fully activate male reproductive behavior in castrated adult male hamsters fail to elicit mounting and intromissions in prepubertal castrates, even when circulating levels of testosterone are equivalent in the two age groups. We hypothesize that this differential responsiveness to testosterone is mediated at least in part by the efficacy with which testosterone in the hypothalamus is aromatized to estradiol, an important hormone mediating male sexual behavior. Therefore, hypothalamic aromatase activity, as measured by the conversion of [3H]testosterone to [3H]estradiol in tissue homogenates, was assessed in four separate experiments: 1) intact prepubertal and adult male golden hamsters, 2 and 3) castrated adult or prepubertal males that received either a 0- or 2.5-mg dose of testosterone, and 4) castrated adult and prepubertal males treated with the 2.5-mg dose oftestosterone. These studies demonstrate that hypothalamic aromatase activity is significantly higher in adult males compared with prepubertal males, and that hypothalamic aromatase activity is increased by testosterone to the same extent in both the adult and prepubertal male hamster. Therefore, the failure of testosterone-treated castrated prepubertal male hamsters to engage in the full suite of male reproductive behaviors is not due to the inability of testosterone to be converted into estradiol in the hypothalamus. Differences in the ability of testosterone to increase aromatase activity in other brain regions, or differences in the action of testosterone and/or estradiol on other cellular processes must account for the inability of testosterone to facilitate male reproductive behavior in juvenile males.

Photoperiodic regulation of prolactin release in male hamsters with hypothalamic knife cuts.

Horizontal knife cuts placed dorsal to the paraventricular nucleus (PVN) of the hypothalamus prevent testicular regression in hamsters kept in short days. We examined the effects of these cuts on the photoperiodic modulation of the postcastration rise in gonadotropins, as well as on the release of prolactin in castrated and gonadally intact animals. The cuts blocked the inhibitory effects of short daylengths on the postcastration rise in circulating levels of gonadotropins. However, the cuts did not prevent the reduction in prolactin levels induced by short daylengths in castrated and gonadally intact animals. We conclude that dorsal connections of the PVN are not required for transduction of photoperiodic information used to regulate prolactin release. The knife cuts may remove tonic inhibitory influences on the release of follicle-stimulating hormone and luteinizing hormone, and thus produce elevated gonadotropin levels that mask the effects of nonstimulatory photoperiods on testicular size.

Immunocytochemical localization of hypothalamic luteinizing hormone-releasing hormone in male ferrets.

LHRH-containing neurons within the hypothalamus were immunocytochemically identified in adult male ferrets that were either gonadally intact, castrated, or castrated and treated with testosterone. The distribution of LHRH-immunopositive neuron cell bodies was similar in the three treatment groups. The majority of these cells was located mediobasally in the retrochiasmatic area, including some within the ventrolateral aspects of the arcuate nucleus. These soma were associated with a dense basal LHRH fiber plexus which extended to the median eminence. A smaller number of cell bodies was found slightly more dorsal and lateral to the major concentration at the base and midline. Isolated LHRH perikarya were occasionally observed in dorsal areas of the hypothalamus. There were no differences in the mean total number of hypothalamic LHRH cell bodies identified in the three treatment groups. These results indicate that the documented negative feedback effects of testosterone on LH secretion in male ferrets are not the result of an alteration in the absolute number of neurons capable of synthesizing LHRH.

Neural pathways involved in the photoperiodic control of reproductive physiology and behavior in female hamsters (Mesocricetus auratus).

Female hamsters received horizontal knife cuts to investigate the role of hypothalamic connections in the photoperiodic control of female reproductive functions. Knife cuts placed ventral to or through the paraventricular nucleus (PVN), but dorsal to the suprachiasmatic nucleus (SCN), prevented photoperiod-induced acyclicity and uterine regression in animals maintained under a nonstimulatory photoperiod for 10 weeks. The animals were then ovariectomized and tested for lordosis behavior following subcutaneous injections of ovarian hormones to investigate the photoperiodic modulation of female sexual behavior. Animals exposed to a nonstimulatory photoperiod were less behaviorally sensitive to treatment with estradiol benzoate (EB) alone, but did not differ from animals maintained under a stimulatory photoperiod when EB was combined with progesterone. The effect of photoperiod and behavioral sensitivity to hormone replacement was independent of the surgical condition. The results are consistent with the hypothesis that dorsal projections from the SCN to the PVN mediate gonadal responses to short photoperiods. They also indicate that photoperiod-induced changes in behavioral sensitivity to gonadal steroids may be mediated by neural pathways distinct from those that mediate the gonadal changes.