Periventricular Preoptic Area Research Articles

Organization of the galaninergic neuronal system in the brain of the gecko Hemidactylus frenatus

Galanin (GAL) is a 29 amino acid peptide present in the central nervous system (CNS) as well as peripheral tissues in vertebrates. However, the brain distribution pattern of GAL is understudied in reptiles. The aim of this study was to determine the organization of galaninergic neuronal system in the brain of the gecko Hemidactylus frenatus, a tropical and sub-tropical lizard, using rabbit anti-galanin antibody. In the telencephalon, GAL-ir perikarya and fibres were found in the lateral septal nucleus, but only GAL-ir fibres were observed in the striatum, nucleus accumbens, anterior commissure, nucleus centralis amygdalae, dorsal and medial septal nuclei, nucleus of the diagonal band of Broca and in the optic chiasma. In the preoptic region, a cluster of GAL-ir cells and fibres was observed in the periventricular preoptic area and lateral preoptic area. GAL-ir perikarya and fibres were observed in hypothalamic areas such as the supraoptic nucleus, suprachiasmatic nucleus, paraventricular nucleus, periventricular nucleus of the hypothalamus, infundibular recess nucleus and in the median eminence, whereas GAL-ir fibres were present in the pars distalis of the pituitary gland. In the thalamus, GAL-ir fibres were observed in the dorsomedial, dorsolateral, and medial thalamic nuclei. GAL-ir fibres were also detected in mesencephalic areas such as the optic tectum, torus semicircularis, ventral tegmental area and substantia nigra, brain stem as well as the spinal cord. The organization of GAL-ir cells and fibres throughout the gecko brain suggests several neuroendocrine, neuromodulatory and behavioural functions for GAL in lizards. The study provides new insights into the evolutionarily conserved nature of GAL peptide in squamate reptiles and forms a valuable basis for future comparative studies.

Social contact elicits immediate-early gene expression in dopaminergic cells of the male prairie vole extended olfactory amygdala

Male prairie voles (Microtus ochrogaster) are a valuable model in which to study the neurobiology of sociality because, unlike most mammals, they pair bond after mating and display paternal behaviors. Research on the regulation of these social behaviors has highlighted dopamine (DA) neurotransmission in both pair bonding and parenting. We recently described large numbers of dopaminergic cells in the male prairie vole principal nucleus of the bed nucleus of the stria terminalis (pBST) and posterodorsal medial amygdala (MeApd), but such cells were very few in number or absent in the non-monogamous species we examined, including meadow voles. This suggests that DA cells in these sites may be important for sociosexual behaviors in male prairie voles. To gain some insight into the function of these DAergic neurons in male prairie voles, we examined expression of the immediate-early genes (IEGs) Fos and Egr-1 in tyrosine hydroxylase (TH)–immunoreactive (TH-ir) cells of the pBST and MeApd after males interacted or not with one of several social stimuli. We found that IEGs were constitutively expressed in some TH-ir neurons under any social condition, but that IEG expression in these cells decreased after a 3.5-h social isolation. Thirty-minute mating bouts (but not 6- or 24-h bouts) that included ejaculation elicited greater IEG expression in TH-ir cells than did non-ejaculatory mating, interactions with a familiar female sibling, or interactions with pups. Furthermore, Fos expression in TH-ir cells was positively correlated with the display of copulatory, but not parental, behaviors. These effects of mating were not found in other DA-rich sites of the forebrain (including the anteroventral periventricular preoptic area, periventricular anterior hypothalamus, zona incerta, and arcuate nucleus). Thus, activity in DAergic cells of the male prairie vole pBST and MeApd is influenced by their social environment, and may be particularly involved in mating and its consequences, including pair bonding.

Sex differences and effects of neonatal aromatase inhibition on masculine and feminine copulatory potentials in prairie voles

Copulatory behaviors in most rodents are highly sexually dimorphic, even when circulating hormones are equated between the sexes. Prairie voles ( Microtus ochrogaster) are monomorphic in their display of some social behaviors, including partner preferences and parenting, but differences between the sexes in their masculine and feminine copulatory behavior potentials have not been studied in detail. Furthermore, the role of neonatal aromatization of testosterone to estradiol on the development of prairie vole sexual behavior potentials or their brain is unknown. To address these issues, prairie vole pups were injected daily for the first week after birth with 0.5 mg of the aromatase inhibitor 1,4,6-androstatriene-3,17-dione (ATD) or oil. Masculine and feminine copulatory behaviors in response to testosterone or estradiol were later examined in both sexes. Males and females showed high mounting and thrusting in response to testosterone, but only males reliably showed ejaculatory behavior. Conversely, males never showed feminine copulatory behaviors in response to estradiol. Sex differences in these behaviors were not affected by neonatal ATD, but ATD-treated females received fewer mounts and thrusts than controls, possibly indicating reduced attractiveness to males. In other groups of subjects, neonatal ATD demasculinized males' tyrosine hydroxylase expression in the anteroventral periventricular preoptic area, and estrogen receptor alpha expression in the medial preoptic area. Thus, although sexual behavior in both sexes of prairie voles is highly masculinized, aromatase during neonatal life is necessary only for females' femininity. Furthermore, copulatory behavior potentials and at least some aspects of brain development in male prairie voles are dissociable by their requirement for neonatal aromatase.

Middle‐Aged Female Rats Lack the Dynamic Changes in GAD67 mRNA Levels Observed in Young Females on the Day of a Luteinising Hormone Surge

The hypothalamic decapeptide gonadotrophin-releasing hormone (GnRH), modulates gonadotrophin synthesis and secretion and is essential for the preovulatory luteinising hormone (LH) surge. As females age, there is a gradual attenuation and eventual loss of the preovulatory LH surge and oestrous cyclicity. Data from previous studies have demonstrated evidence of compromised GnRH neuronal function at this time. The present study begins to explore the hypothesis that the age-related attenuation of the LH surge and decline in GnRH neuronal function are due, in part, to increased inhibitory influences on GnRH neurones. In situ hybridisation (ISH) was used to assess relative levels of mRNA for one isoform of glutamic acid decarboxylase (GAD), the rate-limiting enzyme for GABA synthesis. Ovariectomised young and middle-aged rats were injected with oestradiol benzoate and progesterone in a regimen for LH surge induction. Animals were killed at time points prior to, during the ascending phase, and during the peak and early descending phase of the LH surge. Dynamic changes in GAD(67) mRNA levels were observed in young but not middle-aged females in two regions known to be important for LH surge induction, the rostral proeptic area in the region of the organum vasculosum of the lamina terminalis (OVLT) and in the ventral periventricular preoptic area. Furthermore, GAD(67) mRNA levels were elevated in middle-aged relative to young females in the region of the OVLT at the time of LH surge induction and in the ventral periventricular preoptic area prior to surge induction. Age-related differences were not observed in other brain regions analysed. These data suggest that GABA synthesis may be elevated in middle-aged relative to young females in specific brain regions at critical times in conjunction with the LH surge, and that the lack of dynamic changes in GABA levels in these regions may contribute to the attenuated LH surge observed in middle-aged females.

Evidence of Altered Brain Sexual Differentiation in Mice Exposed Perinatally to Low, Environmentally Relevant Levels of Bisphenol A

Humans are routinely exposed to bisphenol A (BPA), an estrogenic chemical present in food and beverage containers, dental composites, and many products in the home and workplace. BPA binds both classical nuclear estrogen receptors and facilitates membrane-initiated estrogenic effects. Here we explore the ability of environmentally relevant exposure to BPA to affect anatomical and functional measures of brain development and sexual differentiation. Anatomical evidence of alterations in brain sexual differentiation were examined in male and female offspring born to mouse dams exposed to 0, 25, or 250 ng BPA/kg body weight per day from the evening of d 8 of gestation through d 16 of lactation. These studies examined the sexually dimorphic population of tyrosine hydroxylase (TH) neurons in the rostral periventricular preoptic area, an important brain region for estrous cyclicity and estrogen-positive feedback. The significant sex differences in TH neuron number observed in control offspring were diminished or obliterated in offspring exposed to BPA primarily because of a decline in TH neuron number in BPA-exposed females. As a functional endpoint of BPA action on brain sexual differentiation, we examined the effects of perinatal BPA exposure on sexually dimorphic behaviors in the open field. Data from these studies revealed significant sex differences in the vehicle-exposed offspring that were not observed in the BPA-exposed offspring. These data indicate that BPA may be capable of altering important events during critical periods of brain development.

Tyrosine hydroxylase-synthesizing cells in the hypothalamus of prairie voles (Microtus ochrogaster): Sex differences in the anteroventral periventricular preoptic area and effects of adult gonadectomy or neonatal gonadal hormones

The vertebrate hypothalamus and surrounding region contain a large population of cells expressing tyrosine hydroxylase (TH), the rate limiting enzyme for synthesis of dopamine and other catecholamines. Some of these populations are sexually dimorphic in rats. We here examined sex differences in TH-immunoreactive populations in the forebrain of gonadally intact and gonadectomized prairie voles (Microtus ochrogaster), a species that sometimes shows unusual sexual differentiation of brain and behavior. A sex difference was found in the anteroventral periventricular preoptic area (AVPV; likely analogous to the rat rostral A14) only in gonadectomized subjects, which was due to a 50% reduction in the number of TH-immunoreactive cells after castration in males. There was no significant sex difference or effects of gonadectomy on the number of TH-immunoreactive cells in the anteroventral preoptic area (AVP), periventricular anterior hypothalamus (caudal A14), arcuate nucleus (A12), zona incerta (A13), or posterodorsal hypothalamus (A11). In a second experiment, testosterone propionate (TP; 500 microg), diethylstilbestrol (DES; 1 microg), or estradiol benzoate (EB; 30 microg) injected daily during the first week after birth each significantly reduced later TH expression in the AVPV of females by approximately 40-65% compared to oil-treated controls. Unlike rats, therefore, a sex difference in TH expression in the prairie vole AVPV is found only after removal of circulating gonadal hormones in males. Furthermore, unlike our previous findings on the generation of sex differences in extra-hypothalamic arginine-vasopressin expression in prairie voles, TH expression in the AVPV of female prairie voles can be highly masculinized by neonatal exposure to either aromatizable androgens or estrogens.

Estrogen receptor beta messenger ribonucleic acid expression in the forebrain of proestrous, pregnant, and lactating female rats.

Estrogen receptor (ER)beta is present in hypothalamic and limbic neurons of female rat brains, but little is known about its regulation under physiological conditions. To determine whether ER beta expression varies during physiological conditions in which sex steroid hormone profiles are significantly different, we used in situ hybridization to assess ER beta mRNA expression in the periventricular preoptic area, bed nucleus of stria terminalis, paraventricular nucleus, supraoptic nucleus, and the posterodorsal medial amygdala of female rats on proestrus, on d 22 of pregnancy, or on d 10 of lactation (L10). In the periventricular preoptic area, d-22 pregnant females had fewer ER beta-mRNA-expressing cells than did females at proestrus, but the level of ER beta mRNA expression per cell in pregnant females was higher than in the two other groups. In the paraventricular nucleus, no changes in ER beta mRNA expression were observed; whereas in the supraoptic nucleus, proestrous females had fewer ER beta-mRNA-expressing cells than L10 females. In the posterodorsal medial amygdala, proestrous females had a greater number of ER beta-mRNA-expressing cells than did L10 females. These results demonstrate that ER beta mRNA expression is differentially regulated in a brain-region-specific and temporal manner under physiological conditions and suggest that ER beta may participate in the regulation of estrogen-sensitive reproductive functions in female rats.

Coexpression of ER with ER and Progestin Receptor Proteins in the Female Rat Forebrain: Effects of Estradiol Treatment

Estrogen and progestin receptors (ER, PgR) play a critical role in the regulation of neuroendocrine functions in females. The neuroanatomical distribution of the recently cloned, ERβ, overlaps with both ERα and PgR. To determine whether ERβ is found within ERα- or PgR-containing neurons in female rat, we used dual label immunocytochemistry. ERβ-immunoreactivity (ERβ-ir) was primarily detected in the nuclei of cells in the periventricular preoptic area (PvPO), the bed nucleus of the stria terminalis (BNSTpr), the paraventricular nucleus, the supraoptic nucleus, and the medial amygdala (MEApd). Coexpression of ERβ-ir with ERα-ir or PgR-ir was observed in the PvPO, BNSTpr, and MEApd in ovariectomized rats. E2 treatment decreased the number of ERβ-ir cells in the PvPO and BNSTpr and the number of ERα-ir cells in the MEApd and paraventricular nucleus, and therefore decreased the number of cells coexpressing ERβ-ir and ERα-ir in the PvPO, BNSTpr, and MEApd. E2 treatment increased the amount of PgR-ir in cells of the PvPO, BNSTpr, and MEApd, a portion of which also contained ERβ. These results demonstrate that ERβ is expressed in ERα- or PgR-containing cells, and they suggest that E can modulate the ratios of these steroid receptors in a brain region-specific manner.

Coexpression of ER beta with ER alpha and progestin receptor proteins in the female rat forebrain: effects of estradiol treatment.

Estrogen and progestin receptors (ER, PgR) play a critical role in the regulation of neuroendocrine functions in females. The neuroanatomical distribution of the recently cloned, ER beta, overlaps with both ER alpha and PgR. To determine whether ER beta is found within ER alpha- or PgR-containing neurons in female rat, we used dual label immunocytochemistry. ER beta-immunoreactivity (ER beta-ir) was primarily detected in the nuclei of cells in the periventricular preoptic area (PvPO), the bed nucleus of the stria terminalis (BNSTpr), the paraventricular nucleus, the supraoptic nucleus, and the medial amygdala (MEApd). Coexpression of ER beta-ir with ER alpha-ir or PgR-ir was observed in the PvPO, BNSTpr, and MEApd in ovariectomized rats. E2 treatment decreased the number of ER beta-ir cells in the PvPO and BNSTpr and the number of ER alpha-ir cells in the MEApd and paraventricular nucleus, and therefore decreased the number of cells coexpressing ER beta-ir and ER alpha-ir in the PvPO, BNSTpr, and MEApd. E2 treatment increased the amount of PgR-ir in cells of the PvPO, BNSTpr, and MEApd, a portion of which also contained ER beta. These results demonstrate that ER beta is expressed in ER alpha- or PgR-containing cells, and they suggest that E can modulate the ratios of these steroid receptors in a brain region-specific manner.

Parallel declines in Fos activation of the medial anteroventral periventricular nucleus and LHRH neurons in middle-aged rats.

The middle-age decline in reproductive function is manifested by reduced LHRH release, resulting in a decreased magnitude and delay of onset of the LH surge. Earlier studies suggested that the reductions in LHRH neural activation in middle-aged rats resulted from deficits in the afferent drive to the LHRH neurons. One critical afferent to the LHRH neurons lies in the anteroventral periventricular preoptic area (AVPv) nucleus. The neurons of the medial AVPv are synchronously activated to express Fos with LHRH neurons at the time of an LH surge in young adult animals. The present study examined whether, in middle age, reductions in the activation of AVPv neurons accompany the reduction in Fos activation in LHRH neurons. Young (3- to 4-month-old) and middle-aged (10- to 12-month-old) spontaneously cycling and ovariectomized steroid-replaced rats were killed during peak and early descending phase of the LH surge, and their brains were examined for Fos in LHRH and AVPv neurons. Young animals had a characteristic increase in Fos expression in both LHRH and AVPv neurons. In middle-aged rats, the proportion of LHRH neurons expressing Fos at the time of an LH surge was reduced by approximately 50%, irrespective of whether surges were spontaneous or induced by exogenous steroids. A similar reduction in the number of Fos+ cells (by approximately 50%) was noted in the medial AVPv. Linear regression analysis of the relationship between the extent of Fos activation in LHRH and AVPv neurons revealed a strong positive correlation (r(2) = 0.66; P < 0.01), suggesting that changes in the AVPv's drive to LHRH neurons underlie the decrease in LHRH activity in middle age. A second series of experiments examined whether decreased input from the AVPv could account for reduced Fos activation in LHRH neurons seen in middle-aged animals. When the medial AVPv was lesioned, LHRH neurons failed to express Fos on the side ipsilateral to the lesion. Animals with lesioned medial AVPv also had significantly lower LH values than animals with an intact medial AVPv. Taken together, these data suggest that a principal deficit in middle-aged rats is the ability of the medial AVPv to stimulate LHRH neurons.

Androgenic regulation of steroid hormone receptor mRNAs in the brain of whiptail lizards.

Sex and species differences in androgenic regulation of steroid hormone receptor mRNAs were examined in the diencephalon of two species of whiptail lizards: Cnemidophorus inornatus is a sexual species and the direct evolutionary ancestor to Cnemidophorus uniparens, an all-female parthenogenetic species. Lizards were gonadectomized and treated with different doses of either aromatizable testosterone or nonaromatizable dihydrotestosterone. The relative abundances of androgen-, oestrogen-, and progesterone-receptor mRNAs were compared in various nuclei following in situ hybridization with homologous riboprobes. A diversity of patterns in androgenic regulation was observed, with effects differing according to brain region, the steroid-receptor mRNA being considered and, in some cases, between androgens. In the ancestral sexual species, intact males had lower androgen-receptor mRNA abundances than castrated, blank-implanted males in the medial preoptic area. Testosterone significantly decreased androgen-receptor mRNA abundance in the medial preoptic area of castrated males. Males had higher androgen-receptor mRNA levels in the preoptic area than females generally and neither the sexual or parthenogenetic females showed a decrease in androgen-receptor mRNA with androgen treatment. Both testosterone and dihydrotestosterone increased oestrogen-receptor mRNA abundance in the ventromedial hypothalamus of C. inornatus, but no sex differences in this effect were observed. Gonadectomy decreased, whereas androgen treatment increased, progesterone-receptor mRNA abundance in the ventromedial hypothalamus. There was a sex difference in this response to androgen in the sexual species, with males having greater amounts than females in this brain area. The parthenogenetic species exhibited a similar pattern to females of the sexual species, but the levels were higher overall, possibly because Cnemidophorus uniparens is triploid. The periventricular preoptic area showed a different pattern, with testosterone treatment increasing progesterone-receptor mRNA abundance in both sexes of the sexual species and in the parthenogenetic species, while dihydrotestosterone did not. The diversity of patterns in androgen effects indicates that gonadal sex, aromatization of androgen, and perhaps gene dosage all influence the expression of steroid-receptor mRNAs in the lizard brain.

Neural populations in the rat forebrain and brainstem activated by the suckling stimulus as demonstrated by cFos expression

During lactation in the rat, the suckling stimulus plays an important role in mediating alterations in hypothalamic neuroendocrine function associated with lactation. To provide the basis for understanding the neural circuitry that may transmit suckling-induced signals into the hypothalamus, the present study used the expression of the immediate-early gene product, cFos protein, as a marker for neuronal activation to identify neural populations in the brain of lactating female rats activated by the suckling stimulus. In addition, cFos expression induced by the exteroceptive sensory stimuli (olfactory, auditory, visual) associated with pup exposure alone was also determined. Thus, cFos patterns in response to the physical suckling stimulus, which would include exteroceptive sensory stimuli associated with pup exposure, were compared with the patterns induced in response to pup exposure alone, so that neuronal populations specifically activated by the suckling stimulus could be identified. After 90 min of suckling, several forebrain areas, including the lateral septum, medial preoptic area, periventricular preoptic area and supraoptic nucleus of hypothalamus, showed a significant increase in cFos expression, compared with non-suckled controls and pup exposure animals. In addition, in the bed nucleus of stria terminalis, the medial amygdala and several cortical areas, cFos-positive cells were found in both suckling and pup exposure animals. In the brainstem, the suckling stimulus induced a significant increase in cFos expression in the ventrolateral medulla, locus coeruleus, lateral parabrachial nucleus, lateral and ventrolateral portions of the caudal part of the periaqueductal gray, and caudal portion of the paralemniscal nucleus, compared with non-suckled controls and pup exposure animals. As expected, in several areas related with sensory input, such as reticular formation and pontine nucleus, cFos expression was found in both suckling and pup exposure animals. Moreover, when double-label immunocytochemistry was used to identify cFos- and catecholamine-positive neurons in the brainstem, it was found that catecholamine-positive neurons in the ventrolateral medulla and locus coeruleus showed a significant increase in cFos expression in response to suckling compared with non-resuckled and pup-exposure groups. Using cFos expression as a marker for neuronal activation, the present studies identified the neural populations in the brain that are activated by the suckling stimulus. By comparing the pattern of cFos expression observed in response to pup exposure alone or the suckling stimulus, the present studies differentiated the neural populations activated by the physical suckling stimulus from the populations activated by the exteroceptive sensory stimuli associated with pup exposure. These suckling-activated areas are likely candidates for playing an important role in transmitting the effects of the suckling stimulus into the hypothalamus to regulate neuroendocrine alterations associated with lactation.

Hormonal regulation of progesterone receptor mRNA expression in the hypothalamus of whiptail lizards: Regional and species differences

The effects of gonadal steroid hormones on steroid receptor mRNA expression vary across nuclei within the brain, between the sexes, and between species. We report that exogenous estrogen increases progesterone receptor (PR) mRNA levels in the periventricular preoptic area in an ancestor and descendant species pair of whiptail lizards, and also that this effect of estrogen is significantly stronger in females of the descendant species. Second, while progesterone strongly decreases PR mRNA in the ventromedial hypothalamus of whiptail lizards and rodents, we find that there is no discernible effect of progesterone on PR mRNA levels in the periventricular preoptic area in females of the ancestral member of this species pair. These findings are a further demonstration of the variability of steroid effects on steroid receptor mRNA levels across brain nuclei. This variability may be important both in behavioral transitions over the course of the ovarian cycle in this ancestor-descendant species pair of lizards and in the evolution of pseudosexual behavior in the descendant parthenogen species.

Identification of neuronal input to the arcuate nucleus (ARH) activated during lactation: implications in the activation of neuropeptide Y neurons

Activation of the neuropeptide Y (NPY) neuronal system in the arcuate nucleus of the hypothalamus (ARH) during lactation in the rat is likely due to the neural impulses arising from the suckling stimulus. However, the afferent neuronal input to the ARH that is activated during lactation and is responsible for activation of NPY neurons is currently unknown. Previously, using cFos as a marker for neuronal activation, we identified several brain areas in the lactating animals that were activated by the suckling stimulus. Thus, the objective of the present study was to determine if these activated areas observed in the lactating animals project directly into the ARH. The retrograde tracer, fluorogold (FG), was injected into the ARH on day 4 postpartum. Chronically suckled rats were then deprived of their eight-pup litters on day 9 postpartum, and 48 h later, the pups were returned to the females to reinitiate the suckling stimulus for 90 min to induce cFos expression. The animals were then perfused and the brains were subjected to double-label immunohistochemistry to visualize both FG- and cFos-positive cells. Substantial FG/cFos double-labeled cells were found in forebrain regions, including the medial preoptic area, periventricular preoptic area, bed nucleus of the stria terminalis, and the medial amygdala, and in brainstem regions including the lateral parabrachial nucleus, peripeduncular area and ventrolateral medulla. The results of the present study demonstrate that specific areas in the brain are activated during lactation and send direct projections to the ARH. Thus, these areas are potentially important candidates for mediating the activation of the NPY neuronal system in the ARH during lactation.

Periventricular preoptic area neurons coactivated with luteinizing hormone (LH)-releasing hormone (LHRH) neurons at the time of the LH surge are LHRH afferents.

Earlier studies demonstrated coactivation of the periventricular preoptic area (pePOA) with LHRH neurons at the time of an induced or spontaneous LH surge, suggesting that the pePOA might regulate LHRH neurons. To investigate this hypothesis, studies were conducted to determine the temporal pattern of pePOA Fos activation during the rat estrous cycle and establish the connections of the pePOA neurons with LHRH neurons. Fos activation within LHRH and pePOA neurons showed the same temporal pattern. Both were absent during diestrous I, diestrous II, and the morning of proestrus. Fos was induced in the pePOA and LHRH neurons beginning on the afternoon of proestrus (4 h before lights off), with a decline 8 h later on proestrous evening. Tract-tracing studies then established the relationship between LHRH and pePOA neurons. Retrograde labeling with fluorogold determined that a portion of the Fos-positive pePOA neurons present at the time of the LH surge sent a projection to regions that contain LHRH cells. Anterograde tracer (neurobiotin) injections established that the pePOA neurons sent axons to the LHRH cells. Taken together, these data indicate that the pePOA provides direct input to LHRH neurons that is likely to stimulate LHRH neurons at the time of the LH surge.

Periventricular Preoptic Area Neurons Coactivated with Luteinizing Hormone (LH)-Releasing Hormone (LHRH) Neurons at the Time of the LH Surge Are LHRH Afferents

Earlier studies demonstrated coactivation of the periventricular preoptic area (pePOA) with LHRH neurons at the time of an induced or spontaneous LH surge, suggesting that the pePOA might regulate LHRH neurons. To investigate this hypothesis, studies were conducted to determine the temporal pattern of pePOA Fos activation during the rat estrous cycle and establish the connections of the pePOA neurons with LHRH neurons. Fos activation within LHRH and pePOA neurons showed the same temporal pattern. Both were absent during diestrous I, diestrous II, and the morning of proestrus. Fos was induced in the pePOA and LHRH neurons beginning on the afternoon of proestrus (4 h before lights off), with a decline 8 h later on proestrous evening. Tract-tracing studies then established the relationship between LHRH and pePOA neurons. Retrograde labeling with fluorogold determined that a portion of the Fos-positive pePOA neurons present at the time of the LH surge sent a projection to regions that contain LHRH cells. Anterograde tracer (neurobiotin) injections established that the pePOA neurons sent axons to the LHRH cells. Taken together, these data indicate that the pePOA provides direct input to LHRH neurons that is likely to stimulate LHRH neurons at the time of the LH surge.

Distribution and regulation of aromatase activity in the ram hypothalamus and amygdala

Conversion of androgen to estrogen within the brain by cytochrome P450 aromatase is a component of the system controlling the display of normal male reproductive behavior and negative feedback inhibition of LH secretion in rams. In the present study, we used the highly sensitive 3 H 2O assay to measure aromatase activity in microdissected regions of the basal diencephalon and amygdala of intact and castrated rams. We found that aromatase activity was heterogeneously distributed. The highest activity was found in the medial amygdala, cortical amygdala, and bed nucleus of the stria terminalis. Intermediate levels of aromatase activity were found within the medial preoptic area/anterior hypothalamus, periventricular preoptic area, lateral preoptic area/anterior hypothalamus, ventromedial hypothalamus, and lateral hypothalamus. The lowest activity was present in the septum, infundibulum/median eminence, and dorsal medial hypothalamus. After castration, aromatase was significantly reduced in the medial preoptic area/anterior hypothalamus, periventricular preoptic area, lateral preoptic area/anterior hypothalamus, and infundibular nucleus/median eminence. In contrast, levels in castrate rams were unchanged in several other regions, most notably the bed nucleus of the stria terminalis and medial and cortical amygdala. These results provide a quantitative profile of aromatase activity in discrete regions of the ram hypothalamus and limbic system. They also demonstrate that aromatase activity is reduced in some, but not all brain regions after castration which suggests that different regulatory mechanisms control aromatase within different neuronal populations of the ram.

Progesterone blockade of a luteinizing hormone surge blocks luteinizing hormone-releasing hormone Fos activation and activation of its preoptic area afferents

Progesterone is capable of facilitating or blocking the luteinizing hormone (LH) surge, depending on the timing of its administration. However, the precise targets of progesterone's actions are unknown. Since recent studies described the presence of a periventricular preoptic area (pePOA) neuron population afferent to LH-releasing hormone (LHRH) neurons that is co-activated to express c-Fos with LHRH neurons at the time of the LH surge, the present study was designed to determine if the pePOA neurons contain progesterone receptors (PRs) and whether progesterone inhibition is manifested by a failure of LHRH and pePOA neurons to become activated at the time of an LH surge. For progesterone facilitation, a group of immature rats each received a silastic capsule (1.57 mm i.d., 3.18 mm o.d., 1.5 cm long) containing estradiol-17 β (E 2) in peanut oil (150 μg/ml) at 09.00 h on postnatal day 28 followed 24 h later by a progesterone implant (crystalline, 1.57 mm i.d., 3.18 mm o.d., 1.5 cm long). For progesterone inhibition, a second group of rats received the estrogen capsule and a progesterone capsule (3.35 mm i.d., 4.65 mm o.d., 3.0 cm long) together at 09.00 h on day 28, and 24 h later received only a blank capsule. On the afternoon of postnatal day 29, all animals were anesthetized and perfused for localization of c-Fos and LHRH, PRs alone, or c-Fos and PRs. The present studies determined that following a progesterone-inhibition paradigm, along with blockade of the LH surge, both activation of LHRH and pePOA neurons was low or absent. Staining of PRs in progesterone-facilitated and progesterone-inhibited rats indicated that the pePOA neurons contained PRs in similar patterns. Double labeling of c-Fos and PRs in progesterone-facilitated rats indicated that nearly all the c-Fos-positive neurons of the pePOA (80±4.2%) co-expressed PRs; in progesterone-inhibited rats, only 32±12% of few c-Fos-positive neurons also contained PRs. In no instance were LHRH neurons found to contain PRs. Taken together, these data suggest that both progesterone facilitation and inhibition likely involve direct actions of progesterone on the pePOA neurons, and are consistent with a role for the pePOA neurons in transducing steroid effects on LHRH neurons.

Quantitative estimation of estrogen and androgen receptor-immunoreactive cells in the forebrain of neonatally estrogen-deprived male rats

Using quantitative immunocytochemical procedures, the total number of estrogen and androgen receptors was estimated in a large number of hypothalamic and limbic nuclei of male rats, in which brain estrogen formation was inhibited neonatally by treatment with the aromatase inhibitor 1,4,6-androstatriene-3,17-dione. The highest densities of estrogen receptor immunoreactivity were observed in the periventricular preoptic area and the medial preoptic area. Neonatally estrogen-deprived males showed a higher estrogen receptor immunoreactivity than control males in the periventricular preoptic area and the ventrolateral portion of the ventromedial nucleus of the hypothalamus, i.e. those brain areas in which sex differences have been reported, with female rats showing a greater estrogen binding capacity than male rats. The highest densities of androgen receptor immunoreactivity were found in the septohypothalamic nucleus, the medial preoptic area, the posterior division of the bed nucleus of the stria terminalis and the posterodorsal division of the medial amygdaloid nucleus. No significant differences in distribution or total numbers of androgen receptors were found between neonatally estrogen-deprived males and control males. These findings suggest that neonatal estrogens, derived from the neural aromatization of testosterone, are involved in the sexual differentiation of the estrogen receptor system in the periventricular preoptic area and the ventromedial hypothalamus. The role of neonatal estrogens in the development of the forebrain androgen receptor system is less clear.

Sex and the developing brain: suppression of neuronal estrogen sensitivity by developmental androgen exposure.

The developmental effects of androgen play a central role in sexual differentiation of the mammalian central nervous system. The cellular mechanisms responsible for mediating these effects remain incompletely understood. A considerable amount of evidence has accumulated indicating that one of the earliest detectable events in the mechanism of sexual differentiation is a selective and permanent reduction in estrogen receptor concentrations in specific regions of the brain. Using quantitative autoradiographic methods, it has been possible to precisely map the regional distribution of estrogen receptors in the brains of male and female rats, as well as to study the development of sexual dimorphisms in receptor distribution. Despite previous data suggesting that the left and right sides of the brain may be differentially responsive to early androgen exposure, there is no significant right-left asymmetry in estrogen receptor distribution, in either sex. Significant sex differences in receptor density are, however, observed in several regions of the preoptic area, the bed nucleus of the stria terminalis and the ventromedial nucleus of the hypothalamus, particularly in its most rostral and caudal aspects. In the periventricular preoptic area of the female, highest estrogen receptor density occurs in the anteroventral periventricular region: binding in this region is reduced by approximately 50% in the male, as compared to the female. These data are consistent with the hypothesis that androgen-induced defeminization of feminine behavioral and neuroendocrine responses to estrogen may involve selective reductions in the estrogen sensitivity of critical components of the neural circuitry regulating these responses, mediated in part through a reduction in estrogen receptor biosynthesis.