Identification of a sexually dimorphic neural population immunoreactive for calcitonin gene-related peptide (CGRP) in the rat medial preoptic area.

The medial preoptic area (MPOA) of the rat exhibits morphological sex differences and is implicated in sex-specific functioning and behaviour. Using immunocytochemistry, the distribution and numbers of cells containing calcitonin gene-related peptide (CGRP) were examined in the MPOA of adult male and female rats. In the intact female rat, CGRP-immunoreactive (-IR) cells were found in a continuum within the MPOA extending from the caudal aspects of the organum vasculosum of the lamina terminalis through the anteroventral periventricular nucleus (AVPv) to the region of the medial preoptic nucleus (MPN). An additional small group of CGRP-IR cells was noted at the level of the caudal MPNin the ventrolateral (VL) region. Compared with males, the AVPv and MPN regions of the female contained over 25-fold more CGRP-IR cells (P < 0.01). The VL region contained similar numbers of CGRP-IR cells in both sexes. Ovariectomy 1 month earlier, with or without subsequent 17-beta estradiol treatment, had no effect on the numbers or distribution of CGRP-IR cells in the MPOA. Gonadectomy of male rats resulted in a significant increase (P < 0.01) in the numbers of CGRP-IR cells in the AVPv and MPN regions. Subsequent administration of testosterone propionate for 1 week reduced (P < 0.05) numbers of CGRP-IR cells to levels observed in the intact male. Neurones containing CGRP in the VL group were not altered by gonadal steroid manipulation. This study shows that CGRP neurones in the AVPv/MPN region are sexually dimorphic.(ABSTRACT TRUNCATED AT 250 WORDS)     

Distribution of galanin-immunoreactive cells within sexually dimorphic components of the medial preoptic area of the male and female rat

A high percentage of galanin-immunoreactive (GAL-I) cells within sexually dimorphic components of the medial preoptic area (MPOA) of the rat also concentrate estrogen and GAL microinjected within the medial preoptic nucleus (MPN) facilitates masculine sexual behavior after testosterone priming. Thus, we determined the distribution of GAL-I cells within the MPOA and their response to gonadal steroids. We report significantly greater numbers of GAL-I cells within the central division of the medial preoptic nucleus (MPNc) and fewer within the anteroventral periventricular nucleus (AVPv), of the gonadectomized male than the gonadectomized female; that GAL-I cell numbers and densities within the AVPv are increased significantly in the intact, testosterone- or estrogen-treated male compared to the gonadectomized male and that GAL-I cell numbers and densities within the MPNc and GAL-I cell densities within the medial division of the MPN (MPNm), are increased significantly by gonadal steroids in rats of both sexez. The results suggest an involvement of galaninergic cells within the MPOA in the regulation of sexually dimorphic, gonadal steroid-sensitive functions.     

The distribution of neurotransmitter-specific cells and fibers in the anteroventral periventricular nucleus: Implications for the control of gonadotropin secretion in the rat

The anteroventral periventricular nucleus (AVPv), which lies in the periventricular zone of the preoptic region, is critical for normal phasic gonadotropin secretion since lesions of this nucleus abolish the progesterone-induced surge of luteinizing hormone secretion from the anterior pituitary, block ovulation, and induce persistent vaginal estrus in female rats. However, very little is known about the neurotransmitter-specific pathways associated with this nucleus. In the present study we evaluated the distribution of biochemically specific cells and fibers within the AVPv and adjacent regions by using an indirect immunohistochemical method with antisera to serotonin (5-HT), dopamine beta-hydroxylase (DBH), tyrosine hydroxylase (TH), neuropeptide Y (NPY), cholecystokinin-8 (CCK), vasoactive intestinal polypeptide (VIP), substance P (SP), neurotensin (NT), corticotropin-releasing factor (CRF), luteotropin-releasing hormone (LRH), somatostatin (SS), thyrotropin-releasing hormone (TRH), oxytocin (OXY), vasopressin (VAS), adrenocorticotropic hormone (ACTH1-24), alpha-melanocyte-stimulating hormone (alpha-MSH), leucine-enkephalin (L-ENK), and calcitonin gene-related peptide (CGRP). Our findings indicate that both cells and fibers containing these putative neurotransmitters are differentially distributed in and around the AVPv in accordance with the cytoarchitectonic organization of this part of the preoptic region. The AVPv itself appears to receive strong inputs from SP-, VAS-, CCK-, and SS-containing pathways, whereas the highest densities of L-ENK-, NT-, 5-HT-, NPY-, and DBH-immunoreactive fibers were found in the cell-sparse zone just lateral to the AVPv. The suprachiasmatic preoptic nucleus (PSCh), a small group of cells located ventral to the AVPv just dorsal to the optic chiasm, contained high densities of alpha-MSH- and ACTH-immunoreactive fibers, as well as substantial numbers of fibers containing catecholamines or NPY. In contrast, a dense plexus of VAS-stained fibers was distributed fairly evenly throughout the AVPv and PSCh. Numerous L-ENK-immunoreactive cell bodies, and moderate numbers of CCK-, NT-, and CRF-stained cell bodies were found in the AVPv. The PSCh contained many TH-stained cells (presumably dopaminergic), in addition to a moderate number of CCK-containing cell bodies, while a high density of NT- and CRF-stained cells were found in the cell-sparse zone lateral to the AVPv, in addition to several CCK-, SP-, VIP-, and TH-containing cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

Reversal of the sexually dimorphic distribution of serotonin-immunoreactive fibers in the medial preoptic nucleus by treatment with perinatal androgen

A small, discrete nucleus at the rostral end of the third ventricle, the anteroventral periventricular nucleus (AVPv), has been reported to be involved in the control of gonadotropin release. Since monoaminergic neurotransmitter systems have also been implicated in this function we used an indirect immunohistochemical approach to examine the distribution of 3 monoaminergic neurotransmitter systems in this nucleus. Sections through the AVPv of both colchicine and non-colchicine-treated adult male and female Sprague-Dawley rats were processed for immunohistofluorescence with antisera directed against tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DBH), or serotonin (5-HT), and were subsequently counterstained with the fluorescent Nissl stain ethidium bromide. The distributions of TH-, DBH- and 5-HT-immunoreactive neural elements within the AVPv were evaluated and a comparison was made between males and females. In both sexes, few 5-HT-stained fibers were seen within the borders of the AVPv, in contrast to the relatively high 5-HT-stained fiber density of the surrounding region. A dramatic sexual dimorphism was found in the distribution of TH-immunoreactive fibers and cell bodies. Compared to males, the AVPv in the female contained 3-4 times as many TH-stained perikarya, and a 2- to 3-fold greater density of TH-stained fibers. A low to moderate density of DBH-immunoreactive fibers, and no DBH-stained cell bodies, were seen in the nucleus. A clear sex difference was not found in the density of DBH-stained fibers in the AVPv, indicating that the sexual dimorphism in TH-immunoreactive neural elements in this nucleus is due to a greater density of dopaminergic fibers and a greater number of dopaminergic cell bodies in the female. These results suggest that dopamine may participate in the control of gonadotropin secretion at the level of the AVPv.     

Immunoreactive pro-enkephalin and prodynorphin products are differentially distributed within the nucleus of the solitary tract of the rat

In this study we examined the distribution of two different endogenous opioid peptides in the nucleus of the solitary tract of the rat medulla. As a marker for immunoreactive enkephalin, we used an antiserum directed against one of the proenkephalin products, methionine enkephalin-arg-gly-leu (m-Enk). To identify immunoreactive dynorphin we used an antiserum directed against the prodynorphin product, dynorphin B (Dyn B). The PAP method was used on both colchicine and normal animals. Caudal to the obex, within the commissural nucleus, there is extensive overlap of both immunoreactive m-Enk and Dyn B terminals and cells. While the cells are morphologically similar, the immunoreactive dynorphin cells are somewhat larger. Rostral to the obex, there is a marked difference in the distribution of the two compounds. Immunoreactive m-Enk terminals are concentrated medial to the solitary tract; there is minimal staining laterally. In contrast, immunoreactive Dyn B terminals are concentrated lateral to the solitary tract. The rostral cellular distribution of the two opioid peptides follows a similar pattern. The morphology of the medially located m-Enk and laterally located Dyn B cells is also readily distinguished. The former are small, round cells with minimal dendritic labelling; the latter are larger, pyramidal neurons with prominent apical and basal dendrites. Since the medial and lateral nuclei of the solitary tract have been associated with cardiovascular and respiratory control, respectively, these data suggest that different endorphin families have different functional actions within the nucleus of the solitary tract.     

Hormonal control of the development and regulation of tyrosine hydroxylase expression within a sexually dimorphic population of dopaminergic cells in the hypothalamus

In situ hybridization histochemistry was used to examine the development and regulation of tyrosine hydroxylase (TH) mRNA within the sexually dimorphic population of dopaminergic cells in the anteroventral periventricular nucleus (AVPv) of the hypothalamus. The AVPv contains over 3 times as many TH mRNA-containing cells in female rats, compared with males. This sexual dimorphism appears to be dependent on perinatal levels of gonadal steroids since orchidectomy of newborn males increased, and treatment of newborn females with testosterone decreased, the number of TH mRNA-containing cells detected within the AVPv. In addition, circulating gonadal steroids appear to downregulate TH expression within these cells in both adult male and female rats. In adult male animals, gonadectomy increased the number of TH mRNA cells in the AVPv within 7 days. Similarly, estradiol treatment prevented the increase in the number of TH mRNA-containing cells within the AVPv seen in ovariectomized female rats. No sexual differences were detected in the number of TH mRNA-containing cells within the suprachiasmatic preoptic nucleus, located just ventral to the AVPv. These findings indicate that perinatal gonadal steroids influence the number of cells within the AVPv that express TH in detectable amounts by determining the number of cells that are capable of producing sufficient quantities of TH message, as opposed to sex-specific alterations in the post-translational mechanisms. In the adult, circulating gonadal steroids appear to downregulate TH expression within these cells suggesting that testosterone and/or estrogen may exert a sustained influence on the biosynthetic activity of this sexually dimorphic population of dopaminergic cells.     

Coexpression of dynorphin and neurokinin B immunoreactivity in the rat hypothalamus: Morphologic evidence of interrelated function within the arcuate nucleus

Considerable evidence suggests that dynorphin and neurokinin B (NKB) neurons in the hypothalamic arcuate nucleus participate in the sex-steroid regulation of reproduction. In the present study, we used dual-label immunofluorescence to explore the distribution of prodynorphin and proNKB immunoreactivity in the rat hypothalamus. Additionally, we investigated whether arcuate prodynorphin-ir (immunoreactive) neurons expressed the neurokinin 3 receptor (NK3R) or nuclear estrogen receptor-alpha (ERalpha). We found that the majority of prodynorphin-ir neurons in the rat arcuate nucleus expressed proNKB, whereas nearly all (99%) of the proNKB neurons were immunoreactive for prodynorphin. The arcuate nucleus was the only site in the hypothalamus where neuronal somata coexpressing prodynorphin and proNKB-immunoreactivity were identified. A dense plexus of double-labeled prodynorphin/proNKB-ir fibers was found within the arcuate nucleus extending to the median eminence and throughout the periventricular zone of the hypothalamus. Prodynorphin/proNKB fibers were also identified in the paraventricular nucleus, anterior hypothalamic area, medial preoptic area, median preoptic nucleus, anteroventral periventricular nucleus, and bed nucleus of the stria terminalis in a distribution consistent with previously described arcuate nucleus projections. Interestingly, the majority of prodynorphin-ir neurons in the arcuate nucleus expressed NK3R, and nearly 100% of the prodynorphin-ir neurons contained nuclear ERalpha. Our results suggest that there is a close functional relationship between dynorphin and NKB peptides within the arcuate nucleus of the rat, which may include an autofeedback loop mediated through NK3R. The diverse hypothalamic projections of fibers expressing both prodynorphin and proNKB provide evidence that these neurons may participate in a variety of homeostatic and neuroendocrine processes.     

Immunocytochemical evidence for the presence of met-enkephalin and leu-enkephalin distinct neurons in the brain of the elasmobranch fish Scyliorhinus canicula

Immunohistochemical methods have been used to investigate the distribution of various opioid peptides derived from mammalian proenkephalin in the central nervous system of Scyliorhinus canicula. The results indicate that both Leu- and Met-enkephalin-immunoreactive peptides are present in the dogfish brain. In contrast, enkephalin forms similar to Met-enkephalin-Arg-Phe or Met-enkephalin-Arg-Gly-Leu, and mammalian α-neo-endorphin, dynorphin A (1–8), dynorphin A (1–13), and dynorphin A (1–17) were not detected. Met- and Leu-enkephalin immunoreactivities were found in distinct neurons of the telencephalon and hypothalamus. In particular, cell bodies reacting only with the Met-enkephalin antiserum were localized in the preoptic nucleus and in the suprachiasmatic region of the hypothalamus. Conversely, cell bodies reacting only with the Leu-enkephalin antiserum were localized in the pallium and the nucleus lobi lateralis hypothalami. Several areas of the telencephalon and diencephalon exhibited both Met- and Leu-enkephalin-like immunoreactivity, but the two immunoreactive peptides were clearly contained in distinct perikarya. The overall distribution of Met-enkephalin-immunoreactive elements in the dogfish brain exhibited similarities to the distribution of proenkephalin-derived peptides previously reported for the brain of tetrapods. The fact that Met- and Leu-enkephalin-like peptides were detected in distinct neurons, together with the absence of dynorphin-related peptides, suggests the existence of a novel Leu-enkephalin-containing precursor in the dogfish brain. © 1994 Wiley-Liss, Inc.     

Distribution of neuromedin U-like immunoreactivity in the central nervous system of Rana esculenta

The distribution of perikarya and nerve fibers containing neuromedin U-like immunoreactivity in the brain of Rana esculenta was determined with an antiserum directed toward the carboxyl terminus of the peptide. In the telencephalon, immunoreactive perikarya were found in the olfactory bulb, the medial septum, and the diagonal band. In the diencephalon, labeled perikarya were detected in the anterior and posterior preoptic areas, the dorsal nucleus of the hypothalamus, the caudal part of the infundibulum, and the posterior tuberculum. In the mesencephalon, immunoreactive cell bodies were found only in the laminar nucleus of the torus semicircularis and the anterodorsal tegmental nucleus. In the rhombencephalon, labeled perikarya were detected in the secondary visceral nucleus, the cerebellar nucleus, the central gray, and the nucleus of the solitary tract. Immunoreactive nerve fibers were observed in all areas of the brain that contained labeled perikarya. The densest accumulations were found in the nucleus accumbens; the dorsal part of the lateral septum; the periventricular region of the ventral thalamus; the lateral part of the infundibulum; the anterodorsal, anteroventral, posterodorsal, and posteroventral tegmental nuclei; and the periaqueductal region of the tegmentum. The distribution of neuromedin U-like immunoreactivity in the frog brain was substantially different from the distribution described for the rodent brain. © 1996 Wiley-Liss, Inc.     

Estrogen-receptive neurons in the anteroventral periventricular nucleus are synaptic targets of the suprachiasmatic nucleus and peri-suprachiasmatic region

The anteroventral periventricular nucleus (AVPv) in the rat preoptic area is a key site underlying control of the steroid dependent preovulatory gonadotropin surge. Estrogen and progesterone receptor-containing neurons in the preoptic/hypothalamic continuum, particularly those in the AVPv, are believed to transduce steroidal signals and, in turn convey this information to the LHRH system, which lacks steroid receptors. In addition to the influence of the gonadal steroids, the precise timing of the preovulatory gonadotropin surge is believed to be regulated by the hypothalamic suprachiasmatic nucleus (SCN). The SCN and peri-SCN neurons send efferent projections rostrally to the anterior preoptic area suggesting that circadian signals are communicated synaptically to steroid-responsive neurons in the AVPv. To test this hypothesis, ultrastructural double label immunocytochemistry was conducted to determine whether SCN efferents contact estrogen receptor-immunoreactive neurons in the AVPv. Brain sections with SCN injections of phaseolus vulgaris leucoagglutinin (PHA-L) were immunostained for estrogen receptors and PHA-L. Light and electron microscopic data show that the anterior preoptic area received robust PHA-L-immunoreactive efferents from SCN neurons and immediately adjacent subparaventricular zone. In particular, the AVPv contained abundant labeled fibers and terminal boutons. Ultrastructurally, SCN- and subparaventricular zone-derived terminals synaptically contacted the perikaryon of many estrogen receptor-immunoreactive neurons in the AVPv. The perikarya of unlabeled neurons were also contacted, but the majority of the labeled contacts were observed upon neuronal processes. These results demonstrate that estrogen responsive AVPv neurons are regulated by SCN efferents. Furthermore, the present data provide strong support to the idea of collective control of pituitary gonadotropin release by steroid sensitive and circadian signal neural pathways.     

Estrogen-concentrating cells within cell groups of the medial preoptic area: sex differences and co-localization with galanin-immunoreactive cells

Male and female rats have approximately equal numbers of estrogen(E)-concentrating cells within the medial preoptic area (MPOA). Several cell groups within this brain region are sexually dimorphic, however, and these groups may have sexually different numbers of E-containing cells; this, in turn, may reflect sex differences in neural-regulated functions. In order to study this possibility, the distribution of E-concentrating cells was determined using estrogen autoradiography. Except for the lateral portion of the medial preoptic nucleus (MPNl), the density of E-concentrating cells was 3-5-times higher within the most medially situated cell groups of the female than the male, i.e., within the anteroventral periventricular nucleus (AVPv), periventricular preoptic area (PVPO), medial portion of the medial preoptic nucleus (MPNm), and its central portion (MPNc). In addition, we determined whether E-concentrating cells also express the neuropeptide, galanin. An average of 13% of the E-concentrating cells were galanin positive, which represented 15% of the galanin-immunoreactive population. These results demonstrate a frank and dramatic sex difference in the distribution of E-concentrating cells within sexually dimorphic regions of the MPOA, and also suggest that an interaction between galanin and gonadal steroids may be an important means by which cells within the MPOA regulate reproductive function.     

Immunocytochemical localization of peptides and other neurochemicals in the rat laterodorsal tegmental nucleus and adjacent area

The laterodorsal tegmental nucleus (ntdl) contains a cluster of cells located just medial to the locus coeruleus in the pontine brainstem. The ntdl has been shown to project both rostrally to the forebrain and diencephalon and caudally to the spinal cord. In an effort to characterize this region neurochemically, the present study was conducted to identify a variety of neurochemicals localized within perikarya and fibers of the ntdl and surrounding nuclei. Rats were perfused with formalin, and brain sections were processed for fluorescence immunocytochemistry and acetylcholinesterase (AChE). Of the neurochemicals screened, atrial natriuretic factor (ANF), choline acetyltransferase (ChAT), cholecystokinin (CCK), calcitonin gene-related peptide (CGRP), dynorphin B (Dyn B), galanin, somatostatin, substance P, neurotensin (NT), neuropeptide Y (NPY), vasopressin, vasoactive intestinal polypeptide (VIP), serotonin (5HT), glutamic acid decarboxylase (GAD), and tyrosine hydroxylase (TH) were studied. AChE and ChAT staining revealed that the ntdl contains mostly cholinergic neurons. In addition, brightly reactive substance P and galanin and paler staining CRF, ANF, CGRP, NT, VIP, and Dyn B cell bodies were found within the ntdl. Varicose fibers in this nucleus also contained these peptides in addition to CCK, GAD, TH, 5HT, and NPY. The dorsal tegmental nucleus, dorsal raphe nucleus, locus coeruleus, and the parabrachial region contained a dense and varied assortment of peptides with distinct positions and patterns. This multiplicity of neurochemicals within this area suggests a possible influence on a variety of functions modulated by the ntdl and other closely associated tegmental nuclei.     

Prodynorphin peptide distribution in the forebrain of the Syrian hamster and rat: a comparative study with antisera against dynorphin A, dynorphin B, and the C-terminus of the prodynorphin precursor molecule

The neuroanatomical distribution of the prodynorphin precursor molecule in the forebrain of the male Syrian hamster (Mesocricetus auratus) has been studied with a novel antiserum directed against the C-terminus of the leumorphin [dynorphin B (1-29)] peptide product. C-peptide staining in sections from colchicine-treated hamsters is compared to staining in sections from untreated animals. In addition, the pattern of C-peptide immunostaining in hamster brain is compared to that in the rat brain. Finally, the C-peptide immunolabeling patterns in hamsters and rats are compared to those obtained with antisera to dynorphin A (1-17) and dynorphin B (1-13). Areas of heaviest prodynorphin immunoreactivity in the hamster include the hippocampal formation, lateral septum, bed nucleus of the stria terminalis, medial preoptic area, medial and central amygdaloid nuclei, ventral pallidum, substantia nigra, and numerous hypothalamic nuclei. Although this C-peptide staining pattern is similar to dynorphin staining reported previously in the rat, several species differences are apparent. Whereas moderate dentate gyrus granule cell staining and no CA4 cell staining have been reported in the rat hippocampal formation, intense immunostaining in the dentate gyrus and CA4 cell labeling are observed in the hamster. In addition, the medial preoptic area, bed nucleus of the stria terminalis, and medial nucleus of the amygdala stain lightly for prodynorphin-containing fibers and cells in the rat, compared to heavy cell and fiber staining in the hamster in all three of these regions. In the rat there is no differential staining between tissues processed with the C-peptide, dynorphin A, and dynorphin B antisera, but numerous areas of the hamster brain show striking differences. In most hamster brain areas containing prodynorphin peptides, the C-peptide antiserum immunolabels more cells and fibers than the dynorphin B antiserum, which in turn labels more cells and fibers than dynorphin A antiserum. However, exceptions to this hierarchy of staining intensity are found in the lateral hypothalamus, substantia nigra, arcuate nucleus, and habenula. The differences in staining patterns between rat and hamster are greatest when C-peptide antiserum is used; apparent species differences are present, though less pronounced, in dynorphin B- and dynorphin A-immunostained material.     

Characterization of big dynorphins from rat brain and spinal cord

To examine the processing of products of the dynorphin gene in the central nervous system, immunoreactive (ir) dynorphin (Dyn) A, Dyn B, Dyn A-(1-8), alpha- and beta-neo-endorphin (alpha- and beta-Neo) in rat brain and spinal cord were measured, using specific antisera after gel filtration and high-performance liquid chromatography (HPLC). Three peaks of Mr about 8, 4, and 2 kDa for ir-Dyn A and ir-Dyn B, and one peak of Mr less than 2 kDa for ir-Dyn A-(1-8), ir-alpha-, and ir-beta-Neo were found both in the brain and in the spinal cord. The 8 kDa peak was recognized by Dyn A and Dyn B antisera and, after hydrolysis by proline-specific endopeptidase, by beta-Neo antiserum. The 8 kDa peak was recognized by a monoclonal antibody against the amino terminal sequence Tyr-Gly-Gly-Phe of all opioid peptides and by an antiserum directed toward the carboxyl terminus of Dyn B, indicating that it contains, from the amino terminal tyrosine of neo-endorphin to the carboxyl-terminal threonine of Dyn B, all 3 opioid peptide regions in the prodynorphin. By means of proline-specific endopeptidase hydrolysis, we also found a big dynorphin precursor (Mr approximately equal to 26 kDa) in both brain and spinal cord.     

Postnatal development and sex difference in neurons containing estrogen receptor-α immunoreactivity in the preoptic brain,the diencephalon,and the amygdala in the rat

Estrogen has been considered as a key substance that induces sexual differentiation of the brain during fetal and neonatal life in the rat. Thus, to define the brain regions involved in the brain sexual differentiation, we examined the regions where the estrogen receptor (ER) is located in the developing rat brain. We examined immunohistochemical distribution of the cells containing estrogen receptor-α (ER-α) in the preoptic region, the diencephalon, and the amygdala in male and female rats on postnatal days 1–35 (PD1–PD35). The antibody used recognizes ER-α equally well for both occupied and unoccupied forms. ER-α immunostaining was restricted to the cell nuclei of specific cell groups. In PD1 rats, ER-α-immunoreactive (ER-IR) signals were detected in the lateral septum, the organum vasculosum lamina terminalis, the medial preoptic nucleus (MPN), the median preoptic nucleus, the bed nucleus of the stria terminalis, the hypothalamic periventricular nucleus, the lateral habenula, the posterodorsal part of the medial amygdala nucleus, the posterior part of the cortical amygdala nucleus, the hypothalamic ventromedial nucleus (VMH), the hypothalamic arcuate nucleus, and the posterior hypothalamic periventricular nucleus. The distribution pattern of ER-IR cells in the newborn rat was much the same as that in the adult in the preoptic-hypothalamic and amygdala regions. Moreover, the signals in the MPN and the VMH were stronger in the female than in the male, perhaps reflecting the ability of estrogen generated by aromatization of testosterone in the male to down-regulate the ER signal. Thus, the brain regions showing sex differences may be sites of sexual differentiation of the brain by aromatizable androgen during the neonatal period. J. Comp. Neurol. 389:81–93, 1997. © 1997 Wiley-Liss, Inc.     

Semi-quantitative analysis of the effects of estrogen on CGRP- and methionine-enkephalin-immunoreactivity in the periventricular preoptic nucleus and the medial preoptic area of female rats.

The effects of 17 beta-estradiol (E2) on calcitonin gene-related peptide (CGRP)- and methionine-enkephalin (Met-Enk)-immunoreactive (IR) nerve fibers in the periventricular preoptic nucleus (Pe) and the medial preoptic area, including the medial preoptic nucleus (MPN), of the female rat were studied semi-quantitatively by using a computer-based image analysis system. The area occupied by CGRP- and Met-Enk-IR fibers was significantly increased in the Pe and the MPN after 28-day exposure to E2. Computer-based analysis of CGRP- and Met-Enk-IR fibers in an area of 50-microns intervals from the wall of the third ventricle showed a low flat histogram pattern in ovariectomized rats, but E2 treatment caused a diphasic pattern, corresponding to the Pe and the MPN, respectively. Since the Pe and the MPN contain a high population of estrogen receptors, it is suggested that E2 may have an influence on the neuronal configuration of afferent fibers to these areas.     

Sexually dimorphic opioid distribution in the preoptic area: manipulation by gonadal steroids

A striking sexual dimorphism has been found in the density of Met-enkephalin immunoreactive fibers in the periventricular region of the preoptic area in the rat: the enkephalinergic fiber system is much denser in females. The expression of this female-typical fiber plexus is regulated by the actions of gonadal steroids both during development and in adulthood. In light of abundant evidence demonstrating the ability of the opioid peptides to modulate various sexually differentiated neuroendocrine processes and behaviors, this dimorphic system may represent an important anatomical substrate underlying these functions.     

Medial preoptic area delta-opioid receptors inhibit lordosis

Endogenous opioid peptides that activate the delta-opioid receptor (DOR) are thought to facilitate female receptive behavior. This facilitation of lordosis has been demonstrated by intracerebroventricular infusions and injection of DOR-active ligands into the ventromedial hypothalamic nucleus, an area with robust DOR binding. However, DOR binding is distributed throughout the hypothalamus, and the role of DOR in other areas of the hypothalamus has not been examined. In the current study, we demonstrated DOR immunoreactivity in the medial preoptic area (MPO), in particular medial preoptic nucleus (MPN) of the preoptic area. DOR immunoreactive processes were sparsely distributed in the medial and lateral parts of the MPN. Larger DOR immunoreactive fibers were localized in the ventrolateral aspect of the lateral MPN. The MPN is involved in the modulation of female sexual receptivity and the distribution of DOR in this area suggested to us that DOR may regulate lordosis. Ovariectomized rats with unilateral cannulae aimed at the MPN were given 5microg 17beta-estradiol benzoate (EB), once every 4 days and tested for lordosis. [D-Pen(2), D-Pen(5)]-enkephalin (DPDPE), a DOR agonist, microinfused into the MPO, 52-54h after EB-priming, inhibited lordosis when compared with the aCSF (vehicle) control (P <== 0.05). The inhibitory effects of DPDPE were reversed by microinjection of naltrindole, a DOR antagonist (P <== 0.05). Interestingly, the DOR inhibition of lordosis is similar to the micro-opioid receptor inhibition of lordosis in the MPN. These results indicate that DOR in the MPO, particularly in the MPNm, plays an important role in the regulation of lordosis.     

Distribution of GAD-immunoreactive neurons in the diencephalon of the african lungfish Protopterus annectens: colocalization of GAD and NPY in the preoptic area

The distribution of GABAergic neurons was investigated in the diencephalon of the African lungfish, Protopterus annectens, by using specific antibodies directed against glutamic acid decarboxylase (GAD). A dense population of immunoreactive perikarya was observed in the periventricular preoptic nucleus, whereas the caudal hypothalamus and the dorsal thalamus contained only scattered positive cell bodies. Clusters of GAD-positive cells were found in the intermediate lobe of the pituitary. The diencephalon was richly innervated by GAD-immunoreactive fibers that were particularly abundant in the hypothalamus. In the periventricular nucleus, GAD-positive fibers exhibited a radial orientation, and a few neurons extended processes toward the third ventricle. More caudally, a dense bundle of GAD-immunoreactive fibers coursing along the ventral wall of the hypothalamus terminated into the median eminence and the neural lobe of the pituitary. Double-labeling immunocytochemistry revealed that GAD and neuropeptide tyrosine (NPY)-like immunoreactivity was colocalized in a subpopulation of perikarya in the periventricular preoptic nucleus. The proportion of neurons that coexpressed GAD and NPY was higher in the caudal region of the preoptic nucleus. The distribution of GAD-immunoreactive elements in the diencephalon and pituitary of the African lungfish indicates that GABA may act as a hypophysiotropic neurohormone in Dipnoans. The coexistence of GAD and NPY in a subset of neurons of the periventricular preoptic nucleus suggests that GABA and NPY may interact at the synaptic level.     

Sexually dimorphic distribution of neurotensin/neuromedin N mRNA in the rat preoptic area

Neurotensin release from estrogen-responsive neurons in the rostral preoptic area of the female rat may play an important role in triggering preovulatory secretion of gonadotropin-releasing hormone on proestrus. We investigated the possibility of sexually differentiated biosynthesis of neurotensin in the rostral preoptic area, using in situ hybridization histochemistry to detect neurotensin/neuromedin N (NT/N) mRNA in adult male rats and adult female rats at proestrus and the first day of diestrus. In sections through the anteroventral periventricular nucleus (AVPv), the number of labeled cells in proestrous females was four times that in males. Diestrus females exhibited half the number of labeled cells present at proestrus, and there was evidence for a significant correlation between circulating estradiol level and number of labeled cells in the AVPv. In the rostral portion of the medial preoptic nucleus (MPN), two contiguous groups of labeled cells were especially prominent. One group, in the medial half of the MPN, was located closer to the midline in females than in males and displayed greater labeling in males than in females. Furthermore, labeling in the rostral MPN was greater at proestrus than at diestrus. These results indicate that biosynthesis of neurotensin and neuromedin N in the rostral preoptic area may be sexually differentiated and, in the female, may vary across the estrous cycle in parallel with circulating estradiol levels, consistent with the view that neurotensin neurons in this area are involved in the regulation of preovulatory secretion of gonadotropin-releasing hormone. The sex- and region-specific expression of NT/N mRNA in the rostral preoptic area suggests functional heterogeneity of neurotensin neuronal populations in this area and implies complex regulation of NT/N gene expression in the rat brain.