Immunohistochemical colocalization of GABA and insulin in beta-cells of rat islet.

gamma-Aminobutyric acid (GABA) is found in high concentrations in the pancreatic islet. In addition, enzymes regulating the level of GABA (L-glutamate decarboxylase and GABA-alpha-ketoglutarate transaminase) have been immunohistochemically localized in the medullary cells of the islet. In this study, an immunofluorescence and elution/restaining protocol is used to determine the distribution of GABA and either insulin, glucagon, or somatostatin in a tissue section. GABA was not detected within the islet alpha- or delta-cells but was determined to be localized within the insulin-containing beta-cells.

Colocalization of galanin and prolactin within secretory granules of anterior pituitary cells in estrogen-treated Fischer 344 rats.

Galanin is localized within specific cell types of the rat anterior pituitary gland (AP). Immunocytochemical studies at the light microscope level have shown that lactotrophs, somatotrophs, and thyrotrophs contain galanin in the intact female rat, whereas lactotrophs in the male AP do not. We recently reported that galanin and PRL release from estrogen-treated male and female pituitary cells in culture are coregulated by dopamine, TRH, and somatostatin. This suggested that galanin is stored within secretory granules, conceivably with PRL. Using postembedding immunocytochemistry at the ultrastructural level, the objectives of this study were to: 1) determine the subcellular location of galanin in the AP; 2) elucidate if galanin and PRL are colocalized within the same secretory granules; and 3) compare the cellular localization of galanin in the male and female AP. Male and ovariectomized female (OVEX) Fischer 344 rats were implanted with estradiol-containing or empty Silastic capsules for 2 weeks. Postembedding immunogold labeling was performed using rabbit (for galanin) and guinea pig (for PRL) generated antisera. Two different sizes of colloidal gold spheres were used to localize the hormones in the same tissue section. Galanin was primarily localized in secretory granules of adenohypophyseal cells. Based upon immunocytochemical results and morphological criteria, galanin was contained in somatotrophs but not lactotrophs in the male and OVEX AP. The AP of estrogen-treated rats contained more specific immunogold labeling for galanin than untreated rats. The increased immunoreactivity for galanin was notably associated with lactotrophs. After exposure to estrogen, galanin and PRL were colocalized within the same secretory granules of the male and OVEX pituitary cells. We conclude: 1) galanin is localized within secretory granules of the rat AP; 2) galanin and PRL are colocalized within secretory granules of the male and OVEX AP after estrogen treatment; and 3) galanin is localized in similar cell types in the male and OVEX AP, before and after estrogen treatment. These data provide a morphological basis for the coregulation of galanin and PRL secretion by hypothalamic factors.

Colocalization of prolactin and growth hormone within specific adenohypophyseal cells in male, female, and lactating female rats.

The localization of PRL and GH within adenohypophysial cells has been investigated with immunocytochemical methodology using colloidal gold of different sizes. Classically, using morphological criteria at the light and electron microscope levels, two types of individual cells have been described which, it is believed, exclusively produce either PRL or GH, i.e. mammotrophs and somatotrophs. Since it has been reported that some gonadotrophs may colocalize and secrete both FSH and LH, and that some unidentified cells release both PRL and GH, we designed a study to investigate whether PRL and GH are present only in their respective specific cells or may be colocalized in mammotrophs and somatotrophs or possibly other types of cells within the pars distalis. Using immunocytochemistry at the electron microscope level we were able to label the two hormones with different sizes of colloidal gold bound to a second antibody and visualize them within pituitary cells. Two different primary antibodies to PRL and GH as well as a more purified antibody to GH were used. Pituitaries from cycling and lactating female rats and adult males were processed appropriately for electron microscopic studies. After sectioning, individual grids were treated with either primary antibodies for PRL or GH, or the same grid was treated sequentially with these two antibodies. All primary antibodies were absorbed with the heterologous hormone before usage. The second antibodies were bound to colloidal gold particles of either 10 nm (for GH) or 20 nm (for PRL) diameter, so that selective visualization of the two hormones could be achieved within the same cells. It was observed that mammotrophs immunolabeled only for PRL, and somatotrophs labeled only for GH regardless of the source of the antibody. However, an atypical small granule cell, a possible mammosomatotroph, colabeled consistently for both PRL and GH in all types of animals used and with antibodies from all of above sources. This report gives for the first time morphological evidence for the existence of pituitary cells that colocalize both PRL and GH in the normal rat pituitary gland. The possibility that these bihormonal cells represent stem cells which may give rise to both mammotrophs and somatotrophs under appropriate stimulation or that they may release both hormones under the influence of unspecific stimuli is suggested.

Ultrastructural evidence for luteinizing hormone-releasing hormone neuronal control of estrogen responsive neurons in the preoptic area.

Both estrogen receptor (ER) immunoreactive (ir) and LHRH-ir neurons and processes are present in the preoptic area of the guinea pig. This experiment was conducted to determine if LHRH-ir terminals interact synaptically with ER-ir cells. A light microscopic dual chromogen immunocytochemical technique employing diaminobenzidine (DAB) and nickel-enhanced DAB for LHRH and ER localization, respectively, revealed that many varicose LHRH-ir fibers coursed in close proximity to ER-ir cells in the anterior part of the preoptic area at the preventricular periventricular nucleus (Pep), suggesting the likelihood of synaptic interactions. Ultrastructural analysis was performed using DAB and 3,3',5,5'-tetramethylbenzidine (TMB) for LHRH and ER localization, respectively. DAB labeling in LHRH-ir neurons appeared as a dense flocculent product dispersed throughout the cytoplasm. TMB stained ER-ir neurons contained electron dense crystalline spicules located predominantly in their nuclei. Numerous TMB labeled ER-ir neurons were present in the Pep, and occasionally occurred in clusters, closely apposed to one another. Many LHRH-ir terminals made synaptic contact or were apposed to unlabeled dendrites, while fewer contacted perikarya. Most significantly, ER-ir neurons showing clear evidence of intranuclear TMB crystals received synaptic input from LHRH-ir terminals. In addition, LHRH-ir terminals lacking synaptic specializations were also in direct apposition to ER-ir perikarya. These results provide morphological evidence that LHRH-ir neurons can regulate ER-ir neurons in the preoptic area. Since LHRH-ir cells are thought to be regulated by estrogen responsive neurons, interaction of LHRH terminals with ER-ir cells may represent a regulatory feedback circuit between the two systems.

Galanin within the normal and hyperplastic anterior pituitary gland: localization, secretion, and functional analysis in normal and human growth hormone-releasing hormone transgenic mice.

Studies evaluating estrogen-induced anterior pituitary tumors revealed a strong direct correlation between expression of the peptide galanin and tumor growth. To evaluate further the potential roles of galanin in the hyperplastic pituitary, we used a model of estrogen-independent anterior pituitary tumor formation, the male human GH-releasing hormone (hGHRH) transgenic mouse. Pituitaries of hGHRH transgenic mice are characterized by a hyperplasia of somatotrophs and contain markedly elevated levels of galanin. We examined the population of galanin-producing pituitary cells in 4- to 6-month-old male hGHRH transgenic mice and their nontransgenic siblings. The percentage of galanin-containing pituitary cells was significantly increased within the anterior pituitaries of hGHRH transgenic mice. By using the cell immunoblot assay we found that the basal secretion of galanin and GH from individual pituitary cells of hGHRH transgenic mice was significantly greater than that from pituitary cells of nontransgenic mice. By modifying the cell immunoblot assay, we determined that somatotrophs from both hGHRH transgenic and normal mice that were positive for galanin immunoreactivity secreted significantly greater amounts of GH than those somatotrophs devoid of galanin immunoreactivity. Moreover, immunoneutralization of galanin significantly decreased GH secretion from pituitary cells obtained from hGHRH transgenic mice. Thus, we now show that the increased levels of galanin peptide within the hyperplastic pituitaries of hGHRH transgenic mice are due to an increase in the population of cells containing galanin, and that galanin participates in the augmented secretion of GH from hyperplastic proliferating pituitary cells.

Ornithine decarboxylase activity and immunohistochemical location in postischemic brain.

Ornithine decarboxylase, rate-limiting in polyamine formation, has been found to be necessary for the development of vasogenic edema after cryogenic cerebral injury and is postulated to be of importance in late ischemic brain edema formation. Ornithine decarboxylase activity and accompanying edema was studied after transient cerebral ischemia in Mongolian gerbils. Bilateral carotid artery occlusion was utilized to produce dense forebrain ischemia. After 4 h of reperfusion a significant elevation in ornithine decarboxylase activity was present (72.5 +/- 24.7 vs 8.5 +/- 2 pmoles/mg protein/h, p less than 0.05). Immunohistochemical localization of ornithine decarboxylase indicated its presence in cortical neurons of ischemic gerbils. This was typically located in the perinuclear cytoplasm and extended into proximal dendrites. Nonischemic animals did not contain ornithine decarboxylase immunoreactivity. These studies show the presence and location of ornithine decarboxylase in cerebral tissue subjected to transient ischemia. The increase in this marker of polyamine activity paralleled previous studies in this model of cerebral edema formation and reperfusion deficit in blood flow and evoked potential, suggesting that ornithine decarboxylase is a marker for and may be associated with those late metabolic events leading to progressive functional deterioration after incomplete cerebral ischemia.

The ultrastructural localization of enkephalin and substance P immunoreactivities in the nucleus tractus solitarii of the cat.

In the medial and commissural subdivisions of the nucleus tractus solitarii enkephalin and substance P immunoreactivities were localized within synaptic terminals, unmyelinated axons, and neuronal cell bodies. Both enkephalin and substance P immunoreactivities were contained within synaptic terminals which had a mixture of small clear vesicles and dense core vesicles. The presence of dense core vesicles within both the enkephalin- and substance P-immunoreactive terminals was a consistent feature, although they were not associated with the actual synaptic junction. While enkephalin- and substance P-immunoreactive terminals shared a similar morphology, their respective distributions along the dendritic tree were quite distinct. Enkephalin-immunoreactive terminals contacted mainly the cell body and proximal portions of the dendritic tree. In contrast, substance P-immunoreactive terminals synapsed predominantly with spines and shafts of small to medium-sized dendrites. Few substance P-immunoreactive terminals contacted proximal dendrites and they were never presynaptic to the neuronal cell body. This apparent segregation of synaptic terminals on neurons suggests that enkephalin synapses have a more pronounced effect than substance P terminals.

Early development of the inferior olivary complex in pouch young opossums. II. An electron microscopic study.

At birth the inferior olivary complex (IOC) is not present in the caudal ventro-medial brainstem of the opossum. In the 3-7-day-old animal (15-19 days post-conception), this same region does contain neurons of the developing IOC. The immature neurons are characterized by large, centrally placed nuclei surrounded by a thin rim of cytoplasm. The neuropil contains numerous small-diameter profiles which contain bundles of filaments and scattered microtubules. Occasional synaptic endings, containing round clear vesicles, contact large, flocculent profiles. By 10-14 days of age, the olivary complex begins to separate into individual nuclei; however, the olivary cell bodies and the surrounding neuropil exhibit many of the same features as in the 3-7-day-old opossums. In opossums 21-25 days old, there is an increase in varicosities and irregular contours along many of the dendritic shafts. Furthermore, synaptic terminals, possessing round clear vesicles, now contact the soma, dendritic shafts, dendritic varicosities, spines, and large, flocculent profiles. Terminals containing pleomorphic vesicles or a mixture of clear and large granular vesicles are presynaptic only to dendritic spines or large, flocculent profiles. Neuroglial cell bodies have been identified at all ages examined. It is not until days 65-68 that pre- and postsynaptic elements are organized into synaptic clusters (glomeruli), which are typical of the adult. Spiny appendages and small-diameter dendrites comprise the central core of the clusters which are surrounded by synaptic endings containing a variety of vesicle types. Thus it would appear that subsequent to their initial arrival (day 16-17), the synaptic relationships of cerebellar and midbrain afferents are modified to reflect their adult configuration by days 65-68. This extended period of development (postnatal days 3-68) for the olivary complex provides a good model for assessing the effects of experimental manipulations.

Distribution of [3H]quinuclidinyl benzilate, [3H]nicotine, and [125I]alpha-bungarotoxin binding sites in the nucleus tractus solitarii of the cat.

The distribution of muscarinic and nicotinic cholinergic binding sites in the cat nucleus tractus solitarii was studied by the technique of in vitro autoradiography. Using the antagonist [3H]quinuclidinyl benzilate, muscarinic binding sites were differentially located in subdivisions of the nucleus tractus solitarii. The majority of muscarinic binding sites were located predominantly in the caudal half of the nucleus, reaching their greatest amounts at the mid levels of the nucleus tractus solitarii. The medial, dorsolateral, intermediate, and interstitial subdivisions contained the highest densities of quinuclidinyl benzilate binding sites. Nicotinic cholinergic binding sites, using [3H]nicotine and [125I]alpha-bungarotoxin, had unique patterns of distribution. With [3H]nicotine the majority of binding sites were located in rostral levels of the nucleus with very few binding sites present in the caudal half. In contrast, [125I]alpha-bungarotoxin binding sites were present mainly in subdivisions located in the caudal half of the nucleus, i.e., commissural, ventrolateral, dorsolateral, medial, and intermediate subdivisions, and dropped off precipitously at more rostral levels. The differential distribution of [3H]nicotine and [125I]alpha-bungarotoxin suggests the two ligands may be labeling different types of nicotinic binding sites in the nucleus tractus solitarii. The unique distribution of muscarinic and nicotinic cholinergic binding sites in the various subdivisions of the nucleus solitarii suggests that muscarinic and nicotine mechanisms may play an active role in the regulation of the diverse autonomic functions at the level of the nucleus tractus solitarii.

A comparison of GABA- and GAD-like immunoreactivity within the area postrema of the rat and cat.

The immunohistochemical localization of gamma-aminobutyric acid (GABA) was compared to that of glutamate decarboxylase (GAD) in the rat and cat area postrema with the aid of a polyclonal antibody produced in rabbits directed against GABA. In both the rat and cat, dense and very dense accumulations of GABA-like immunoreactive (GABA-LI) varicosities were present throughout the area postrema. GABA-LI cell bodies were present in both species and were evenly distributed throughout the area postrema's extent. However, the rat area postrema contained more GABA-LI cell bodies and varicosities than the cat area postrema. In the cat area postrema, a range of cell sizes were immunostained with the GABA antibody. The GAD antibody, however, failed to reveal cell bodies in the area postrema of the cat, thus indicating that the GABA polyclonal antibody may be a better indicator of GABA-containing somata. Although the mechanism of action of GABA in the area postrema is not understood, it is possible that GABA may play a role in the different functions of the area postrema in emetic and nonemetic species.

The synaptic terminations of certain midbrain-olivary fibers in the opossum.

The nuclear origin and distribution of midbrain-olivary fibers has been described in a previous study utilizing axonal transport techniques (Linauts and Martin, '78a). The present report extends their results to the electron microscopic level and details the postsynaptic distribution of such fibers. Lesions within the ventral periaqueductal grey and adjacent tegmentum, the red nucleus or the nucleus subparafascicularis result in electron dense axon terminals within the olive at survival times of 48, 72 and 96 hours. At 72 hours, many degenerating presynaptic profiles shrink, become irregular in shape and are totally or partially surrounded by glial processes. The principal olivary nucleus contains the majority of these profiles. However, the subparafascicular terminals are more abundant in the rostral and intermediate parts of the medial accessory nucleus and the rubral terminals are concentrated within the dorsal lamella of the principal nucleus. The nuclear location of the degenerating terminals was determined by examination of 1 micrometer plastic sections cut in the transverse plane from each block face prior to thin sectioning. Degenerating terminals were counted in three cases, one from each of the three lesion sites described above. When taken together these cases show that just over 50% of the degenerating terminals are presynaptic to spiny appendages and are located within the synaptic clusters (glomeruli) described previously (King, '76). The percentage of degenerating terminals in the glomeruli increases to 70% when the lesion is in the ventral periaqueductal grey and adjacent tegmentum. The remaining degenerating terminals contact dendritic shafts outside the astrocytic boundaries of the synaptic clusters. The synpatic vesicle populations within the degenerating terminals vary with the location of the lesion. Lesions in the ventral periaqueductal grey and the adjacent tegmentum result in the degeneration of terminals with either clear spherical vesicles or endings with both clear spherical vesicles and a variable number of large dense core vesicles. In contrast, the primary degenerative changes that occur after destruction of the red nucleus or the nucleus subparafascicularis are in terminals with clear spherical vesicles. When the synaptic complex was present in the plane of section, regardless of the site of the lesion, the degenerating terminals could be classified as Gray's type I. Thus, we have demonstrated that afferents from the mesencephalon terminate within synpatic clusters located in the principal and medial accessory (part A) subnuclei of the inferior olive. Although the mesencephalic afferents have multiple origins (Linauts and Martin, '78a), many of their synaptic terminals contact spiny appendages within the synaptic clusters. This postsynaptic site also receives cerebellar terminals (King et al., '76). The origin of presynaptic profiles within the synaptic clusters that contain clear pleomorphlic vesicles is yet to be determined.

Immunohistochemical localization of substance P and enkephalin in the nucleus tractus solitarii of the rhesus monkey, Macaca mulatta.

The nucleus tractus solitarii in the monkey Macaca mulatta was found to have several subdivisions based upon cytoarchitectonics and immunohistochemistry. Subdivisions that could be identified included commissural, medial, parvicellular, dorsolateral, ventrolateral, intermediate, and interstitial. Substance P and enkephalin immunoreactivity was localized within discrete regions of the nucleus tractus solitarii, by means of the peroxidase-antiperoxidase technique. Substance P immunoreactivity occurred most frequently in the interstitial subdivision of the nucleus tractus solitarii. Moderate accumulations of substance P immunoreactivity were present in the commissural, medial, parvicellular, dorsolateral, and intermediate subdivisions, but very little was present in the ventrolateral subdivision. Enkephalin immunoreactivity followed the staining patterns of substance P; however, the amounts of enkephalin immunoreactivity were less than amounts for substance P. Following colchicine treatment, large numbers of enkephalin-immunoreactive neurons were distributed throughout all subdivisions, many being located in the parvicellular and medial subdivisions. The few substance P-immunoreactive neurons found were restricted to the parvicellular subdivision. The distribution of substance P and enkephalin immunoreactivity in M. mulatta is very similar to that described in the cat and rat. In addition, the extensive overlap of the distribution of these two putative neurotransmitters provides morphological evidence for their possible participation in the autonomic regulation within the nucleus tractus solitarii.

Vasoactive intestinal polypeptide mRNA and peptide levels are decreased in the anterior pituitary of the human growth hormone-releasing hormone transgenic mouse.

We recently reported that galanin gene expression is markedly increased in the hyperplastic anterior pituitary gland of the human growth hormone-releasing hormone (hGHRH) transgenic mouse. To determine if another pituitary peptide hormone with putative growth-promoting activity is similarly affected, or if this effect is specific to the peptide galanin, we examined vasoactive intestinal polypeptide (VIP) gene expression in the hypothalamic-pituitary axis of male hGHRH transgenic and non-transgenic mice. The objectives were to: 1) assess VIP peptide concentrations, 2) estimate relative differences in VIP mRNA levels, 3) determine the effects of acute treatment with 17beta-estradiol on VIP peptide and mRNA levels, and 4) quantify the density of immunoreactive VIP pituitary cells by immunohistochemistry. Four to five month old male hGHRH transgenic mice and their non-transgenic siblings were identified by PCR. Immunoreactive VIP concentrations were decreased by 50% in the anterior pituitary glands of hGHRH transgenic mice as compared to non-transgenic siblings. In contrast, no differences in immunoreactive VIP concentrations were observed in the hypothalamus or frontal cerebral cortex of transgenic and non-transgenic mice. Treatment with 17beta-estradiol significantly increased VIP concentrations in the anterior pituitary gland of both transgenic and non-transgenic mice; however, VIP peptide concentrations in the anterior pituitary glands of hGHRH transgenic mice remained 50% lower. Relative differences in VIP mRNA levels were estimated by RT-PCR, and were found to be 2.5-fold higher in the anterior pituitary glands of non-transgenic mice. In contrast, no differences in VIP mRNA levels in the cerebral cortex were detected between transgenic and non-transgenic mice. Treatment with 17beta-estradiol increased VIP mRNA levels in the anterior pituitary, but not in the cerebral cortex. In concert with the changes in VIP peptide and mRNA, the density of immunoreactive VIP pituitary cells was decreased approximately 50% in hGHRH transgenic mice. In conclusion, unlike galanin gene expression, VIP peptide and mRNA levels are significantly decreased in the anterior pituitary gland of hGHRH transgenic mice. Moreover, these changes appear to be tissue-specific and are likely due, in part, to the decrease in the density of VIP-containing pituitary cells in the hyperplastic pituitary. Although the pituitary cell type(s) synthesizing VIP remains unclear, these data suggest that VIP in the anterior pituitary is not stimulating pituitary tumor development in hGHRH transgenic mice.

Distribution of neurotensin-like immunoreactivity in rat and cat area postrema.

The distribution of neurotensin-like immunoreactive (NT-LI) fibers and cell bodies in the area postrema (AP) of rat and cat, utilizing the peroxidase, antiperoxidase (PAP) technique, is described. In the rat, the greatest accumulation of NT-LI fibers were present along the borders of the AP, while there were very few NT-LI fibers in central regions. In the cat, scattered NT-LI fibers occupied the majority of the central AP, while moderate numbers of NT-LI fibers were present at the ventromedial border. In more rostral cat AP levels, the number of NT-LI fibers decreased. NT-LI somata were present in rat AP, but were lacking in cat AP. The localization of NT within the AP suggests that the cardiovascular and gastric effects attributed to NT may be mediated, in part, through the AP.

Substance P binding sites in the nucleus tractus solitarius of the cat.

Substance P binding sites in the nucleus tractus solitarius were visualized with receptor autoradiography using Bolton-Hunter [125I]substance P. Substance P binding sites were found to have distinct patterns within the cat nucleus tractus solitarius. The majority of substance P binding sites were present in the medial, intermediate and the peripheral rim of the parvocellular subdivisions. Lower amounts of substance P binding sites were present in the commissural, ventrolateral, interstitial and dorsolateral subdivisions. No substance P binding sites were present in the central region of the parvocellular subdivision or the solitary tract. The localization of substance P binding sites in the nucleus tractus solitarius is very similar to the patterns of substance P immunoreactive fibers previously described for this region. Results of this study add further support for a functional role of substance P in synaptic circuits of the nucleus tractus solitarius.

Cholecystokinin octapeptide immunoreactivity in the nucleus tractus solitarius of the cat.

The nucleus tractus solitarius possessed distinct patterns of cholecystokinin immunoreactive fibers and cell bodies within its various subdivisions. The commissural, medial, intermediate, parvocellular, dorsolateral and interstitial subdivisions contained relatively dense amounts of CCK immunolabelled fibers. In contrast, CCK immunoreactivity within the ventrolateral subdivision consisted of a few scattered fibers and small neurons. The commissural, intermediate, medial, dorsolateral and parvocellular subdivisions contained CCK immunoreactive neurons following colchicine treatment. The presence of CCK in the NTS suggest that it may be involved as a neuromodulator and/or neurotransmitter in circuitry that mediate cardiovascular, respiratory, gastrointestinal and taste functions.

Cholecystokinin-octapeptide like immunoreactivity in the area postrema of the rat and cat.

The distribution of cholecystokinin-8 (CCK-8)-like immunoreactivity in the area postrema of the rat and cat was visualized using the peroxidase, antiperoxidase technique. In the rat the greatest amount of immunostaining occurred in peripheral regions of the area postrema at intermediate and rostral levels. Caudally, scattered immunoreactivity predominated. After colchicine treatment, numerous immunoreactive somata were observed throughout the area postrema. The cat area postrema had a different and more complex pattern of immunostaining than the rat. Moderate to dense accumulations of immunostaining occurred in the ventromedial region of the area postrema bordering the solitary tract and dorsal vagal nuclei. The central region of the area postrema possessed scattered amounts of immunoreactivity at rostral levels. Following colchicine treatment, no visible CCK-8-like immunoreactive cell bodies were observed in the cat area postrema. Results of the present investigation provide morphological evidence for the role of CCK-8 in cardiovascular regulation and satiety. The difference in the distribution of CCK-8 in the rat and cat suggest a possible role in the emetic reflex.

Enkephalin-immunoreactive neurons in the nucleus tractus solitarius project to the parabrachial nucleus of the cat.

Enkephalin immunoreactive neurons within the nucleus tractus solitarius (NTS) were found to project to the parabrachial nucleus of the cat with the use of a combination of immunohistochemistry and retrograde transport of horseradish peroxidase. Double labelled neurons were located in the medial, parvocellular and commissural subdivisions of the NTS and were present predominantly ipsilateral to the injection site within the parabrachial nucleus. Only a few double labelled neurons were found in the contralateral NTS. The presence of neurons containing enkephalin immunoreactivity suggests that the role of enkephalin in the regulation of autonomic functions may be, in part, by circuits between the NTS and the parabrachial nucleus.

Estrous cycle-associated axosomatic synaptic plasticity upon estrogen receptive neurons in the rat preoptic area.

This study examined the hypothesis that synaptic relationships change on a daily basis in the anterior preoptic area of the intact, cycling female rat. Specifically, the anteroventral periventricular nucleus (AVPv) was chosen for analysis due to its abundant estrogen- and progesterone-receptive neurons and its critical role in the control of gonadotropin secretion. Ultrastructural analysis revealed that in the 24 h between proestrus and estrus, there was a 39% increase in axosomatic synapses upon AVPv neurons. In the subsequent 24 h to metestrus, the number of synapses decreased by 22%. Additional data showed that ovariectomy resulted in more axosomatic synapses in the AVPv relative to proestrus (46%) and metestrus (35%). Another component of the study investigated synaptic relationships on estrogen receptor-immunoreactive (ER-ir) and non-ER-ir neurons across the estrous cycle. ER-ir neurons received significantly more synaptic input at proestrus and estrus than did non-ER-ir neurons. At metestrus and following ovariectomy, no significant differences were present. The present study indicates that estrous cycle-associated synaptic plasticity occurs in the AVPv and lends further support to the critical role of this nucleus in regulation of gonadotropin secretion. Moreover, it provides the first evidence that ER- and non-ER-ir neurons are differentially innervated.

Immunohistochemical demonstration of serotonin neurons in autonomic regions of the rat spinal cord.

The immunohistochemical distribution of serotonin neurons in normal and transected spinal cords of rats was examined. Intraspinal serotonin neurons were immunostained as far rostral and caudal as T3 and Co1, respectively. All serotonin neurons were located in lamina VII and X, and most were located in spinal autonomic areas. Both bipolar and multipolar neurons were observed with many of the neurons oriented longitudinally to the long axis of the cord. Spinal neurons immunostained for serotonin were visible with and without L-tryptophan and monoamine oxidase inhibitor pretreatment.