A Golgi study of the monkey paraventricular nucleus: neuronal types, afferent and efferent fibers

The neuronal organization of the paraventricular nucleus (PVN) was examined in Golgi impregnations of adult monkey. Results showed that at least six types of neurons could be identified in the nucleus on the basis of morphological features of the somata, dendrites, and axons. Four types of neurons with sparse to densely spined cell bodies and dendrites exhibited long axons and included large neurons (types I and II), medium-sized to large neurons (type III), and small to medium-sized cells (type IV). Axons of type I, III, and IV neurons had different diameters and were followed out of the PVN. Axon collaterals that arborized within the PVN were seen on the axons of types III and IV cells. Two types of interneurons with small somata were also found. One (type V) exhibited varicose dendrites and a profusely arborizing local axon. The other cell (type VI) had recurved dendrites with long appendages and no impregnated axon. Afferent fibers were also identified. Type 1 was a fine-caliber axon that coursed long distances in the PVN and exhibited numerous short branches. Additional observations suggested that type 1 afferents originated from the stria terminalis. The other afferent axon (type 2) was thicker and gave rise to terminal arborizations containing clusters of small swellings. The efferent fibers of the PVN were also examined in impregnations of the paraventriculosupraopticohypophysial tract. Fibers formed an extensive plexus as they coursed ventrally and passed through the lateral hypothalamus. Axons coursing more laterally in the tract were much larger than those more medially located. Our findings show a diverse organization of neuronal types within the monkey PVN with evidence for intrinsic connections through axon collaterals of efferent neurons and the locally arborizing axons of interneurons. Correlations are proposed between morphological subtypes of neurons seen in this Golgi study and the known functional output pathways of the PVN.     

Matrix metalloproteinases MMP2 and MMP9 are upregulated by noradrenaline in the mouse neuroendocrine hypothalamus

Magnocellular neurons of the hypothalamic supraoptic nuclei (SON) are involved in the synthesis and release of two major neuropeptides: oxytocin (OT) and arginine-vassopressin (AVP). Neurochemical plasticity in this system is induced by physiological conditions such as lactation, parturition and dehydration, and may be accompanied by reversible structural plasticity affecting neurons, astrocytes and the extracellular matrix (ECM). The noradrenergic system plays a critical role in triggering this chemical plasticity associated with structural plasticity. Matrix metalloproteinases (MMPs) are good candidates for involvement in the ECM remodelling observed in structural plasticity. We investigated the possible regulation of the two gelatinases, MMP2 and MMP9, by noradrenaline (NA) in the mouse neuroendocrine hypothalamus. We looked for the presence, location and activity of MMP2 and MMP9 in the SON, using an ex vivo experimental model of mouse hypothalamic slices incubated for 4 h with 10⁻⁴  m NA. We showed that: (i) immunoreactivity for MMP2 and MMP9 was detected not only in AVP-positive and OT-positive magnocellular neurons, but also in astrocyte processes in control and NA-treated slices; (ii) the number of MMP2- and MMP9-positive cells increased after incubation with NA; (iii) MMP2 and MMP9 displayed markedly higher levels of gelatinolytic activity after NA treatment. These results suggest that both MMP2 and MMP9 are regulated by NA, and could therefore also be involved in structural plasticity within the SON.     

Distribution of galanin receptor 1 immunoreactivity in the rat stomach and small intestine

Galanin affects gastrointestinal functions by activating different G protein-coupled receptors. Here, we identified the sites of expression of the galanin receptor 1 (GAL-R1) subtype in the rat stomach and small intestine by using immunohistochemistry with an antibody raised to the third intracellular loop of rat GAL-R1 (GAL-R1(Y225-238)) and confocal microscopy. Antibody specificity was confirmed by (1) the detection of a band at approximately 70 kDa in Western blot of membranes from GAL-R1 transfected cells, (2) the cell surface staining of GAL-R1 transfected cells, which was not detected in control cells, and (3) the abolition of Western signal and tissue immunostaining by preadsorbing the antibody with the peptide used for immunization. GAL-R1 immunoreactivity was localized to the cell surface of enterochromaffin-like cells, and of myenteric and submucous neurons, and to fibers distributed to the plexuses, interconnecting strands, muscle layers, vasculature, and mucosa. A dense network of GAL-R1 immunoreactivity was observed in the deep muscular plexus in very close association with interstitial cells of Cajal visualized by c-kit immunostaining. In the ileum, 81.6% of GAL-R1 myenteric neurons and 70.7% of GAL-R1 submucosal neurons were substance P immunoreactive. Vasoactive intestinal polypeptide immunoreactivity was found in 48.3% of GAL-R1 submucosal neurons, but not in GAL-R1 myenteric neurons. These findings support the hypothesis that GAL-R1 mediates galanin actions on gastrointestinal motility and secretion by modulating the release of other neurotransmitters and contributes to galanin-induced inhibition of gastric acid secretion by means of the suppression of endogenous histamine release.     

Distribution of sensorin immunoreactivity in the central nervous system of Helix pomatia: functional aspects

Land snails belonging to the genus Helix are commonly used to study several behaviors and their plasticity at the cellular level. Because the knowledge of sensory neurons in these species is far from being complete, we have investigated the presence and distribution in Helix pomatia central nervous system of the immunoreactivity for sensorin, a peptide specific for mechanosensory neurons in Aplysia. We found that the majority of immunopositive cells were grouped in clusters located in all the central ganglia, except for the pedal ganglion, where only a single large neuron was stained. A symmetrical cluster of stained cells in the cerebral ganglia showed homology with the cerebral J clusters in Aplysia. Most of the somata of these Helix cerebral clusters send their axons in the ipsilateral cerebropedal connective and lip nerves and make monosynaptic connections with cells located in a medial adjacent cluster. This monosynaptic circuit can be reestablished in culture, where it shows homosynaptic depression as it does in the ganglionic preparation.     

Characterization of neuropeptide Y2 receptor protein expression in the mouse brain. I. Distribution in cell bodies and nerve terminals

Neuropeptide Y (NPY), a 36-amino-acid peptide, mediates biological effects by activating Y1, Y2, Y5, and y6 receptors. NPY neurons innervate many brain regions, including the hypothalamus, where NPY is involved in regulation of a broad range of homeostatic functions. We examined, by immunohistochemistry with tyramide signal amplification, the expression of the NPY Y2 receptor (Y2R) in the mouse brain with a newly developed rabbit polyclonal antibody. Y2R immunoreactivity was specific with its absence in Y2R knockout (KO) mice and in adjacent sections following preadsorption with the immunogenic peptide (10(-5) M). Y2R-positive processes were located in many brain regions, including the olfactory bulb, some cortical areas, septum, basal forebrain, nucleus accumbens, amygdala, hippocampus, hypothalamus, substantia nigra compacta, locus coeruleus, and solitary tract nucleus. However, colchicine treatment was needed to detect Y2R-like immunoreactivity in cell bodies in many, but not all, areas. The densest distributions of cell bodies were located in the septum basal forebrain, including the bed nucleus, and amygdala, with lower density in the anterior olfactory nucleus, nucleus accumbens, caudal striatum, CA1, CA2, and CA3 hippocampal fields, preoptic nuclei lateral hypothalamus, and A13 DA cells. The widespread distribution of Y2R-positive cell bodies and fibers suggests that NPY signaling through the Y2R is common in the mouse brain. Localization of the Y2R suggests that it is mostly presynaptic, a view supported by its frequent absence in cell bodies in the normal mouse and its dramatic increase in cell bodies of colchicine-treated mice.     

Distribution of the neuropeptide FF1 receptor (hFF1) in the human hypothalamus and surrounding basal forebrain structures: immunohistochemical study

Neuropeptides with C-terminal RFamide and their receptors NPFF1 (FF1) and NPFF2 (FF2) have been implicated in a wide variety of functions, including nociception and autonomic and neuroendocrine regulation. Recent studies indicate that the FF1, but not FF2, mRNA is highly expressed in the human hypothalamus. In the present study, localization of FF1 in the human hypothalamus and surrounding regions was studied immunohistochemically by using an antibody against human FF1 (hFF1). Brain sections from healthy 30-50-year-old individuals were used for hFF1 immunohistochemistry. The highest density of hFF1-stained cells was found in the posterior division of the bed nucleus of the stria terminalis and in the zona incerta. A moderate density of cells was observed in the perifornical nucleus, infundibular nucleus, tuberomammillary nucleus, and lateral tuberal nucleus. A lesser density was revealed in the dorsomedial hypothalamic nucleus, basal nucleus of Meynert, and anterior amygdaloid area. Only scattered hFF1 cells were found in the suprachiasmatic nucleus and hypothalamic paraventricular nucleus. hFF1 cells and fibers were absent in the supraoptic and mammillary nuclei. Single and double strands of hFF1-immunopositive punctate varicosities marked cellular processes of different caliber. The density of hFF1-immunostained fiber networks did not always coincide with that of hFF1-immunostained cells. hFF1 immunoreactivity was also found in the wall of blood vessels within most brain areas studied. Localization of hFF1 in discrete regions of the hypothalamus and extended amygdala may provide important insights into the role of amidated neuropeptides in central autonomic and neuroendocrine control in the human brain.     

Highly regionalized distribution of stromal cell-derived factor-1/CXCL12 in adult rat brain: constitutive expression in cholinergic, dopaminergic and vasopressinergic neurons

The stromal cell-derived factor-1 (SDF-1)/CXCL12 and its receptor CXCR4 are key modulators of immune functions. In the nervous system, SDF-1/CXCL12 is crucial for neuronal guidance in developing brain, intercellular communication and the neuropathogenesis of acquired immunodeficiency syndrome. However, cerebral functions of SDF-1/CXCL12 in adult brain are poorly understood. The understanding of its role in the adult brain needs a detailed neuroanatomical mapping of SDF-1/CXCL12. By dual immunohistochemistry we demonstrate that this chemokine is constitutively expressed not only in astrocytes and microglia but also in neurons, in discrete neuroanatomical regions. Indeed, neuronal expression of SDF-1/CXCL12 is mainly found in cerebral cortex, substantia innominata, globus pallidus, hippocampus, paraventricular and supraoptic hypothalamic nuclei, lateral hypothalamus, substantia nigra and oculomotor nuclei. Moreover, we provide the first evidence that SDF-1/CXCL12 is constitutively expressed in cholinergic neurons in the medial septum and substantia innominata and in dopaminergic neurons in substantia nigra pars compacta and the ventral tegmental area. Interestingly we also show, for the first time, a selective co-localization of SDF-1/CXCL12 with vasopressin-expressing neurons in the supraoptic and paraventricular hypothalamic nuclei. In addition, in the lateral hypothalamic area, SDF-1/CXCL12 was found to be located on melanin concentrating hormone-expressing neurons. Altogether, these original data suggest that SDF-1/CXCL12 could be a modulatory neuropeptide regulating both central cholinergic and dopaminergic systems. In addition, a key role for SDF-1/CXCL12 in neuroendocrine regulation of vasopressin-expressing neurons represents an exciting new field of research.     

Dendritic Release of Vasopressin and Oxytocin

In addition to the release of neurotransmitters from their axon terminals, several neuronal populations are able to release their products from their dendrites. The cell bodies and dendrites of vasopressin- and oxytocin-producing neurones are mainly located within the hypothalamic supraoptic and paraventricular nuclei and neuropeptide release within the magnocellular nuclei has been shown in vitro and in vivo . Local release is induced by a range of physiological and pharmacological stimuli, and is regulated by a number of brain areas; locally released peptides are mainly involved in pre- and postsynaptic modulation of the electrical activity of magnocellular neurones. Spatial and temporal differences between peptide release within the nuclei and that from the distant axonal varicosities indicate that the release mechanisms are at least partially independent, supporting the hypothesis of locally regulated dendritic release of vasopressin and oxytocin. In this respect, magnocellular neurones show similarities to other neuronal populations and thus autoregulation of neuronal activity by dendritic neuromodulator release may be a general phenomenon within the brain.     

Distribution of immunoreactivity for the NK1 receptor on different subpopulations of sympathetic preganglionic neurons in the rat

The distribuktion of immunoreactivity to the recptor for substance P, the neurokinin 1 (NK1) receptor, was ezamined in preganglionic sympathetic neurons of the rat by usingimmunohistochemistry and retroggrade neuronal tracing abour one-third of all sympathetic preganglionic neurons were NK1 receptor immunoreactiv, and most of the NK1 receptor immunoractive neurons were also nitric oxide synthase immunoreactive the proportions ofsympathetic preganglionic neurons projecting to the superior and inferior mesenteric ganglia, addrenal glnd, and lumbar sympathetic hain which werre nK1 rdeptor-immunoreactive were determined. Most (89%) of the preganglionic neurons projecting to the addrenal glands were NK1 rceptor immunoreactive few (17%) of the preganglionic neurons projecting to the L5 sympathetic hain ganglion were immunoreactive for the receptor, while preganglionic neurons projecting tothe prevertebral ganglia were NK1 receptor immunoractive at intermediate frequencies (61–64%). Thus, substance P acting on NK1 receptors in likely to be importnt inthe pregnglionic pathways to the addrenal medulla and viscera via the preveertebral ganglia, but the NK1 receptor withthe enzyme nitric oxide synthase was also examined. The co-localisation of NK1 receptor-immunoreactive neurons were also nitric oxide synthase immunoreactiv. Thus NK1 receptors occur onpreganglionic neurons over many spinal segments and in a range of prganglionic pathways, as well as in a range of combinations with nitric oxide shnthase. The heterogeneity ofjpreganglionic neurons showing NK1 receeptor immunoreactivity may reglect the involvement of NK1-mediated trnsmkssion in a variety of functional pathways, most notably thepreganglionic projections to the adrenal medulla and to the viscera. © 1996 Wiley-Liss, Inc.     

Neuron and stimulus typologies in the rat gustatory system

Although gustatory neurons may be categorized in terms of one or a few characteristics (e.g. ‘best stimulus’), such typologies are essentialistic and inconsistent with modern taxonomic methods¹⁵. If polythetic taxonomic criteria are used and the variability among neuronal responses is closely analyzed, neuronal ‘types’ are found to disappear, at least within the acid-salt range. This applies to both the primary nerve level (chorda tympani nerve) and secondary level (nucleus tractus solitarius) of the taste system in the rat. In the same context, taste stimuli may fall into different groups if several very similar stimuli are used (e.g. sodium and lithium salts). This is not surprising, and may depend on the choice of stimulus arrays rather than a differentiation of a few stimulus types by the taste system. Finally, it should be noted that the arguments regarding neuron and stimulus typologies presented here for the taste system are also valid for other sensory systems, although the conclusions may be different.     

Hypervascularization in the magnocellular nuclei of the rat hypothalamus: relationship with the distribution of aquaporin-4 and markers of energy metabolism

In the magnocellular nuclei of the hypothalamus, there is a rich vascular network for which the function remains to be established. In the supraoptic nucleus, the high vascular density may be one element, which together with the water channel aquaporin-4 expressed in the astrocytes, is related to a role in osmoreception. We tested the osmoreception hypothesis by studying the correlation between vascular and cellular densities in the paraventricular nucleus and the supraoptic nucleus. Whether aquaporin-4 is likely to contribute to osmoreception was tested by studying the distribution in the magnocellular nuclei of the hypothalamus. The high vascular density may also reflect a high metabolic activity due to the synthesis of vasopressin and oxytocin. This metabolic hypothesis was tested by studying the regional cytochrome oxidase histochemistry, the local cerebral blood flow, and the density of glucose transporter type-1 in the supraoptic and paraventricular nuclei. All the magnocellular nuclei were characterized by an extended and intense aquaporin-4 labelling and a weak cytochrome oxidase histochemistry. The highest vascular density was found in the supraoptic nucleus and the magnocellular regions of the paraventricular nucleus. The local cerebral blood flow rates were surprisingly low in the paraventricular nucleus and the supraoptic nucleus in comparison to the cerebral cortex. Furthermore in these nuclei, the antibody for glucose transporter type-1 revealed two populations of vessels differing by their labelling intensity. The similarities observed between the different nuclei suggest that, in the hypothalamus, all magnocellular regions sense the plasma osmolarity. The low local cerebral blood flow, and the patterns of glucose transporter type-1 labelling and cytochrome oxidase histochemistry suggest that the high vascularization of these hypothalamic nuclei is not related to a high metabolic capacity in basal conditions.     

Opioid receptor subtypes involved in the regulation of prolactin secretion during pregnancy and lactation

Afferent endogenous opioid neuronal systems facilitate prolactin secretion in a number of physiological conditions including pregnancy and lactation, by decreasing tuberoinfundibular dopamine (TIDA) inhibitory tone. The aim of this study was to investigate the opioid receptor subtypes involved in regulating TIDA neuronal activity and therefore facilitating prolactin secretion during early pregnancy, late pregnancy and lactation in rats. Selective opioid receptor antagonists nor-binaltorphimine (kappa-receptor antagonist, 15 micro g/5 micro l), beta funaltrexamine (mu-receptor antagonist, 5 microg/5 microl) and naltrindole (delta-receptor antagonist, 5 microg/5 microl) or saline were administered intracerebroventricularly (i.c.v.) on day 8 of pregnancy during a nocturnal prolactin surge, on day 21 of pregnancy during the ante partum prolactin surge or on day 7 of lactation before the onset of a suckling stimulus. Serial blood samples were collected at regular time intervals, via chronic indwelling jugular cannulae, before and after drug administration and plasma prolactin was determined by radioimmunoassay. TIDA neuronal activity was measured using the 3,4-dihydroxyphenylacetic acid (DOPAC) : dopamine ratio in the median eminence 2 h 30 min after i.c.v. drug injection. In each experimental condition, plasma prolactin was significantly inhibited by both kappa- and mu-receptor antagonists, whereas the delta-receptor antagonist had no effect compared to saline-injected controls. Similarly, nor-binaltorphimine and beta funaltrexamine significantly increased the median eminence DOPAC : dopamine ratio during early and late pregnancy, and lactation whereas naltrindole had no effect compared to saline-injected controls. These data suggest that TIDA neuronal activity, and subsequent prolactin secretion, is regulated by endogenous opioid peptides acting at both kappa- and mu-opioid receptors during prolactin surges of early pregnancy, late pregnancy and lactation.     

Regulation of cardiac sympathetic afferent reflex by GABA(A) and GABA(B) receptors in paraventricular nucleus in rats

The aim of the present study was to determine the role of GABA_A and GABA_B receptors in paraventricular nucleus (PVN) in regulating cardiac sympathetic afferent reflex (CSAR). Under urethane (800 mg/kg) and α-chloralose (40 mg/kg) anesthesia, renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP) and heart rate (HR) were recorded in sinoaortic-denervated and cervical-vagotomized rats. CSAR was evaluated based in the response of RSNA to epicardial application of capsaicin (0.3 nmol) or bradykinin (1 nmol). Bilateral PVN microinjection of the GABA_A receptor agonist isoguvacine (10 nmol) attenuated CSAR, while the GABA_B receptor agonist baclofen (1 nmol) abolished CSAR. Both isoguvacine and baclofen greatly decreased baseline RSNA and MAP. The GABA_A receptor antagonist gabazine (0.1 nmol) had no significant effect on CSAR, but the GABA_B receptor antagonist CGP-35348 (10 nmol) enhanced CSAR. Gabazine caused greater increases in baseline RSNA and MAP than CGP-35348. Vigabatrin (10 nmol), a selective GABA-transaminase inhibitor which increases endogenous GABA level, abolished CSAR, and decreased baseline RSNA, MAP and HR. The effects of vigabatrin were antagonized by combined gabazine (0.1 nmol) and CGP-35348 (10 nmol). The results indicate that activation of either GABA_A or GABA_B receptors in the PVN inhibits CSAR, while blockage of GABA_B receptors in the PVN enhances CSAR. Endogenous GABA in the PVN could have an important role in regulating CSAR.     

Cortical regulation of striatal projection neurons and interneurons in a Parkinson's disease rat model

Striatal neurons can be either projection neurons or interneurons, with each type exhibiting distinct susceptibility to various types of brain damage. In this study, 6-hydroxydopamine was injected into the right medial forebrain bundle to induce dopamine depletion, and/or ibotenic acid was injected into the M1 cortex to induce motor cortex lesions. Immunohistochemistry and western blot assay showed that dopaminergic depletion results in significant loss of striatal projection neurons marked by dopamine- and cyclic adenosine monophosphate-regulated phosphoprotein, molecular weight 32 k Da, calbindin, and μ-opioid receptor, while cortical lesions reversed these pathological changes. After dopaminergic deletion, the number of neuropeptide Y-positive striatal interneurons markedly increased, which was also inhibited by cortical lesioning. No noticeable change in the number of parvalbumin-positive interneurons was found in 6-hydroxydopamine-treated rats. Striatal projection neurons and interneurons show different susceptibility to dopaminergic depletion. Further, cortical lesions inhibit striatal dysfunction and damage induced by 6-hydroxydopamine, which provides a new possibility for clinical treatment of Parkinson's disease.     

Expression of corticotropin releasing factor (CRF), urocortin and CRF type 1 receptors in hypothalamic-hypophyseal systems under osmotic stimulation

The expression of corticotropin releasing factor (CRF) and urocortin in hypothalamic magnocellular neurones increases in response to osmotic challenge. To gain a better understanding of the physiological roles of CRF and urocortin in fluid homeostasis, CRF, urocortin and CRF type 1 receptor (CRFR-1) gene expression was examined in the hypothalamic-hypophyseal system usingin situ and double-label in situ hybridization following chronic salt loading. CRFR-1 expression was further examined by immunohistochemistry and receptor binding. Ingestion of hypertonic saline by Sprague-Dawley rats for 7 days induced CRF mRNA exclusively in the oxytocin neurones of the magnocellular paraventricular nucleus (PVN) and the supraoptic nucleus (SON), but induced CRFR-1 mRNA in both oxytocin and vasopressin-containing magnocellular neurones. Hypertonic saline treatment also increased urocortin mRNA expression in the PVN and the SON. In the SON, urocortin was localized to vasopressin and oxytocin neurones but was rarely seen in CRF-positive cells. Changes in CRFR-1 mRNA expression in magnocellular neurones by hypertonic saline treatment were accompanied by changes in CRFR-1 protein levels and receptor binding. Hypertonic saline treatment increased CRFR-1-like immunoreactivity in the magnocellular PVN and SON, and decreased it in the parvocellular PVN. CRF receptor binding in the PVN and SON was also increased in response to osmotic stimulation. Finally, hypertonic saline treatment increased CRFR-1 mRNA, CRFR-1-like immunoreactivity and CRF receptor binding in the intermediate pituitary. These results demonstrate that the increase in the expression of CRF and urocortin message in magnocellular neurones induced by salt loading is accompanied by an increase in CRF receptor levels and binding in the hypothalamus and intermediate pituitary. Thus, CRF and urocortin may exert modulatory effects locally within magnocellular neurones as well as at the pituitary gland in response to osmotic stimulation.     

Role of galanin receptor 1 in gastric motility in rat

Galanin actions are mediated by distinct galanin receptors (GAL-R), GAL-R1, -R2 and -R3. We investigated the role of GAL-R1 in gastric motility and the expression of GAL-R1 in the rat stomach. In vivo, in urethane-anaesthetized rats, galanin (equipotent for all GAL-Rs) induced a short inhibition of gastric motility, followed by increase in tonic and phasic gastric motility; the latter was significantly reduced by the GAL-R1 antagonist, RWJ-57408. Galanin 1-16 (high affinity for GAL-R1 and -R2) induced a long-lasting decrease of intragastric pressure, which was not modified by RWJ-57408. In vitro, galanin and galanin 1-16 induced increase of intragastric pressure that was not affected by RWJ-57408. Tetrodotoxin (TTX) did not suppress the galanin excitatory effect, whereas the effect of galanin 1-16 on gastric contraction was increased by TTX- or N-nitro-L-arginine, an inhibitor of nitric oxide synthase. GAL-R1 immunoreactivity was localized to cholinergic and tachykinergic neurons and to neurons immunoreactive for nitric oxide synthase or vasoactive intestinal polypeptide. This study suggests that an extrinsic GAL-R1 pathway mediates the galanin excitatory action, whereas an extrinsic, non GAL-R1 pathway is likely to mediate the galanin inhibitory effect in vivo. GAL-R1 intrinsic neurons do not appear to play a major role in the control of gastric motility.     

Estrogen receptor-beta in oxytocin and vasopressin neurons of the rat and human hypothalamus: Immunocytochemical and in situ hybridization studies

Topographical distribution of estrogen receptor-beta (ER-beta)-synthesizing oxytocin (OT) and vasopressin (VP) neurons was studied in the hypothalamic paraventricular and supraoptic nuclei (PVH; SO) of ovariectomized rats. In distinct subregions, 45-98% of OT neurons and 88-99% of VP neurons exhibited ER-beta immunoreactivity that was confined to cell nuclei. Neuronal populations differed markedly with respect to the intensity of the ER-beta signal. Magnocellular OT neurons in the PVH, SO, and accessory cell groups typically contained low levels of the ER-beta signal; in contrast, robust receptor labeling was displayed by OT cells in the ventral subdivision of medial parvicellular subnucleus and in the caudal PVH (dorsal subdivision of medial parvicellular subnucleus and lateral parvicellular subnucleus). Estrogen receptor-beta signal was generally more intense and present in higher proportions of magnocellular and parvicellular VP vs. OT neurons of similar topography. Immunocytochemical observations were confirmed via triple-label in situ hybridization, an approach combining use of digoxigenin-, fluorescein-, and 35S-labeled cRNA hybridization probes. Further, ER-beta mRNA was also detectable in corticotropin-releasing hormone neurons in the parvicellular PVH. Finally, double-label immunocytochemical analysis of human autopsy samples showed that subsets of OT and VP neurons also express ER-beta in the human. These neuroanatomical studies provide detailed information about the topographical distribution and cellular abundance of ER-beta within subsets of hypothalamic OT and VP neurons in the rat. The variable receptor content may indicate the differential responsiveness to estrogen in distinct OT and VP neuronal populations. In addition, a relevance of these findings to the human hypothalamus is suggested.     

Low temperature stimulates alpha-melanophore-stimulating hormone secretion and inhibits background adaptation in Xenopus laevis

It is well-known that alpha-melanophore-stimulating hormone (alpha-MSH) release from the amphibian pars intermedia (PI) depends on the light condition of the animal's background, permitting the animal to adapt the colour of its skin to background light intensity. In the present study, we carried out nine experiments on the effect of low temperature on this skin adaptation process in the toad Xenopus laevis, using the skin melanophore index (MI) bioassay and a radioimmunoassay to measure skin colour adaptation and alpha-MSH secretion, respectively. We show that temperatures below 8 degrees C stimulate alpha-MSH secretion and skin darkening, with a maximum at 5 degrees C, independent of the illumination state of the background. No significant stimulatory effect of low temperature on the MI and alpha-MSH plasma contents was noted when the experiment was repeated with toads from which the neurointermediate lobe (NIL) had been surgically extirpated. This indicates that low temperature stimulates alpha-MSH release from melanotrope cells located in the PI. An in vitro superfusion study with the NIL demonstrated that low temperature does not act directly on the PI. A possible role of the central nervous system in cold-induced alpha-MSH release from the PI was tested by studying the hypothalamic expression of c-Fos (as an indicator for neuronal activity) and the coexistence of c-Fos with the regulators of melanotrope cell activity, neuropeptide Y (NPY) and thyrotrophin-releasing hormone (TRH), using double fluorescence immunocytochemistry. Upon lowering temperature from 22 degrees C to 5 degrees C, in white-adapted animals c-Fos expression decreased in NPY-producing suprachiasmatic-melanotrope-inhibiting neurones (SMIN) in the ventrolateral area of the suprachiasmatic nucleus (SC) but increased in TRH-containing neurones of the magnocellular nucleus. TRH is known to stimulate melanotrope alpha-MSH release. We conclude that temperatures around 5 degrees C inactivate the SMIN in the SC and activate TRH-neurones in the magnocellular nucleus, resulting in enhanced alpha-MSH secretion from the PI, darkening the skin of white-adapted X. laevis.