Embryonic Ethanol Exposure Affects the Early Development, Migration, and Location of Hypocretin/Orexin Neurons in Zebrafish.

BACKGROUND: Embryonic ethanol (EtOH) exposure is known to increase alcohol drinking later in life and have long-term effects on neurochemical systems in the brain. With zebrafish having marked advantages for elucidating neural mechanisms underlying brain disorders, we recently tested and showed in these fish, similar to rodents, that low-dose embryonic EtOH stimulates voluntary consumption of EtOH while increasing expression of hypocretin/orexin (hcrt) neurons, a neuropeptide that promotes consummatory and reward-related behaviors. The goal of the present study was to characterize how embryonic EtOH affects early development of the hcrt system and produces persistent changes at older ages that may contribute to this increase in EtOH consumption. METHODS: We utilized live imaging and Imaris software to investigate how low-dose embryonic EtOH (0.5%), administered from 22 to 24 hours postfertilization, affects specific properties of hcrt neurons in hcrt:EGFP transgenic zebrafish at different ages. RESULTS: Time-lapse imaging from 24 to 28 hpf showed that embryonic EtOH increased the number of hcrt neurons, reduced the speed, straightness, and displacement of their migratory paths, and altered their direction early in development. At older ages up to 6 dpf, the embryonic EtOH-induced increase in hcrt neurons was persistent, and the neurons became more widely dispersed. These effects of embryonic EtOH were found to be asymmetric, occurring predominantly on the left side of the brain, and at 6 dpf, they resulted in marked changes in the anatomical location of the hcrt neurons, with some detected outside their normal position in the anterior hypothalamus again primarily on the left side. CONCLUSIONS: Our findings demonstrate that low-dose embryonic EtOH has diverse, persistent, and asymmetric effects on the early development of hypothalamic hcrt neurons, which lead to abnormalities in their ultimate location that may contribute to behavioral disturbances, including an increase in EtOH consumption.

Brief daily suckling shifts locomotor behavior and induces PER1 protein in paraventricular and supraoptic nuclei, but not in the suprachiasmatic nucleus, of rabbit does.

Nursing in the rabbit is a circadian event during which mother and pups interact for a period of < 5 min every day. Here we explored behavioral and neuronal changes in the mother by analyzing the suprachiasmatic nucleus (SCN), and oxytocinergic (OT) neurons in the paraventricular nucleus (PVN) and the supraoptic nucleus (SON). We maintained lactating does in a light-dark cycle (lights on at 07 : 00 hours; ZT0); they were scheduled to nurse during either the day (ZT03) or the night (ZT19). Groups of intact and nursing females was perfused, one at each 4-h point through a 24-h cycle. We explored, by immunohistochemistry, the PER1 expression and double-labeling, with OT antibody, of neurons in the PVN and SON at lactation on day 7. In the SCN, intact and lactating groups had peak PER1 expression at ZT11; however, there was a reduction in PER1 at peak time in the nursing groups. There was a locomotor activity rhythm with increased activity around the time of lights-on in intact subjects and around the time of suckling in lactating does. There was an induction of PER1 in OT cells in the PVN and SON that shifted in phase with timing of nursing. We further explored the maintenance of the PER1 expression in OT cells in nursing-deprived does and found a significant decrease at 24 and 48 h after the last nursing. We conclude that suckling induced PER1 in the PVN and SON, but not in the SCN, in nursing does, and also shifted their locomotor behavior.

The central vasopressinergic system: examining the opportunities for psychiatric drug development.

Arginine vasopressin (AVP) is a cyclic nonapeptide synthesized exclusively by neurosecretory cells of the central nervous system (CNS). Two functionally distinct vasopressinergic systems can be defined based on differences in the sites of action and release of AVP. The peripheral vasopressinergic system encompasses the sites of action for AVP released into peripheral circulation (e.g. vascular smooth muscle, liver, kidney) from nerve terminals in the posterior pituitary. Peripherally circulating AVP is responsible for the classic endocrine functions ascribed to this neurohormone (e.g. vasoconstriction, glycogen metabolism, antidiuresis). The central vasopressinergic system, on the other hand, includes the sites of AVP synthesis and release within the CNS, where AVP acts as a neuromodulator/neurotransmitter regulating an array of CNS-mediated functions (e.g. learning and memory, neuroendocrine reactivity, social behaviors, circadian rhythmicity, thermoregulation, and autonomic function). Historically, pharmaceutical interest has focused on drug development efforts that sought to exploit the peripheral effects of AVP. Evidence, however, from clinical studies and animal models of CNS disorders has directly implicated disturbances in vasopressinergic activity in the pathophysiology of a number of human psychiatric disorders (mood, anxiety, and cognitive disorders). This review will examine the available evidence of central vasopressinergic system involvement in psychiatric disorders, and the potential opportunities for development of novel psychopharmaceuticals around this system will be discussed. Specific lines of evidence will be presented which rationalize each AVP receptor subtype (V(1)R or V(1a), V(2)R, V(3)R or V(1b)) as a molecular target for particular psychiatric indications.

The effects of anterior hypothalamic lesions on short-day responses in Siberian hamsters given timed melatonin infusions.

The suprachiasmatic nucleus (SCN) of the hypothalamus is an area of dense 2-[125I]Iodomelatonin binding in Siberian hamsters (Phodopus sungorus sungorus) that is suggestive of a possible role in the reception and/or relaying of melatonin (MEL) signals. Indeed, in pinealectomized male Siberian hamsters given short day (SD) MEL signals (long-duration MEL infusions), lesions of the SCN (SCNx) block testicular regression and decreases in body and fat pad masses seen in identically treated hamsters with sham lesions (SCNs). In similar studies using Syrian hamsters (Mesocricetus auratus), anterior hypothalamic lesions (AHx), but not SCNx, blocked SD MEL signal-induced gonadal regression despite the similarity in the 2-[125I]Iodomelatonin binding pattern between the two species. The discrepancy between the ability of SCNx to block the reception of SD MEL signals between the two species is puzzling, given the similarity in the reproductive status of the Syrian and Siberian hamsters to systemically administered and timed MEL infusions. One possible way of reconciling the differences between these studies was that ancillary damage to areas neighboring the SCN, including the AH, may have occurred in our attempt to achieve complete SCNx in Siberian hamsters. Therefore, the purpose of the present study was to challenge AHx Siberian hamsters with SD MEL signals. Adult male hamsters were pinealectomized, fitted with subcutaneous catheters, and given daily timed infusions of MEL for 5 or 10 h (long day-like and short day-like, respectively) or the saline vehicle for 6 wk following bilateral electrolytic, or sham (AHs) lesions of the AH. Hamsters receiving 10 h MEL infusions that lacked evidence of anatomical or functional damage to the SCN showed SD-like gonadal regression, decreases in body and fat pad mass, and food intake similar to that observed in AHs animals. In contrast, 10 h MEL-infused SCNx hamsters did not exhibit SD-like responses, a finding confirming our previous report. These data suggest that interspecies differences exist between Syrian and Siberian hamsters in central nervous system sites and pathways involved in the reception/transmission of SD MEL signals.

Prostaglandins and hypothalamic neurotransmitter receptors involved in hyperthermia: a critical evaluation.

The role of a prostaglandin of the E series (PGE) in the hypothalamic mechanisms underlying a fever continues to be controversial. This paper reviews the historical literature and current findings on the central action of the PGEs on body temperature (Tb). New experiments were undertaken to examine the local effect of muscarinic, nicotinic, serotonergic, alpha-adrenergic, or beta-adrenergic receptor antagonists at hypothalamic sites where PGE1 caused a rise in Tb of the primate. Guide tubes for microinjection were implanted stereotaxically above sites in and around the anterior hypothalamic, preoptic area (AH/POA) of male Macaque monkeys. Following postoperative recovery, 30-100 ng of PGE1 was micro-injected unilaterally in a volume of 1.0-1.5 microliter at sites in the AH/POA to evoke a rise in Tb, and once identified, pretreated with a receptor antagonist. PGE1 hyperthermia was significantly reduced by microinjections of the muscarinic and nicotinic antagonists, atropine, or mecamylamine, at PGE1 reactive sites in the AH/POA. The serotonergic antagonist, methysergide, injected at PGE1 sensitive sites in the ventromedial hypothalamus also attenuated the rise in Tb. However, the 5-HT reuptake blocker, fluoxetine, and the beta-adrenergic receptor antagonist, propranolol, injected in the AH/POA failed to alter the PGE1 hyperthermia. In contrast, the alpha-adrenergic antagonist, phentolamine, potentiated the increase in Tb at all PGE1 reactive sites in the hypothalamus. An updated model is presented to explain how the concurrent actions of aminergic neurotransmitters acting on their respective receptors in the hypothalamus can interact with a PGE to elicit hyperthermia. Finally, an evaluation of the current literature including recent findings on macrophage inflammatory protein (MIP-1) supports the conclusion that a PGE in the brain is neither an obligatory nor essential factor for the expression of a pyrogen fever.

The volume of a sexually dimorphic nucleus in the ovine medial preoptic area/anterior hypothalamus varies with sexual partner preference.

Sheep are one of the few animal models in which natural variations in male sexual preferences have been studied experimentally. Approximately 8% of rams exhibit sexual preferences for male partners (male-oriented rams) in contrast to most rams, which prefer female partners (female-oriented rams). We identified a cell group within the medial preoptic area/anterior hypothalamus of age-matched adult sheep that was significantly larger in adult rams than in ewes. This cell group was labeled the ovine sexually dimorphic nucleus (oSDN). In addition to a sex difference, we found that the volume of the oSDN was two times greater in female-oriented rams than in male-oriented rams. The dense cluster of neurons that comprise the oSDN express cytochrome P450 aromatase. Aromatase mRNA levels in the oSDN were significantly greater in female-oriented rams than in ewes, whereas male-oriented rams exhibited intermediate levels of expression. Because the medial preoptic area/anterior hypothalamus is known to control the expression of male sexual behaviors, these results suggest that naturally occurring variations in sexual partner preferences may be related to differences in brain anatomy and capacity for estrogen synthesis.

The efferent connections of the anterior hypothalamic area of the rat, cat and monkey.

The general morphology and topographic relations of the anterior hypothalamic area (AHA) in the rat, cat and squirrel monkey have been described, and its efferent connections analyzed autoradiographically, after small injections of 3H-labeled amino acids into, or around, the area. In all three species the AHA is rather poorly separated from the surrounding preoptic and hypothalamic areas and nuclei but shows three distinct cellular condensations, located rostrally, centrally, and posterodorsally. Closely associated with the AHA are the retrochiasmatic area, the anterior periventricular nucleus and the scattered neurons usually referred to as the accessory supraoptic nucleus. The AHA has primarily short connections to the adjoining medial preoptic area, the lateral hypothalamic area, the periventricular nucleus, the dorsomedial nucleus, and to the "capsule" of the ventromedial nucleus. However, it also has certain more distant projections, rostrally to a narrow zone centered in the ventral part of the lateral septal nucleus, and caudally to the dorsal premammillary nuclei, the posterior hypothalamic area and the central gray. There is some evidence to suggest that the various subdivisions of the AHA have different efferent connections. Thus the posterodorsal cell condensation appears to give rise to the bilateral projection to the dorsal premammillary nuclei, while the projections to the septum, the posterior hypothalamic area and the central gray seem to have their origin in the central condensation. Similarly, the retrochiasmatic area sends its efferents through the ventral supraoptic commissure to the amygdala, the anterior periventricular nucleus contributes to the periventricular fiber system and to the external lamina of the median eminence, and the accessory supraoptic neurons project to the internal lamina of the median eminence.

Three-dimensional structure of the mammalian suprachiasmatic nuclei: a comparative study of five species.

The suprachiasmatic nuclei from five mammalian species (rat, hamster, cat, rhesus, and squirrel monkey) were reconstructed in three-dimensions by use of computer graphics and conventional histological techniques. The gross nuclear structures of the suprachiasmatic nuclei have complex three-dimensional geometries in every plane of orientation, and between the five species there are marked differences in the three-dimensional morphology of the suprachiasmatic nuclei. These dimensionally accurate reconstructions are discussed in relation to previous data suggesting morphological and/or functional specialization within specific regions of the suprachiasmatic nuclei.

Retinohypothalamic projection and suprachiasmatic nucleus of the house sparrow, Passer domesticus.

The distribution of retinohypothalamic projections and the organization of the suprachiasmatic region of the hypothalamus was investigated in the house sparrow (Passer domesticus). Retinohypothalamic projections (RHT) were studied by two anterograde tracing methods, and hypothalamic organization was investigated immunohistochemically with antisera against a number of substances known to be present in the mammalian suprachiasmatic nucleus (SCN): bombesin (BBS), glutamic acid decarboxylase (GAD), 5-hydroxytryptamine (5HT), neuropeptide Y (NPY), neurotensin (NT), somatostatin (SS), substance P (SP), vasoactive intestinal polypeptide (VIP), and arginine vasopressin (AVP). Observations from these experiments were analysed within the framework of a cytoarchitectural study using Nissl-stained material. From this study, we have identified an area in the anterior hypothalamus which we believe is an avian homologue of the mammalian SCN. This area contains a nucleus located in close apposition to the optic chiasm between the dorsal supraoptic decussation (DSD) and the ventral lateral geniculate body (GLv) for much of its rostrocaudal extent. The central portion of this nucleus contains neurons that exhibit GAD- and BBS-like immunoreactivity and is the terminal field for the RHT. For this reason, we term this nucleus the visual SCN. It also contains axon plexuses exhibiting 5HT-like, SP-like, and NPY-like immunoreactivity and is bordered ventrally by AVP-like, SP-like, and NT-like immunoreactive cells and medially by VIP-like and SS-like immunoreactive cells. Although it is not established that these cell groups together compose a single suprachiasmatic nucleus, the organization in the avian brain of a nuclear complex with a retinorecipient area surrounded by nonvisual components would be very similar to that of the mammalian SCN.

A cytoarchitectonic atlas of the mouse hypothalamus.

A description of the organization, areas, and cell groups within the hypothalamus of the mouse is presented in detail. Photomicrographs of cell-stained serial sections through the hypothalamus in frontal, sagittal and horizontal planes are included. The hypothalamus has been divided basically into medial and lateral parts with most well-defined cell groups or nuclei lying within the medial subdivision and surrounded by diffuse collections of cells referred to as areas. The heterogenetiy of cell types within most hypothalamic nuclei and areas has been emphasized with the consequent implications for heterogeneity of neuronal connections and of functions. Recently introduced neuroanatomical techniques permitting increased attention to the cellular level of organization have demonstrated precise connections and functional localization of cells within the hypothalamus. While cytoarchitectonic distinctions imply functional distinctions, morphological and experimental evidence suggest the existence also of systems of cells which transcend conventional cytoarchitectonic boundaries, the cells within each system being interconnected functionally or neuronally.

The structure of the inferior lobe of the teleost hypothalamus.

Electron microscopic and Golgi studies on the inferior lobes of sunfish and goldfish are described. The inferior lobe consists primarily of a nucleus ventricularis of densely packed cells surrounding the lateral recess of the third ventricle, and a peripherally situated nucleus diffusus consisting mostly of scattered neurons. A cell-sparse zone of dense neuropil is located between the two cellular areas. Neurons of both nuclei have spiny dendrites and axons which originate from basal dendrites. In some cases axons are found to send a collateral into the cell-sparse zone. Neurons of the nucleus diffusus possess collaterals that extend a considerable distance within the nucleus itself. The ultrastructure of cells of both nuclei reveals cytoplasmic organelles typical of most neurons. Synapses containing dense-cored and clear vesicles are present on the spines and shafts of the dendrites of both neuronal types. In only rare cases synapses were observed on the soma of neurons of the nucleus ventricularis. Possible anatomical substrates involved in the control of feeding and aggression in teleosts are considered in light of the present findings. Morphological similarities of the inferior lobes and related areas in various fishes and amphibians are discussed and their possible significance for the understanding of the evolution of hypothalamic mechanisms is considered.

Comparative distribution of bombesin/GRP- and substance-P-like immunoreactivities in rat hypothalamus.

Immunohistochemical localization of bombesin/gastrin-releasing peptide ( GRP )-like immunoreactivity (BN/ GRP -LI) and substance P-like immunoreactivity (SP-LI) in consecutive sections of rat hypothalamus was studied. Bombesin/ GRP -like immunoreactivity in the hypothalamus was partially characterized by gel filtration chromatography followed by radioimmunoassay. In the hypothalamus, SP-LI was more widely distributed than BN/ GRP -LI. Only the anterior and medial parvocellular parts of the nucleus paraventricularis and the nucleus suprachiasmaticus contained numerous cell bodies which exhibited BN/ GRP -LI. Neurons in these areas did not exhibit SP-LI. In contrast, cell bodies exhibiting SP-LI were numerous in the nucleus preopticus medialis and lateralis, nucleus anterior, nucleus ventromedialis and dorsomedialis, nucleus lateralis, nucleus arcuatus, and nucleus premamillaris ventralis and dorsalis. Only occasional cell bodies in these areas exhibited BN/ GRP -LI. It is concluded that the neuronal systems in the hypothalamus containing BN/ GRP -LI and SP-LI are separate, though the terminal fields in many areas overlap. Two peaks of BN/ GRP -LI were detected after gel filtration chromatography from extracts of the rat nucleus paraventricularis. The high molecular weight form coeluted with synthetic GRP (1-27), and the small molecular weight form eluted after synthetic bombesin. Thus, the endogenous BN/ GRP -LI is probably not authentic bombesin.

Distribution of substance P-immunoreactive elements in the preoptic area and the hypothalamus of the rat.

The localization and morphology of neurons, processes, and neuronal groups in the rat preoptic area and hypothalamus containing substance P-like immunoreactivity were studied with a highly selective antiserum raised against synthetic substance P. The antiserum was thoroughly characterized by immunoblotting; only substance P was recognized by the antiserum. Absorption of the antiserum with synthetic substance P abolished immunostaining while addition of other hypothalamic neuropeptides had no effect on the immunostaining. The specificity of the observed immunohistochemical staining pattern was further confirmed with a monoclonal substance P antiserum. The distribution of substance P immunoreactive perikarya was investigated in colchicine-treated animals, whereas the distribution of immunoreactive nerve fibers and terminals was described in brains from untreated animals. In colchicine-treated rats, immunoreactive cells were reliably detected throughout the preoptic area and the hypothalamus. In the preoptic region, labeled cells were found in the anteroventral periventricular and the anteroventral preoptic nuclei and the medial and lateral preoptic areas. Within the hypothalamus, immunoreactive cells were found in the suprachiasmatic, paraventricular, supraoptic, ventromedial, dorsomedial, supramammillary, and premammillary nuclei, the retrochiasmatic, medial hypothalamic, and lateral hypothalamic areas, and the tuber cinereum. The immunoreactive cell groups were usually continuous with adjacent cell groups. Because of the highly variable effect of the colchicine treatment, it was not possible to determine the actual number of immunoreactive cells. Mean soma size varied considerably from one cell group to another. Cells in the magnocellular subnuclei of the paraventricular and supraoptic nuclei were among the largest, with a diameter of about 25 microns, while cells in the supramammillary and suprachiasmatic nuclei were among the smallest, with a diameter of about 12 microns. Immunoreactive nerve fibers were found in all areas of the preoptic area and the hypothalamus. The morphology, size, density, and number of terminals varied considerably from region to region. Thus, some areas contained single immunoreactive fibers, while others were innervated with such a density that individual nerve fibers were hardly discernible. During the last decade, knowledge about neural organization of rodent hypothalamic areas and mammalian tachykinin biochemistry has increased substantially. In the light of these new insights, the present study gives comprehensive morphological evidence that substance P may be centrally involved in a wide variety of hypothalamic functions. Among these could be sexual behavior, pituitary hormone release, and water homeostasis.

Autoradiographic evidence for pathways from the medial preoptic area to the midbrain involved in the drinking response to angiotensin II.

The 3H-amino acid autoradiographic method was used to localize intracerebral sites from which angiotensin II (AII) elicits drinking and to identify their efferent neural pathways. Small injections (0.02-0.1 mul) of AII and 3H-amino acid mixtures were injected together or separately into widespread regions of the forebrain of adult rats in normal food and water balance. From an analysis of 39 positive and negative injection sites it was concluded that the caudal half of the medial preoptic area and the adjacent rostral part of the anterior hypothalamic area are sensitive to AII. Two anatomically defined pathways arising from neurons within this region were identified. One descends through the medial forebrain bundle and appears to terminate in the lateral hypothalamic area, the ventromedial nucleus, the mammillary body, and the ventral tegmental area. The other descends through the periventricular region and posterior hypothalamic area to end in the midbrain central gray. Additional widespread connections with the amygdala, septum, habenula, and pons appear to arise in the lateral preoptic area (Swanson, '76). Combined AII-3H-amino acid injections centered in the subfornical organ only elicited drinking in those cases in which injected label diffuse through the third ventricle to the medial preoptic area. No efferent pathways were identified in experiments in which a small injection (0.02 mul) heavily labeled cells strictly confined to the subfornical organ and there was no ventricular spread of label.

RANTES: a new prostaglandin dependent endogenous pyrogen in the rat.

Fever, a hallmark of disease, is a highly complex process initiated by the action of a number of endogenous pyrogens on the thermosensitive cells of the brain. We describe the activity of RANTES, a chemotactic cytokine, as intrinsically pyrogenic in the rat, when it is delivered directly to the thermosensitive region of the rat's anterior hypothalamic, pre-optic area (AH/POA). RANTES, microinjected into the AH/POA in a dose of 1, 5, 10, 15, 25 or 50 pg, produces an immediate and intense dose-related fever following injection. Increasing the dose to 100 pg did not result in a further increase in the febrile response. No significant change in body temperature was produced by heat-inactivated RANTES. The intrahypothalamic injection of antibodies against RANTES (2.0 microg, 15 min prior to RANTES) significantly blocked the fever induced by this chemokine. Pretreatment with ibuprofen blocked the fever induced by RANTES. In order of potency, the magnitude of the febrile response induced by RANTES was greater than that produced with equipotent doses of either macrophage inflammatory protein-1beta or interleukin-6. The results thus demonstrate that RANTES is the most potent endopyrogen discovered thus far and exerts its action directly on pyrogen-sensitive cells of the AH/POA through a prostaglandin-dependent pathway.

A direct pretectosuprachiasmatic projection in the rat.

The major afferent projections of the suprachiasmatic nuclei originate in the retina and the intergeniculate leaflet of the lateral geniculate nucleus and are important in the entrainment of endogenous circadian rhythms. A characteristic feature of the suprachiasmatic nucleus and the intergeniculate leaflet of the thalamus is that they are bilaterally innervated from the retina. However, parts of the olivary and posterior pretectal nuclei have been shown to be bilaterally innervated from the retina as well. We therefore aimed to explore whether these two nuclei, in the rat, were anatomically related to the suprachiasmatic nucleus. The anterograde neuronal tract-tracer, Phaseolus vulgaris-leucoagglutinin, was injected iontophoretically into different pretectal nuclei. Pretectal injections centered only in the medial part of the pretectum, i.e. involving the olivary and posterior pretectal nuclei, gave rise to a substantial bilateral innervation of the suprachiasmatic nucleus. From the site of injection, Phaseolus vulgaris-leucoagglutinin-immunoreactive nerve fibers coursed laterally and rostrally into the optic tract, and within the optic tract and chiasm, under the diencephalon to penetrate dorsally into the suprachiasmatic nucleus. Varicose Phaseolus vulgaris-leucoagglutinin-labeled nerve fibers were found exclusively in the ventrolateral part of the suprachiasmatic nucleus, mostly on the ipsilateral side. To determine the precise location of the projecting neurons, the retrograde tracer Cholera toxin, subunit B, was iontophoretically injected into the suprachiasmatic nucleus. The presence of of labeled neurons scattered in both the posterior and olivary pretectal nuclei was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Visceral inputs to neurons in the anterior hypothalamus including those that project to the periaqueductal gray: a functional anatomical and electrophysiological study.

The present study was designed to examine peripheral, in particular noxious visceral, inputs to neurons in the hypothalamus that project to the midbrain periaqueductal gray. The induction of Fos protein was used to localize hypothalamic neurons that were activated by noxious visceral stimulation. This was combined with retrograde transport of fluorescent latex microspheres from identified "pressor" and "depressor" sites in the dorsolateral/lateral or ventrolateral columns of the periaqueductal gray. A second series of electrophysiological experiments examined the receptive field characteristics, including the incidence of viscerosomatic convergence, of neurons in the ventral part of the anterior hypothalamus. Noxious visceral stimulation (intraperitoneal acetic acid) induced Fos-like immunoreactivity in significantly more neurons in the hypothalamus than control stimuli (intraperitoneal saline and intravenous phenylephrine). Particularly high numbers of Fos-positive neurons were found in the paraventricular nucleus, the supraoptic nucleus and ventral regions of the anterior hypothalamus. When combined with retrograde tracing from "depressor" sites in the ventrolateral periaqueductal gray, the highest numbers of double-labelled neurons were localized in the paraventricular nucleus and the lateral area of the anterior hypothalamus. However, the regions that contained the greatest proportions of Fos-positive neurons that projected to "depressor" sites in the ventrolateral periaqueductal gray were the lateral area of the anterior hypothalamus and its rostral extension, the lateral preoptic area. Fewer double-labelled neurons were localized in the hypothalamus after retrograde transport from sites in the dorsolateral/lateral periaqueductal gray compared to the results obtained from injections of tracer in the ventrolateral periaqueductal gray. Furthermore, the numbers of Fos-positive hypothalamic neurons that projected to the dorsolateral/lateral periaqueductal gray were very similar in experimental and control animals. The electrophysiological study confirmed that a large proportion of neurons in and around the lateral area of the anterior hypothalamus can be driven by noxious visceral stimulation and demonstrated a high incidence of viscerosomatic convergence in these cells (66% of cells driven from somatic structures were also driven by electrical stimulation of the splanchnic nerve). Somatic receptive fields of these neurons were generally large, often including all four limbs and the face. The results of the functional anatomical and electrophysiological studies have identified neurons in an area of the ventral anterior hypothalamus that are a focus of nociceptive visceral input and which project to the midbrain periaqueductal gray, in particular to its ventrolateral column. These results are discussed in relation to the roles of the anterior hypothalamus and the different longitudinal columns of the periaqueductal gray in co-ordinating autonomic and sensory functions in response to visceral pain.

Facilitative role of prolactin-releasing peptide neurons in oxytocin cell activation after conditioned-fear stimuli.

Emotional stress activates oxytocin neurons in the hypothalamic supraoptic and paraventricular nuclei and stimulates oxytocin release from the posterior pituitary. Oxytocin neurons in the hypothalamus have synaptic contact with prolactin-releasing peptide (PrRP) neurons. Intracerebroventricular administration of PrRP stimulates oxytocin release from the pituitary. These observations raise the possibility that PrRP neurons play a role in oxytocin response to emotional stress. To test this hypothesis, we first examined expression of Fos protein, an immediate early gene product, in the PrRP neurons in the medulla oblongata after conditioned-fear stimuli. Conditioned-fear stimuli increased the number of PrRP cells expressing Fos protein especially in the dorsomedial medulla. In order to determine whether PrRP cells projecting to the supraoptic nucleus are activated after conditioned-fear stimuli, we injected retrograde tracers into the supraoptic nucleus. Conditioned-fear stimuli induced expression of Fos protein in retrogradely labeled PrRP cells in the dorsomedial medulla. Finally we investigated whether immunoneutralization of endogenous PrRP impairs oxytocin release after emotional stimuli. An i.c.v. injection of a mouse monoclonal anti-PrRP antibody impaired release of oxytocin but not of adrenocorticotrophic hormone or prolactin and did not significantly change freezing behavior in response to conditioned-fear stimuli. From these data, we conclude that PrRP neurons in the dorsomedial medulla that project to the hypothalamus play a facilitative role in oxytocin release after emotional stimuli in rats.

A comparative analysis of the neuroendocrine mechanisms regulating ovulation, affected by a unilateral implant of atropine in the preoptic-anterior hypothalamic area, in intact and hemiovariectomized adult rats.

The effects were analysed of a unilateral implant of atropine on ovulation in intact and hemiovariectomized adult rats, together with the response of the atropine-implanted rats to hormone replacement. An outer cannula directed to the left or right preoptic (POA)-anterior-hypothalamic area (AHA) was implanted into cyclic adult rats. A group of animals in oestrus was hemiovariectomized and some were also implanted with a cannula. After two consecutive 4-day cycles, the hemiovariectomized animals were implanted with atropine (23 +/- 4 micrograms) or cholesterol (25 +/- 2 micrograms) on the day of oestrus. Atropine implanted into the left side of the POA-AHA blocked ovulation and compensatory ovarian hypertrophy, whilst implants in the right side had no effects. Administration of gonadotrophin-releasing hormone (GnRH; 3.7 micrograms/kg) at 13.00 h on the expected day of pro-oestrus induced ovulation in six out of seven treated animals. Of 19 rats with an implant of atropine in the left side of the POA-AHA, one ovulated after treatment with pregnant mare serum gonadotrophin (PMSG) on oestrus, or oestradiol benzoate or human chorionic gonadotrophin (hCG) on day 2 of dioestrus. The effects on ovulation of a unilateral implant of atropine into the POA-AHA of cyclic adult rats and the responses of such rats to GnRH, PMSG, hCG and oestradiol benzoate replacement were also studied. Ovulation was induced in rats with a unilateral implant of atropine and which had been treated with GnRH or hCG at 13.00 h on the expected day of pro-oestrus after the implant.(ABSTRACT TRUNCATED AT 250 WORDS)

Serotonin regulation of aggressive behavior in male golden hamsters (Mesocricetus auratus).

These studies examined the neurochemistry and neuroanatomy of the serotonin (5-HT) system innervating the anterior hypothalamus (AH) and the interaction of 5-HT receptor agonists with arginine vasopressin (AVP) in the regulation of offensive aggression in golden hamsters. Because specific 5-HT1A, 5-HT1B, and AVP V1A binding sites were observed within the AH by in vitro autoradiography, the hamsters were tested for offensive aggression after microinjections of AVP in combination with either the 5-HT1A agonist 8-hydroxy-2-(di-n-propylamino) tetraline (DPAT) or the 5-HT1B agonist CGS-12066A (CGS) directly within the AH. Though treatment with DPAT resulted in a dose-dependent inhibition of AVP-facilitated offensive aggression, CGS was ineffective. In addition, a retrograde tracer was injected within the AH to localize the distribution of 5-HT neurons projecting to the area. Retrogradely labeled 5-HT neurons were found within the dorsal, median, and caudal linear raphe nuclei and are suspected to inhibit AVP-facilitated offensive aggression by an activation of 5-HT1A receptors in the AH.