The median preoptic nucleus participates in the control of paraventricular vasopressin neurons by the subfornical organ in the rat

Extracellular single-unit activity was recorded from phasically firing neurohypophyseal neurons (n = 41) in the hypothalamic paraventricular nucleus (PVN) of urethane-anesthetized male rats. Electrical stimulation of the subfornical organ (SFO) produced orthodromic long-duration (n = 18) or short-duration (n = 10) excitation or inhibition (n = 8) of the activity of PVN neurons. The long-duration excitatory response of about half (n = 7) the neurons (n = 15) tested was reversibly abolished by microinjection of the local anesthetic lidocaine into the median preoptic nucleus (MnPO), whereas neither the short-duration excitatory (n = 7) nor inhibitory (n = 6) responses were affected. These results suggest that the SFO efferents through the MnPO to the PVN may transmit the neuromodulatory signals which evoke long-duration increases in the excitability of putative vasopressin (VP)-secreting neurons in the PVN.     

Vasopressin-sensitive neurons in the lateral paraventricular nucleus area in a guinea pig slice preparation

The effects of vasopressin (VP) on hypothalamic neurons located in the region of the paraventricular nucleus (PVN) were analyzed using intracellular techniques in slices of guinea pig brains. Two different classes of neurons were electrophysiologically identified in the magnocellular lateral part of the PVN and in the adjacent area. In the former area, vasopressinergic neurons were identified according to their phasic activity and their endogenous properties. These neurons were not responsive to VP, applied through the perfusion medium or locally by pressure. On the other hand, nonmagnocellular neurons exhibiting low-threshold Ca2+ spikes (LTS) were recorded in the area adjacent to the lateral part of the PVN. LTS were deinactivated at hyperpolarized membrane levels and induced short bursts of action potentials. On these neurons, VP evoked depolarizations accompanied by increases in firing, without modification of membrane resistance. VP effects were not blocked by TTX, suggesting a postsynaptic action of the peptide. These data indicate that VP controls the firing pattern of LTS neurons and suggest that this action may involve collaterals of axons originating from neighbouring vasopressinergic neurons.     

Central nesfatin‐1 influences the excitability of ghrelin‐responsive gastric distension neurons in the arcuate nucleus and reduces gastric motility in rats

Although the novel satiety peptide nesfatin‐1 has been shown to regulate gastric motility, the underlying mechanisms have yet to be elucidated. The study aimed to explore the effects of nesfatin‐1 on ghrelin‐responsive gastric distension (GD) neurons in the arcuate nucleus (Arc), and potential regulation mechanisms of gastric motility by the paraventricular nucleus (PVN). Single‐unit discharges in the Arc were recorded extracellularly, and gastric motility in conscious rats was monitored during the administration of nesfatin‐1 to the Arc or electrical stimulation of the PVN. Retrograde tracing and fluo‐immunohistochemistry staining were used to determine NUCB2/nesfatin‐1 neuronal projections. Nesfatin‐1 inhibited most of the ghrelin‐responsive GD‐excitatory neurons, but excited ghrelin‐responsive GD‐inhibitory neurons in the Arc. Gastric motility was significantly reduced by nesfatin‐1 administration to the Arc in a dose‐dependent manner. The firing activity in the Arc and changes to gastric motility were partly reduced by SHU9119, an antagonist of melanocortin 3/4 receptors. Electrical stimulation of PVN excited most of the ghrelin‐responsive GD neurons in the Arc and promoted gastric motility. Nonetheless, pretreatment with an anti‐NUCB2/nesfatin‐1 antibody in the Arc further increased the firing rate of most of the ghrelin‐responsive GD‐excitatory neurons and decreased the ghrelin‐responsive GD‐inhibitory neurons following electrical stimulation of the PVN. Gastric motility was enhanced by pretreatment with an anti‐NUCB2/nesfatin‐1 antibody in the Arc following PVN stimulation. Furthermore, NUCB2/nesfatin‐1/fluorogold double‐labeled neurons were detected in the PVN. These results suggest that nesfatin‐1 could serve as an inhibitory factor in the Arc to regulate gastric motility via the melanocortin pathway. The PVN could be involved in the regulation of the Arc in gastric activity.     

Characterization of the parabrachial nucleus input to the hypothalamic paraventricular nucleus in the rat

The brainstem parabrachial nucleus (PBN) is viewed as an increasingly important site for the transfer of autonomic-related information to more rostral structures in the forebrain including the hypothalamus. In this study, we examined electrophysiologically in vivo and anatomically the nature of PBN input to the hypothalamic paraventricular nucleus (PVN) and particularly to the vasopressin-and oxytocin-secreting magnocellular neurosecretory cells within this nucleus. In urethane-anaesthetized rats, extracellular recordings from 108 antidromically identified neurosecretory PVN cells revealed an excitatory (37/43 cells) and less frequently an inhibitory (6/43 cells) response consequent to electrical stimulation in the PBN. Both vasopressin (12/37 cells)-and oxytocin (9/37 cells)-secreting neurons appear to respond to the PBN stimulus. Four cells projecting to the neurohypophysis could also be antidromically activated from PBN, and this observation may be indicative of collateral branching in some PVN neurosecretory neurons. In addition, recordings from 60 non-magnocellular (i.e. non-neurohypophysially-projecting) PVN cells revealed a facilitatory response (43/60 cells) following PBN stimulation, Iontophoretic injections of the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) were made within the rat lateral PBN and brains prepared for immunocytochemical examination of projections to the PVN region. PHA-L-labelled fibres and terminals were visualized within both the parvocellular and magnocellular divisions of the PVN. In addition, labelled fibres were also seen in a region immediately dorsal to the PVN. PHA-L-labelled fibres with axonal varicosities and boutons were visualized over immunocyto-chemically-identified vasopressin and oxytocin neurons within the magnocellular PVN. These convergent electrophysiological and anatomical data provide evidence for a PBN projection to the PVN that is predominantly excitatory to both magnocellular neurosecretory and non-magnocellular cells. Moreover, with respect to vasopressin-and oxytocin-secreting cells, the PBN input appears to be directed at both populations of peptidergic neurons.     

Connections of the hypothalamic paraventricular necleus with the neurohypophysis, median eminence, amygdala, lateral septum and midbrain periaqueductal gray: An electrophysiological study in the rat

Extracellular recordings were obtained from 555 paraventricular (PVN) nucleus neurons in pentobarbital-anesthetized male rats. Cells were examined for their spontaneous activity patterns and response to single 1-Hz electrical stimulation of the neurohypophysis, median eminence, amygdala, lateral septum (LS) and midbrain periaqueductal gray (PAG).Neurohypophyseal stimulation evokedantidromic activation from 109 neurons. Among spontaneouly active neurohypophyseal neurons, evidence of a recurrent inhibitory pathway usually required pituitary stimulus intensities twice threshold for antidromic activation. Orthodromic excitatory or inhibitory responses followed amygdala and LS stimulation, but not PAG stimulation. The amygdala influence was predominantly inhibitory to ‘phasic’ (putative vasopressin-secreting) PVN neurohypophyseal neurons. Neurohypophyseal stimulation evokedorthodromic responses from 124 PVN cells; some of these neurons were also responsive to stimulation in other sites.Median eminence stimulation evoked antidromic responses from 37 PVN neurons; some of these cells also displayed phasic activity but not evidence for recurrent inhibition. Twelve cells in this group were also activated antidromically from both the median eminence and the neurohypophysis; collision tests suggest that the median eminence innervation may be an axon collateral of a neurohypophyseal pathway. Amygdala stimulation was inhibitory to some cells in this category.Amygdala, LS and PAG stimulation evoked antidromic activation from a small number of PVN cells, but none of these cells appeared to innervate more than one area, including the neurohypophysis, and none displayed phasic activity. Orthodromic responses were recorded among other PVN neurons after stimulation in these sites; however, PAG stimulation was the least effective stimulation area.These observations provide additional electrophysiological data that confirm efferent PVN connections to all areas tested, afferent connections from amygdala and LS but not PAG, and the possibility for coordinated activity among PVN neurons through local recurrent or common afferent connections.     

Electrophysiological evidence that circulating angiotensin II sensitive neurons in the subfornical organ alter the activity of hypothalamic paraventricular neurohypophyseal neurons in the rat

Electrical stimulation of the rat A1 noradrenergic region produced excitation (77%) of the activity of putative vasopressin (VP)-secreting neurons in the paraventricular nucleus (PVN) and produced excitation (4%), inhibition (26%) and excitation-inhibition (11%) of the activity of PVN neurons that were not antidromically identified by neurohypophysial stimulation. The excitatory response of putative VP-secreting neurons was blocked by microiontophoretically applied phentolamine, an α-adrenoceptor antagonist, but not by timolol, a β-adrenoceptor antagonist. The inhibitory response of unidentified PVN neurons, on the other hand, was blocked by timolol, but not by phentolamine.     

Activation of gastric afferents increases noradrenaline release in the paraventricular nucleus and plasma oxytocin level

Effects of electrical stimulation of the gastric vagal nerves on plasma levels of oxytocin (OXT) and arginine vasopressin (AVP) were examined in rats anesthetized with urethane. Electrical stimulation of the gastric vagal nerves increased the plasma levels of OXT, but not AVP. The concentrations of extracellular noradrenaline (NA) in the paraventricular nucleus (PVN) were measured by in vivo microdialysis in rats anesthetized with urethane. Electrical stimulation of the gastric vagal nerves evoked an increase followed by a slight decrease in the concentrations of NA. The responses of spontaneous firing magnocellular neurosecretory neurons in the PVN to both electrical stimulation of the gastric vagal nerves and intravenous (i.v.) administration of CCK-8 were examined. Most of the putative OXT-secreting cells recorded were excited by both electrical stimulation of gastric vagal nerves and i.v. administration of CCK-8. These results suggest that gastric vagal afferents activate the central noradrenergic system from the brainstem to the PVN and secretion of OXT.     

Cholinergic inputs to rostral ventrolateral medulla pressor neurons from hypothalamus

The rostral ventrolateral medulla (RVLM) has cholinergic mechanisms responsible for pressor responses. Stimulation of the hypothalamic paraventricular nucleus (PVN) causes an increase of arterial pressure via activation of neurons in the RVLM. In this study, we examined whether PVN stimulation causes a pressor response via activation of cholinergic mechanisms in the RVLM. Male Wistar rats were used and they were anesthetized, paralyzed and artificially ventilated. Electrical stimulation of the PVN produced a pressor response. Microinjection of the muscarinic receptor antagonist scopolamine and the cholinesterase inhibitor physostigmine into the RVLM inhibited and potentiated, respectively, the pressor response induced by PVN stimulation. PVN stimulation also increased the firing rate of RVLM barosensitive neurons and the increase in the firing rate was inhibited and potentiated by scopolamine and physostigmine, respectively, iontophoretically applied on neurons. Microinjection of L-glutamate into the PVN produced a release of ACh in the RVLM. The inhibitory amino acid gamma-aminobutyric acid injected into the lateral parabrachial nucleus (LPBN) inhibited the pressor response induced by PVN stimulation. These results suggest that PVN stimulation causes an increase in arterial pressure via activation of cholinergic inputs in the RVLM. It appears that the pressor response is mediated, at least in part, via cholinergic inputs from the LPBN.     

Central Endogenous Vasopressin Induced by Central Salt-Loading Participates in Body Fluid Homeostasis through Modulatory Effects on Neurones of the Paraventricular Nucleus in Conscious Rats

Peripherally secreted arginine vasopressin (AVP) plays a role in controlling body fluid homeostasis, and central endogenous AVP acts as a neurotransmitter or neuromodulator. The limbic system, which appears to exert an inhibitory effect on the endocrine hypothalamus, is also innervated by fibres that contain AVP. We examined whether central endogenous AVP is also involved in the control of body fluid homeostasis. To explore this possibility, we examined neuronal activity in the paraventricular nucleus of the hypothalamus (PVN), periventricular parts of the PVN and limbic brain areas, as well as AVP mRNA expression in the PVN and the peripheral secretion of AVP after central salt-loading in rats that had been pretreated i.c.v. with the AVP V₁ receptor antagonist OPC-21268. Neuronal activity in the PVN evaluated in terms of Fos-like immunoreactivity (FLI), especially in the parvocellular subdivisions, was suppressed. On the other hand, FLI was enhanced in the lateral septum, the bed nucleus of the stria terminalis and the anterior hypothalamic area. Similarly, AVP mRNA expression was enhanced in the magnocellular subnucleus of the PVN, despite the lack of a significant difference in the peripheral AVP level between OPC-21268- and vehicle-pretreated groups. We recorded renal sympathetic nerve activity (RSNA) as sympathetic nerve outflow during central salt-loading. The suppression of RSNA was significantly attenuated by i.c.v. pretreatment with OPC-21268. These results suggest that the suppression of RSNA during central salt-loading might be the result of a decrease in neuronal activity in the parvocellular subdivisions of the PVN via the inhibitory action of central endogenous AVP. The parvocellular and magnocellular neurones in the PVN might show different responses to central salt-loading to maintain body fluid homeostasis as a result of the modulatory role of central endogenous AVP.     

A comparison of the hemodynamic effects produced by electrical stimulation of subnuclei of the paraventricular nucleus

The paraventricular nucleus of the hypothalamus (PVN) is composed of magnocellular and parvocellular subdivisions. Magnocellular neurosecretory neurons project to the neurohypophysis while parvocellular neurons send monosynaptic axonal projections to autonomic regulatory areas in the brainstem and spinal cord. In the present study, we investigated the hemodynamic effects produced by selective magnocellular or parvocellular stimulation. In urethane anesthetized rats with intact baroreflexes, magnocellular and parvocellular stimulation produced only slight differences in hemodynamic responses, however, following acute sinoaortic denervation a clear difference was observed. Parvocellular stimulation produced an increase in arterial pressure and vasoconstriction in gut, kidney and skeletal muscle. Magnocellular stimulation produced little effect on arterial pressure and marked vasodilation in the hindquarters. Blockade of peripheral vasopressin vascular receptors did not affect the vasoconstrictor response produced by stimulation of PVN. These data are consistent with the hypothesis that the long descending neural projections of the parvocellular PVN subserve a selective vasoconstrictor function.     

Electrophysiological evidence that noradrenergic afferents selectively facilitate the activity of supraoptic vasopressin neurons

The functional role of the ascending projection from A1 noradrenergic neurons of the caudal ventrolateral medulla to the supraoptic nucleus of the hypothalamus was investigated by examining the effects of electrical stimulation of the A1 region on the activity of supraoptic neurons deemed to be vasopressinergic or oxytocinergic on the basis of basal firing patterns and responsivity to baroreceptor activation. A1 stimulation enhanced the activity of all putative vasopressin-secreting supraoptic neurons tested. This effect appeared to be selective in that no putative oxytocin-secreting neurons were excited by A1 stimulation. Destruction of the supraoptic noradrenergic terminal plexus by local application of the neurotoxin 6-hydroxydopamine abolished the facilitatory effects of A1 stimulation but did not noticeably alter basal activity patterns, nor the influence of baroreceptor inhibitory pathways. These findings suggest a facilitatory role for noradrenergic afferents in regulating the activity of neurohypophysially-projecting vasopressin neurons of the supraoptic nucleus.     

Characterization of hypothalamic noradrenaline receptors in the supraoptic nucleus and periventricular region of the paraventricular nucleus of mice in vitro

In an attempt to determine the basis for apparently conflicting reports of the effects of noradrenaline (NA) on the neurohypophyseal system and its effects on the parvocellular periventricular region of the paraventricular nucleus (PVN), recordings were made from the neurons in the supraoptic nucleus (SON) and the periventricular region in the mouse hypothalamic slice preparation. Of 47 SON neurons, 43 (91%) were excited and two (4%) were inhibited by NA. Seven SON neurons increased the firing rate with increase of NA concentration (10(-7)-10(-4) M). Both the alpha 1-agonists phenylephrine and methoxamine also increased the activity of all SON neurons tested whereas application of the alpha 2-agonist clonidine and the beta-agonist isoproterenol had weak and inconsistent effects. While the alpha 2-antagonist yohimbine had no consistent influence, the alpha 1-antagonist prazosin blocked or reversed the effects of NA. Another group of 37 neurons in the periventricular region of the PVN was also tested; 13 (35%) were excited and 22 (59%) inhibited by application of NA (10(-5) M). When tested with phenylephrine or methoxamine, 6 of the 7 neurons were excited and one inhibited but all the 4 neurons tested were excited by isoproterenol. Clonidine strongly depressed the activity of all 12 neurons tested. The NA-induced excitatory effects were suppressed or reversed by pre-application of prazosin and the beta-antagonist propranolol while the inhibitory ones were suppressed or reversed by yohimbine. Synaptic blockade did not affect the excitatory responses of SON cells to NA nor the inhibitory responses of periventricular neurons to NA or clonidine. We conclude that SON neurons receive adrenergic excitatory effects mainly through alpha 1-receptors. The periventricular neurons receive the excitatory effects through alpha 1- or beta-receptors and receive the inhibitory effects through alpha 2-receptors.     

Lateral hypothalamic area orexin-A influence the firing activity of gastric distension-sensitive neurons and gastric motility in rats

The orexins system consists of two G-protein coupled receptors (the orexin-1 and the orexin-2 receptor) and two neuropeptides, orexin-A and orexin-B. Orexin-A is an excitatory neuropeptide that regulates arousal, wakefulness and appetite. Recent studies have shown that orexin-A may promote gastric motility. We aim to explore the effects of orexin-A on the gastric -distension (GD) sensitive neurons and gastric motility in the lateral hypothalamic area (LHA), and the possible regulation by the paraventricular nucleus (PVN). Extracellular single unit discharges were recorded and the gastric motility was monitored by administration of orexin-A into the LHA and electrical stimulation of the PVN. There were GD neurons in the LHA, and administration of orexin-A to the LHA could increase the firing rate of both GD-excitatory (GD-E) and GD-inhibited (GD-I) neurons. The gastric motility was significantly enhanced by injection of orexin-A into the LHA with a dose dependent manner, which could be completely abolished by pre-treatment with orexin-A receptor antagonist SB334867. Electrical stimulation of the PVN could significantly increase the firing rate of GD neurons responsive to orexin-A in the LHA as well as promote gastric motility of rats. However, those effects could be partly blocked by pre-treatment with SB334867 in the LHA. It is suggested that orexin-A plays an important role in promoting gastric motility via LHA. The PVN may be involved in regulation of LHA on gastric motility.     

Activation of supraoptic magnocellular neurons by gamma 2-melanocyte stimulating hormone (gamma 2-MSH)

The effects of intracarotid infusions of the peptide gamma₂-melanocyte stimulating hormone (γ₂-MSH) on electrophysiologically and immunohistochemically identified supraoptic nucleus (SON) units were investigated Over a wide dose range this agent always excited SON units, while control infusions of vehicle had no effect. Because neural responses invariably preceded blood pressure elevation, it appears that γ₂-MSH excitation of the magnocellular system was due to a direct effect on the central nervous system and was not a result of systemic cardiovascular responses. These results suggest a forebrain γ₂-MSH sensitive site in the activation of SON magnocellular neurons.     

Differential regulation of parvocellular neuronal activity in the paraventricular nucleus of the hypothalamus following single vs. repeated episodes of water restriction-induced drinking

Acute activation of the hypothalamic-pituitary-adrenal (HPA) axis releases glucocorticoids to maintain homeostasis, whereas prolonged exposure to elevated glucocorticoids has deleterious effects. Due to the potential benefits of limiting stress-induced glucocorticoid secretion, the present study uses drinking in dehydrated rats as a model to delineate mechanisms mobilized to rapidly inhibit HPA activity during stress. Using Fos expression as an indicator of neuronal activation, the effect of a single or repeated episode of dehydration-induced drinking on the activity of magnocellular and parvocellular neurons in the paraventricular nucleus (PVN) of the hypothalamus was examined. Adult male rats underwent a single episode or repeated (six) episodes of water restriction and were sacrificed before or after drinking water in the AM. Plasma osmolality, vasopressin (AVP), adrenocorticotropic hormone (ACTH) and corticosterone were elevated by water restriction and reduced after drinking in both models. Fos expression was elevated in AVP-positive magnocellular PVN neurons and AVP- and corticotropin releasing hormone (CRH)-positive parvocellular PVN neurons after water restriction. Fos expression was reduced in magnocellular AVP neurons after both models of restriction-induced drinking. In contrast, Fos expression did not change in AVP and CRH parvocellular neurons after a single episode of restriction-induced drinking, but was reduced after repeated episodes of restriction-induced drinking. These data indicate that drinking-induced decreases in glucocorticoids in dehydrated rats involve multiple factors including reduction in magnocellular release of vasopressin and reduction in parvocellular neuronal activity. The differential inhibition of PVN parvocellular neurons after repeated rehydration may reflect a conditioned response to repeated stress reduction.     

Prolactin and corticosterone secretion in response to acute stress after paraventricular nucleus lesion by ibotenic acid

The cellular organization of the paraventricular nucleus (PVN) is complex and eight distinct regions have been identified by Nissl staining. Three consist of magnocellular neurons and five of parvocellular neurons. Ibotenic acid, a glutamate analogue, is a toxin with neuroexcitatory properties which acts on N-methyl-D-aspartate and metabotropic receptors. Depending on the dose used, ibotenic acid causes extensive damage of parvocellular neurons of the paraventricular nucleus but preserves magnocellular neurons and passage fibers, in contrast to electrolytic lesions, which causes diffuse and nonspecific destruction. We studied the prolactin (PRL) and corticosterone secretion in response to acute stress induced by exposure to the ether, 3 weeks after selective neurotoxic lesion of parvocellular neurons of the paraventricular nucleus by microinjection of ibotenic acid. There was no significant difference in the basal levels of PRL and corticosterone between control and lesioned animals. The plasma PRL increased in the sham and lesioned groups after stress of similar manner. However, the increase in plasma corticosterone in response to stress was significantly higher in lesioned animals. In conclusion, the selective lesion of parvocellular neurons of the PVN did not change basal or stress induced PRL secretion but it caused hypersensitivity of the hypothalamus-pituitary-adrenal axis 3 weeks later, probably by corticotropin releasing hormone (CRH) from hypothalamic areas others than parvocellular neurons of the PVN; hypersensitivity of corticotropes to the secretagogues others than CRH; or hyperresponsiveness of AVP receptors in the adenohypophysis. Furthermore, we cannot rule out a putative inhibitory factor of the hypothalamus-pituitary axis produced by parvocellular neurons of the PVN. This factor modulator of corticotropin secretion could be absent after recuperation of the response of the hypothalamus-pituitary axis to the stress.     

Catecholaminergic projections to tuberoinfundibular neurones of the paraventricular nucleus: I. Effects of stimulation of A1, A2, A6 and C2 cell groups

Extracellular electrical activity was recorded from 203 paraventricular nucleus (PVN) neurones antidromically identified as projecting to the median eminence. Spontaneous activity and the effects of stimulation of the A1, A2, A6 and C2 catecholaminergic cell groups upon the PVN neurones were examined. Cells were located at a mean height 2.29 +/- 0.03 mm above the base of the brain, corresponding with the corticotropin-releasing factor (CRF) rich component of the nucleus. The mean firing rate was 3.2 +/- 0.3 Hz and antidromic invasion latency was 9.9 +/- 0.3 msec. Seventy-six % of cells tested were activated by painful somatosensory stimuli. Electrical stimulation of the A1 or A2 region evoked excitatory responses from the majority of cells tested (76% and 85%, respectively), whilst stimulation of the A6 and C2 regions evoked more inhibitory responses (43% and 59%, respectively). Most responses (56%), whether excitatory or inhibitory, were not clearly defined in terms of latency, and were only observed following delivery of 5-10 single shocks at 0.5 Hz. Excitation recorded following A1 and A2 stimulation suggests a facilitatory role for noradrenaline in the regulation of PVN activity. Inhibitory responses following C2 stimulation indicate that adrenaline may serve to inhibit such activity, whilst the more mixed responses following A6 stimulation suggest that the projections of this region differ in some way from those of the A1 and A2 cells. Response reversals were observed, after delivery of higher frequency stimulation, for a substantial proportion (20%) of the cells tested.     

Role of Anterior Peri-Third Ventricular Structures in the Regulation of Supraoptic Neuronal Activity and Neurohypophysical Hormone Secretion in the Rat

Neurohypophysical hormone release, and the electrical activity of single neurons of the supraoptic nucleus, were monitored in urethane-anaesthetized rats. Immediately after electrolytic lesions of the region anterior and ventral to the third ventricle (AV3V region), supraoptic neurons showed little spontaneous activity and their responses to ip injection of hypertonic saline were severely impaired; corresponding deficits were found in the secretion of both oxytocin and vasopressin. Similar deficits in oxytocin secretion were also found in rats following electrolytic lesions which destroyed all or part of the subfornical organ; however the effects of the lesions were not additive: rats with lesions of both the AV3V region and the subfornical organ region showed a similar degree of impairment of osmotically stimulated oxytocin secretion to rats with lesions of either site alone. Such deficits might occur either as a result of destruction of osmoresponsive projections to the magnocellular nuclei, or as a result of destruction of an afferent input which is essential for the full expression of the innate osmosensitivity of supraoptic neurons. To test the latter possibility, supraoptic neurons in AV3V-lesioned rats were activated by continuous application of glutamate, and then tested with ip injection of hypertonic saline. Five of seven cells tested responded significantly to the hyperosmotic stimulus, though the responses were significantly weaker than observed in sham-lesioned rats. We suggest that the innate osmosensitivity of supraoptic neurons does contribute to their responses to systemic osmotic stimulation, but that expression of this innate osmosensitivity requires inputs from the AV3V region and/or the subfornical organ, some of which may also be osmoresponsive. Electrical stimulus pulses applied to the AV3V region influenced the electrical activity of most supraoptic neurons strongly: the predominant response was a short-latency, short-duration inhibition followed by long-latency, long-duration excitation. Whereas intracerebroventricular administration of the angiotensin II antagonist saralasin reduced spontaneous or osmotically induced activity of supraoptic neurons, the neuronal responses to AV3V stimulation were impaired only with relatively high doses of saralasin. We conclude that angiotensin ll-sensitive neurons are an important component of the afferent pathways that sustain the excitability of supraoptic neurons, but that angiotensin is probably not the major transmitter of the projection from the AV3V region to the supraoptic nucleus.     

Nociceptin/orphanin FQ and [Phe(1)psi(CH2-NH)Gly2]nociceptin(1-13)NH2 modulates the activity of hypothalamic paraventricular nucleus neurons in vitro

Nociceptin, also known as orphanin FQ (N/OFQ), an endogenous ligand for the orphan opioid receptor-like(1) (ORL(1)) receptor, is moderately expressed in the hypothalamic paraventricular nucleus (PVN) involved in the integrative control of the function of the endocrine and autonomic nervous systems. Our previous study demonstrated that intracerebroventricular administration of N/OFQ elicits an inhibitory action on the function of the cardiovascular and sympathetic nervous systems in conscious rats. However, the effects of N/OFQ on PVN neurons have not been examined. We investigated the effects of N/OFQ on PVN neurons using a whole-cell patch-clamp recording technique in rat brain slices. N/OFQ (30-1000 nM) hyperpolarized membrane potentials in type 1 and type 2 neurons of the PVN classified by the electrophysiological property. [Phe(1)psi(CH2-NH)Gly2]nociceptin(1-13)NH2 (Phepsi) (1-9 microM), a presumed competitive antagonist of the ORL(1) receptor, also hyperpolarized membrane potential in both types of neurons. In voltage clamp studies, N/OFQ (3-3000 nM) activated a K+ current concentration-dependently in 69.7% of PVN neurons with an EC(50) of 72.4+/-12 nM. Phepsi (100-9000 nM) also activated a K+ current with an EC(50) of 818+/-162 nM in PVN neurons, and significantly reduced the amplitude of the N/OFQ-stimulated current. The N/OFQ-induced current was not antagonized by the classical opioid receptor antagonist naloxone and putative antagonist nocistatin. These findings suggest that N/OFQ may have a functional role in the PVN.     

Involvement of hypothalamic paraventricular nucleus non-N-methyl-d-aspartate receptors in the pressor response to noradrenaline microinjected into the bed nucleus of the stria terminalis of unanesthetized rats

Microinjection of noradrenaline into the bed nucleus of the stria terminalis (BST) has been reported to cause a pressor response in unanesthetized rats, which was shown to be mediated by acute vasopressin release into the systemic circulation. In the present study we verified the involvement of magnocellular neurons of the hypothalamic paraventricular (PVN) or supraoptic (SON) nuclei and the local neurotransmitter involved in the pressor response to noradrenaline microinjection into the BST. The PVN pretreatment with the non-selective neurotransmission blocker CoCl₂ (1 nmol/100 nL) inhibited the noradrenaline-evoked pressor response. However, responses were not affected by SON treatment with CoCl₂. Further experiments were carried out to test if glutamatergic neurotransmission in the PVN mediates the pressor response evoked by noradrenaline microinjection into the BST. Pretreatment of the PVN with the selective N -methyl- d -aspartate (NMDA) receptor antagonist LY235959 (2 nmol/100 nL) did not affect the noradrenaline-evoked pressor response. However, PVN pretreatment with the selective non-NMDA receptor antagonist NBQX (2 nmol/100 nL) significantly reduced the pressor response to noradrenaline microinjection into the BST. In conclusion, our results suggest that pressor responses to noradrenaline microinjection into the BST are mediated by PVN magnocellular neurons without involvement of SON neurons. They also suggest that a glutamatergic neurotransmission through non-NMDA glutamate receptors in the PVN mediates the response.