Vasopressin Cells Research Articles

Population Dynamics in Vasopressin Cells

Most neurons sense and code change, and when presented with a constant stimulus they adapt, so as to be able to detect a fresh change. However, for some things it is important to know their absolute level; to encode such information, neurons must sustain their response to an unchanging stimulus while remaining able to respond to a change in that stimulus. One system that encodes the absolute level of a stimulus is the vasopressin system, which generates a hormonal signal that is proportional to plasma osmolality. Vasopressin cells sense plasma osmolality and secrete appropriate levels of vasopressin from the neurohypophysis as needed to control water excretion; this requires sustained secretion under basal conditions and the ability to increase (or decrease) secretion should plasma osmolality change. Here we explore the mechanisms that enable vasopressin cells to fulfill this function, and consider how coordination between the cells might distribute the secretory load across the population of vasopressin cells.

Immunoreactive vasoactive intestinal polypeptide and vasopressin cells after a protein malnutrition diet in the suprachiasmatic nucleus of the rat

The aim of the present study was to evaluate the effects of prenatal and postnatal protein deprivation on the morphology and density of vasopressin (VP) and vasoactive intestinal polypeptide (VIP) immunoreactive neurons in the suprachiasmatic nucleus (SCN) of young rats. Female Wistar rats were fed either 6% (malnourished group) or 25% (control group) casein diet five weeks before conception, during gestation and lactation. After weaning, the pups were maintained on the same diet until sacrificed at 30 days of age. The major and minor axes, somatic area and the density of VP- and VIP-immunoreactive neurons were evaluated in the middle sections of the SCN. The present study shows that chronic protein malnutrition (ChPM) in VP neurons induces a significant decrease in number of cells (-31%,) and a significant increase in major and minor axes and somatic area (+12.2%, +21.1% and +15.0%, respectively). The VIP cells showed a significant decrease in cellular density (-41.5%) and a significant increase in minor axis (+13.5%) and somatic area (+10.1%). Our findings suggest that ChPM induces abnormalities in the density and morphology of the soma of VP and VIP neurons. These alterations may be a morphological substrate underlying circadian alterations previously observed in malnourished rats.

Sexual Differentiation of Vasopressin Innervation of the Brain: Cell Death Versus Phenotypic Differentiation

In most vertebrates studied, males have more vasopressin (VP) cells in the bed nucleus of the stria terminalis, or homologous vasotocin cells in nonmammalian species, than females. Previous research excluded differential cell birth and migration as likely mechanisms underlying this difference, leaving just differential cell death and phenotypic differentiation of existing cells. To differentiate between these remaining possibilities, we compared VP cell number in wild-type mice vs. mice overexpressing the anti-cell death factor, Bcl-2. All animals were gonadectomized in adulthood and given testosterone capsules. Three weeks later, brains were processed for in situ hybridization to identify VP cells. Bcl-2 overexpression increased VP cell number in both sexes but did not reduce the sex difference. We repeated this experiment in mice with a null mutation of the pro-cell death gene, Bax, and obtained similar results; cell number was increased in Bax(-/-) mice of both sexes, but males had about 40% more VP cells, regardless of Bax gene status. Taken together, cell death is unlikely to account for the sex difference in VP cell number, leaving differentiation of cell phenotype as the most likely underlying mechanism. We also used immunocytochemistry to examine VP projections in Bcl-2-overexpressing mice. As expected, males showed denser VP-immunoreactive fibers than females in the lateral septum, a projection area of the bed nucleus of the stria terminalis. However, even though Bcl-2 overexpression increased VP cell number, it did not affect fiber density. Thus, a compensatory mechanism may control total septal innervation regardless of the number of contributing cells.

Absence of progestin receptors alters distribution of vasopressin fibers but not sexual differentiation of vasopressin system in mice

Perinatal estrogens increase the number of vasopressin-expressing cells and the density of vasopressin-immunoreactive fibers observed in adult male rodents. The mechanism of action of estrogens on sexual differentiation of the extra-hypothalamic vasopressin system is unknown. We hypothesized that the sexually dimorphic expression of progestin receptors (PRs) during development would masculinize vasopressin expression in mice. We compared the number of vasopressin-expressing cells in the bed nucleus of the stria terminalis (BNST) and medial amygdala and the density of vasopressin-immunoreactive fibers in several brain regions of male and female wild type and PRKO mice using in situ hybridization and immunohistochemistry. As expected, sex differences in vasopressin cell number were observed in the BNST and medial amygdaloid nucleus. Vasopressin-immunoreactive fiber density was sexually dimorphic in the lateral septum, lateral habenular nucleus, medial amygdaloid nucleus, and mediodorsal thalamus. Sex differences were also observed in the principal nucleus of the BNST and medial preoptic area but not in the dorsomedial hypothalamus, which are thought to receive vasopressin innervation from the suprachiasmatic nucleus. Deletion of PRs did not alter the sex difference in vasopressin mRNA expression and vasopressin fiber immunoreactivity in any area examined. However, deletion of PRs increased the density of vasopressin fiber immunoreactivity in the lateral habenular nucleus. Our data suggest that PRs modulate vasopressin levels, but not sexual differentiation of vasopressin innervation in mice.

Effects of selective Bcl-2 inhibitor HA14-1 treatments on functional activity of magnocellular vasopressinergic neurons of rat hypothalamus

In this study we examined whether in vivo treatments with Bcl-2 inhibitor HA14-1 can affect the function of vasopressinergic system of rat. HA14-1 is a novel organic compound that has micromolar affinity for Bcl-2 and Bcl-xL and acts as a mimetic of BH3-only proteins by antagonizing the anti-apoptotic Bcl-2 proteins and triggering Bax-dependent apoptosis. We found that intrahypothalamic injections of HA14-1 did not induce apoptosis of vasopressin (VP) cells of supraoptic nucleus, but led to activation of VP synthesis and release, resulting in decreased diuresis. Our data has also demonstrated that injections of HA14-1 increased phospho-MEK1/2, phospho-CREB and phospho-Elk-1 levels in magnocellular neurons. Thus we propose that injections of HA14-1 into the hypothalamus do not lead to neuronal death, but change the functional activity of VP neurons of hypothalamus centres.

Vasopressin and neuronal nitric oxide synthase in hypothalamic paraventricular nucleus (PVN) of pregnant rats.

Decreased baseline GABAergic sympathoinhibition within the PVN of pregnant (P) rats is associated with decreased neuronal nitric oxide synthase (nNOS) activity (NADPH Diaphorase), and presumably decreased GABA release. nNOS is colocalized with vasopressin (VP) in both parvo‐ and magno‐cellular PVN neurons. Our goal was to determine if pregnancy changed the number of spinally projecting PVN neurons (presympathetic) containing nNOS or VP. The retrograde tracer fluorogold (FG) was injected unilaterally into the spinal cord of female nonpregnant (NP, n=3) and P (n=3) rats. After 7 days, ipsilateral FG and immunohistochemical staining for VP and nNOS in PVN sections (30 ìm) was counted at 3 levels: rostral (lev‐1) and intermediate (lev‐2) PVN contained the posterior magnocellular cell group, while lev‐3 (caudal) contained the majority of spinally projecting (FG) neurons. In both groups, distribution of VP cells was: lev2 > lev 1 > lev 3. Approximately 7% of all VP cells were spinally projecting, with the majority at level 3 in both groups (~ 19%). Of spinally projecting cells the % also containing nNOS tended to be less in P (16 ±2.9 %) compared to NP rats (24 ±2.4 %). Although preliminary, these data are consistent with decreased nNOS in presympathetic neurons in the PVN of P rats which could contribute to the previously reported decrease in GABAergic sympathoinhibition in the PVN in pregnancy. (HL36245; HL55306)

Anti-hypothalamus and anti-pituitary antibodies may contribute to perpetuate the hypopituitarism in patients with Sheehan's syndrome

While anti-pituitary antibodies (APAs) were detected in some patients with Sheehan's syndrome (SS) suggesting an autoimmune pituitary involvement in the development of their hypopituitarism, hypothalamic cell anti-hypothalamus antibodies (AHAs) have not been investigated so far. The aim of this study was to evaluate the presence of AHA and APA in SS patients to verify whether an autoimmune hypothalamic-pituitary process can contribute to their late hypopituitarism. Twenty women with SS with a duration of disease ranging from 3 to 40 years (median 25.5 years) were enrolled into the study. Out of 20 patients, 12 (60%) had panhypopituitarism and the others had partial hypopituitarism well corrected with appropriate replacement therapy. None of them had clinical central diabetes insipidus. AHA and APA were investigated by immunofluorescence method in all patients. In addition, a four-layer immunofluorescence method was used to verify whether AHA immunostained vasopressin-secreting cells (AVP-c) or not. AHAs were found in 8 out of 20 (40%) and APAs in 7 out of 20 (35%) patients with titers ranging from 1:32 to 1:128 and 1:16 to 1:32 respectively; however, in none of these positive patients AHA immunostained vasopressin cells. None of controls resulted positive for both antibodies. Patients with SS, even many years after the onset of SS, can show antibodies to pituitary and/or hypothalamic but not AVP-secreting cells. Antibodies to unknown hypothalamic cells (releasing factor-secreting cells) other than APAs suggest that an autoimmune process involving both the hypothalamus and pituitary gland may contribute to late pituitary dysfunction in SS patients.

Adiponectin selectively inhibits oxytocin neurons of the paraventricular nucleus of the hypothalamus

Adiponectin is an adipocyte derived hormone which acts in the brain to modulate energy homeostasis and autonomic function. The paraventricular nucleus of the hypothalamus (PVN) which plays a key role in controlling pituitary hormone secretion has been suggested to be a central target for adiponectin actions. A number of hormones produced by PVN neurons have been implicated in the regulation of energy homeostasis including oxytocin, corticotropin releasing hormone and thyrotropin releasing hormone. In the present study we investigated the role of adiponectin in controlling the excitability of magnocellular (MNC--oxytocin or vasopressin secreting) neurons within the PVN. Using RT-PCR techniques we have shown expression of both adiponectin receptors in the PVN. Patch clamp recordings from MNC neurons in hypothalamic slices have also identified mixed (27% hyperpolarization, 42% depolarization) effects of adiponectin in modulating the excitability of the majority of MNC neurons tested. These effects are maintained when cells are placed in synaptic isolation using tetrodotoxin. Additionally we combined electrophysiological recordings with single cell RT-PCR to examine the actions of adiponectin on MNC neurons which expressed oxytocin only, vasopressin only, or both oxytocin and vasopressin mRNA and assess the profile of receptor expression in these subgroups. Adiponectin was found to hyperpolarize 100% of oxytocin neurons tested (n = 6), while vasopressin cells, while all affected (n = 6), showed mixed responses. Further analysis indicates oxytocin neurons express both receptors (6/7) while vasopressin neurons express either both receptors (3/8) or one receptor (5/8). In contrast 6/6 oxytocin/vasopressin neurons were unaffected by adiponectin. Co-expressing oxytocin and vasopressin neurons express neither receptor (4/6). The results presented in this study suggest that adiponectin plays specific roles in controlling the excitability oxytocin secreting neurons, actions which correlate with the current literature showing increased oxytocin secretion in the obese population.

Heterogeneous expression of γ‐aminobutyric acid and γ‐aminobutyric acid‐associated receptors and transporters in the rat suprachiasmatic nucleus

The hypothalamic suprachiasmatic nucleus (SCN) is the primary mammalian circadian clock that regulates rhythmic physiology and behavior. The SCN is composed of a diverse set of neurons arranged in a tight intrinsic network. In the rat, vasoactive intestinal peptide (VIP)- and gastrin-releasing peptide (GRP)-containing neurons are the dominant cell phenotypes of the ventral SCN, and these cells receive photic information from the retina and the intergeniculate leaflet. Neurons expressing vasopressin (VP) are concentrated in the dorsal and medial aspects of the SCN. Although the VIP/GRP and VP cell groups are concentrated in different regions of the SCN, the separation of these cell groups is not absolute. The inhibitory neurotransmitter gamma-aminobutyric acid (GABA) is expressed in most SCN neurons irrespective of their location or peptidergic phenotype. In the present study, immunoperoxidase labeling, immunofluorescence confocal microscopy, and ultrastructural immunocytochemistry were used to examine the spatial distribution of several markers associated with SCN GABAergic neurons. Glutamate decarboxylase, a marker of GABA synthesis, and vesicular GABA transporter were more prominently observed in the ventral SCN. KCC2, a K(+)/Cl(-) cotransporter, was highly expressed in the ventral SCN in association with VIP- and GRP-producing neurons, whereas VP neurons in the dorsal SCN were devoid of KCC2. On the other hand, GABA(B) receptors were observed predominantly in VPergic neurons dorsally, whereas, in the ventral SCN, GABA(B) receptors were associated almost exclusively with retinal afferent fibers and terminals. The differential expression of GABAergic markers within the SCN suggests that GABA may play dissimilar roles in different SCN neuronal phenotypes.

SCN Organization Reconsidered

The SCN has long had organizational schemas imposed on it. In most, the SCN is dichotomized, with one region typically associated with the presence of vasopressin cells and the other associated with cells containing vasoactive intestinal polypeptide and certain afferent terminal fields. If assumed to be accurate, the schemas that have been intended to simplify and conceptually organize the known anatomy may actually interfere with the understanding of how various cell types and input pathways contribute to circadian rhythm regulation. This review describes inadequacies of existing schemas and notes several practical difficulties that undermine their usefulness. These include "static" versus "dynamic" anatomy, generalizations about SCN organization in relation to the plane or level of section, and the concept of differential density, all of which contribute to a view in which the SCN is substantially more complex than typically depicted in oversimplified line drawings. The need for accurate topographical description is emphasized.

Laser Capture Microscopy (LCM) and real time RT‐PCR: Gene expression in PVN of transgenic vasopressin‐enhanced green fluorescent protein (VP‐eGFP) rats

Water deprivation decreases plasma volume and increases plasma osmolality and vasopressin (VP) release. Immunohistochemistry and in situ hybridization indicate that osmotic stimuli deplete magnocellular neurons of VP content, while increasing VP gene expression. The current experiments tested feasibility of using LCM and real time RT-PCR to evaluate mRNA expression in hypothalamic paraventricular nucleus (PVN) VP cells. Transgenic VP-eGFP Wistar rats were water deprived for 48 hrs. VP-eGFP expressing cells in the magnocellular PVN from euhydrated (E, n= 3) and dehydrated (D, n= 3) rats were identified in frozen hypothalamic sections (5 μm) with fluorescent microscopy and laser dissected (spot size~7.5 μm; power = 75mW) onto LCM caps (Arcturus). RNA was extracted, cDNA synthesized and real time RT-PCR was performed for mRNA’s of the ribosomal protein L-32 (housekeeper), and VP. Relative expression of VP was increased 3.5 X in the PVN of D compared to E rats. In contrast, expression of GAPDH (another housekeeping gene) relative to L-32 in PVN VP neurons was not different between D and E rats. Thus, the use of brain tissue from VP-eGFP rats circumvents potential RNA degradation during standard post-sectioning immunohistochemistry and allows for evaluation of mRNA expression of target genes in identified VP neurons. (NIH R01 HL36245, CMH and R01 HL68725, JES)

Bistability with Hysteresis in the Activity of Vasopressin Cells

Magnocellular vasopressin neurones generate distinctive 'phasic' patterns of electrical activity during which periods of spiking activity (bursts) alternate with periods of no spikes or occasional spikes. The mechanisms of burst termination in vivo are still not clearly understood. We recorded from single phasic vasopressin cells in vivo and here we show that burst terminations in some phasic cells is preceded by transient increases in activity, consistent with bursts ending as a result of activity-dependent inhibition. We show that extrinsically imposed increases in activity, evoked by brief stimulation of the organum vasculosum of the lamina terminalis, can either trigger bursts if given when a cell is silent, or stop bursts if given when a cell is active. Thus, the magnocellular vasopressin system is a population of independent bistable oscillators. The population as a whole is insensitive to transient changes in input level, whether these are excitatory or inhibitory. The vasopressin cell population thus acts like a 'low-pass filter'; although brief large changes in input rate have little overall effect, the population responds very effectively to small, sustained changes in input rate by evolving a pattern of discharge activity that efficiently maintains secretion. We note that these filtering characteristics are the opposite of the filtering characteristics that are typically associated with neurones.

Distribution of vasopressin in the forebrain of spotted hyenas

The extreme virilization of the female spotted hyena raises interesting questions with respect to sexual differentiation of the brain and behavior. Females are larger and more aggressive than adult, non-natal males and dominate them in social encounters; their external genitalia also are highly masculinized. In many vertebrates, the arginine vasopressin (VP) innervation of the forebrain, particularly that of the lateral septum, is associated with social behaviors such as aggression and dominance. Here, we used immunohistochemistry to examine the distribution of VP cells and fibers in the forebrains of adult spotted hyenas. We find the expected densely staining VP immunoreactive (VP-ir) neurons in the paraventricular and supraoptic nuclei, as well as an unusually extensive distribution of magnocelluar VP-ir neurons in accessory regions. A small number of VP-ir cell bodies are present in the suprachiasmatic nucleus and bed nucleus of the stria terminalis; however, there are extensive VP-ir fiber networks in presumed projection areas of these nuclei, for example, the subparaventricular zone and lateral septum, respectively. No significant sex differences were detected in the density of VP-ir fibers in any area examined. In the lateral septum, however, marked variability was observed. Intact females exhibited a dense fiber network, as did two of the four males examined; the two other males had almost no VP-ir septal fibers. This contrasts with findings in many other vertebrate species, in which VP innervation of the lateral septum is consistently greater in males than in females.

Activity-dependent feedback modulation of spike patterning of supraoptic nucleus neurons by endogenous adenosine

Neuropeptide secretion from the dendrites of hypothalamic magnocellular supraoptic nucleus (SON) neurons contributes to the regulation of neuronal activity patterning, which ultimately determines their peptide output from axon terminals in the posterior pituitary gland. SON dendrites also secrete a number of other neuromodulators, including ATP. ATP degrades to adenosine in the extracellular space to complement transported adenosine acting on pre- and postsynaptic SON A1 receptors to reduce neuronal excitability, measured in vitro. To assess adenosine control of electrical activity in vivo, we made extracellular single-unit recordings of the electrical activity of SON neurons in anesthetized male rats. Microdialysis application (retrodialysis) of the A1 receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT) increased phasic vasopressin cell intraburst firing rates progressively over the first 5 s by 4.5 +/- 1.6 Hz (P < 0.05), and increased burst duration by 293 +/- 64% (P < 0.05). Hazard function plots were generated from interval interspike histograms and revealed that these effects were associated with increased postspike excitability. In contrast, CPT had no effect on the firing rates and hazard function plot profiles of continuously active vasopressin and oxytocin cells. However, CPT significantly increased clustering of spikes, as quantified by the index of dispersion, in oxytocin cells and continuously active vasopressin cells (by 267 +/- 113% and 462 +/- 67%, respectively, P < 0.05). Indeed, in 4 of 5 continuously active vasopressin cells, CPT induced a pseudophasic activity pattern. Together, these results indicate that endogenous adenosine is involved in the local control of SON cell activity in vivo.

Influence of Dehydration on the Expression of Neuropeptide Y Y1 Receptors in Hypothalamic Magnocellular Neurons

Regulation of vasopressin (VP) and oxytocin (OT) secretion involves integration of neural signals from hypothalamic osmoreceptors, ascending catecholaminergic and peptidergic cell groups in the brain stem, and local and autoregulatory afferents. Neuropeptide Y (NPY) is one factor that stimulates the release of VP and OT from the supraoptic (SON) and paraventricular nuclei of the hypothalamus via activation of Y1 receptors (Y1R). The current studies were designed to assess the regulation and distribution of NPY Y1R expression in the SON of male rats that were either given 2% NaCl drinking water (24-72 h) or water deprived (48 h). Subjecting male rats to these conditions resulted in significant increases in both the number of cells expressing Y1R immunoreactivity (ir) and the amount of Y1R protein per cell within the SON. Y1R immunoreactivity was increased in the magnocellular but not medial parvocellular paraventricular nuclei, and Y1R mRNA levels were increased in the SON of salt-loaded rats. Subpopulations of both VP and OT cells in the hypothalamus express Y1R immunoreactivity and a greater percentage of VP-ir cells express Y1R after salt loading. To control for potential effects of dehydration-induced anorexia, a group of euhydrate animals was pair fed with animals consuming 2% NaCl. No detectable change in Y1R expression was observed in the SON of pair-fed animals, even though body weights were significantly lower than controls. These data demonstrate that NPY Y1R gene and protein expression are increased in the SON of salt-loaded and water-deprived animals and provide a mechanism whereby NPY can support VP/OT release during prolonged challenges to fluid homeostasis.

Bistability in the activity of vasopressin cells

Bistability in the activity of vasopressin cells

Endogenous Activation of Supraoptic Nucleus κ-Opioid Receptors Terminates Spontaneous Phasic Bursts in Rat Magnocellular Neurosecretory Cells

Phasic activity in magnocellular neurosecretory vasopressin cells is characterized by alternating periods of activity (bursts) and silence. During phasic bursts, action potentials (spikes) are superimposed on plateau potentials that are generated by summation of depolarizing after-potentials (DAPs). Burst termination is believed to result from autocrine feedback inhibition of plateau potentials by the kappa-opioid peptide, dynorphin, which is copackaged in vasopressin neurosecretory vesicles and exocytosed from vasopressin cell dendrites during phasic bursts. Here we tested this hypothesis, using intracellular recording in vitro to show that kappa-opioid receptor antagonist administration enhanced plateau potential amplitude to increase postspike excitability during spontaneous phasic activity. The antagonist also increased postburst DAP amplitude in vitro, indicating that endogenous dynorphin probably reduces plateau potential amplitude by inhibiting the DAP mechanism. However, the kappa-opioid receptor antagonist did not affect the slow depolarization that follows burst termination, suggesting that recovery from endogenous kappa-opioid inhibition does not contribute to the slow depolarization. We also show, by extracellular single-unit recording, that that there is a strong random element in the timing of burst initiation and termination in vivo. Administration of a kappa-opioid receptor antagonist eliminated the random element of burst termination but did not alter the timing of burst initiation. We conclude that dendritic dynorphin release terminates phasic bursts by reducing the amplitude of plateau potentials to reduce the probability of spike firing as bursts progress. By contrast, dendritic dynorphin release does not greatly influence the membrane potential between bursts and evidently does not influence the timing of burst initiation.

Localization and osmotic regulation of vesicular glutamate transporter-2 in magnocellular neurons of the rat hypothalamus

In this report we present immunocytochemical and in situ hybridization evidence that magnocellular vasopressin and oxytocin neurons in the hypothalamic supraoptic and paraventricular nuclei express type-2 vesicular glutamate transporter, a marker for their glutamatergic neuronal phenotype. To address the issue of whether an increase in magnocellular neuron activity coincides with the altered synthesis of the endogenous glutamate marker, we have introduced a new dual-label in situ hybridization method which combines fluorescent and autoradiographic signal detection components for vasopressin and vesicular glutamate transporter-2 mRNAs, respectively. Application of this technique provided evidence that 2% sodium chloride in the drinking water for 7 days produced a robust and significant increase of vesicular glutamate transporter-2 mRNA in vasopressin neurons of the supraoptic nucleus. The immunocytochemical labeling of pituitary sections, followed by the densitometric analysis of vesicular glutamate transporter-2 immunoreactivity in the posterior pituitary, revealed a concomitant increase in vesicular glutamate transporter-2 protein levels at the major termination site of the magnocellular axons. These data demonstrate that magnocellular oxytocin as well as vasopressin cells contain the glutamatergic marker vesicular glutamate transporter-2, similarly to most of the parvicellular neurosecretory neurons examined so far. The robust increase in vesicular glutamate transporter-2 mRNA and immunoreactivity after salt loading suggests that the cellular levels of vesicular glutamate transporter-2 in vasopressin neurons are regulated by alterations in water–electrolyte balance. In addition to the known synaptic actions of excitatory amino acids in magnocellular nuclei, the new observations suggest novel mechanisms whereby glutamate of endogenous sources can regulate magnocellular neuronal functions.

Expression of the long form of the prolactin receptor in magnocellular oxytocin neurons is associated with specific prolactin regulation of oxytocin neurons

Magnocellular neurons of the supraoptic (SON) and paraventricular nuclei (PVN) show considerable plasticity during pregnancy and lactation. Prolactin receptors (PRL-R) have been identified in both these nuclei. The aim of this study was to investigate the cell type(s) expressing mRNA for the long form of prolactin receptor (PRL-R(L)) and to determine whether patterns of expression change during pregnancy and lactation. In addition, we examined effects of prolactin on excitability of oxytocin and vasopressin neurons. Sections from brains of nonpregnant, pregnant, and lactating rats were hybridized with an 35S-labeled probe to label PRL-R(L) mRNA together with digoxigenin-labeled probes to detect either oxytocin or vasopressin mRNA. In the SON, PRL-R(L) mRNA was predominantly colocalized with oxytocin mRNA, with over 80% of oxytocin neurons positive for PRL-R(L) mRNA. Very few (<10%) vasopressin neurons expressed PRL-R(L) mRNA. In the PVN, PRL-R(L) mRNA was also predominantly found in oxytocin neurons, and the proportion of PRL-R(L)-positive oxytocin neurons increased significantly during pregnancy and lactation. As in the SON, relatively few vasopressin cells contained PRL-R(L) mRNA. For in vivo electrophysiology, nonpregnant rats were anesthetized, and then extracellular single neuron activity was recorded in identified oxytocin and vasopressin neurons. After a period of baseline recording, the effect of prolactin (1 microg i.c.v.) on firing rate was examined. Prolactin treatment of nonpregnant rats induced a significant decrease in firing rates of oxytocin neurons. There was no effect of prolactin on the activity of vasopressin neurons. Together, these data provide strong evidence that prolactin directly and specifically regulates activity of oxytocin neurons.

Progesterone Treatment of Adult Male Rats Suppresses Arginine Vasopressin Expression in the Bed Nucleus of the Stria Terminalis and the Centromedial Amygdala

The steroid sensitive vasopressin cells of the bed nucleus of the stria terminalis (BST) and centromedial amygdala (CMA) are involved in numerous behavioural and physiological functions. These cells are known to be greatly influenced by gonadal steroids. Castration reduces and testosterone replacement restores arginine vasopressin (AVP)-immunoreactive (-ir) labelling and AVP mRNA expression in the BST and CMA. Gonadal steroids appear to act directly in AVP-expressing cells within the BST and CMA, because the majority of AVP-ir cells in these areas contain oestrogen and androgen receptor immunoreactivity. Recently, we have localised progestin receptor immunoreactivity in virtually all of the AVP-ir cells in the BST and CMA. To understand the role played by progestin receptors in AVP cells within the BST and CMA, we treated male rats with 1 mg of progesterone or oil for 5 days, and then examined AVP immunoreactivity within the brain. We found that progesterone decreased AVP-ir labelling within the BST and CMA, as well as in two of the projection sites of these cells, the lateral septum and lateral habenula. Progesterone treatment did not alter testosterone secretion from the testes, nor did it alter adult male sexual behaviour. These data illustrate an additional mechanism by which the AVP cells in the BST and CMA can be regulated. These data also suggest that progesterone may act in the male brain to influence behaviours that are AVP-dependent.