Showing papers in "Journal of Neuroendocrinology in 1992"

Repeated immobilization stress alters tyrosine hydroxylase, corticotropin-releasing hormone and corticosteroid receptor messenger ribonucleic Acid levels in rat brain.

Abstract: In situ hybridization histochemistry was used to localize and quantify the effects of acute and repeated immobilization stress on mRNA levels of tyrosine hydroxylase (TH) in catecholaminergic neurons in the locus ceruleus and substantia nigra and on mRNA levels of relevant markers of the hypothalamic-pituitary-adrenal axis, namely corticotropin-releasing hormone (CRH) in the hypothalamic paraventricular nucleus (PVN), proopiomelanocortin in the pituitary, and mineralocorticoid receptors (MR, type I) and glucocorticoid receptors (GR, type II) in the hippocampus, PVN and pituitary. Control, acutely stressed (1 × lMO, sacrificed immediately after 2 h of immobilization), and repeatedly stressed (6 × IMO plus delay, sacrificed 24 h after 6 daily 2-h immobilizations and 6 × lMO plus challenge, sacrificed immediately after the seventh daily 2-h immobilization) male Sprague-Dawley rats were examined. TH mRNA expression was increased in the locus ceruleus in the acutely stressed and repeatedly stressed animals. The increase in TH mRNA levels was greatest in the repeatedly stressed (6 × IMO plus challenge) group. TH mRNA levels were not altered in the substantia nigra. CRH mRNA levels in the PVN were significantly increased in the three stressed groups and the increase was greatest in the 6 × IMO plus challenge group. CRH mRNA levels were increased in the central nucleus of the amygdala only after acute stress. Proopiomelanocortin mRNA levels were elevated in the anterior pituitary during acute and repeated stress, but the magnitude of the effect was largest after acute stress. The changes in the hypothalamic-pituitary-adrenal axis were accompanied by an acute stress-induced increase in MR mRNA levels in the hippocampus, MR and GR mRNA levels in the PVN and GR mRNA levels in the pituitary. MR mRNA levels continued to be elevated in the PVN in the 6 × IMO plus challenge animals. Plasma corticosterone levels were elevated in the acute and repeated stress conditions. The results show that repeated immobilization stress produces a rapid and persistent increase in mRNA expression of TH in the locus ceruleus, CRH in the PVN, and proopiomelanocortin in the anterior pituitary. The TH-containing neurons in the locus ceruleus and the CRH-containing neurons in the PVN appear to preserve the capability to respond to repeated stimulation (6 × IMO plus challenge) indicating altered feedback mechanisms under repeated stress conditions. GR and MR mRNA levels are differentially regulated in the hippocampus, PVN and pituitary by acute and repeated stress. It is of interest that the central nervous system systems which are activated during repeated stress, namely the locus ceruleus-norepinephrine system and hypothalamic-pituitary-adrenal axis, are dysregulated in melancholic depression. Further studies of the central nervous system effects of prolonged exposure to stress may help elucidate the mechanisms underlying dysregulation of the locus ceruleus-norepinephrine system and hypothalamic-pituitary-adrenal axis in depression and other stress-related psychiatric diseases.

Estradiol and progesterone modulation of norepinephrine neurotransmission: implications for the regulation of female reproductive behavior.

Abstract: A long range objective of research in behavioral neuroendocrinology is to elucidate the mechanisms by which hormones modify behavior. Significant progress in understanding hormone-brainbehavior relationships is most likely to be achieved if one studies a behavior: 1) which can be measured with a high degree of validity and reliability, 2) whose occurrence can be manipulated in a predictable fashion by hormone administration, and 3) whose neural circuitry, including the necessary and sufficient sensory inputs and motor outputs and the sites of hormone action, has been established. One of the few mammalian behaviors which fits these criteria is the lordosis response of female rodents (for detailed descriptions of the lordosis posture and underlying neural circuitry, see 1). In rodents, the expression of this component of female reproductive (sexual, estrous, mating) behavior is strictly dependent on sequential exposure of neurons in specific hypothalamic sites to the ovarian steroids, estradiol (E,) and progesterone (P) (1-3). It should be emphasized that hormones do not ‘elicit’ or ‘activate’ lordosis; rather, the hormonal milieu determines the probability that animals display the behavior in response to appropriate sensory stimulation (in this case, flank and perineal stimulation). The lordosis response is also of great physiological importance; unless female rodents assume the lordosis posture when mounted by males, penile insertion and hence fertilization cannot take place. In addition, the hormonal control of reproduction in female vertebrates is an elegant example of neuroendocrine integration. Ovarian E2 and P act in extensively interconnected neuronal populations (4) to ensure that the release of pituitary gonadotropins which trigger ovulation (especially luteinizing hormone; LH) coincides with the expression of behavioral receptivity (5) . This neuroendocrine coordination of physiology and behavior maximizes the probability that a female will contact and be inseminated by a conspecific male at the optimal time for achieving fertilization. If one accepts the view that behavior is the product of neuronal activity, then it is reasonable to examine the influence of hormones on brain cells in the neural circuits that mediate hormoneregulated behaviors. Indeed, there is compelling evidence that E2 facilitation of lordosis behavior in rats requires increased excitability of neurons in the ventromedial hypothalamus (VMH) which project to the midbrain central gray, a critical site of sensory and motor integration of lordosis (1, 4, 6, 7). Similarly, it is rational to propose that hormone-dependent changes in chemical neurotransmission in specific neural circuits are likely to produce behavioral changes. In keeping with this perspective, numerous reports of alterations in neurotransmitter metabolism, release, and receptor binding as a function of estrous cycle stage and/or experimental manipulation of circulating E, and P have appeared (5, 8). Likewise, pharmacological manipulation of a variety of neurotransmitter systems can either facilitate or inhibit hormonedependent lordosis (810). Nevertheless, we still have only a rudimentary knowledge of which molecular components of specific neurotransmitter systems are regulated by E, and/or P in brain regions that control reproductive behavior, and of how the regulated molecules alter neuronal function such that stimuli which have a low probability of eliciting lordosis responses in hormone-deprived rodents have a high probability of doing so in hormone-exposed animals. Moreover, the development over the past decade of sensitive new methods for monitoring transmitter release in vivo and for mapping the qualitative and quantitative distribution of putative neurotransmitters, neuromodulators and their receptors in discrete neuronal populations, has produced an explosive increase in the number of potential molecular targets for hormonal regulation. In an attempt to begin developing a coherent picture of the neural mechanisms that mediate ovarian steroid regulation of lordosis, we have elected to focus on the monoamine neurotransmitter norepinephrine (NE). As summarized below, consideration of a variety of neuroendocrine, behavioral, neuroanatomical and neurophysiological observations which had accumulated by the mid 1980s led us to this choice.

Fos-Like Immunoreactivity in the Brainstem of the Rat Following Peripheral Administration of Cholecystokinin

Abstract: Ninety min after intraperitoneal (ip) injection of Cholecystokinin (CCK, 50 μg/kg body wt) Fos-like protein was expressed in cells throughout the nucleus of the tractus solitarii (NTS) and area postrema, and also in scattered cells in the lateral reticular area of the brainstem. Using dual fluorescent immunocytochemistry for Fos-like protein and tyrosine hydroxylase (TH), catecholaminergic cells in the A2 region of the NTS and the A1 region of the lateral reticular area were shown to be activated.

NADPH-Diaphorase (Nitric Oxide Synthase) Staining in the Rat Supraoptic Nucleus is Activity-Dependent: Possible Functional Implications.

Abstract: NADPH-diaphorase has recently been shown to be the enzyme nitric oxide (NO) synthase, and to be present in the rat supraoptic nucleus (SON) and posterior pituitary. Investigations were carried out to assess whether there is any difference in the extent to which this enzyme is present, as assessed by light-microscopic histochemistry, in SON of normal and dehydrated male Wistar rats. In normal rats there was clear cellular heterogeneity; cells located in the ventral and caudal areas of the SON stained only weakly or not at all, while cells in the rostro-dorsal areas of the nucleus stained strongly. Dehydration of rats for 12 h caused a large and rapid increase in staining intensity of the nucleus, particularly of cells in its ventral and caudal parts. On the basis of its known biological actions, and the kinetics of its induction, it is suggested that NO would be a strong candidate as a modulator of SON and posterior pituitary morphology and function, with the potential to rapidly modulate blood flow, neuronal activity, and possibly astrocyte morphology, in response to changes in neuronal activity.

Further Evidence that Most Luteinizing Hormone-Releasing Hormone Neurons are not Directly Estrogen-Responsive: Simultaneous Localization of Luteinizing Hormone-Releasing Hormone and Estrogen Receptor Immunoreactivity in the Guinea-Pig Brain.

Abstract: Gonadotropin secretion from the pituitary is regulated in large part by steroid action on the brain. An important question concerns whether luteinizing hormone-releasing hormone (LHRH) neurons themselves transduce steroid signals, or whether, alternatively, steroid-sensitive interneuronal populations regulate their activity. A previous study in the rat employing steroid autoradiography combined with LHRH immunocytochemistry revealed that only an exceedingly small percentage of LHRH-immunoreactive (ir) neurons was estrogen concentrating. This study has examined the relationship of estrogen receptive and LHRH-ir cells in the male and female guinea-pig brain with double label immunocytochemistry. Since estrogen receptor-ir, as demonstrated with antibody H222, is known to be confined predominantly to the cell nucleus, whereas LHRH-ir is localized mainly in the cytoplasm, single cells can be double-labeled. Diaminobenzidine tetrahydrochloride was used for localization of LHRH-ir while nickel-enhanced diaminobenzidine tetrahydrochloride was used for localization of estrogen receptor-ir. The results revealed that there were many brain nuclei that contained both LHRH and estrogen receptor-positive cells, including the preventricular periventricular nucleus, the anterior subcompact nucleus of the medial preoptic nucleus (MPNa), the remainder of the medial preoptic nucleus, the retrochiasmatic area, and the anterior, dorsomedial, ventrolateral and arcuate nuclei. However, of a total of 2,604 LHRH-ir cells that were examined, we observed only 5 double-labeled cells (<0.2%). The double-labeled cells were not restricted to a single nucleus; they were present in the MPNa, the retrochiasmatic area and the arcuate nucleus. Moreover, at the light microscopic level, LHRH cells quite frequently appeared to be apposed to estrogen receptor-positive cells (8.8% in the female), especially in the MPNa. These results lend further support to the notion that estrogen-mediated regulation of the LHRH system is achieved primarily through estrogen receptive interneurons. However, due to the existence of LHRH-LHRH synaptic interactions, the possibility also exists that a small population of estrogen-sensitive LHRH neurons could contribute to generalized activation of the LHRH system.

Novel hypothalamic and preoptic sites of prepro-melanin-concentrating hormone messenger ribonucleic Acid and Peptide expression in lactating rats.

Abstract: The deduced structure of the rat melanin-concentrating hormone (MCH) precursor predicted the existence of at least two peptides that may be processed from it, one similar to teleost MCH and a second novel neuropeptide, NEI. Cellular localization studies confirmed that prepro-MCH (ppMCH) mRNA and the MCH and NEI peptides are expressed predominantly in cells in the zona incerta and caudal lateral hypothalamic area with minor contingents seen in the olfactory tubercle and pons. A moderate MCH-and NEI-immunoreactive axonal projection to the median eminence and, particularly, to oxytocin-rich regions of the posterior pituitary suggested some anatomical heterogeneity of ppMCH-expressing neurons in the hypothalamus, and an involvement in neuroendocrine function. In the present study, immunohistochemical and hybridization histochemical methods were used to follow MCH gene and peptide expression as a function of reproductive status in female rats. Nursing dams sacrificed after 8 to 21 days of lactation consistently displayed ppMCH mRNA and MCH and NEI immunoreactivity in a discrete and contiguous band, encompassing the ventral aspect of the medial part of the medial preoptic nucleus, the periventricular preoptic nucleus, and the most rostral aspects of the paraventricular nucleus of the hypothalamus (PVH). Combined immunohistochemical (for oxytocin) and hybridization histochemical (for ppMCH mRNA) staining failed to reveal a significant degree of congruence in the two chemically-specified cell populations in the PVH of lactating dams. The apparent induction of ppMCH-derived peptides and mRNA in the preoptic area and PVH was not apparent in animals sacrificed 4 to 8 days after weaning, during late pregnancy, or at any point in the estrous cycle. Moreover, no frank alterations in ppMCH mRNA were evident in the principal sites at which ppMCH is expressed at constitutively high levels, i.e. in the lateral hypothalamic area and zona incerta, as a function of reproductive status. The loci and apparent state-dependency of the induction of ppMCH mRNA and peptide expression suggests a role for these gene products in the control of lactation.

Glucocorticoids are Required for Food Deprivation-Induced Increases in Hypothalamic Neuropeptide Y Expression

Abstract: Neuropeptide Y (NPY), a 36 amino-acid peptide found within the hypothalamus, is thought to be an important regulator of food intake. Hypothalamic NPY gene expression, synthesis and secretion are all known to be increased in models of increased metabolic demand in which serum glucocorticoids are also elevated. The present studies were designed to test the hypothesis that glucocorticoids are required for increased hypothalamic preproNPY mRNA levels induced by food deprivation (FD). First, animals underwent bilateral sham-adrenalectomy (sham) or not (control), and were subjected to 72 h FD, or not. Total RNA was isolated from hypothalamic tissue blocks and the content of preproNPY mRNA was measured by solution hybridization/RNase protection analysis. This study revealed that there was no significant difference in hypothalamic preproNPY mRNA content between shamfed and control-fed groups, or between sham-FD and control-FD groups. In the second experiment, animals underwent bilateral adrenalectomy (ADX), were allowed to feed ad libitum and were sacrificed 1 day, 4 days and 7 days after ADX. Nuclease protection analysis revealed no significant effect of ADX on hypothalamic preproNPY mRNA levels over this time-course. Finally, we examined the role of glucocorticoids in regulating NPY gene expression following FD. Animals underwent bilateral ADX, or not. At the time of surgery, ADX animals received placebo, or corticosterone (B) replacement in the form of constant release pellets, at one of two doses. Food was removed from half of the animals in each group 24 h after surgery; all animals were sacrificed 72 h thereafter. There was no difference in preproNPY mRNA content between the ADX-FD and ADX-fed groups, relative to the fed controls. Replacement with corticosterone [ADX(B)] did not alter preproNPY mRNA content in fed animals, however preproNPY mRNA content in FD animals was increased 2.5-fold. These studies demonstrate that glucocorticoids are necessary and serve a stimulatory role in the increase in hypothalamic preproNPY mRNA levels observed under conditions of FD, and suggest that hypothalamic NPY gene expression may be directly responsive to peripheral metabolic and hormonal signals.

cFos activity identifies recruitment of luteinizing hormone-releasing hormone neurons during the ascending phase of the proestrous luteinizing hormone surge

Abstract: The proto-oncogene product of the c-fos gene, cFos, is a useful marker for luteinizing hormone-releasing hormone (LHRH) neuronal activation. While recent data indicate that in the rat, an LHRH surge plays an active role in stimulating the proestrous luteinizing hormone (LH) surge, the mechanics of the LHRH surge remain unknown. The aim of this study was to determine whether LHRH neuronal activation occurs entirely at the beginning of the LH surge or whether the number of LHRH neurons activated increases during the ascending phase of the surge. To accomplish this aim, we determined the relationship between the number of LHRH neurons expressing cFos and LH concentrations during the ascending limb of the proestrous LH surge. During the estrous cycle in the rat, on the afternoon of proestrus, the number of LHRH neurons expressing cFos increased as plasma LH levels increased to reach peak concentrations. The regression line describing these two variables had a very highly significant correlation coefficient, indicating a linear relationship. Treatment with RU486 to block progesterone's action on the afternoon of proestrus significantly reduced both the number of LHRH neurons expressing cFos and the magnitude of LH secretion during the entire ascending phase of the LH surge. An analysis of covariance with comparison of regression lines from untreated and RU486-treated animals revealed that while both sets of data established significant linear relationships between the degree of activation of LHRH neurons and plasma LH values, the slopes of the two lines were different (P = 0.031) with no statistical difference in the two intercepts. These data, together with the demonstration of an overall reduction of cFos intensity following removal of progesterone's actions, suggest progesterone alters the dynamics of LHRH neuronal activation by significantly reducing the recruitment of LHRH neurons and suppressing the level of activation of individual LHRH neurons. The results of our study support the hypothesis that the ascending phase of the LH surge results from the gradual recruitment of LHRH neurons into the active state.

Cellular levels of messenger ribonucleic acids encoding vasoactive intestinal Peptide and gastrin-releasing Peptide in neurons of the suprachiasmatic nucleus exhibit distinct 24-hour rhythms.

Abstract: There is strong evidence supporting the view that the Suprachiasmatic nucleus (SCN) functions as a circadian clock; however, the neural and molecular events underlying SCN function remain unclear. A specific subpopulation of neurons within the ventrolateral aspect of the SCN that contains three peptides, vasoactive intestinal peptide (VIP), peptide histidine isoleucine (PHI) and gastrin-releasing peptide (GRP), play an important role in SCN function. VIP-containing neurons of the SCN receive synapses from photic projections, and co-injection of all three peptides mimics the phase-delaying effects of light on circadian activity rhythms. In principle, the signaling potential of a neuron containing several transmitters may be affected by the concentration ratio of co-released factors; hence, one mechanism by which VIP/PHI/GRP-containing neurons could influence SCN function is by changing the concentration ratio of these peptides throughout the light-dark cycle. The present study was performed to examine this possibility. Relative cellular levels of mRNA encoding both VIP/PHI and GRP were determined within the SCN every 4 h in rats housed in a 14 h light: 10 h dark cycle. Quantitative in situ hybridization revealed a statistically significant (P<0.005) 24-h profile of changes in VIP/PHI mRNA that peaked during the dark phase, and a significant (P<0.005) 24-h profile of changes in GRP mRNA that peaked during the light phase. These data support the interpretation that cellular levels of mRNAs encoding VIP/PHI and GRP within the SCN exhibit distinct profiles of changes throughout the light-dark cycle. Further, these findings are consistent with the hypothesis that the concentration ratio of VIP and PHI to GRP changes over the light-dark cycle, and that this may be an important mechanism by which circadian rhythms are generated or entrained.

Melatonin binding sites in the ovine brain and pituitary: characterization during the oestrous cycle.

Abstract: The distribution of putative melatonin receptors in the sheep has been investigated using in vitro autoradiography and the high affinity, high specific activity ligand 2-[(125) l]iodomelatonin. A wide distribution of specific labelling was found in both the ovine brain and pituitary gland as previously reported. Several novel areas of binding were also identified in the present study, including a fine layer of labelling at the medial edge of the diagonal band of Broca, the trigeminal nucleus, laminae II and III of the substantia gelatinosa, the molecular layer of the cerebellum as well as a scattered labelling in the pars distalis of the pituitary. There was no evidence of specific labelling in any of the peripheral tissues examined. Characterization studies performed on both neuronal and pituitary melatonin binding sites revealed that binding was time- and temperature-dependent and reversible on addition of 1 μM melatonin. The binding of 2-[(125) l]iodomelatonin was also competitively inhibited by increasing concentrations of 2-iodomelatonin and melatonin. The inhibition constants (K(i) ) estimated for each of these substances were similar for both neuronal and pituitary sites. Saturation studies also revealed similarities between neuronal and pituitary tissues with 2-[(125) l]iodomelatonin binding specifically to a single class of high affinity binding sites. Values for equilibrium constants (K(d) ) were within a range of 28 to 48 pM, and values were found to be not significantly different amongst the four regions of the brain investigated and the pars tuberalis of the pituitary. In contrast, the concentration of 2-[(125) l]iodomelatonin binding sites (B(max) ) ranged from 3 to 218fmol/mg protein and were maximal for the pars tuberalis. Saturation studies on brain and pituitary tissues taken from ewes killed either on the day of oestrus or during the luteal phase of the oestrous cycle, indicated that no differences exist in the affinity or concentration of 2-[(125) l]iodomelatonin binding in any region between the two times of the cycle investigated.

Multiplex solution hybridization-ribonuclease protection assay for quantitation of different ribonucleic Acid transcripts from snap-frozen neuroendocrine tissues of individual animals.

Abstract: We have compiled a protocol for simultaneous quantitation of a variety of gene transcripts in multiple individual brain and pituitary gland dissections for studying pretranslational regulation of neuroendocrine systems in vivo, using experimental designs compatible with meaningful statistical power. To facilitate collection of many samples at a time, the tissue was snap-frozen in chilled liquid Freon and stored at -80 °C until further processing. In this way, as many as five different brain and pituitary gland dissections per rat could be collected from eight rats in about an hour. The snap-frozen tissue was suitable for isolation of separate cytoplasmic and nuclear RNA fractions using homogenization in the presence of detergents. To facilitate homogenization of many samples at a time, we devised a method in which the tissue was repeatedly expelled through a 22 gauge hypodermic needle attached to a 1-ml plastic syringe used as a disposable, ready-to-use homogenizer. In order to promote dissolution of lipid membrane structures which are prevalent in the brain, the lysis buffer has been optimized to include the detergent sodium deoxycholate in addition to Nonidet P-40. Specific RNA transcripts were analyzed using a quantitative solution hybridization-ribonuclease protection assay coupled with polyacrylamide gel electrophoresis. The value of this highly sensitive assay has been expanded by including several molecular probes against a variety of neuroendocrine mRNA sequences simultaneously (e.g. progonadotropin-releasing hormone, proopiomelanocortin, tyrosine hydroxylase, dopamine D2 receptor, prolactin), thus increasing the amount of information obtained from each sample in one assay. Furthermore, each sample was routinely co-assayed for cyclophilin mRNA, an abundant, generally non-regulated mRNA whose levels reflected the individual variability in sample processing, thus serving as an internal reference. Once stored in hybridization solution, as many as 100 samples could be analyzed simultaneously for several different RNA transcripts in one assay. This protocol provides a powerful tool for studying regulation of neuroendocrine systems at the molecular level in vivo, using sample sizes suitable for applying statistical analysis of meaningful statistical power.

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

Abstract: 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.

Central Amygdaloid Involvement in Neuroendocrine Correlates of Conditioned Stress Responses

Abstract: The purpose of this study was to examine the effects of bilateral electrolytic lesions of the central nucleus of the amygdala (CEA) in comparison with sham lesions on neuroendocrine responses during conditioned emotional stress in male Wistar rats. Lesions in the CEA, made either before or after the single learning trial of inescapable footshock, failed to affect the conditioned response of plasma epinephrine levels. Plasma levels of norepinephrine showed neither a conditioned stress effect nor were influenced by lesioning. Pre-training CEA lesions, but not post-training intervention, abolished the conditioned elevations of circulating plasma corticosterone and prolactin. These results suggest that the CEA is involved in the conditioning rather than the retention of neuroendocrine stress responses. The effects of pre-training lesioning of the CEA can possibly be explained by a reduced feedback of all these neuroendocrine factors during or shortly after acquisition. In addition, there is a remarkable differentiation between various hormonal correlates of conditioned stress following CEA lesioning. Only corticosterone and prolactin, that appear to be correlates of a passive behavioural stress response, were abolished. The lesions failed to affect the sympatho-active stress parameters (epinephrine and norepinephrine). Relations between coping strategy-active and passive behaviour-and physiology in connection with CEA functioning are discussed.

Expression of the Oxytocin and Vasopressin Genes

Abstract: The nonapeptides oxytocin (OT) and vasopressin (VP) have held prominent roles in the fields of endocrinology and neuroscience. By the early 1900's, the effects of neurohypophysial substances on blood pressure, rapidity of labor, and milk secretory activity were described (1-4). The concept of neurosecretion arose from the studies of these peptides (5). The early determination of their amino-acid sequences (Fig. 1) (6) permitted more refined studies of their biological effects and distributions. VP and O T were both determined to exist in the magnocellular neurons of the paraventricular (PVN) and supraoptic (SON) nuclei of the hypothalamus (7, 8) and processed from their precursor forms along the axonal projection to the neural lobe (9). The orders of the nonapeptide, neurophysin, and glycopeptide (VP precursor only) components in the preprohormones were subsequently determined (10). The functions of the neurophysins and glycopeptide remain essentially unknown, although the neurophysins can bind their respective nonapeptides during axonal transport. Excellent discussions of the physiologic roles of VP and OT are found elsewhere (1 1-13) and a recent review discusses the physiology of magnocellular neurons of the PVN and SON (14).

Sexual behavior triggers the appearance of non-neuronal cells containing gonadotropin-releasing hormone-like immunoreactivity.

Abstract: Gonadotropin-releasing hormone (GnRH) regulates the secretion of pituitary gonadotropins and facilitates the display of sexual behavior. We report that in doves, following a brief period of courtship, non-neuronal cells containing GnRH-like immunoreactivity (ir) are seen in the habenula of both male and female doves. These cells appear to be translocated from either the cerebrospinal fluid or from capillaries in the pia or choroid plexus into the parenchyma of the brain. Immunoreactive cells are virtually absent in the habenula in control animals housed alone. The identity of the ir cells that enter the habenula is unknown but they would appear to be of either the macrophage or the mast cell lineage. Both of these blood-derived cells have heterochromatic nuclei and irregular cell surfaces with many filamentous processes, as do the GnRH-ir cells. The vacuolated granules of the ir cells could be indicative of mast cell degranulation or of endocytic vesicles of a phagocytosing cell. These data suggest that there is a population of cells within the habenula that are of similar size and morphology to the GnRH-ir ceils and are metachromatic when stained with toluidine blue. The latter is a property of heparin-containing mast cells. Lineage specific markers that permit a double-label study will be required to determine the exact nature of the GnRH-ir cells. Whatever their lineage, the translocation of non-neuronal cells into the undamaged adult central nervous system has not been described previously, and may provide a means of delivering biologically active substances into specific brain regions.

Immunocytochemical Localization of 11 Beta‐Hydroxysteroid Dehydrogenase in Hippocampus and Other Brain Regions of the Rat

Abstract: The dehydrogenase form of 11 β-hydroxysteroid dehydrogenase (11-DH) which catalyzes the oxidation of the biologically active steroid, corticosterone, to its inactive metabolite, 11-dehydrocorticosterone, is found in rat brain. The distribution and localization of 11-DH-like labeling in the rat brain was examined by immunocytochemistry. 11-DH-like immunostaining was found in all subfields of the hippocampus and in many other parts of the brain, including the preoptic area (POA), central nucleus of the amygdala, bed nucleus of the stria terminalis (NIST) and the cerebral cortex. Percentages of 11-DH-positive cells ranged from 10% in the POA and NIST to 50% to 60% in the hippocampus. When combined with neuronal or glial markers, 11-DH-like immunostaining was found to be predominantly localized within neurons, ranging from 10% or less glial labeling in hippocampus, amgydala and cortex to 22% glial labeling in the POA and NIST. Immunostaining was present in both the cytoplasmic and nuclear components of some cells in addition to their projections. In the kidney, 11-DH has been postulated to be a key component in a mechanism by which aldosterone gains access to renal Type I receptors despite the presence of much higher concentrations of glucocorticoids. The present data is consistent with a similar mechanism occurring in at least some parts of the brain, although the hippocampus appears to be an important exception because it does not appear to be differentially responsive to aldosterone in spite of its high 11-DH activity and immunoreactivity. However, the hippocampus is not implicated in neural control of salt appetite and fluid balance, whereas some of the other brain regions like the POA, NIST and amygdala are believed to be involved. Other aspects of 11-DH localization must therefore be examined in future studies, including the co-presence of mineraiocorticoid receptors and 11-DH in the same or adjacent cells and the possible significance of the relatively high glial localization of 11-DH immunoreactivity in the POA and NIST.

Effects of Cyclic AMP and Andre‐gens on in vitro Brain Aromatase Enzyme Activity During Prenatal Development in the Rat

Abstract: In the rat, there is a marked but transient increase in hypothalamic aromatase activity during the last week of fetal life. The present study was undertaken to gain insight into the regulation of this developmental pattern. Hypothalamic fragments comprising the medial basal hypothalamus and the suprachiasmatic region (henceforth referred to as preoptic area) were explanted and cultured in serum-free medium for 2 to 5 days. Aromatase activity was measured by the formation of (3) H(2) O, utilizing either [1s-(3) H]androstene-dione or [1s-(3) H]testosterone as substrate. Maximal rates of activity were obtained at a saturating concentration of 0.3 μM [1s-(3) H]testosterone. Confirmation of the identity of the [(3) H]estradiol formed was demonstrated by recrystallization of the derivatized estradiol to constant specific activity following incubation with [1,2,6,7-(3) H]testosterone. In agreement with previous reports, in vivo hypothalamic aromatase activity was negligible before gestational day (GD) 16, increased strikingly by GD19 (>5.0 pmol/h/mg protein) and decreased, thereafter, to low levels at GD22 (∼1.0 pmol/h/mg protein). Medial basal hypothalamus-preoptic area fragments explanted before GD17 failed to develop aromatase activity in vitro. If the tissue was explanted on GD17 or 18 (i.e. when the in vivo rate of activity was increasing), the enzyme activity did not continue to increase, but it was rather maintained for 2 days before decreasing in a manner that closely mimicked the decline observed in vivo. A similar, butimmediate decline was observed when the tissue was explanted on GD19 (i.e. at the time when theactivity peaks in vivo). Exposure of explants to either growth factors (insulin-like growth factor II, epidermal growth factor, and basic or acidic fibroblast growth factor), or steroids (estradiol-17s, progesterone, testosterone, dihydrotestosterone and corticosterone) failed to either increase aromatase activity before the peak at GD19 or ameliorate its perinatal decline. Increase of Ca(2+) fluxes with the ionophore A23187 or activation of the cyclic AMP, cyclic GMP, or protein kinase C pathways were similarly ineffective, as was angiotensin II, a recently proposed stimulator of neural aromatase. In contrast, aromatase activity was suppressed 2- to 4-fold by activation of the cyclic AMP pathway (with either forskolin or 8-bromo-cyclic AMP) or by the androgens, testosterone and dihydrotestosterone. These results suggest that: 1) the appearance of aromatase activity in the rat hypothalamus before GD17 requires the unfolding of extrahypothalamic events, 2) the increase in aromatase activity that occurs before GD19 also requires extrahypothalamic inputs and does not involve any of the known intracellular signal transduction pathways, and 3) the decline in activity observed after GD19 is regulated within the hypothalamus, and appears to be determined, at least in part, by the activation of cyclic AMP formation. A potential role for androgens is discussed.

Melatonin Regulates the Synthesis and Secretion of Several Proteins by Pars Tuberalis Cells of the Ovine Pituitary

Abstract: The pars tuberalis (PT) of the pituitary may be an important target for melatonin action, but the secretory output of the melatonin-responsive cells is unknown. Using [35S]methionine, protein synthesis and secretion have been studied in primary cultures of ovine PT cells, and analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Only 4% of the labelled proteins appeared in the medium with the majority retained in the cells. Stimulation of the cells with 10μM forskolin increased the accumulation of several labelled proteins in the medium without corresponding changes in the cell (72, 62, 44, 39, 29, 24, 23, 18 and 14 kd). Two-dimensional gel electrophoresis showed the proteins to have mildly acidic isoelectric points. Melatonin (1 μM) counteracted the stimulatory effect of forskolin on all but one (23 kd) of these secreted proteins. Immunoprecipitation showed this to be prolactin. Furthermore, melatonin alone appeared to have an inhibitory effect on the synthesis and release of proteins into the medium. The synthesis and secretion of the melatonin-responsive proteins was not inhibited by actinomycin D (1 μg/ml), indicating control at the translational level. This contrasts with the regulation of prolactin which is actinomycin D-sensitive. Pulse-chase experiments demonstrated that it requires 30 min for the secretory proteins to appear in the medium, consistent with intracellular processing and packaging prior to secretion. The secretory proteins labelled in the ovine PT, and responsive to melatonin, did not appear to be specific to the PT, as a similar profile of labelled secretory proteins was produced in primary cultures of pars distalis cells. However, melatonin had no effect on the synthesis and secretion of proteins by the pars distalis. These results demonstrate that in the ovine PT melatonin regulates the synthesis and export of several secretory proteins. These are possibly packaging proteins of secretory granules, similar to the granin family of proteins. Thus, the results confirm that melatonin-responsive cells are secretory cells and further imply that the PT-specific product is not a protein.

Steroid regulation and sex differences in [(3) h]muscimol binding in hippocampus, hypothalamus and midbrain in rats.

Abstract: The gonadal steroids estradiol and progesterone have previously been shown to modulate the specific binding of the GABA(A) agonist, [(3) H]muscimol, in the CA1 region of the hippocampus, the ventromedial nucleus of the hypothalamus and the midbrain central gray of ovariectomized female rats. In this report we show a sex difference in the level of binding in the very caudal ventromedial nucleus of the hypothalamus. In contrast to females, there is no steroid modulation of [(3) H]muscimol binding in the ventromedial nucleus of the hypothalamus and midbrain central gray of males. These effects may be functionally related to GABAergic control of female sexual behavior. In contrast, steroid modulation of [(3) H]muscimol binding in the CA1 region of the hippocampus occurred to the same degree in males and females, and there was no difference in the level of binding in any region of the hippocampus between gonadectomized males and females. Incubation of brain slices with progesterone or its metabolite 5α-3α-pregnanolone dissolved in ethanol, produced a significant increase in [(3) H]muscimol binding in most brain regions as compared to control brain slices treated with ethanol alone. Moreover, there was also a marked increase in [(3) H]muscimol binding in all brain areas in the control condition which contained 100 mM ethanol, as compared to brain slices not preincubated with ethanol. The increase in binding after in vitro treatment with either progesterone or 5α-3α-pregnanolone is notably different from that seen after progesterone given in vivo 4 h prior to assay in that it is not site-specific, does not depend on prior treatment with estradiol and shows no sex difference. These results suggest different mechanisms for progesterone effects on the GABA(A) receptor when administered in vivo as compared to in vitro.

Inhibition of luteinizing hormone secretion in the ewe by progesterone: associated changes in the release of gamma-aminobutyric Acid and noradrenaline in the preoptic area as measured by intracranial microdialysis.

Abstract: Progesterone inhibits the pulsatile release of luteinizing hormone (LH) in sheep by an action in the brain to suppress the release of LH-releasing hormone (LHRH). In addition, progesterone blocks the preovulatory surge of LH in this species. The neural basis of this inhibitory action is unknown, but as LHRH cells do not appear to contain progestin receptors other neural systems must mediate the action of this ovarian steroid on LH release. This study focuses on a possible role for the inhibitory amino-acid GABA and the monoamines (noradrenaline, adrenaline, dopamine and serotonin). The technique of microdialysis was used to monitor changes in these substances in the vicinity of the LHRH cell bodies (in the preoptic area) both before and following the administration of progesterone. Levels of this steroid, similar to those measured during the mid-luteal phase of the oestrous cycle, inhibited LH release and this was associated with significant alterations in the release of GABA and noradrenaline (but not adrenaline, dopamine or serotonin). Specifically, progesterone augmented GABA while noradrenaline release was depressed. Whether steroid actions on these neurotransmitters were mediated by opioids was also investigated. This possibility arises because of the reported involvement of opioids in progesterone negative feedback in the ewe. The long-acting opioid antagonist, naltrexone, was administered and GABA and noradrenaline release monitored for a further period both in the presence and absence of progesterone. Naltrexone significantly depressed GABA release in steroid-treated (but not untreated) ewes suggesting that the actions of progesterone on GABA are mediated by the endogenous opioid peptides. However, noradrenaline release was unaltered. In an earlier study we demonstrated that GABA release fell prior to the LH surge while noradrenaline release increased. These data, in conjunction with those from the present study, suggest that the mechanism by which progesterone is able to inhibit the preovulatory surge of LH in the ewe is by enhancing GABA and depressing noradrenaline release in the vicinity of the LHRH cell bodies. As opioid tone is also reported to fall prior to the surge, the interaction between opiate and GABAergic systems in the regulation of gonadotrophin secretion warrants further investigation.

Corticosterone Reduces Synaptic Inhibition in Rat Hippocampal and Neocortical Neurons in vitro

Abstract: We examined the effect of corticosterone (10-7 to 10-5 M) on membrane properties and postsynaptic potentials, by means of intracellular recordings from neocortical and hippocampal CA1 pyramidal neurons of the intact adult rat in vitro. Corticosterone reduced both the early and the late components of the orthodromically-evoked inhibitory postsynaptic potential in both structures. The glucocorticoid receptor antagonist RU 38486 (10-6 M) prevented this effect in the hippocampus. In hippocampal, but not in neocortical pyramidal neurons, corticosterone reduced a depolarizing membrane transient evoked by a depolarizing current step and increased the threshold for eliciting action potentials evoked by depolarizing current pulses. Corticosterone did not detectably alter the afterhyperpolarization following repetitive neuronal discharges evoked by current injection, in either the neocortex or in the hippocampus. Excitatory postsynaptic potentials, action potentials, membrane potential and membrane input resistance were also unchanged. The decrease in synaptic inhibition together with the reduction of electrical excitability in the hippocampus, would imply a modulation of response characteristics in pyramidal neurons such that repeated synaptic inputs become more efficient and low frequency input is blunted.

The chicken vasotocin gene.

Abstract: cDNA clones corresponding to the vasotocin precursor polypeptide were isolated from a chicken hypothalamic library and sequenced. The derived amino-acid sequence indicates a precursor of comparable structural organization to that described for members of the vasotocin/vasopressin gene family from other species. Unlike in mammals the C-terminal glycopeptide moiety appears not be cleaved off from the neurophysin. Subsequent screening of a chicken genomic library permitted an analysis also of the vasotocin gene structure and exonic composition. The 5'region upstream of the first exon was sequenced and revealed an unusual pattern of 49 repetitive -YYCYCYAAAYY- motifs, together with a polyadenyl region supporting a bend in the DNA, and a long pyrimidine-rich sequence. Three AP2-like elements, identified in the mammalian vasopressin gene, were also observed in the immediate upstream region. There was no obvious homology to the promoter regions of the known oxytocin genes, nor to any other sequence deposited in available databases, nor to other known cis-elements.

Pineal photoreceptor cells in culture: fine structure, and light/ dark control of cyclic nucleotide levels and melatonin secretion.

Abstract: Trout pineal photoreceptor cells were dissociated by trypsin-DNase digestion and further purified by a Percoll gradient centrifugation Total cells or purified photoreceptor cells were then embedded in a collagen gel, or layered on culture-treated polycarbonate membranes, or maintained in suspension, with RPMI 1640 medium or BGjb medium It has been shown that cells maintain a rhythmic production of melatonin for at least seven 24 h light/dark cycles under these conditions In this complementary study, the morphofunctional state of the photoreceptor cells was examined 1) by electron (transmission, scanning) microscopy, and 2) by pharmacological tests under different lighting conditions Using polycarbonate membranes together with RPMI 1640 medium appeared the most suitable The segmented organization of photoreceptor cells was well preserved when using the culture-treated membranes It tended to disappear in cells embedded in the collagen gel and was lost after passage through the Percoll gradient However, this one allowed obtention of an homogeneous population of photoreceptors, as recognized by their intracellular components Intracellular organelles were rather well preserved in the embedded photoreceptors The study also provides novel information on the nature of second messengers involved in the photoperiodic control of melatonin production in photoreceptor cells From the effects of an adenylyl cyclase activator and a phosphodiesterase inhibitor it appeared that 1) total cells and Percoll-selected cells behaved similarly, 2) the nocturnal rise in melatonin secretion was associated with an increase in cAMP content, and 3) a fall in cAMP may be a mechanism through which light reduces melatonin secretion by photoreceptor cells Cyclic GMP, the metabolism of which also appeared to be controlled by light, did not seem involved in the photoperiodic control of melatonin production The method proposed herein offers interesting perspectives for the study of the photoneuroendocrine properties of isolated photoreceptor cells

Functionally and anatomically distinct populations of vasopressinergic magnocellular neurons in the female golden hamster.

Abstract: The present study was done to determine whether the vasopressinergic neurons in the hypothalamus controlling flank marking behavior are distinct from the magnocellular neurons comprising the hypothalamo-neurohypophysial system. Animals were either hypophysectomized or injected with a suicide transport lectin, volkensin, into the neurohypophysis. Both procedures resulted in a pronounced loss of vasopressin-immunoreactive perikarya throughout the hypothalamus concomitant with increases in water intake and urine output and decreases in circulating levels of vasopressin. The loss of the hypothalamo-neurohypophysial system was most pronounced in volkensin-treated animals that presented with frank diabetes insipidus and exceedingly low levels of plasma vasopressin. However, the vasopressinergic fibers and magnocellular neurons in and around the anterior hypothalamus implicated in the control of flank marking survived the volkensin treatment. Volkensin-treated animals exhibited levels of flank marking typical of untreated animals. These data suggest the presence of anatomically and functionally distinct populations of vasopressinergic magnocellular neurons in the hypothalamus of the golden hamster.

Effects on plasma luteinizing hormone levels of microinjection of noradrenaline and adrenaline into the septo-preoptic area of the brain of the ovariectomized ewe: changes with season and chronic oestrogen treatment.

Abstract: We have investigated the effects on luteinizing hormone (LH) secretion of noradrenaline (NA) and adrenaline (A) microinjected (1 μl) into the septo-preoptic area of ovariectomized (OVX) ewes with or without oestrogen (E) treatment and across the breeding and non-breeding seasons. Guide tubes (19 gauge) were placed into the septo-preoptic area of OVX ewes using lateral ventriculograms for localization of the target area. The sheep were tamed so that injections could be made into conscious animals during blood sampling procedures. Jugular venous blood was collected at 10-min intervals for 3 h, an injection of NA or A (10 μg) or saline was given and samples collected for a further 3 h. The plasma samples were assayed for LH. On completion of the experiments the brains were sectioned to locate the site of injection. In the non-breeding season of the first year, 9 ewes were used of which 3 had correct guide tube placement; in these 3 ewes NA and A had no effect in OVX ewes. In OVX ewes treated with 0.5 cm Silastic ®implants of E for 1 week, plasma LH levels were reduced from 9.1 ± 1.96 nglrnl before E treatment to 2.8±0.95 ng/ml after E treatment. In these E-treated ewes NA and A caused a robust increase in plasma LH levels. In the breeding season, 9 ewes were used of which 7 had correct guide tube placement; in these 7 ewes NA and A had no effect in OVX ewes. When OVX ewes were treated with 0.5cm E implants, NA or A injection decreased LH interpulse interval. In OVX ewes which received 1.0 cm E, NA caused a pronounced but transient suppression of plasma LH secretion due to an increase in interpulse interval. When 3.0 cm E implants were given to OVX ewes there was a strong suppression of plasma LH secretion with pulsatility abolished; NA injection had no effect in these sheep. In the second year, in the non breeding-season, 17 ewes were used of which 14 had correct guide tube placement although a number of injections were above the target region. NA injection had no effect on plasma LH levels in OVX ewes but had a variable effect on OVX ewes treated with 0.5 cm E implants, depending upon the degree of suppression of plasma LH secretion by E. When plasma LH was fully suppressed by E, injection of 1Opg NA provoked a profound and sustained increase in plasma LH levels. When plasma LH secretion was pulsatile after E treatment, NA injection decreased LH interpulse interval. Similar responses were obtained with 0.5cm E-treated sheep when injected with 1.Opg NA. When OVX ewes were given 3.0cm E implants a small and non-significant (P = 0.09) rise in plasma LH levels occurred, following 10 pg NA injection. These results provide further evidence of involvement of NA/A systems in the regulation of gonadotrophin-releasing hormone (GnRH) secretion at the level of the GnRH cell bodies in the septo-preoptic area, with clear influences of season and E status on this regulation. In the OVX sheep the GnRH pulse generation system is probably subserved by endogenous permissive NA/A input rendering exogenous input ineffective. In the breeding season NAlA can inhibit GnRH/LH secretion in the presence of physiological doses of E whereas in the non-breeding season, E profoundly suppresses GnRH/LH secretion, possibly by the removal of permissive NA inputs, which can be overcome by the injection of NAlA into the septo-preoptic area.

Simultaneous Display of Sexual and Ingestive Behaviour by Rats

Abstract: Intraoral infusion of sucrose activates consummatory ingestive behaviour in rats selectively, i.e. the rat only emits the responses used to ingest food. Activation of consummatory ingestive behaviour in this way had no effect on the subsequent display of sexual behaviour by male or female rats and vice versa. Rats infused intraorally with sucrose and presented with a sexual partner showed ingestive and sexual behaviour simultaneously. Pretreatment with cholecystokinin octapeptide inhibited the ingestion of sucrose in both males and females but had no effect on the simultaneous display of sexual behaviour. Ingestion of sucrose from a drinking spout, a test in which the rat has to emit responses to obtain food, i.e. show appetitive ingestive behaviour, was inhibited by the presentation of a sexual partner in rats of both sexes. These results show that the mechanisms controlling consummatory sexual and ingestive behaviour operate independently and that the presentation of a sexual partner inhibits appetitive ingestive behaviour. Daily intraoral infusion of sucrose reduced pellet intake in ovariectomized rats while the rats maintained their body weight. Implantation of an oestradiol-filled implant reduced body weight and inhibited daily intake of pellets but had no effect on the intake of intraorally administered sucrose. Subsequent removal of the oestradiol implant increased sucrose intake and body weight but did not have a marked effect on pellet intake. Thus, rats respond to a lowering of the set point for body weight by decreasing their intake of the least preferable kind of food and increase their intake of the most preferable kind of food in response to an elevation of the set point for body weight. Ovariectomized rats infused intraorally once daily with a highly nutritive milk diet in the absence of food pellets ingested very large amounts and reduced their intake in response to oestradiol implantation. Thus, although oestradiol can inhibit consummatory ingestive behaviour, its suppressive effect on ingestion cannot be described in terms of selective effects on appetitive and/or consummatory aspects of the behaviour nor in terms of an alteration in the preference for a sweet solution. Inhibition of ingestive behaviour occurred within 24 h after oestrogen treatment as opposed to stimulation of sexual behaviour which had a longer latency, suggesting that oestradiol affects ingestive and sexual behaviour via different mechanisms. While the mechanisms controlling consummatory ingestive and sexual behaviour must be different, there is evidence for a common mechanism mediating the incentive motivation and reward aspects of these behaviours. The mechanisms which enable rats to select between two, possibly equally rewarding courses of action, i.e. display of sexual or ingestive responses, however, are unknown.

Ibotenate lesions of the diagonal band of broca attenuate baroreceptor sensitivity of rat supraoptic vasopressin neurons.

Abstract: Previous electrophysiological studies in the rat suggest that neurons in the diagonal band of Broca participate in baroreceptor-induced suppression of the spontaneous activity of vasopressin-secreting neurons in the hypothalamic supraoptic nucleus. In order to test this hypothesis, extracellular recordings were obtained from phasically-active vasopressin neurons in the supraoptic nucleus of anesthetized rats injected at least 3 days previously with ibotenic acid (1.25 μg/250 nl) in the diagonal band of Broca, the medial and lateral septum, or the median preoptic nucleus. In normal rats, brief increases in blood pressure produced by injections of metaraminol (10 μg/10 μl iv) that were sufficient to activate peripheral baroreceptors, suppressed the activity of a majority (21 tested, 19 suppressed) of phasically-active vasopressin-secreting neurons. In rats with ibotenic acid lesions of the diagonal band of Broca, the number of phasically-active neurons that were baroreceptor-sensitive was significantly reduced (21 tested, 8 suppressed) while lesions of the medial and lateral septum (17 tested, 16 suppressed) or the median preoptic nucleus (21 tested, 20 suppressed) had no effect. The results support the hypothesis that diagonal band of Broca neurons participate in a central pathway mediating the inhibitory effects of peripheral baroreceptor stimulation on the activity of vasopressin-secreting neurons in the rat supraoptic nucleus.

Arginine vasotocin gene expression during osmotic challenge in the chicken.

Abstract: The avian hypothalamic nonapeptide arginine vasotocin (AVT) is released from axon terminals in the neural lobe upon the application of osmotic stimuli We have investigated whether, and to what extent, hormone secretion from the neurohypophysis is related to gene expression in the hypothalamus Results from hybridization experiments with an AVT-specific cDNA probe indicate that in adult chickens stimulated by water deprivation or by hypertonic saline (2% w/v) drinking water, an upregulation of the AVT mRNA pool takes place, since consistently higher AVT mRNA levels compared to controls were monitored in osmotically challenged birds This stimulatory effect was even visible at the transcriptional level after 19 h of water deprivation when osmolality was still near the basal value In hens osmotically challenged by hypertonic saline drinking water for 5 days, a dissociation between osmolality and AVT plasma concentration was visible: extremely high plasma osmolality was accompanied by only moderately increased plasma AVT concentration This might be caused either by exhaustion of stored hormone, or by downregulation of the system after chronic challenge The latter suggestion is supported by the fact that the AVT mRNA concentration after 5 days of hypertonic saline challenge was well below the AVT mRNA levels of the groups with the more short-term stimuli of water deprivation for 19 or 48 h In 30-day-old chicks the hypothalamic AVT mRNA concentration hardly reached 70% of the adult value, although AVT plasma concentrations were similar to those in the mature bird We conclude that osmotic challenge of the hypothalamo-neurohypophysial system not only causes secretion of AVT from stores in the neural lobe but is accompanied by upregulation of AVT gene expression Upregulation already occurs after marginal increase in plasma osmolality, as seen after 19 h of water deprivation in hens In 30-day-old chicks gene expression is only slightly upregulated after short-term water deprivation while increase in plasma AVT is even greater compared to hens

Interactions between the hypothalamo-pituitary adrenal axis and the thymus in the rat: a role for corticotrophin in the control of thymulin release.

Abstract: Our recent observations in man suggested that the secretion of the thymic peptide, thymulin, is influenced by hormones of the pituitary-adrenal axis. In the present study, we have used the rat as a model in order to examine 1) the effects of corticotrophin (ACTH) and glucocorticoids on the release of thymulin in vivo and in vitro, and 2) the influence of an acute rise in plasma thymulin on the secretion of corticosterone and luteinizing hormone. Immunoreactive thymulin was readily detectable in plasma from male Sprague-Dawley rats(≃200 g). Chronic bilateral adrenalec-tomy, which effectively removed endogenous corticosterone, produced highly significant (P 0.2) from that in sham-operated controls. In vitro, two non-specific depolarizing agents, K(+) (56 mM) and veratridine (10 ≃M), caused significant (P<0.01) Ca(2+) -dependent increases in thymulin release from segments of rat thymic tissue. Their effects were mimicked by ACTH(1-39) . The secretory responses to ACTH (0.025 to 1 ng/ml) were concentration-dependent but a very high concentration (2 ng/ml) of the peptide was without effect. Dexamethasone (0.1 μM) reduced (P<0.05) the spontaneous release of thymulin in vitro but potentiated markedly (P<0.01) the secretory responses to ACTH (0.5 to 1.0 ng/ml). Administration of thymulin (0.1 and 10 μg/kg ip) produced, within 10 min, striking increases in the plasma thymulin concentration which were still evident at 30 min. The peptide concentration then declined rapidly and, within 24 h, was lower than that in the corresponding vehicle-treated controls. The serum concentrations of corticosterone and luteinizing hormone were unaffected by the thymulin treatment. The saline vehicle (2.0 ml/kg ip) also produced a small increase in plasma thymulin concentration which was maximal at 10 min; a further small rise was evident 6 h after the injection but thereafter the thymulin values were indistinguishable from those in uninjected controls. A similar biphasic profile of serum corticosterone was apparent after the saline injection but the serum luteinizing hormone was unaffected. The results suggest that ACTH is a physiological enhancer of thymulin release and that, in certain circumstances, its effects may be potentiated by glucocorticoids.