Inhibition of release of neurohypophysial hormones by endogenous opioid peptides in pregnant and parturient rats

Naloxone, an opiate receptor antagonist, was used to determine whether opioid peptides modulate release of oxytocin (OT) or vasopressin (AVP) in the rat after expulsion of the fetus, i.e. parturition. We measured the concentrations of AVP and OT in plasma and in the neurointermediate lobe of the pituitary of pregnant rats given naloxone (5 mg/kg, s.c.) or saline on day 20 of gestation, and on day 21 either before or during the expulsive stage of labor. Non-pregnant rats in diestrus were given naloxone for comparison. On days 20 and 21 of gestation, before the onset of parturition, plasma [AVP] but not [OT] was elevated, compared to the non-pregnant controls. After delivery of the first two pups, plasma [OT] approximately doubled, whereas plasma [AVP] remained unchanged. Blocking the action of endogenous opioid peptides with naloxone caused an elevation of plasma [OT] in pregnant animals on days 20 and 21 of gestation and during parturition. Naloxone, however, did not alter plasma [AVP] in either parturient or preparturient animals. In contrast, [AVP], but not [OT], was increased in plasma of non-pregnant rats given naloxone. The content of OT in the neuro-intermediate lobe was similar in pregnant and non-pregnant rats and was unaffected by delivery of the first two pups. However, AVP content and the ratio of AVP/OT in the pituitary were lower in pregnant animals before and during delivery than in the non-pregnant controls. The content of neither hormone was altered by naloxone. Thus, AVP release apparently increases and pituitary stores of this peptide are decreased by day 20 of gestation, when labor has not yet begun.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential effects on naloxone on the release of neurohypophysial hormones in normotensive and spontaneously hypertensive rats

The hypothalamo-neurohypophysial system is altered in the spontaneously hypertensive rat (SHR). We hypothesized that an aberrant regulation of vasopressin (VP) and oxytocin (OT) release by endogenous opiod peptides alters this neuroendocrine system in the SHR. Concentrations of the neurohypophysial hormones in plasma and the pituitary were measured in 17-week-old SHRs and two strains of normotensive controls, Wistar Kyoto (WKY) and Sprague-Dawley rats. Animals were decapitated 20 min after s.c. injection of saline (1 ml/kg) or naloxone hydrochloride (1 or 10 mg/kg). In addition, neurohypophysial hormones excreted during the day (08.00–17.30 h) and night (17.30–08.00 h) were determined in urine from 16-week-old animals kept in metabolic cages for 5 days. VP at extrahypothalamic sites was also measured as [VP] in acid extracts of the subfornical organ area, hippocampal commissure-fornix and choriod plexus. Hormones were quantified by radioimmunoassay. The pituitary content, plasma concentration, and urinary excretion of OT were reduced (P < 0.05) in SHRs, whereas VP content was increased (P < 0.05) in the pituitary and plasma, but unchanged in urine, of hypertensive animals. In extrahypothalamic tissues, [VP] in the hippocampal commissure-fornix was increased in the SHR. Naloxone elevated (P < 0.05) the plasma concentration of OT in WKY animals and VP in SHRs. Neither [VP] nor [OT] in plasma was changed by naloxone in Sprague-Dawley rats. Pituitary stores of the neurohypophysial hormones were not altered by naloxone in either hypertensive or normotensive rats. In conclusion, endogenous opioid peptides tonically inhibit OT release in WKY rats, whereas VP releas is decreased by opioid peptides in SHRs, 16–17 weeks of age. The neuromodulatory role of opioid peptides in the release of neurohypophysial hormones appears to be altered in the SHR such that VP release is suppressed and OT release is augmented.     

A functional role for opioid peptides in the differential secretion of vasopressin and oxytocin

The presence of opioid peptides and opiate receptors in the hypothalamo-neurohypophysial system, as well as the inhibitory effects of enkephalins and beta-endorphin on release of oxytocin and vasopressin have been well documented. The physiological importance of opioid peptides in this classical neurosecretory system, however, has remained illusive. In the present study we tested the effects of naltrexone on the plasma concentrations of oxytocin and vasopressin during dehydration, hemorrhage and suckling in the conscious rat. We obtained evidence supporting the hypothesis that opioid peptides inhibit oxytocin release and thereby promote the preferential secretion of vasopressin when it is of functional importance to maintain homeostasis during dehydration and hemorrhage. Our data support the concept that the coexistence of a neuromodulator and a neurohormone in the same neuron, as demonstrated for vasopressin with dynorphin or leucine-enkephalin, serves to regulate the differential release of two biologically different, yet evolutionarily-related, neurohormones, e.g. oxytocin and vasopressin, from the same neuroendocrine system.     

Serotonin stimulates hypothalamic mRNA expression and local release of neurohypophysial peptides

The neurotransmitter serotonin (5-HT) stimulates the secretion of vasopressin and oxytocin, and 5-HT is involved in the mediation of the vasopressin and oxytocin response to stress. In male Wistar rats, we investigated the 5-HT receptors involved in the 5-HT-induced increase of mRNA expression of vasopressin and oxytocin in the hypothalamic paraventricular nucleus (PVN) and supraoptic nucleus (SON). The 5-HT precursor, 5-hydroxytryptophan, injected in combination with the 5-HT reuptake inhibitor, fluoxetine, increased oxytocin mRNA expression in the PVN, and the concentration of vasopressin and oxytocin in plasma, whereas mRNA in the SON was not affected. Intracerebroventricular infusion of 5-HT agonists selective for the 5-HT1A, 5-HT1B, 5-HT2A and 5-HT2C receptor increased oxytocin mRNA in the SON and PVN. Infusion of agonists selective for the 5-HT2A + 2C receptor increased vasopressin mRNA in the PVN, whereas none of the 5-HT agonists affected vasopressin mRNA in the SON. All the 5-HT agonists infused increased peripheral oxytocin concentration and vasopressin was increased by stimulation of the 5-HT2A, 5-HT2C and 5-HT3 receptor. Intracerebroventricular infusion of 100 nmol 5-HT increased the extracellular hypothalamic concentration of vasopressin as measured by microdialysis in the PVN. To evaluate the involvement of hypothalamic-pituitary system in the 5-hydroxytryptophan and fluoxetine-induced vasopressin secretion, rats were immunoneutralized with a specific anti-corticotropin-releasing hormone antiserum. This treatment reduced plasma vasopressin and oxytocin responses. We conclude that stimulation with 5-hydroxytryptophan or 5-HT agonists increases mRNA expression of oxytocin in the PVN and the SON via stimulation of at least 5-HT1A, 5-HT1B, 5-HT2A and 5-HT2C receptors. Vasopressin mRNA in the PVN was increased only via the 5-HT2 receptor, whereas vasopressin mRNA in the SON does not seem to be affected by 5-HT stimulation. Corticotropin-releasing hormone appears to be partly involved in the mediation of 5-HT induced vasopressin and oxytocin secretion.     

Enkephalin inhibition of angiotensin-stimulated release of oxytocin and vasopressin

The effect of leucine5 -enkephalin on angiotensin II (AII)-stimulated release of oxytocin and vasopressin (VP) was investigated in the conscious male rat. Changes in the plasma concentration ([]) of both oxytocin and VP were measured in animals: (1) 60 s after intracerebroventricular (i.v.t.) administration of either artificial cerebrospinal fluid (CSF) or CSF with AII (10, 50, 100 ng/5 microliter); (2) 30, 60, 90 and 300 s after single injection of AII (50 ng/5 microliter; i.v.t.) or CSF and (3) 60 s after AII (50 ng/5 microliter) or CSF in animals pretreated with leucine5 -enkephalin (100 ng/5 microliter; i.v.t.) or CSF (5 microliter). Oxytocin and VP were quantified by radioimmunoassay and values corrected for 100% recovery. The development of a sensitive radioimmunoassay for oxytocin is described. The antiserum for oxytocin enabled detection of greater than or equal to 0.8 pg/ml oxytocin with cross-reactivity of 0.01% with arginine vasopressin and 0.10% with arginine vasotocin. The inter- and intra-assay coefficients of variation were 3-9% and 3.2%, respectively. The hypotheses being tested were that i.v.t. injection of AII stimulates release of both neurohypophysial hormones non-selectively and that leucine5 -enkephalin inhibits both AII-stimulated oxytocin and VP release. Angiotensin II at doses ranging from 10 to 100 ng/5 microliter, i.v.t. increased the plasma concentration of both oxytocin and VP. Plasma levels of both neurohypophysial hormones were elevated 30 s after AII administration i.v.t. and remained elevated 300 sec later.(ABSTRACT TRUNCATED AT 250 WORDS)     

Contrasting effects of neurohypophysial peptides on pyramidal and non-pyramidal neurones in the rat hippocampus

Oxytocin and vasopressin increased the rate of firing of a class of presumed non-pyramidal neurones located in the CA1 area of rat hippocampal slices. This excitatory effect persisted in conditions of synaptic uncoupling. In contrast, pyramidal neurones were either unaffected by neurohypophysial peptides or showed one or several of the following effects: (a) a decrease in firing rate in cells which were spontaneously active; (b) a slight membrane hyperpolarization; and (c) an increase in the rate of occurrence os spontaneous inhibitory postsynaptic potentials. We therefore propose that oxytocin and vasopressin excite directly a class of non-pyramidal inhibitory interneurones, whereas their observed effect on pyramidal neurones is indirect and inhibitory.     

Immunocytochemical localization of neuropeptide FF (FMRF amide-like peptide) in the hypothalamo-neurohypophyseal system of Wistar and Brattleboro rats by light and electron microscopy

Neuropeptide FF (F8Famide, FMRFamide-like, or morphine modulating peptide) immunoreactivity was localized by light and electron microscopy in the hypothalamo-neurohypophyseal system of Wistar and Brattleboro rats. In Wistar rats neuropeptide FF was present in part of the magnocellular neurones of the paraventricular and supraoptic nuclei in which it was coexpressed with vasopressin. Neuropeptide FF containing fibers were present in the paraventricular and the supraoptic nuclei, and in the central part of the neural lobe. At the electron microscopic level, neuropeptide FF containing nerve terminals in the neural lobe formed synaptoid contacts exclusively with pituicytes. No neuropeptide FF containing neurovascular contacts or contacts with other neuronal structures were observed. In contrast with Wistar rats, neuropeptide FF was almost completely absent in cell bodies of the paraventricular and supraoptic nuclei, and in fibres of the neural lobe in Brattleboro rats. Only a few solitary cells could be observed in these structures. The present results demonstrate that neuropeptide FF coexists with vasopressin within the hypothalamo-neurohypophyseal system. As we did not observe neuropeptide FF containing neurovascular contacts, neuropeptide FF containing nerve terminals probably have a local function within the neural lobe. Neuropeptide FF may be involved in the modulation ofoxytocin and vasopressin release, with the pituicyte as an intermediate cell. © 1993 Wiley-Liss, Inc.     

Enhanced Neurohypophyseal Vasopressin Release is Associated with Increased Opioid Inhibition of Oxytocin Release

We tested the hypothesis of a cross-inhibition of oxytocin (OT) release by endogenous opioid peptides co-released with vasopressin (VP). This opioid cross-inhibition resulted in a selective block of OT release and hence in preferential release of VP. The effects of the opiate receptor antagonist naloxone were tested on neurohypophyseal VP release during dehydration, ethanol administration and sulphated cholecystokinin octapeptide (CCK-8S) application, assuming that the inhibition of pituitary OT release by endogenous opioids increases as neurohypophyseal VP output increases. A high VP output was found to coincide with increased inhibition of OT release: Subcutaneous injection of graded doses of naloxone (30 min prior to decapitation), augmented OT plasma levels significantly more in 24 h water-deprived male rats than in normally hydrated rats. Naloxone had no effect on VP release. Ethanol (10% in saline) administered intragastrically 50 min prior to decapitation and 20 min before subcutaneous naloxone (5 mg/kg) resulted in the inhibition of VP output. The ethanol treatment resulted in a rise in plasma OT levels that was additional to the effect of naloxone. These features were present in normally hydrated as well as in 24 h water-deprived animals, but were more pronounced in the latter group. Peripheral CCK-8S administration induces an abrupt and selective secretion of OT. Blocking the opioid inhibition of OT release with naloxone resulted in a significant rise of OT compared to that with CCK-8S alone. The magnitude of the opioid inhibition coincided with the activity of the VP system, and a higher dose of naloxone was needed to potentiate the CCK-8S effect on OT release in the water-deprived group than in euhydrated rats. No effect of CCK-8S and/or naloxone was found on VP plasma levels. The data indicate that opioid peptides co-released with VP (like dynorphin) may be responsible for cross-inhibition of OT release during dehydration. This suggests that dynorphin acts in a paracrine way, making it a strong candidate for this role.     

Cellular interactions between axon terminals containing endogenous opioid peptides or corticotropin-releasing factor in the rat locus coeruleus and surrounding dorsal pontine tegmentum

Recent evidence suggests that certain stressors release both endogenous opioids and corticotropin-releasing factor (CRF) to modulate activity of the locus coeruleus (LC)-norepinephrine (NE) system. In ultrastructural studies, axon terminals containing methionine(5)-enkephalin (ENK) or CRF have been shown to target LC dendrites. These findings suggested the hypothesis that both neuropeptides may coexist in common axon terminals that are positioned to have an impact on the LC. This possibility was examined by using immunofluorescence and immunoelectron microscopic analysis of the rat LC and neighboring dorsal pontine tegmentum. Ultrastructural analysis indicated that CRF- and ENK-containing axon terminals were abundant in similar portions of the neuropil and that approximately 16% of the axon terminals containing ENK were also immunoreactive for CRF. Dually labeled terminals were more frequently encountered in the "core" of the LC vs. its extranuclear dendritic zone, which included the medial parabrachial nucleus (mPB). Triple labeling for ENK, CRF, and tyrosine hydroxylase (TH) showed convergence of opioid and CRF axon terminals with noradrenergic dendrites as well as evidence for inputs to TH-labeled dendrites from dually labeled opioid/CRF axon terminals. One potential source of ENK and CRF in the dorsal pons is the central nucleus of the amygdala (CNA). To determine the relative contribution of ENK and CRF terminals from the CNA, the CNA was electrolytically lesioned. Light-level densitometry revealed robust decreases in CRF immunoreactivity in the LC and mPB on the side ipsilateral to the lesion but little or no change in ENK immunoreactivity, confirming previous studies of the mPB. Degenerating terminals from the CNA in lesioned rats were found to be in direct contact with TH-labeled dendrites. Together, these data indicate that ENK and CRF may be colocalized to a subset of individual axon terminals in the LC "core." The finding that the CNA provides, to dendrites in the area examined, a robust CRF innervation, but little or no opioid innervation, suggests that ENK and CRF axon terminals impacting LC neurons originate from distinct sources and that terminals that colocalize ENK and CRF are not from the CNA.     

Central blockade of oxytocin receptors during late gestation disrupts systemic release of oxytocin during suckling in rats

There is evidence that the central oxytocin system is activated and undergoes reorganization before parturition. The present study was designed to determine the effects of central oxytocin receptor blockade during late gestation on parturition, pup growth, and oxytocin release during suckling. Female Sprague-Dawley rats were implanted on gestation day 12-14 with Alzet osmotic minipumps containing an oxytocin receptor antagonist (d(CH2)5, Tyr(Me)(2), Orn(8)-vasotocin; OT-X) or artificial cerebrospinal fluid (VEH), which was infused into the third cerebral ventricle. Pumps were removed within 24 h of parturition. Daily maternal body weight and food intake were monitored during gestation and lactation. The length of gestation, duration of parturition, pup number, litter weight and interbirth interval were recorded. Subsequently, pup number and litter weights were recorded daily until lactation day 10 or 11, when maternal and pup behaviour, and plasma oxytocin concentration before and during suckling were measured. Central oxytocin blockade had no effect on the timing of parturition, maternal behaviour, litter size, still births, or litter weights at birth. However, beginning on day 3 of lactation, average weights of litters of OT-X females were significantly lower than litters of VEH-treated females. Furthermore, while basal plasma oxytocin concentrations, oxytocin increases in response to suckling and dam/pup interactions did not differ between groups, a significant delay in suckling-induced systemic oxytocin release was observed in OT-X females. Finally, OT-X dams weighed less than VEH dams during the postpartum observation period, although food intakes were similar. These data suggest that central actions of oxytocin during late gestation are necessary for the normal timing of systemic release of oxytocin during suckling, normal pup weight gain, and maintenance of maternal body weight.     

Beta-endorphin cells in the arcuate nucleus: projections to the supraoptic nucleus and changes in expression during pregnancy and parturition

Supraoptic nucleus oxytocin neurone activity and secretion are inhibited in late pregnancy and parturition by endogenous opioids. Here, we investigated alterations in the projections and gene expression of beta-endorphin/pro-opiomelanocortin neurones in the arcuate nucleus in the pregnant rat. All regions of the arcuate nucleus were found to contain cells immunoreactive for beta-endorphin fluorescent microbeads retrogradely transported from the supraoptic nucleus, and double-labelled neurones (beta-endorphin plus microbeads), showing that beta-endorphin neurones throughout the arcuate nucleus project to the supraoptic nucleus. There was an increase in the number of beta-endorphin-immunoreactive cells in the arcuate nucleus and an increase in the density of beta-endorphin fibres within the supraoptic nucleus and peri-supraoptic region in late pregnancy and parturition, suggesting enhanced expression of beta-endorphin and increased beta-endorphin innervation of the supraoptic nucleus. Pro-opiomelanocortin mRNA expression in the arcuate nucleus increased in late compared to early pregnancy: the number of positive neurones significantly increased in the caudal region. Fos expression (an indicator of neuronal activation) in the arcuate nucleus was colocalized in beta-endorphin neurones in both proestrus and parturient rats, but the number of positive cells did not increase during parturition, suggesting lack of activation of beta-endorphin neurones at birth. Thus, beta-endorphin cells in the arcuate nucleus project to the supraoptic nucleus and increased innervation during pregnancy may explain the enhanced endogenous opioid inhibition of oxytocin neurones.     

Plasticity of neurohypophysial terminals with increased hormonal release during dehydration: ultrastructural and biochemical analyses

Arginine vasopressin- (AVP) and oxytocin- (OXT) secreting magnocellular neurons undergo gross structural changes with chronic physiological stimulation. Here, we investigated subcellular aspects of plasticity in rat neurohypophysial terminals during dehydration. Ultrastructural analyses demonstrated that chronic dehydration by 2% NaCl drinking for 7 days significantly decreased the numbers of neurosecretory granules and microvesicles but not the numbers of mitochondria. Moreover, in dehydrated rats, terminals making neurovascular contacts enlarged, whereas terminals in apposition to astrocytes, i.e., neuroglial contacts, became smaller. Western blot analyses demonstrated significant decreases in the levels of F3 and Thy-1 together with those of AVP- and OXT-neurophysin, but the levels of synaptophysin, SNAP-25, and GAP-43 were unchanged. Both F3 and Thy-1 were recovered in the buffer-insoluble pellet, and phosphatidyl inositol-specific phospholipase C treatment released both molecules from the crude membrane fraction, indicating that they are attached to terminal membranes by glycosylphosphatidyl inositol anchors. Confocal microscopic observations demonstrated that F3 colocalized with Thy-1 in the same terminals of magnocellular neurons. In contrast, the level of calretinin, a Ca(2+) binding protein was significantly increased with chronic dehydration. Thus, the present results suggest that enhancement of neurovascular contacts results from rearrangement of terminal-astrocyte and terminal-vessel contacts rather than enlargement or sprouting of magnocellular terminals themselves. The down-regulation of F3 and Thy-1 may contribute to enhancement of neurovascular contacts that accompany increased peptide release during dehydration.     

Nitric oxide acutely modulates hypothalamic and neurohypophyseal carbon monoxide and hydrogen sulphide production to control vasopressin,oxytocin and atrial natriuretic peptide release in rats

Nitric oxide (NO) negatively modulates the secretion of vasopressin (AVP), oxytocin (OT) and atrial natriuretic peptide (ANP) induced by the increase in extracellular osmolality, whereas carbon monoxide (CO) and hydrogen sulphide (H₂S) act to potentiate it; however, little information is available for the osmotic challenge model about whether and how such gaseous systems modulate each other. Therefore, using an acute ex vivo model of hypothalamic and neurohypophyseal explants (obtained from male 6/7‐week‐old Wistar rats) under conditions of extracellular iso‐ and hypertonicity, we determined the effects of NO (600 μmol L^‐1 sodium nitroprusside), CO (100 μmol L^‐1 tricarbonylchloro[glycinato]ruthenium [II]) and H₂S (10 mmol L^‐1 sodium sulphide) donors and nitric oxide synthase (NOS) (300 μmol L^‐1 N_ω‐methyl‐l ‐arginine [LNMMA]), haeme oxygenase (HO) (200 μmol L^‐1 Zn(II) deuteroporphyrin IX 2,4‐bis‐ethylene glycol [ZnDPBG]) and cystathionine β‐synthase (CBS) (100 μmol L^‐1 aminooxyacetate [AOA]) inhibitors on the release of hypothalamic ANP and hypothalamic and neurohypophyseal AVP and OT, as well as on the activities of NOS, HO and CBS. LNMMA reversed hyperosmolality‐induced NOS activity, and enhanced hormonal release by the hypothalamus and neurohypophysis, in addition to increasing CBS and hypothalamic HO activity. AOA decreased hypothalamic and neurohypophyseal CBS activity and hormonal release, whereas ZnDPBG inhibited HO activity and hypothalamic hormone release; however, in both cases, AOA did not modulate NOS and HO activity and ZnDPBG did not affect NOS and CBS activity. Thus, our data indicate that, although endogenous CO and H₂S positively modulate AVP, OT and ANP release, only NO plays a concomitant role of modulator of hormonal release and CBS activity in the hypothalamus and neurohypophysis and that of HO activity in the hypothalamus during an acute osmotic stimulus, which suggests that NO is a key gaseous controller of the neuroendocrine system.     

Osmotic Inhibition of Prolactin Secretion in Rats

Chronic hyponatremia is known to cause inhibition of pituitary vasopressin (AVP) and oxytocin (OT) secretion in response to most physiological stimuli, as well as a marked inhibition of synthesis of these peptides. Because many studies have implicated neurohypophyseal peptides in the regulation of pituitary prolactin (PRL) secretion, we investigated the effects of chronic hyponatremia on basal and stimulus-induced PRL secretion in rats. Hyponatremia was induced by subcutaneous infusion of 1-deamino-[8-D-arginine]-vasopressin (dDAVP) (5ng/h) to rats fed a nutritionally balanced liquid diet, and plasma [Na+] was maintained ≤115 mmol/l for 10–12 days. After this period, hyponatremic rats and normonatremic controls fed the same diet without dDAVP were subjected to one of the following stimuli known to stimulate PRL release in rats: 3 min exposure to ether, hemorrhage (20 ml/kg), intravenous injection of 5-hydroxytryptophane (5-HTP, 10 mg/kg), or intravenous injection of estradiol (5 μg/kg). A baseline blood sample was collected before each stimulus, and 3–6 additional blood samples were collected at selected intervals after the stimulus. Baseline levels of plasma PRL were not different between normonatremic and hyponatremic rats. However, PRL responses induced by ether or estradiol, but not those induced by hemorrhage or 5-HTP, were very significantly blunted in the chronically hyponatremic rats. Plasma AVP and OT responses were measured as an index of magnocellular secretion, but did not correlate with the PRL responses for any of the stimuli tested. Our results therefore demonstrate that ether- and estradiol-induced PRL release can be osmotically inhibited, but the mechanisms underlying this inhibition appear to be relatively independent of effects on magnocellular AVP and OT secretion. This allows the possibility that either some parvocellular systems regulating PRL secretion are osmosensitive, or alternatively that other substances released from the neural lobe may selectively modulate pituitary PRL release in response to some, but not all, stimuli.     

Vasopressin and oxytocin release in the brain - a synaptic event

In order to obtain evidence for a central release of vasopressin and oxytocin, the release of these peptides was demonstrated in various extrahypothalamic areas of the brain. It proved that in those areas where these peptidergic fibers terminate synaptically a vasopressin and/or oxytocin calcium-dependent release, similar to that in the neurohypophysis, could be evoked by potassium or veratridine. Such release was not found in areas in which these fibers do not exhibit synaptic specialization.     

Effects of pinealectomy on neurohypophysial hormones in the SFO and plasma of dehydrated rats exposed to 12 hours of light

Magnocellular neurons in the supraoptic and paraventricular nuclei synthesize and release vasopressin and oxytocin in response to dehydration. Pinealectomy has been observed to decrease the distribution in the supraoptic nuclei of thiamine diphosphate-phosphohydrolase, an enzyme specific for the Golgi apparatus that correlates positively with neurosecretory activity. Based upon these studies we postulated that pinealectomy would alter the concentration of neurohypohysial hormones in plasma elevated by 48 hr of water deprivation. In addition, we investigated the possibility that pinealectomy would affect vasopressin concentration in another circumventricular organ, the subfornical organ (SFO) and in a adjacent fiber tract of the limbic system, the hippocampal commissure-fornix (HC-F). Adult, male, Sprague-Dawley rats exposed to a 12 hr light/dark cycle were either unoperated (controls; C), sham-operated (Sham; S) or pinealectomized (PX) three weeks prior to testing. Food and water consumption and urinary excretion of Na and K were measured for 7 days. On the fifth day, half of the animals in each treatment group (C, S, PX) were deprived of water for 48 hr. Animals were decapitated on day 8. Vasopressin and oxytocin in plasma were extracted using bentonite and acetone-ether, respectively, then quantified by radioimmunoassay. The SFO and HC-F were microdissected from each brain. Like tissues from 4 rats were pooled, homogenized in 0.1 N HCl, and centrifuged. The supernatant was neutralized and vasopressin was quantified by radioimmunoassay. Dehydration resulted in antidiuresis, increased urine concentrations of Na and K, a decreased ratio of Na:K in urine, and reduced food consumption of similar magnitudes in all groups (C, S, PX; p greater than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Stress-induced changes in the analgesic and thermic effects of opioid peptides in the rat

Stress (e.g. restraint) potentiates analgesia and alters changes in body temperature induced by morphine administered either systemically or intracerebroventricularly (i.c.v.) in rats. In order to extend the generality of this phenomenon to opioid peptides, we determined whether the analgesic and thermic effects of i.c.v. D-Ala2-D-Leu5-enkephalin (DADLE) or D-Ala2-N-MePhe4-Gly5(ol)-enkephalin (DAGO), agonists selective for delta- and mu-opioid receptors, respectively, were affected by restraint stress. Analgesia was measured in the tail-flick test and core body temperature by rectal probe. The unstressed rats exhibited a dose-dependent increase in tail-flick latencies after administration of either DAGO or DADLE. Restrained rats treated with DAGO or DADLE had a greater analgesic response to each dose of peptide than did unstressed rats; both the magnitude and duration of the drug effect were increased. The unstressed group of rats responded to all doses of DAGO and DADLE with an increase of core temperature. In contrast, restrained rats showed a decrease of core temperature following injection with either DAGO or DADLE. Thus, restraint stress can significantly modify the effects of DAGO and DADLE on analgesia and body temperature in a manner that is qualitatively and quantitatively similar to that observed previously for morphine administered by the i.c.v. route.     

Effects of centrally administered endogenous opioid peptides on drinking behavior, increased plasma vasopressin concentration and pressor response to hypertonic sodium chloride

Intracerebroventricular (i.v.t.) administration of beta-endorphin or leucine5-enkephalin inhibited drinking behavior, the pressor response and increased plasma vasopressin concentration stimulated by an acute elevation in CSF sodium chloride concentration (10 microliter, 1 M NaCl i.v.t.). These effects of endogenous opioid peptides were prevented by naloxone, indicating opiate receptors were required for the biologic response. Drinking behavior associated with regulatory stimuli operant during dehydration was also inhibited by opioid peptides. beta-Endorphin (i.v.t.) delayed the onset and/or reduced the volume of water consumed in response to hypertonic sodium chloride (relative cellular dehydration), polyethylene glycol (hypovolemia) and food-associated drinking behavior. Inhibition of drinking did not appear related to sensory-motor dysfunction as another motivated behavior, eating (onset, amount consumed) was unaffected by beta-endorphin. It is concluded from these results that centrally administered endogenous opioid peptides inhibit sodium chloride-stimulated cerebral mechanisms affecting blood pressure and hydration.     

Oestradiol potentiates hormone secretion and neuronal activation in response to hypertonic extracellular volume expansion in ovariectomised rats

Secretion of vasopressin (VP), oxytocin (OT) and atrial natriuretic peptide (ANP) is an essential mechanism for the maintenance of hydromineral homeostasis. Secretion of these hormones is modulated by several circulating factors, including oestradiol. However, it remains unclear how oestradiol exerts this modulation. In the present study we investigated the participation of oestradiol in the secretion of VP, OT and ANP and in activation of vasopressinergic and oxytocinergic neurones of the supraoptic (SON) and paraventricular (PVN) nuclei of the hypothalamus in response to extracellular volume expansion (EVE). For this purpose, ovariectomised (OVX) rats treated for 7 days with vehicle (corn oil, 0.1 ml/rat, OVX+O group) or oestradiol (oestradiol cypionate, 10 μg/kg, OVX+E group) were subjected to either isotonic (0.15 m NaCl, 2 ml/100 g b.w., i.v.) or hypertonic (0.30 m NaCl, 2 ml/100 g b.w., i.v.) EVE. Blood samples were collected for plasma VP, OT and ANP determination. Another group of rats was subjected to cerebral perfusion, and brain sections were processed for c‐Fos‐VP and c‐Fos‐OT double‐labelling immunohistochemistry. In OVX+O rats, we observed that both isotonic and hypertonic EVE increased plasma OT and ANP concentrations, although no changes were observed in VP secretion. Oestradiol replacement did not alter hormonal secretion in response to isotonic EVE, but it increased VP secretion and potentiated plasma OT and ANP concentrations in response to hypertonic EVE. Immunohistochemical data showed that, in the OVX+O group, hypertonic EVE increased the number of c‐Fos‐OT and c‐Fos‐VP double‐labelled neurones in the PVN and SON. Oestradiol replacement did not alter neuronal activation in response to isotonic EVE, but it potentiated vasopressinergic and oxytocinergic neuronal activation in the medial magnocellular PVN (PaMM) and SON. Taken together, these results suggest that oestradiol increases the responsiveness of vasopressinergic and oxytocinergic magnocellular neurones in the PVN and SON in response to osmotic stimulation.