Foot shock stress-induced release of vasopressin in adenohypophysectomized and hypophysectomized rats.

The effect of inescapable electric foot shock stress on vasopressin (VP) release was studied in conscious rats. In sham-operated animals, foot shock stress markedly increased plasma beta-endorphin-like immunoreactivity whereas plasma VP levels (RIA) remained unchanged. However, after selective ablation of the anterior lobe of the hypophysis, foot shock stress produced an 8-fold increase in plasma VP concentrations. Injection of hypertonic saline did not change plasma VP levels in adenohypophysectomized (Ahx) rats, while in sham-operated rats VP levels increased in response to this osmotic challenge. Neurointermediate lobes or medial basal hypothalami (MBH; containing the median eminence region) were superfused in vitro and stimulated electrically; as compared to sham operations, the evoked release of VP from the neurointermediate lobes was abolished, whereas that from the MBH was markedly (13-fold) enhanced when the tissues were taken from Ahx rats. The VP-like immunoreactivity released from the MBH of Ahx rats comigrated with synthetic arginine-VP on Sephadex G-15 column chromatography. Similarly, as found in Ahx rats, foot shock stress markedly raised plasma VP levels in totally hypophysectomized rats, whereas after selective ablation of the neurointermediate lobe of the hypophysis VP levels increased only slightly after stress. In conclusion, our data indicate that 1) foot shock stress induces the release of VP from some site other than the neurohypophysis, probably from the median eminence region, in Ahx rats; and 2) an increase in plasma osmolality is a powerful stimulus for the release of VP from the neurohypophysis but cannot release VP from the median eminence region. Thus, our results support the concept of a morphological and functional differentiation of the vasopressinergic neurosecretory system.

In vitro adrenocorticotropin/beta-endorphin-releasing activity of vasopressin analogs is related neither to pressor nor to antidiuretic activity.

The ability of vasopressin and related analogs to induce ACTH, beta-endorphin, and beta-lipotropin release was studied in vitro using incubated rat anterior pituitary quarters or a perifused rat isolated anterior pituitary cell column. Vasopressin and its analogs exhibited corticotropin-releasing factor (CRF)-like activity in a rank order which was different from those for vasopressor or antidiuretic activity. Two dissimilar antagonists with antivasopressor activity showed different effects: one possessed CRF-like activity itself, the other blocked the CRF-like activity of vasopressin. Another antagonist with antipressor and also antiantidiuretic activity had no effect when given alone and also didn't block the CRF-like activity of vasopressin. Some analogs were also tested for their effects on cAMP accumulation. Analogs, which possessed CRF-like activity or blocked CRF-like activity of vasopressin, stimulated cAMP accumulation or inhibited vasopressin-stimulated cAMP accumulation in anterior pituitary quarters, respectively. These results imply that the structural requirements of the CRF-like activity of vasopressin differ from those of the pressor and antidiuretic activity. Therefore, it is possible that the pituitary receptors responsible for CRF-like activity of vasopressin represent a separate category of vasopressin receptors which may be linked to an adenylate cyclase.

Inhibition by prostaglandin E2 of the release of vasopressin and beta-endorphin from rat pituitary neurointermediate lobe or medial basal hypothalamus in vitro.

The present study was performed to examine the effect of the cyclo-oxygenase inhibitor, indomethacin, and that of various prostaglandins on the release of vasopressin and beta-endorphin-like immunoreactivity (beta-EI) from the rat neurointermediate lobe of the hypophysis, which was superfused in vitro. Indomethacin (2.8 and 28 mumol/l) changed neither basal secretion of vasopressin nor that evoked by electrical stimulation, whereas the resting release of beta-EI was enhanced by indomethacin (28 mumol/l). Prostaglandin (PG) E2 did not influence resting release of vasopressin but markedly inhibited (by about 50%) electrically induced release of vasopressin (least effective concentration: 300 nmol/l) as well as spontaneous secretion of beta-EI (least effective concentration: 100 nmol/l) in the presence of indomethacin (28 mumol/l). Prostaglandin F2 alpha (5 mumol/l) also inhibited the evoked release of vasopressin, whereas PGD2 (5 mumol/l) did not. Prostaglandin F2 alpha (5 mumol/l), D2 and I2 (1.5 mumol/l each) produced no effects on beta-EI release. As observed in the neurohypophysis, PGE2 inhibited the electrically induced release of vasopressin from the medial basal hypothalamus in vitro. We conclude that prostaglandins (especially PGE2) can inhibit (1) the stimulated release of vasopressin when acting on vasopressin-containing nerve terminals of either neurosecretory system (neurohypophysis, median eminence region), and (2) the secretion of beta-EI and, as can be inferred, alpha-MSH, by a direct action on intermediate lobe cells.

beta-Endorphin controls vasopressin release during foot shock-induced stress in the rat.

This study tested the possibility that beta-endorphin is involved in the regulation of vasopressin release during stress induced by inescapable electric foot shock. To this end, a specific anti-beta-endorphin antiserum or a control serum lacking the specific anti-beta-endorphin antibodies was administered to male rats. Plasma vasopressin concentrations, measured by radioimmunoassay, were not affected by brief foot shock stress in control rats, but were raised significantly by the stress in animals which had received an intracerebroventricular (i.c.v.) injection of the anti-beta-endorphin antiserum. In contrast, when the same volume of the anti-beta-endorphin antiserum was injected into a tail vein, foot shock stress produced only a slight effect on vasopressin release. I.c.v. injection of the antiserum changed neither basal nociceptive threshold nor stress-induced analgesia as revealed by the tail-flick latency. Vasopressin release induced by an osmotic stimulus was not influenced by the anti-beta-endorphin antiserum given i.c.v. The opiate antagonist naloxone or the glucocorticoid dexamethasone raised plasma vasopressin concentration in stressed rats which had received the control serum (i.c.v.); however, after i.c.v. injection of the anti-beta-endorphin antiserum neither naloxone nor dexamethasone elevated the plasma vasopressin concentration beyond the level reached by the anti-beta-endorphin antiserum (i.c.v.) alone. These results suggest that beta-endorphin inhibits the release of vasopressin during foot shock-induced stress in the rat.

Effect of transection of subfornical organ efferent projections on vasopressin release induced by angiotensin or isoprenaline in the rat.

Transection of subfornical organ efferents in the rat prevented the vasopressin release in response to intravenous angiotensin II infusion or following a small dose of the beta-sympathomimetic amine isoprenaline (30 micrograms/kg i.m.). In contrast, this lesion had no effect on vasopressin release after hypertonic saline injection or a high dose of isoprenaline (480 micrograms/kg i.m.). We conclude that blood-borne angiotensin II induces vasopressin release by acting on the subfornical organ; depending on the dose of isoprenaline, activation of the endogenous renin-angiotensin system may mediate isoprenaline-induced vasopressin release.

Evidence that vasopressin is involved in the isoprenaline-induced beta-endorphin release.

We investigated in conscious rats, whether vasopressin, whose release was stimulated by isoprenaline (i.m.), caused the simultaneous increase in plasma beta-endorphin-like immunoreactivity (beta-EI). The increase in plasma beta-EI following isoprenaline administration was diminished by about 50% in rats pretreated with a vasopressin antagonist and also in rats with a hereditary absolute lack of vasopressin. We conclude that the isoprenaline-induced release of beta-EI is mediated in part by vasopressin.

Vasopressin and beta-endorphin release after osmotic and non-osmotic stimuli: effect of naloxone and dexamethasone.

The purpose of this study was to determine whether or not vasopressin release in response to various stimuli in the conscious rat is controlled by endogenous opioid peptides, in particular beta-endorphin. Naloxone (1 mg.kg-1 i.m.) promoted vasopressin release in response to both an angiotensin II infusion (500 ng . kg-1 . min-1) or an isosmolar, nonhypotensive hypovolaemia achieved by polyethylene glycol injection (PEG, 20% solution i.p.); however, naloxone was without effect when vasopressin release was induced by hypertonic saline injection (2.5% solution i.p.) or a severe fall in arterial blood pressure following trimethidinium (10 mg . kg-1 i.m.) induced ganglionic blockade. Vasopressin release was accompanied by an increase in plasma beta-endorphin-like immunoreactivity (beta-EI) following an angiotensin II infusion of PEG administration, but not after hypertonic saline or trimethidinium injection. Dexamethasone pretreatment (0.5 mg . kg-1 twice i.p.) prevented the increase in plasma beta-EI following an angiotensin II infusion or PEG administration. The simultaneous angiotensin II- or PEG-induced increase in vasopressin release was unaffected or potentiated, respectively, by the glucocorticoid. In contrast, vasopressin release in response to hypertonic saline or trimethidinium injection was significantly inhibited by dexamethasone. We conclude that an inhibitory control by endogenous opiates is involved in some, but not all of the different pathways leading to vasopressin release. The results obtained do not prove but can be reconciled with the proposal that hypophyseal beta-endorphin is the compound responsible.

Vasopressin release from rat medial basal hypothalamus induced in vitro by angiotensin.

Rat medial basal hypothalami were superfused in vitro. The effect of angiotensin II on vasopressin outflow was investigated. Angiotensin II (10 nM or 1 microM, added to the superfusion medium) increased the veratridine-evoked vasopressin release. The higher concentration also slightly elevated the basal outflow. The effect of angiotensin II was blocked by saralasin. We conclude that angiotensin II can act on the median eminence and/or on the stump of the pituitary stalk to promote the release of vasopressin, which then may influence anterior pituitary hormone secretion.

Vasopressin release from rat medial basal hypothalamus in vitro after adrenalectomy or lesions of the paraventricular nuclei.

Rat medial basal hypothalami (MBH) or neurointermediate lobes of the hypophysis (NIL) were superfused in vitro and stimulated electrically. The evoked release of vasopressin from the MBH was enhanced to 700% of controls when the tissue was taken from adrenalectomized rats. By contrast, the evoked release of vasopressin from the NIL was not changed after adrenalectomy. After bilateral lesions of the paraventricular nuclei, the evoked release of vasopressin from the MBH was reduced in subsequent in vitro experiments. The marked changes in vasopressin release occurred in spite of no or only small changes in the total tissue content of vasopressin. These data support the view that vasopressin may be released from the external layer of the median eminence into the hypophysial portal blood after activation of the hypothalamo-pituitary-adrenal axis.

Phosphorylated derivatives of phloretin inhibit cyclic AMP accumulation in neuronal and glial tumor cells in culture.

The potencies of polyphloretin phosphate, di-4-phloretin phosphate, 4-phloretin phosphate and phloretin to inhibit the stimulation of cAMP accumulation by prostaglandins, isoproterenol and adenosine were studied in 2 clonal cell lines of CNS origin. The sequence of potency to inhibit PGE1 effects was the same in neuroblastoma (N4TG3) and human astrocytoma cells (1321N1): di-4-phloretin phosphate greater than polyphloretin phosphate greater than phloretin greater than 4-phloretin phosphate. The inhibition of PGE1 stimulated cAMP accumulation by the most prostaglandin-specific inhibitor di-4-phloretin phosphate was rapidly established after its addition, fully reversible after a 30 min preincubation period and independent of the presence of calcium. Kinetic studies of the inhibition of PGE1 effects by di-4-phloretin-phosphate suggest a different type of inhibition in 1321N1 and N4TG3 cells.

The effect of isoprenaline on plasma concentrations of immunoreactive beta-endorphin and beta-lipotropin in the conscious rat.

The effect of the beta-adrenoceptor agonist isoprenaline on the plasma concentrations of beta-endorphin (beta-E) and beta-lipotropin (beta-LPH) was investigated in conscious rats. Isoprenaline (i.m.) elevated plasma beta-endorphin-like immunoreactivity (beta-EI) as measured by radioimmunoassay of unextracted plasma, with peak values 24 min after drug administration. This effect was dose-dependent. The lowest effective dose of isoprenaline was 15 micrograms kg-1; 240 micrograms kg-1 exerted a maximum effect, raising plasma beta-EI about ten-fold above control values. Plasma vasopressin concentrations also increased in response to isoprenaline following a time-course identical to that of plasma beta-EI. (+/-)-Propranolol (1 mg kg-1) but not phentolamine (10 mg kg-1) rendered isoprenaline (240 micrograms kg-1) injections almost ineffective. Because of the cross-reactivity of beta-LPH in the radioimmunoassay used, plasma was extracted by means of a cation exchange resin and subjected to gel chromatography on a Sephadex G-50 column, avoiding artefactual degradation of the peptides. In isoprenaline-treated rats about 50% of the beta-EI behaved similar to human beta-LPH, whereas 45% co-migrated with human beta-E; immunoreactivity corresponding to beta-LPH or beta-E comprised about 70% or 30%, respectively, in the plasma extract of vehicle-treated rats. Dexamethasone pretreatment reduced the isoprenaline-induced increase in plasma beta-EI by 87%, but left the simultaneous elevation of plasma vasopressin concentrations unchanged. These data demonstrate that isoprenaline stimulates beta-LPH and beta-E release in vivo. The possibility of an interrelationship between vasopressin and beta-E release is discussed.

Vasopressin stimulates release of beta-lipotropin and beta-endorphin in conscious rats as measured by radioimmunoassay of unextracted plasma.

Using a newly developed radioimmunoassay to determine the beta-endorphin-like immunoreactivity (beta-EI) in unextracted plasma, the effect of vasopressin injections on plasma beta-EI was investigated in conscious rats. Arginine vasopressin caused a dose-dependent increase of plasma beta-EI from 34.5 to 7.8 fmol ml--1 (n = 6) in vehicle-treated animals to 205.0 +/- 36.1 fmol ml--1 (n = 7) after injection of the highest vasopressin dose employed (486 ng/100 g b.w.). In view of the appreciable cross-reactivity of beta-lipotropin (beta-LPH) in the radioimmunoassay used, plasma was extracted and subjected to gel chromatography on a Sephadex G-50 column. On average, about 70% of the beta-EI co-eluted with human beta-LPH and about 30% with human beta-endorphin in plasma extracts obtained from both control and vasopressin-treated rats. No peripheral conversion of human beta-LPH occurred under the experimental conditions, since after i.v. bolus injection of human beta-LPH 97% of the beta-EI comigrated with human beta-LPH during gel filtration. A similar blood pressure increase to that induced by the vasopressin injections, when elicited by noradrenaline or angiotensin II i.v., was not followed by an elevation of plasma beta-EI. These data indicate that vasopressin stimulates beta-lipotropin and beta-endorphin release into the systemic circulation in vivo.

Effect of neonatal treatment with monosodium glutamate on vasopressin release during foot shock stress in the rat.

Several lines of evidence indicate that beta-endorphin inhibits the release of vasopressin during foot shock-induced stress in the rat. This study was to evaluate the relative importance of the hypothalamic versus the pituitary pool of beta-endorphin. Neonatal treatment with monosodium glutamate (MSG) reduced drastically the content of beta-endorphin-like immunoreactivity (beta-EI) of hypothalamus but not the beta-EI concentration in the pituitary; the content of vasopressin in the hypothalamus and the pituitary was not altered by MSG treatment. MSG treatment had no effect on the plasma vasopressin response to inescapable electric foot shock stress, when compared to controls. Naloxone enhanced vasopressin release during stress both in MSG-treated rats and in controls. These results suggest that hypothalamic beta-endorphin is not involved in the control of vasopressin release during foot shock-induced stress in the rat.

Naloxone increases vasopressin secretion from the neurointermediate lobe of the hypophysis of the rat: search for the endogenous agonist.

Naloxone increases the electrically induced vasopressin release from rat pituitary neurointermediate lobes under appropriate stimulation conditions. In order to examine a possible role of hypophyseal opioid peptides we studied in vitro the effect of opioid peptides and of naloxone on the electrically induced vasopressin release from the rat neurointermediate lobe or isolated neural lobe of the hypophysis. (D-Ala2,D-Leu5)-enkephalin (5 microM), dynorphin-(1-13) (Dyn; 0.2 microM), beta-endorphin (beta-End; 0.02 and 0.2 microM) and also naloxone (1 or 10 microM) increased the evoked vasopressin release from the neurointermediate lobe, but in higher concentrations (2 microM) Dyn or beta-End had no effect. After removal of the intermediate lobe, beta-End 2 microM inhibited, while naloxone 10 microM did not change the evoked vasopressin release from the isolated neural lobe. These results demonstrate that hypophyseal opioid peptides can influence vasopressin release in several ways and suggest that endogenous opioids predominantly provide inhibitory influences which depend on the presence of the intermediate lobe.

Evidence for the involvement of a GABA-mediated inhibition in the hypovolaemia-induced vasopressin release.

The influence of GABA and of drugs, known to alter GABA-metabolism, on the hypovolaemia-provoked vasopressin release was investigated in rats. Blood volume was decreased without altering plasma osmolality or arterial blood pressure by i.p. injection of polyethylene glycol and the resulting plasma vasopressin concentration was measured using a radioimmunoassay. I.c.v. injections of GABA (0.4-2 mg) markedly suppressed the hypovolaemia-induced vasopressin release. The central inhibitory effect of GABA could not be related to appropriate changes in peripheral parameters believed to regulate vasopressin release (arterial blood pressure, renin-angiotensin system). Aminooxyacetic acid (9-81 mg kg-1, i.m.) and gamma-vinyl-GABA (1.5 g kg-1, i.p.), two potent inhibitors of GABA aminotransferase and known to increase brain GABA content, reduced vasopressin release to a comparable degree as did GABA (i.c.v.). On the other hand, 3-mercaptopropionic acid (10-90 mg kg-1, i.p.), an inhibitor of the GABA synthetizing enzyme glutamic acid decarboxylase, promoted the release of vasopressin when the rats were killed prior to the onset of convulsions. These results, on the whole, intimate the existence of a GABA-mediated inhibition in the central control of vasopressin release.

Evidence for inhibition by beta-endorphin of vasopressin release during foot shock-induced stress in the rat.

This study was to ascertain the effect of naloxone and dexamethasone on vasopressin and beta-endorphin release in the rat during inescapable electric foot shock stress. Plasma vasopressin concentrations were not affected by electric foot shock in vehicle-treated rats, but were raised significantly by the stress in animals pretreated with naloxone. The stress-induced increase in plasma beta-endorphin-like immunoreactivity (beta-EI) was similar whether the rats had received naloxone or not. Plasma beta-EI consisted of equal amounts of beta-endorphin-like and beta-lipotropin-like material as revealed by gel filtration. Dexamethasone almost abolished the foot shock-induced increase in plasma beta-EI and, in the presence of dexamethasone, stress was now effective in elevating plasma vasopressin concentrations. These results are consistent with the hypothesis that beta-endorphin, released from the anterior pituitary, inhibits the release of vasopressin from the posterior lobe of the pituitary gland during foot shock-induced stress.

On the mechanism of the vasopressin-induced inhibition of renin release.

The isoprenaline-induced renin release was used to study both in vivo and in vitro and mechanism of the inhibitory effect of vasopressin on renin secretion. (1) The vasopressin analogue [1-deaminopenicillamine, 2-(0-methyl)-tyrosine]-arginine-vasopressin, which antagonizes the vasopressor response to vasopressin but possesses antidiuretic potency, abated the increase in plasma renin concentration following isoprenaline injection in the conscious rat without altering the fall in arterial blood pressure, (2) In a continuous superfusion system of rat kidney cortical slices, vasopressin in concentrations up to 10(-6) M failed to affect the isoprenaline-induced increase in renin secretion, which was evidently beta-adrenoceptor-mediated. In contrast, angiotensin II (10(-6) M) prevented the renin release in response to isoprenaline. These results led us to conclude that the inhibition of renin release caused by vasopressin is not necessarily related to its vasoconstrictor potency. The mechanism, through which vasopressin works, appears to differ from that of angiotensin II and is non functional in renal cortical slices.

Effect of isoprenaline on the plasma concentrations of angiotensin III in rats.

1. A new column-chromatographic method is described for the simple and reproducible determination of the concentration of [des-Asp1]angiotensin II (angiotensin III) in rat plasma. 2. The method uses the different abilities of the ion-exchange resins Dowex 1 (X8) and Bio Rex 70 to bind angiotensin II and angiotensin III. Under the conditions used, Bio Rex 70 binds only angiotensin III. Angiotensin II and its hexapeptide metabolite [des-Asp1,des-Arg2]angiotensin II pass the resin with the effluent. Dowex 1 (X8) binds angiotensin II and the hexapeptide metabolite, whereas it does not extract angiotensin III. It does not separate angiotensin II from the hexapeptide. Therefore the sum of both peptides is expressed as angiotensin II-like activity. The ratio of the concentrations, angiotensin II/hexapeptide, was 5:1. 3. In normal rats the plasma concentration of angiotensin III was 20 fmol/ml (SD 15; n = 8), and angiotensin II-like activity was 60 fmol/ml (SD 35; n = 8). 4. The beta-sympathomimetic amine isoprenaline caused a time- and dose-dependent increase in plasma angiotensin III and angiotensin II-like activities. 5. Under the conditions studied angiotensin III contributed approximately 25% to the total amount of angiotensins in plasma.