Osmotic control of vasopressin release by rat hypothalamo-neurohypophyseal explants in organ culture

The rat hypothalamo-neurohypophyseal system (HNS) in organ culture has been used as an in vitro system for studying the osmotic control of vasopressin (VP) release. The HNS retains osmotically sensitive components as demonstrated by changes in the rate of VP release following alterations in the osmolality of the culture medium. Increasing the osmolality from 295 to 305 mosmol/kg H2O by the addition of NaCl resulted in a 2.5-fold increase in VP release. VP release was significantly decreased subsequent to reducing the osmolality from 295 to 280 mosmol/kg H2O by the addition of distilled water. Also, VP release was stimulated when the osmolality was increased to 300 mosmol/kg H2O by the addition of mannitol, but not by additions of urea or glucose which resulted in comparable increases in the tonicity of the culture medium. These studies demonstrate that the HNS in organ culture responds appropriately to osmotic challenges within the physiological range, and support Verney's concept of an osmoreceptor inasmuch as both NaCl and mannitol were effective osmotic agents.     

Angiotensin stimulation of vasopressin release from the rat hypothalamo-neurohypophyseal system in organ culture.

Angiotensin II (AII) stimulated vasopressin (VP) release from the rat hypothalamo-neurohypophyseal system (HNS) in organ culture in a concentration-dependent manner. Exposure to AII at 10(-8) M for 1 hr yielded a 1.8-fold increase in VP release over control release (P less than 0.01), while a 1-h exposure to 10(-5) M AII resulted in a 4-fold increment over control VP release by HNS explants maintained in organ culture for 3 days (P less than 0.01). Saralasin, an AII antagonist, blocked AII stimulation of VP release without significantly altering basal VP release by the HNS explants. Saralasin did not interfere with stimulation of VP release by acetylcholine or nicotine. Tetrodotoxin (10(-7) g/ml) also blocked AII stimulation of VP release. These findings suggest that action potentials are generated in response to AII stimulation of specific receptors in the HNS and are requisite for VP release in response to this stimulus.     

Secretion of vasopressin, oxytocin, and two neurophysins from rat hypothalamo-neurohypophyseal explants in organ culture

Secretion of the neurohypophyseal principles arginine vasopressin, oxytocin, vasopressin-neurophysin, and oxytocin-neurophysin by hypothalamic-neurohypophyseal explants in organ culture was studied. The viability of explant cultures was tested on day 6 by their ability to incorporate 3H-amino acid into acid-precipitable protein, specifically vasopressin-neurophysin. The explants released all four neurosecretory products in a controlled manner during 8 successive days of culture. In addition, these substances were all present in neural lobes and hypothalami at the end of the culture period. Although the tissue retained a 1:1 molar ratio between the hormone and corresponding neurophysin, the levels of neurophysins in culture media were much higher than the levels of hormones. Through immunoprecipitation techniques we determined that both arginine vasopressin and oxytocin were unstable in culture media containing explants or in medium containing 10% heat-inactivated fetal calf serum. This suggests that neurophysins are more stable indicators of secretion by explants than are the corresponding hormones. An increased release of arginine vasopressin, oxytocin, vasopressin-neurophysin, and oxytocin-neurophysin occurred in response to elevated K+ and to increased sodium ions in the medium, and these four substances were released in different molecular proportions. The results indicate that explant cultures may be useful for comparisons of secretion of the two neurophysins and, to a more limited extent, of the two hormones in response to such stimuli.     

Alteration of endotoxin fever and release of arginine vasopressin by dehydration in the guinea pig.

Arginine vasopressin (AVP), synthesized in hypothalamic neurons, is transported in axons either to the pituitary for release into the circulation or to different brain areas. In our previous experiments we documented central antipyretic AVP pathways from the hypothalamus to the ventrolateral septal area in the limbic system. In the present study we investigated if osmotic stimulation is able to activate peripheral and central release of AVP concurrently and if the antipyretic pathways are influenced by this kind of stimulation. In dehydrated animals (24 h water deprivation) the arterial blood plasma level of AVP doubled causing antidiuretic effects. Also the concentration of AVP in push-pull perfusates of the limbic septal area was significantly higher in dehydrated (5.6 pg AVP/ml perfusate) than in control animals (2.6 pg AVP/ml perfusate). The febrile response to bacterial endotoxin was reduced by 50% in dehydrated guinea pigs compared to controls, statistically significant between 30 and 180 min after pyrogen application. A microinfusion of AVP antiserum into the limbic septal area enhanced the fever reaction of dehydrated guinea pigs compared to the effects of a microinfused preimmune serum, in this case statistically significant between 180 and 360 min after application. From these data we assume a simultaneous activation of peripheral and central release of AVP with antidiuretic and antipyretic effects by dehydration.     

Osmotic stimulation in DOC-salt hypertension: vasopressin and blood pressure

To determine whether there is a change in the sensitivity of the osmotic control of vasopressin release in deoxycorticosterone (DOC)-salt hypertension, experiments were performed in unilaterally nephrectomized rats that were either normotensive or were made hypertensive with DOC and given 1% saline to drink. After 3 weeks of treatment, 2.5 mol/l NaCl was infused i.v. into conscious normotensive and hypertensive rats. Increases in both plasma osmolality and plasma vasopressin concentration were similar throughout the course of this infusion in the two groups of rats. Hypertonic saline infusion increased the mean arterial blood pressure in the two groups of rats, but this increase was partially attenuated by the i.v. injection of a vasopressin pressor antagonist. In conclusion, vasopressin release in response to osmotic stimulation was similar in normotensive and hypertensive rats. The pressor response to hypertonic saline in both groups of rats could be partially attributed to the increased plasma vasopressin concentration.     

A comparison between the effects of angiotensin II and angiotensin III injected into the third cerebral ventricle on vasopressin secretion in conscious rats (author's transl)

As the greater part of the immunoreactive angiotensin II in cerebrospinal fluid has been suggested to be angiotensin III, a comparison was made between the effects on vasopressin release of angiotensin II and angiotensin III administered into the third cerebral ventricle in conscious male rats. The blood samples were collected 90 seconds after the injection of angiotensin II or angiotensin III by means of decapitation. Plasma vasopressin (microU/ml) extracted and determined by radioimmunoassay were 2.3 +/- 0.8, 6.7 +/- 5.0, 14.0 +/- 2.2, 16.3 +/- 4.3 and 20.7 +/- 2.5 (mean +/- SEM), respectively following the injection of 0, 10, 25, 50 and 100ng of angiotensin II. The increases in plasma vasopressin produced by angiotensin II 25, 50 and 100ng were statistically significant (p less than 0.05). On the other hand, plasma vasopressin following the injection of 22.7 and 45.4ng of angiotensin III, which are equimolar to 25 and 50ng of angiotensin II each, were 14.9 +/- 2.7 and 16.3 +/- 5.6, respectively. No significant difference was found between the effect on plasma vasopressin of angiotensin II and that of angiotensin III at the dose level of 24.3 or 48.6 p. mol. These data indicate that angiotensin III is equipotent to angiotensin II in terms of vasopressin release when administered into the third cerebral ventricle. The possible role of angiotensin III in the brain on vasopressin secretion is discussed.     

Neurohypophysial peptides in guinea pig hypophysial portal blood: equimolar release of the carboxyl terminal glycopeptide with arginine vasopressin

A method has been devised for collecting hypophysial portal blood from the anaesthetised guinea pig in order to measure the release in vivo of the neurohypophysial peptides, oxytocin (OT), vasopressin (AVP), neurophysin (NP), and the glycopeptide (GP) found at the carboxyl terminus of the AVP precursor. These peptides were measured in samples of portal and peripheral venous plasma by specific radioimmunoassays. The concentration of OT and AVP was 50- to 100-fold higher in hypophysial portal blood than in peripheral blood, with more OT than AVP usually present. There were correspondingly large amounts of NP and GP also present in portal blood. In particular, GP levels paralleled AVP levels over a wide range of concentrations and in virtually equimolar proportions. These results provide the first in vivo evidence which shows that, as for the magnocellular neurohypophysial system, GP is synthesised, processed and released in equal amounts with AVP from their common precursor in the subpopulation of parvocellular AVP neurons which project from the paraventricular nucleus to the median eminence.     

Intimal permeability evaluated in a short-term organ culture of diabetic guinea pig aorta

A novel short-term organ culture system was used to evaluate intimal permeability changes by measuring aortic [14C]methylated albumin accumulation. Aortic plugs were removed from the upper thoracic aorta of male guinea pigs and maintained in serum-free media. The accumulation of [14C]albumin in the intimal-medial layer was determined after a 5 h incubation. In preliminary studies, albumin recovered from intimal-injured aortic plugs was significantly greater than those from non-injured plugs. Aortic plugs from streptozotocin-treated guinea pigs, diabetic for 3 weeks, also accumulated significantly more [14C]albumin than plugs from nondiabetic controls. Histological changes were not observed in the aorta of either the diabetic or control group. A strong significant inverse correlation was found between plasma ascorbic acid levels and [14C]-activity recovered from aortic plugs. This study demonstrates a simple and rapid method for assessing aortic permeability changes under a well-defined in vitro system, and suggests that vascular permeability changes in the streptozotocin-diabetic guinea pig may be associated with an ascorbic acid deficit.