Further Studies on the Role of Angiotensin in the Osmotic Control of Vasopressin Release by the Organ-Cultured Rat Hypothalamo-Neurohypophyseal System*

Abstract

Previous studies with organ-cultured hypothalamo-neurohypophyseal (HNS) explants suggested that angiotensin (A) II may either mediate or modulate osmotically stimulated vasopressin (VP) release by HNS explants and also suggested that All is generated in the culture system. The former was investigated by examining the effect of simultaneous exposure of explants to All and osmotic stimulation. VP release was significantly greater (P < 0.005) when All (10-8 M) was combined with a 5 mosmol/kg increment in osmolality than with either stimulus alone. Similar augmentation of osmotically stimulated VP was not observed when acetylcholine (10-7 M or 10-9 M) was presented simultaneously with a 5 mosmol/kg increment in osmolality. Thus these data suggest that a specific effect of All is to potentiate VP release to an osmotic stimulus. Generation of AH in the culture system was evaluated using the converting enzyme inhibitor, SQ 14,255. Converting enzyme activity in HNS cultures was demonstrated by evaluating the effectiveness of AI as a stimulus for VP release. AI increased VP release (P < 0.005), but the response was blocked by the simultaneous addition of SQ14,255 (10-5 M), suggesting that AI must be converted to All in order to stimulate VP release and that this conversion takes place in the culture system. The role of culture system-generated All in the osmotic stimulation of VP release was evaluated by assessing the response of explants exposed to SQ14,255 for various periods before the application of an osmotic stimulus. There was a significant increase in VP release by explants exposed simultaneously to a 20 mosmol increase in osmolality and SQ 14,255 (10-5M), but explants with prior exposure (4 h or 24 h periods) to the converting enzyme inhibitor did not show a significant response to the osmotic stimulus. Since SQ14,255, saralasin, and All antiserum do not alter unstimulated VP release, these observations suggest that All is being generated in the culture system in amounts which potentiate the osmotic response, but which alone are insufficient to stimulate VP release. In order to evaluate the source of All in the culture system, the potential contribution of the fetal calf serum component of the culture medium to the generation of All during culture was assessed. The fetal calf serum was evaluated for renin-AII components and was demonstrated to contain renin, renin substrate, AI, and AIL In order to eliminate the possibility that the All necessary for osmotic stimulation of VP release was generated as a result of the intermingling of the renin-AII components of the fetal calf serum with converting enzyme from the explant, explants were maintained in medium supplemented with heatinactivated serum which had no demonstrable renin activity. These explants maintained osmosensitivity, indicating that exogenous renin is not required for All generation in the culture system.