Evidence for two different stimulatory adenylate cyclase coupling mechanisms in rat renal papilla.

Abstract

Adenylate cyclase activity in renal papillary membranes was stimulated by both vasopressin and the adenosine agonist 5'-N-ethylcarboxamidoadenosine (NECA). The stimulations mediated by the two receptors were additive at all concentrations and interacted differently with other AC-stimulatory factors viz cholera toxin, pertussis toxin and fluoride ion. Treatment of papillary tubules with cholera toxin increased cyclase activity from 4.5 +/- 1.5 to 110 +/- 9.1 (SE, n = 5) pmol/min/mg protein. Maximally effective concentrations of vasopressin increased activity in control preparations to 10.3 +/- 2.8 (an increase of 5.8 +/- 1.3). In cholera toxin treated preparations, vasopressin increased activity to 138.9 +/- 14.5 (an increase of 28.9 +/- 5.4, n = 5; p less than .01). Pertussis toxin increased activity to 9.1 +/- 3.0. The response to vasopressin was enhanced such that the absolute maximum increase in activity was 12.6 +/- 3.9 (n = 5; p less than .01). Addition of the two toxins together produced a greater than additive stimulation to 145 +/- 36. Maximum increase in activity caused by vasopressin was further enhanced to 48 +/- 13 (n = 5; p less than .01). In contrast, cyclase stimulation by NECA was additive with stimulations by the two toxins, separately and in combination. The NECA stimulation however, was enhanced in the presence of fluoride ion while the vasopressin stimulation was additive at all concentrations. Papillary membranes contained two different cyclase-stimulatory coupling proteins with alpha-subunits of MW's 46,600 +/- 450 (SE, n = 6) and 41,500 +/- 480 (SE, n = 6) as identified on SDS-polyacrylamide gel electrophoresis following cholera toxin labeling. Taken together, these data suggest that two adenylate cyclase-stimulatory coupling mechanisms with different properties are operative in renal papillary membranes.