ROLE OF G-PROTEINS IN THE REGULATION OF ORGANIC OSMOLYTE EFFLUX FROM ISOLATED RAT RENAL INNER MEDULLARY COLLECTING DUCT CELLS

Hypotonic shock (change of osmolality from 600 mosmol to 300 mosmol by lowering NaCl concentration) increases the release of organic osmolyt es from isolated inner medullary collecting duct (IMCD) cells in the f ollowing sequence: taurine > betaine > sorbitol > myo-inositol > glyce rophosphorylcholine (GPC). The role of G-proteins in regulating the hy potonicity-induced efflux was analysed by exposing cells to various co ncentrations of a G-protein inhibitor, pertussis toxin (PTX; 20-200 ng /ml), and a G(i alpha)-protein stimulator, mastoparan (10-50 mu M). PT X diminished the hypotonic release of sorbitol and betaine by 43.2+/-9 .5% and 32.2+/-7.8% (n = 5), respectively. Efflux of GPC, myo-inositol and taurine was not significantly altered. Mastoparan (10 mu M) incre ased osmolyte release under isotonic conditions such that release of b etaine was increased 3.8-fold and that of sorbitol 2.1-fold, while GPC , myo-inositol and taurine effluxes were only slightly augmented. Unde r hypotonic conditions, mastoparan stimulated betaine release (1.86+/- 0.2-fold, n = 5) but not that of sorbitol. As tested in connection wit h sorbitol and betaine release, the effect of mastoparan was abolished by PTX, but not the A23187-evoked sorbitol release. Like mastoparan, arachidonic acid increased the release of sorbitol and betaine under i sotonic conditions, but under hypotonic conditions it only increased t he release of betaine. As to the role of intracellular Ca2+, hypotonic shock evoked an intracellular Ca2+ peak which could be prevented by P TX. Mastoparan increased intracellular Ca2+ under isotonic conditions, whether the extracellular Ca2+ concentration was low or high. The res ults indicate that G-proteins are involved in regulating sorbitol and betaine efflux from IMCD cells. The G-proteins regulating sorbitol rel ease are probably involved in generating the proper intracellular Ca2 signal. Betaine efflux, which is independent of intracellular Ca-2+, might be regulated by a G-protein stimulated release of arachidonic ac id. Thus, probably several G-proteins are involved in controlling orga nic osmolyte efflux from IMCD cells.