MECHANISM AND REGULATION OF SWELLING-ACTIVATED INOSITOL EFFLUX IN BRAIN GLIAL-CELLS

Rat C6 glioma cells chronically acclimated to hypertonic media accumul ate large quantities of inositol. When returned to isotonic conditions , the cells swell and lose inositol slowly via a four- to fivefold inc rease in the rate of passive inositol efflux. The inositol efflux path way is a Na+-independent transport mechanism with low affinity for ino sitol and is inhibited by quinidine, quinine, various anion transport blockers, and cis-unsaturated fatty acids. Ionomycin-induced elevation of intracellular Ca2+ (Ca(i)2+) had no effect on basal or swelling-in duced inositol efflux. Inositol efflux was not inhibited by chelation of Ca(i)2+ with 1,2-bis(2-aminophenoxy)-N,N',N'-tetraacetic acid. In a ddition, Ca(i)2+ measured with fura 2 did not change during cell swell ing, indicating that increases in Ca(i)2+ do not regulate inositol eff lux. Exposure of C6 cells to 20 nM phorbol 12-myristate 13-acetate, 0. 5 mM adenosine 3',5'-cyclic monophosphate (cAMP), or 50 muM forskolin had no effect on basal inositol efflux but stimulated swelling-induced inositol loss by 2.6-, 2.2-, and 3.4-fold respectively. Exposure to t he protein kinase inhibitors 1-(5-isoquinolinylsulfonyl)-2-methylpiper azine or staurosporine or downregulation of protein kinase C (PKC) act ivity, however, had no inhibitory effect on inositol efflux, and cellu lar cAMP levels were not altered by cell swelling. Taken together, the se results indicate that stimulation of PKC and protein kinase A modul ates the activity of the efflux pathway but is not required for swelli ng-induced activation. Ketoconazole, cinnamyl-3,4-dihydroxy-alpha-cyan ocinnamate, and gossypol, inhibitors of lipoxygenase enzymes, blocked both basal and swelling-induced inositol efflux, suggesting indirectly that lipoxygenase metabolites may be responsible for swelling-induced activation of the efflux mechanism. The characterisitics of inositol efflux in C6 cells are similar to those described for volume regulator y sorbitol and taurine efflux in a number of cell types, suggesting th e existence of a common transport mechanism.