CHOLERA AND PERTUSSIS TOXINS MODIFY REGULATION OF GLUCOSE-TRANSPORT ACTIVITY IN RAT ADIPOSE-CELLS - EVIDENCE FOR MEDIATION OF A CAMP-INDEPENDENT PROCESS BY G-PROTEINS

Adenylyl cyclase in rat adipose cells is stimulated by ligands for R(s) receptors (e.g. isoproterenol) and inhibited by ligands for R(i) receptors (e.g. adenosine). In contrast, R(s) receptors mediate inhibition and R(i) receptors mediate augmentation of insulin-stimulated glucose transport activity by a process independent of changes in cellular cAMP-dependent protein kinase activity [Kuroda M., Honnor R. C., Cushman S. W., Londos C. and Simpson I. A. (1987) J. biol. Chem. 262, 245-253]. The present study examines the possible role of G-proteins in the regulation of insulin-stimulated glucose transport activity by R(s) and R(i) receptors. First, conditions were established that permit intoxication of isolated rat adipocytes by cholera and pertussis toxins without compromising cell integrity. Effectiveness of toxin treatment was monitored by examining adenylyl cyclase activity in isolated plasma membranes. Secondly, neither toxin interfered with the ability of a maximal concentration insulin to initiate the glucose transport response. Thirdly, pertussis toxin eliminated the augmenting effects of adenosine on insulin-stimulated glucose transport activity, but enhanced the inhibitory effects of isoproterenol. Findings with ligands for other R(i) receptors (nicotinic acid and prostaglandin E2) mirrored those with adenosine. Finally, cholera toxin elicted a modest depression of transport activity, and only in the absence of an R(i) ligand (e.g. adenosine). Furthermore, in contrast to the enhanced stimulation of adenylyl cyclase by isoproterenol and GTP, cholera toxin eliminated the inhibitory effect of isoproterenol on transport activity. The augmentative effects of adenosine on transport activity were unchanged. Measurements of (-/+ cAMP) cAMP-dependent protein kinase activity ratios reinforce the notion that modulation of glucose transport activity is independent of changes in cAMP. We conclude that regulation of glucose transport activity by R(s) and R(i) receptors is mediated by the G-proteins, G(s) and G(i) (or other toxin substrates), respectively. Inasmuch as such regulation occurs at the plasma membrane and appears to be cAMP-independent, it is suggested that glucose transporters may be direct targets for receptor: G-protein interactions.