Membrane depolarization-mediated survival of sympathetic neurons occurs through both phosphatidylinositol 3-kinase- and CaM kinase II-dependent pathways

It has been well established that the NGF-mediated survival of sympathetic neurons in culture occurs through the phosphatidylinositol (PI) 3-kinase/Ak t-dependent pathway. In contrast, the mechanism by which membrane depolariz ation promotes neuronal survival independently of NGF remains unresolved. H ere we show that LY294002, a specific inhibitor of PI 3-kinase, induced cel l death of sympathetic neurons under depolarizing conditions with elevated K- (IC50=similar to 30 mu M) Interestingly, lower concentrations of this ag ent (less than or equal to 10 mu M) were sufficient to suppress Akt phospho rylation at Ser-473, a putative downstream target of PI 3-kinase, under the se conditions. We also show that KN-62, a specific inhibitor of Ca2+/calmod ulin-dependent protein kinase II (CaMKII) suppressed depolarization-mediate d survival in a does-dependent manner (IC50=similar to 2 mu M) that paralle led attenuation of sustained levels of intracellular Ca2+ evoked by depolar ization. This IC50 value is greater than that for CaMKII (similar to 0.8 mu M). These findings led us to hypothesize that depolarization-mediated surv ival occurs through both the PI 3-kinase/Akt and the CaMKII pathways. Indee d, combined treatment with LY294002 (25 mu M) and KN-62 (0.5 mu M) dramatic ally abolished depolarization-mediated survival, whereas each alone did not significantly attenuate it. Under these conditions, KN-62 neither impaired sustained levels of intracellular Ca2+, nor inhibited the phosphorylation of Akt. It is thus likely that PI 3-kinase and CaMKII independently promote the membrane depolarization-mediated survival of sympathetic neurons in cu lture. (C) 2000 Elsevier Science B.V. All rights reserved.