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/Akt-dependent pathway. In contrast, the mechanism by which membrane depolarization promotes neuronal survival independently of NGF remains unresolved. Here we show that LY294002, a specific inhibitor of PI 3-kinase, induced cell death of sympathetic neurons under depolarizing conditions with elevated K(+) (IC(50)= approximately 30 microM). Interestingly, lower concentrations of this agent (< or =10 microM) were sufficient to suppress Akt phosphorylation at Ser-473, a putative downstream target of PI 3-kinase, under these conditions. We also show that KN-62, a specific inhibitor of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) suppressed depolarization-mediated survival in a does-dependent manner (IC(50)= approximately 2 microM) that paralleled attenuation of sustained levels of intracellular Ca(2+) evoked by depolarization. This IC(50) value is greater than that for CaMKII ( approximately 0.8 microM). These findings led us to hypothesize that depolarization-mediated survival occurs through both the PI 3-kinase/Akt and the CaMKII pathways. Indeed, combined treatment with LY294002 (25 microM) and KN-62 (0.5 microM) dramatically abolished depolarization-mediated survival, whereas each alone did not significantly attenuate it. Under these conditions, KN-62 neither impaired sustained levels of intracellular Ca(2+), 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 culture.
Ikegami
K
K
Molecular Neurobiology Laboratory, Division of Biological Sciences, Graduate School of Science, Hokkaido University, North Ward N10 W8, 060-0810, Sapporo, Japan.
Koike
T
T
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects* / Q000502:physiology*; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005728:Ganglia, Sympathetic / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects* / Q000502:physiology*; D009025:Morpholines / Q000494:pharmacology; D020932:Nerve Growth Factor / Q000187:drug effects / Q000378:metabolism / Q000494:pharmacology; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000187:drug effects / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D051381:Rats
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D002118:Calcium; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002867:Chromones; D004791:Enzyme Inhibitors; D009025:Morpholines; D020932:Nerve Growth Factor; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C063302:KN 62