Differential alteration of catecholamine release during chemical hypoxia is correlated with cell toxicity and is blocked by protein kinase C inhibitors in PC12 cells

Release of neurotransmitters, including dopamine and glutamate, has been im plicated in hypoxia/ischemia-induced alterations in neuronal function and i n subsequent tissue damage. Although extensive studies have been done on th e mechanism underlying the changes in glutamate release, few have examined the mechanism that is responsible for the changes in catecholamines. Rat ph eochromocytoma-12 (PC12) cells synthesize, store, and release catecholamine s including DA and NE. Therefore, we used HPLC and ED to evaluate extracell ular DA and NE concentrations in a medium during chemical hypoxia in PC12 c ells. Chemical hypoxia produced by KCN induced differential release of DA a nd NE. Under normal glucose conditions, KCN induced release of NE, but not DA. Under glucose-free conditions, KCN-induced release of DA was elevated t ransiently, whereas the release of NE increased progressively. Under parall el conditions, KCN biphasically elevated the level of cytosolic free calciu m ([CA(2+)](i)) in glucose-free DMEM, peaking at 95 +/- 18 nM at 1,107 +/- 151 s, followed by a new plateau level at 249 +/- 24 nM sustained from 4,24 3 +/- 466 to 5,263 +/- 440 s. Cell toxicity, as measured by LDH release, wa s increased significantly by KCN in glucose-free DMEM but was diminished in the presence of glucose, and was correlated with DA release by chemical hy poxia. The protein kinase C (PKC) inhibitor GO6976 or staurosporine inhibit ed KCN-induced LDH release as well as the release of NE and DA. Taken toget her, selective activation of DA but not NE was correlated with the LDH rele ase by chemical hypoxia, and was diminished with CO6976 or staurosporine. T hese results suggest that selective activation of PKC isoforms is involved in the chemical hypoxia-induced DA release, which may lead to neuronal cell toxicity. (C) 2000 Wiley-Liss. Inc.