Abstract
Sympathetic neurons deprived of nerve growth factor (NGF) die by apoptosis. Chronic depolarization with elevated concentrations of extracellular potassium ([K+](E)) supports long-term survival of these and other types of neurons in culture. While depolarization has long been used to support neuronal cultures, little is known about the mechanism. We explored how chronic depolarization of NGF-deprived mouse sympathetic neurons in culture blocks apoptotic death. First, we determined the effects of elevated [K+](E) on proapoptotic BH3-only proteins reported to be upregulated in sympathetic neurons after NGF withdrawal. Upregulation of BIMEL was blocked by depolarization while upregulation of PUMA was not. BMF levels did not increase after NGF withdrawal, and elevated [K+](E) had no effect on its expression. dp5/HRK was not detectable. A large increase in production of mitochondria-derived reactive species (RS), including reactive oxygen species (ROS), occurs in NGF-deprived sympathetic neurons. Suppressing these RS prevents cytochrome c release from mitochondria and apoptosis. The addition of high [K+](E) to cultures rapidly blocked increased RS and cytochrome c release. Elevated [K+](E) caused an increase of the cellular antioxidant glutathione (GSH). Preventing this increase prevented the elevated [K+](E) from blocking cytochrome c release and death. While suppression of BIMEL upregulation by elevated [K+](E) may contribute to high [K+](E) pro-survival effects, we conclude that elevated [K+](E) prevents apoptotic death of NGF-deprived sympathetic neurons primarily via an antioxidant mechanism.