Dopamine release in PC12 cells is mediated by Ca2+-dependent production of ceramide via sphingomyelin pathway

Abstract

A presynaptic membrane disturbance is an essential process for the release of various neurotransmitters. Ceramide, which is a tumor suppressive lipid, has been shown to act as a channel-forming molecule and serve as a precursor of ceramide-1-phosphate, which can disturb the cellular membrane. This study found that while permeable ceramide increases the rate of dopamine release in the presence of a Ca2+-ionophore, A23187, permeable ceramide-1-phosphate provoked its release even without the ionophore. The treatment of PC12 cells with the ionophore at concentrations < 2 mu m produced ceramide via the sphingomyelin (SM) pathway with a concomitant release of dopamine, and no cell damage was observed. The addition of a Ca2+ chelator, EGTA, to the medium inhibited the increase in the release of both the ceramide and dopamine. This suggests that ceramide might be produced by Ca2+ and is implicated in the membrane disturbance associated with the release of dopamine as a result of its conversion to ceramide-1-phosphate. Consistent with these results, this study detected a membrane-associated and neutral pH optimum sphingomyelinase (SMase) whose activity was increased by Ca2+. Together, these results demonstrate that ceramide can be produced via the activation of a neutral form of SMase through Ca2+, and is involved in the dopamine release in concert with Ca2+.