Effect of voltage-dependent calcium channel blockers on ethanol-induced β-endorphin release from hypothalamic neurons in primary cultures

The voltage-dependent calcium channel (VDCC) has been shown to mediate calcium entry into neurons that regulates neurotransmission in many neuronal cells. Four major types of VDCCs (three high-voltage-activated L-, N-, and P- types and one low-voltage-activated T-type) have been identified in neurons. Involvement of the VDCC in ethanol-stimulated β-endorphin (β-EP) release from hypothalamic neurons has not been studied. In the present study, the role of VDCC on basal and ethanol-induced β-EP release was determined by using rat fetal hypothalamic cells in primary cultures. Treatments with a 50 mM dose of ethanol for 3 hr increased immunoreactive β-EP (IR-β-EP) release from hypothalamic cells maintained in cultures for 9 days. Ethanol-induced IR-β-EP release was inhibited by a P/Q-type channel blocker ω-agatoxin TK (0.1-1 μM), an N-type channel blocker ω-conotoxin (0.1-1 μM), an L-type blocker nifedipine (1-10 μM), and a T-type blocker flunarizine (1-10 μM). The minimal effective doses of these blockers that blocked the ethanol response produced no significant effects on basal release of IR-β-EP; neither did these doses of the blockers produce any significant effects on cell viability. These results suggest that ethanol-stimulated IR-β-EP release is regulated by extracellular calcium involving P-, N-, L- and T- type channels.