THE DIHYDROPYRAZOLE RH-5529 BLOCKS VOLTAGE-SENSITIVE CALCIUM CHANNELS IN MAMMALIAN SYNAPTOSOMES

The effect of the dihydropyrazole RH-5529 on depolarization-induced rise in free [Ca2+] has been characterized in synaptosomes prepared from mouse central nervous system. At high concentrations RH-5529 had no effect on the concentration of free [Ca2+] in resting synaptosomes. RH-5529 inhibited the rise in synaptosomal [Ca2+] elicited by the sodium channel activator veratridine with an IC50 of approximately 3 μM. Tetrodotoxin completely suppressed the rise in intraterminal [Ca2+] produced by veratridine. Inhibitory effects of tetrodotoxin on potassium-stimulated Ca2+ accumulation in this preparation were not detected. In contrast, RH-5529 in the low micromolar range was effective in blocking the rise in intrasynaptosomal [Ca2+] induced by elevated potassium (IC50 approximately 3.5 μM. Moreover, when synaptosomes were exposed to RH-5529 and tetrodotoxin the dihydropyrazole was still capable of fully inhibiting the potassium-stimulated rise in intraterminal [Ca2+]. When synaptosomes were suspended in saline to which no calcium was added, elevated potassium failed to raise intraterminal [Ca2+] significantly. The inhibition by RH-5529 of the rise in the concentration of Ca2+ induced by veratridine is consistent with its known effects on sodium channel blockade. However, since we demonstrate that RH-5529 suppresses a potassium-induced rise in synaptosomal [Ca2+], and also does so in the presence of tetrodotoxin, our results provide strong evidence that RH-5529 is directly inhibiting Ca2+ entry into the nerve ending through voltage-sensitive calcium channels. This investigation therefore provides evidence for a new mechanism by which RH-5529 may interact with the nervous system. © 1993 Academic Press.