Pharmacological identification of a novel Ca 2+ channel in chicken brain synaptosomes

Abstract

Ca 2+ influx was measured in rat and chicken brain synaptosomes in the presence of a number of pharmacological tools which have recently been used to define voltage-sensitive Ca 2+-channel (VSCC) types. In chicken brain synaptosomes, VSCCs which, because of their sensitivity to inhibition by omega-conotoxin (ω-CgTx), are thought to be exclusively N-type, the P-type VSCC polyamine inhibitor FTX (from Agelenopsis operta venom; 1 μl/ml), its synthetic analogue, sFTX (1–5 mM) and the polypeptides AgaIVA (IC 50 0.29 μM) and ω-CgTx MVIIC (IC 50 0.0022 μM) inhibited 70–100% of the measurable K + stimulated Ca 2+ influx. The prototypical N-channel VSCC inhibitor ω-CgTx GVIA (IC 50 0.014 μM), Cd 2+ (50 μM) and diluted venom from Hololena curta (1:2,500) also caused complete or almost complete, inhibition of Ca 2+ influx. In comparable studies using rat brain synaptosomes, sFTX (1–10 mM) caused a dose-dependent reduction of Ca 2+ influx, while FTX (1 μl/ml) and AgaIVA (IC 50 0.02 μM) completely inhibited Ca 2+ influx. Similar to the findings in chicken synaptosomes, Cd 2+ (50 μM) and H. curta (1:2,500 dilution) both inhibited K + stimulated influx by > 80% whereas ω-CgTx (1 μM) only caused a maximum 25% inhibition. Both sFTX and its congener spermine, inibited [ 125I]ω-CgTx binding to rat and chicken synaptosomal membranes. These results strongly implicate P-type channels as the major VSCC in rat brain. The results also clearly demonstrate a herefore unrecognized, novel, FTX/AgaIVA/ω-CgTx GVIA/ω-CgTx MVIIC-sensitive VSCC in chicken brain.