Sodium-dependent action potentials induced by brevetoxin-3 trigger both IP3 increase and intracellular Ca2+ release in rat skeletal myotubes

Summary Brevetoxin-3 (PbTx-3), described to increase the open probability of voltage-dependent sodium channels, caused trains of action potentials and fast oscillatory changes in fluorescence intensity of fluo-3-loaded rat skeletal muscle cells in primary culture, indicating that the toxin increased intracellular Ca 2+ levels. PbTx-3 did not elicit calcium transients in dysgenic myotubes (GLT cell line), lacking the α1 subunit of the dihydropyridine receptor (DHPR), but after transfection of the α1DHPR cDNA to GLT cells, PbTx-3 induced slow calcium transients that were similar to those of normal cells. Ca 2+ signals evoked by PbTx-3 were inhibited by blocking either IP 3 receptors, with 2-aminoethoxydiphenyl borate, or phospholipase C with U73122. PbTx-3 caused a tetrodotoxin-sensitive increase in intracellular IP 3 mass levels, dependent on extra-cellular Na + . A similar increase in IP 3 mass was induced by high K + depolarization but no action potential trains (nor calcium signals) were elicited by prolonged depolarization under current clamp conditions. The increase in IP 3 mass induced by either PbTx-3 or K + was also detected in Ca 2+ -free medium. These results establish that the effect of the toxin on both intracellular Ca 2+ and IP 3 levels occurs via a membrane potential sensor instead of directly by Na + flux and supports the notion of a train of action potentials being more efficient as a stimulus than sustained depolarization, suggesting that tetanus is the physiological stimulus for the IP 3 -dependent calcium signal involved in regulation of gene expression.