Role of Hyperpolarization Attained by Linoleic Acid in Chick Myoblast Fusion

Abstract

Our previous report has suggested that hyperpolarization generated by reciprocal activation of calcium-activated potassium (KCa) channels and stretch-activated channels induces calcium influx that triggers myoblast fusion. Here we show that linoleic acid is involved in the process of generating hyperpolarization in cultured chick myoblasts and hence in promotion of the cell fusion. Linoleic acid dramatically hyperpolarized the membrane potential from −14 ± 3 to −58 ± 5 mV within 10 min. This effect was partially blocked by 1 mM tetraethylammonium (TEA) or 30 nM charybdotoxin, a selective KCa channel inhibitor, and completely abolished by 10 mM TEA. Single-channel recordings revealed that linoleic acid activates TEA-resistant potassium channels as well as KCa channels. Furthermore, linoleic acid induced calcium influx from extracellular solution, and this effect was partially blocked by 1 mM TEA and completely prevented at 10 mM, similar to the effect of TEA on linoleic acid-mediated hyperpolarization. Since the valinomycin-mediated hyperpolarization promoted calcium influx, hyperpolarization itself appears capable of inducing calcium influx. In addition, gadolinium prevented the valinomycin-mediated increase in intracellular calcium level under hypotonic conditions, revealing the involvement of stretch-activated channels in calcium influx. Furthermore, linoleic acid stimulated myoblast fusion, and this stimulatory effect could completely be prevented by 10 mM TEA. These results suggest that linoleic acid induces hyperpolarization of membrane potential by activation of potassium channels, which induces calcium influx through stretch-activated channels, and thereby triggers myoblast fusion.