Cytoplasmic [Ca²⁺] ([Ca²⁺]_i ) variation is a key event in myoblast differentiation but the mechanism by which it occurs is still debated. Here we show that increases of extracellular Ca²⁺ concentration ([Ca²⁺]_o ) produced membrane hyperpolarization and a concentration-dependent increases of [Ca²⁺]_i due to Ca²⁺ influx across the plasma membrane. Responses were not related to inositol-phosphate turnover and calcium sensing receptor. [Ca²⁺]_o induced [Ca²⁺]_i increase was inhibited by Ca²⁺ channel inhibitors and appeared to be modulated by several kinase activities. [Ca²⁺]_i increase was potentiated by depletion of intracellular Ca²⁺ stores and depressed by inactivation of the Na⁺/Ca²⁺ exchanger. The response to vasopressin (AVP), which induces inositol-1,4,5-trisphosphate-dependent [Ca²⁺]_i increase in L6-C5 cells, was not modified by high [Ca²⁺]_o. On the contrary, AVP potentiated [Ca²⁺]_i increase in the presence of elevated [Ca²⁺]_o. Other clones of the L6 line as well as the rhabdomyosarcoma RD cell line and the satellite cell-derived C2-C12 line, expressed similar responses to high [Ca²⁺]_o and the amplitude of the responses was correlated with the myogenic potential of the cells.