Heat shock proteins (HSPs) are a conserved family of cytoprotective polypeptides, synthesized by cells in response to stress. Hsp70 and hemoxygenase 1 (Hmox-1) are induced by a variety of cellular stressors in skeletal muscle, playing a role in long term adaptations and muscle fibers regeneration. Though HSPs expression after exercise has been intensely investigated, the molecular mechanisms concerning Hsp70 and Hmox-1 induction are poorly understood. The aim of this work was to investigate the involvement of calcium in Hsp70 and Hmox-1 expression upon depolarization of skeletal muscle cells. We observed that depolarization of myotubes increased both mRNA levels and protein expression for Hsp70 and Hmox-1. Stimulation in the presence of intracellular calcium chelator BAPTA-AM resulted in a complete inhibition of Hsp70 induced expression. It is known that inositol-1,4,5-triphophate (IP₃) mediated slow Ca²⁺ transients, evoked by membrane depolarization, are involved in the regulation of gene expression. Here we demonstrated that inhibition of IP₃-dependent calcium signals decreased both Hsp70 mRNA induction and Hsp70 and Hmox-1 protein expression. Inhibitors of calcium dependent PKCs also abolished Hsp70 mRNA induction. Our results provide evidence that membrane depolarization increases Hsp70 and Hmox-1 expression in cultured skeletal muscle cells, that the effect is critically dependent on Ca²⁺ released from IP₃ sensitive intracellular stores and that it involves PKC as an upstream effector in Hsp70 mRNA induced expression.