In skeletal muscle myogenesis, precursor cells or myoblasts fuse to form multinucleated cells (myotubes), which then further develop into functional muscle. We investigated if the inhibition of myogenesis by transforming growth factor beta-1 (TGF-1) and bone morphogenetic protein-2 (BMP-2) involve regulation of voltage-dependent Ca²⁺ channels. Primary cultured myoblasts were kept in fusion medium (0-6 days) in either the absence (control conditions) or the presence of 40 pM TGF-1 or 5 nM BMP-2. Subsequently, the developing myotubes were transferred to a growth factor-free recording solution, and subjected to whole cell patch-clamp experiments. At day 0, 14% of non-fusing myoblasts exhibited T-current, whereas the L-current was practically absent. Under control conditions however, the percentage of T- and L-channel-expressing myotubes increased sharply, from 25% at day 1 to ~100% at days 2-6. In addition, parallel increases were determined for Ca²⁺-currents density and cell membrane capacitance or Cm, which is proportional to the size of myotubes. Interestingly, TGF-1 and BMP-2 eliminated the initial 14% of T-channel-expressing myoblasts. Moreover, the growth factors significantly reduced the maximal values of both T-current density (80%) and Cm (60%). The effect of BMP-2 was selective on T-channels, whereas TGF-1 decreased also the L-current density (90%). A similar reduction in Ca²⁺-channels maximal conductance was determined, in the absence of significant alterations in other essential properties of the channels, including the time course and voltage-dependence of activation and inactivation. The results suggest these growth factors markedly reduce the number of functional T- (both TGF-1 and BMP-2) and L-channels (only TGF-1) in the surface of the plasma membrane, and contribute to explain the associated effects on myogenesis.