The effects of changing the intracellular concentrations of either free Mg2+ ions ([Mg2+]i) or Mg(2+)-bound adenosine triphosphate ([Mg.ATP]i) on Ca2+ channel currents were assessed in cultured rat cerebellar granule neurones using the whole-cell patch-clamp technique. Raising [Mg2+]i from 0.06 mM to 1.0 mM inhibited Ca2+ channel currents by approximately 50%. The action of omega-conotoxin GVIA (omega-CgTX), a selective inhibitor of "N"-type Ca2+ channels was also investigated. With increasing [Mg2+]i, the proportion of current irreversibly blocked by omega-CgTX was reduced, and was negligible (approximately 5 pA of current) in the presence of [Mg2+]i values of 0.5 mM or greater. Block of the omega-CgTX-sensitive current accounted for the reduction in total current by concentrations of [Mg2+]i to 0.5 mM. Raising [Mg2+]i had no effect on the rate of decay of Ca2+ currents, but did produce a negative shift in current activation, possibly due to a non-specific interaction with negative surface charge. Altering [Mg.ATP]i from 0.3 to 5.0 mM caused a twofold increase in the size of currents without affecting the proportion of current sensitive to omega-CgTX. [Mg2+]i was also effective in inhibiting the Ca2+ channel current following potentiation by increasing [Mg.ATP]i. These data suggest that omega-CgTX-sensitive current in these cells is selectively inhibited by internal Mg2+ whereas both omega-CgTX-sensitive and -resistant components of current are potentiated by internal Mg.ATP. The mechanism by which Mg2+ inhibits "N"-type channels is unclear, but may involve an open channel block.