We examined the concentration-dependent blocking effects of intracellular Mg²⁺ on L-type Ca²⁺ channels in cardiac myocytes using the whole cell patch-clamp technique. The increase of L-type Ca²⁺ channel current (I_Ca) (due to relief of Mg²⁺ block) occurred in two temporal phases. The rapid phase (runup) transiently appeared early (<5 min) in dialysis of the low-Mg²⁺ solution; the slow phase began later in dialysis (>10 min). Runup was not blocked by intracellular GTP (GTP_i). The late phase of the I_Ca increase (late I_Ca) was suppressed by GTP_i (0.4 mM) and was observed in myocytes of the guinea pig or frog at higher (32 or 24°C, respectively) rather than lower temperatures (24 or 17.5°C, respectively). At pMg = 6.0, raising the temperature from 24 to 32°C evoked late I_Ca with a Q₁₀ of 14.5. Restoring the temperature to 24°C decreased I_Ca with a Q₁₀ of only 2.4. The marked difference in the Q₁₀ values indicated that late I_Ca (pMg = 5-6) is an irreversible phenomenon. Phosphorylation suppressed the intracellular [Mg²⁺] dependency of late I_Ca. This effect of phosphorylation together with the inhibitory action of GTP_i on Mg²⁺-dependent blocking of I_Ca are common properties of mammalian and amphibian cardiomyocytes.