Phosphorylation-dependent modulation of cardiac calcium current by intracellular free magnesium

We compared the effects of cytosolic free magnesium (Mg[Formula: see text]) on L-type Ca2+current ( ICa,L) in patch-clamped guinea pig ventricular cardiomyocytes under basal conditions, after inhibition of protein phosphorylation, and after stimulation of cAMP-mediated phosphorylation. Basal ICa,Ldensity displayed a bimodal dependence on the concentration of Mg[Formula: see text]([Mg2+]i; 10−6–10−2M), which changed significantly as cell dialysis progressed due to a pronounced and long-lasting rundown of ICa,Lin low-Mg2+dialysates. Ten minutes after patch breakthrough, ICa,Ldensity (at +10 mV) in Mg[Formula: see text]-depleted cells ([Mg2+]i∼1 μM) was elevated, increased to a maximum at ∼20 μM [Mg2+]i, and declined steeply at higher [Mg2+]i. Treatment with the broad-spectrum protein kinase inhibitor K252a (10 μM) reduced ICa,Ldensity and abolished these effects of Mg[Formula: see text] except for a negative shift of ICa,L-voltage relations with increasing [Mg2+]i. Maximal stimulation of cAMP-mediated phosphorylation occluded the Mg[Formula: see text]-induced stimulation of ICa,Land prevented inhibitory effects of the ion at [Mg2+]i<1 mM but not at higher concentrations. These results show that the modulation of ICa,Lby Mg[Formula: see text] requires protein kinase activity and likely originates from interactions of the ion with proteins involved in the regulation of protein phosphorylation/dephosphorylation. Stimulatory effects of Mg[Formula: see text] on ICa,Lseem to increase the cAMP-mediated phosphorylation of Ca2+channels, whereas inhibitory effects of Mg[Formula: see text] appear to curtail and/or reverse cAMP-mediated phosphorylation.