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

Abstract

We compared the effects of cytosolic free magnesium (Mg[Formula: see text]) on L-type Ca2+ current ( I Ca,L) in patch-clamped guinea pig ventricular cardiomyocytes under basal conditions, after inhibition of protein phosphorylation, and after stimulation of cAMP-mediated phosphorylation. Basal I Ca,L density displayed a bimodal dependence on the concentration of Mg[Formula: see text]([Mg2+]i; 10−6–10−2 M), which changed significantly as cell dialysis progressed due to a pronounced and long-lasting rundown of I Ca,L in low-Mg2+ dialysates. Ten minutes after patch breakthrough, I Ca,L density (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 I Ca,L density and abolished these effects of Mg[Formula: see text] except for a negative shift of I Ca,L-voltage relations with increasing [Mg2+]i. Maximal stimulation of cAMP-mediated phosphorylation occluded the Mg[Formula: see text]-induced stimulation of I Ca,L and prevented inhibitory effects of the ion at [Mg2+]i <1 mM but not at higher concentrations. These results show that the modulation of I Ca,L by 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 I Ca,L seem 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.