AJP - Cell Physiology, Vol 264, Issue 6 C1473-C1479, Copyright © 1993 by American Physiological Society

ARTICLES

Regulation of cardiac L-type Ca channels in planar lipid bilayers by G proteins and protein phosphorylation

Y. Wang, C. Townsend and R. L. Rosenberg
Department of Pharmacology, University of North Carolina, Chapel Hill 27599.

We have studied the effects of activated G proteins (Gs alpha and Gi1 alpha), adenosine 3',5'-cyclic monophosphate-dependent protein kinase (PKA), and okadaic acid on L-type Ca channels incorporated from porcine ventricular sarcolemma into planar lipid bilayers. Channel activity evoked by membrane depolarizations diminished to extremely low levels within 2 min of incorporation (channel "rundown"). When Gs alpha [activated with guanosine 5'-O-(3-thiotriphosphate)] was present in the intracellular chamber, the initial level of channel activity was increased and rundown was delayed, so that channel activity was sustained for longer times after incorporation. The effect was specific for activated Gs alpha; activated Gi1 alpha, heat-denatured, activated Gs alpha, and unactivated Gs alpha did not augment channel activity. Activated Gi1 alpha inhibited the stimulation of Ca channel activity by Gs alpha. Treatment of the sarcolemmal membranes with PKA and Mg-ATP also increased the initial channel open probability and delayed their rundown. Addition of intracellular Gs alpha to PKA-treated channels increased the initial level of activity above that seen with PKA or Gs alpha alone, suggesting different nonocclusive pathways for the channel stimulation. This was also supported by the observation that activated Gi1 alpha had no effect on PKA-treated channels. Okadaic acid (100 nM) increased the level of Ca channel activity, suggesting that dephosphorylation by endogenous phosphatases participated in the downregulation of the channels in cell-free membranes.(ABSTRACT TRUNCATED AT 250 WORDS)

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