title: Modulation of cardiac Na+ channels expressed in a mammalian cell line and in ventricular myocytes by protein kinase C.
abstract: Cardiac rH1 Na+ channel alpha subunits were expressed in cells of the Chinese hamster lung 1610 cell line by transfection, and a stable cell line expressing cardiac Na+ channels (SNa-rH1) was isolated. Mean Na+ currents of 2.2 +/- 1.0 nA were recorded, which corresponds to a cell surface density of approximately 1-2 channels active at the peak of the Na+ current per micron2. The expressed cardiac Na+ current was tetrodotoxin resistant (Kd = 1.8 microM) and had voltage-dependent properties similar to those of the Na+ current in neonatal ventricular myocytes. Activation of protein kinase C by 1-oleoyl-2-acetyl-sn-glycerol (OAG) (10 microM) decreased this current approximately 33% at a holding potential of -114 mV and 56% at -94 mV. This reduction in peak current was caused in part by an 8- to 14-mV shift of steady-state inactivation in the hyperpolarized direction. Na+ channel activation was unchanged. Effects of OAG in SNa-rH1 cells and in neonatal rat cardiac myocytes were similar, except that the time course of inactivation was slowed either transiently or persistently when protein kinase C was activated in myocytes bathed in low-Ca2+ (1 microM) or Ca(2+)-free solution but was unaffected in SNa-rH1 cells. The effects of OAG on cardiac Na+ current were blocked in cells that had been previously microinjected with a peptide inhibitor of protein kinase C but not with a peptide inhibitor of cAMP-dependent protein kinase, indicating that protein kinase C is responsible for the effect of OAG. Single-channel recordings from SNa-rH1 cells showed that the probability of channel opening was reduced by OAG, but the conductance was unaffected. OAG did not induce the late Na+ channel openings observed with PKC modulation of neuronal and skeletal muscle Na+ channels. Thus, the substantial reduction in Na+ current at normal diastolic depolarizations with 10 microM OAG is due to failure of channel opening in response to depolarization. Such Na+ current reductions may have profound effects on cardiac cell excitability.
authors.affiliation: Department of Pharmacology, University of Washington, Seattle 98195.
authors.forename: Y
authors.initials: Y
authors.lastname: Qu
authors.forename: J
authors.initials: J
authors.lastname: Rogers
authors.forename: T
authors.initials: T
authors.lastname: Tanada
authors.forename: T
authors.initials: T
authors.lastname: Scheuer
authors.forename: W A
authors.initials: WA
authors.lastname: Catterall
mesh_terms: D000200:Action Potentials; D000818:Animals; D002460:Cell Line; D004075:Diglycerides / Q000494:pharmacology; D004789:Enzyme Activation; D066298:In Vitro Techniques; D008297:Male; D008564:Membrane Potentials; D009206:Myocardium / Q000378:metabolism*; D011493:Protein Kinase C / Q000502:physiology*; D051381:Rats; D012964:Sodium / Q000502:physiology; D015222:Sodium Channels / Q000502:physiology*
chemical_list: D004075:Diglycerides; D015222:Sodium Channels; C038237:1-oleoyl-2-acetylglycerol; D012964:Sodium; D011493:Protein Kinase C