PHOSPHORYLATION BY PROTEIN-KINASE-A ENHANCES DELAYED RECTIFIER K-MUSCLE CELLS( CURRENT IN RABBIT VASCULAR SMOOTH)

The effect of adenosine 3',5'-cyclic monophosphate-dependent protein k inase (PKA) activity on 4-aminopyridine (4-AP)-sensitive delayed recti fier current (I-dK) in isolated rabbit portal vein smooth muscle cells was studied via whole cell voltage clamp (20-22 degrees C). A threefo ld increase in 4-AP-sensitive (5 mM) I-dK was recorded after gaining c ell access during dialysis with 5 mM intracellular ATP and internal Ca 2+ buffered to a low level with 5 mM ethylene glycol-bis(beta-aminoeth yl ether)-N,N,N',N'-tetraacetic acid. Dialysis with the nonhydrolyzabl e ATP analogue 5'-adenylylimidodiphosphate (5 mM) or the specific pept ide inhibitor of PKA (PKI; 10 mu M) reduced current runup by 50 and 70 %, respectively. Delayed dialysis with PKI reversed runup and inhibite d I-dK to below initial levels. Forskolin (1 mu M) caused a reversible increase in I-dK, which was inhibited by 4-AP (5 mM). Isoproterenol ( 1 mu M) reversibly enhanced I-dK; the increase was sensitive to propra nolol (2 mu M) and 4-AP (5 mM) and was prevented by dialysis with PKI (10 mu M) I-dK was enhanced over the entire voltage range of activatio n and associated with a negative shift in reversal potential of net wh ole cell current, consistent with hyperpolarization of resting membran e potential. The data provide the first evidence for a signal transduc tion mechanism involving beta-adrenoceptors, adenylate cyclase, and a phosphotransferase reaction mediated by PKA for the regulation of dela yed rectifier K+ channels in vascular smooth muscle.