Effect of anoxic preconditioning on ATP-sensitive potassium channels in guinea-pig ventricular myocytes

Ischemic or hypoxic preconditioning in experimental animals and humans is d escribed. The mechanism of preconditioning may involve several endogenous s ubstances released from ischemic or hypoxic tissues (such as adenosine, nor adrenaline and bradykinin) that stimulate protein kinase C (PKC), which the n phosphorylates ATP-sensitive potassium channels (K-ATP, channels). Howeve r, the effect of hypoxic preconditioning, on K-ATP channels in guinea-pig v entricular myocytes is unclear. The uncoupler carbonyl cyanide p-(trifluoro methoxy)phenylhydrazone (FCCP) has been shown to activate KATP channels in isolated cardiac cells. In the present study we rested whether anoxic preco nditioning (APC) could affect the opening of K-ATP channels activated by me tabolic inhibition (MI) induced by FCCP in cell-attached and inside-out pat ches from guinea-pig ventricular myocytes. We measured the channel activity as NP,I and calculated it using the formula P-o=I/(Ni), where P-o is open- state probability, I is the mean patch current carried by all K-ATP, channe ls activated in a particular patch for a certain period of time, N is the n umber of functioning channels in the patch, and i is the unitary current of the KATP channels. In cell-attached membrane patches, after about 5 min of initiating MI, KATP channels were activated at a holding potential of +40 mV (NP(o)i=3.7+/-0.9 pA); APC pretreatment (3 min of anoxia followed by 7 m in of reoxygenation) before MI (APC+MI group) shortened the time to activat e KATP channels by MI (2.3+/-0.5 min) and increased the activity of KATP cu rrents (NP(o)i=8.4+/-0.5 pA). This effect of APC was eliminated by administ ration of a PKC blocker, chelerythrine (5 mu M), for 5 min before the APC p retreatment. In the inside-out patches, the IC,, of intracellular ATP again st the K-ATP channels in the APC+MI group was significantly increased to 64 2 mu M compared to that in the MI group (IC50 of intracellular ATP=252 mu M . Chelerythrine inhibited the effect of APC on the sensitivity of K-ATP cha nnels to the intracellular ATP concentration (IC50 of [ATP](i)=301 mu M). O ur results demonstrate that APC can increase and accelerate the opening of K-ATP channels induced by MI, and decrease the sensitivity of K-ATP channel s to [ATP],, which is mediated by promoting the activation of PKC induced b y APC.