Protein kinase C modulation of recombinant ATP-sensitive K+ channels composed of Kir6.1 and/or Kir6.2 expressed with SUR2B

The molecular identity of smooth muscle ATP-sensitive K⁺ channels (K_ATP) is not established with certainty. Patch clamp methods were employed to determine if recombinant K_ATP channels composed of Kir6.1 and SUR2B subunits expressed by human embryonic kidney (HEK293) cells share an identical modulation by protein kinase C (PKC) with the vascular K_NDP subtype of K_ATP channel. The open probability of Kir6.1/SUR2B channels was determined before and after sequential exposure to pinacidil (50 μM) and the combination of pinacidil and phorbol 12,13-dibutyrate (PdBu; 50 n m ). Treatment with PdBu caused a decline in channel activity, but this was not seen with an inactive phorbol ester, 4α-phorbol 12,13-didecanoate (PdDe; 50 n m ). Angiotensin II (0.1 μM) induced a similar inhibition of Kir6.1/SUR2B channels in cells expressing angiotensin AT₁ receptors. The effects of PdBu and angiotensin II were blocked by the PKC inhibitor, chelerythrine (3 μM). Purified PKC inhibited Kir6.1/SUR2B activity (in 0.5 m m ATP/ 0.5 m m ADP), and the inhibition was blocked by a specific peptide inhibitor of PKC, PKC(19-31). In contrast, PdBu increased the activity of recombinant K_ATP channels composed of Kir6.2 and SUR2B, or the combination of Kir6.1, Kir6.2 and SUR2B subunits. The results indicate that the modulation by PKC of Kir6.1/SUR2B, but not Kir6.2/SUR2B or Kir6.1-Kir6.2/SUR2B channel gating mimics that of native vascular K_NDP channels. Physiological inhibition of vascular K_ATP current by vasoconstrictors which utilize intracellular signalling cascades involving PKC is concluded to involve the modulation of K_NDP channel complexes composed of four Kir6.1 and their associated SUR2B subunits.