Involvement of voltage‐dependent potassium channels in the EDHF‐mediated relaxation of rat hepatic artery
Authors: Zygmunt Peter M , Edwards Gillian , Weston Arthur H , Larsson Bengt , Högestätt Edward D 
Published: August 2006

In the rat hepatic artery, the acetylcholine‐induced relaxation mediated by endothelium‐derived hyperpolarizing factor (EDHF) is abolished by a combination of apamin and charybdotoxin, inhibitors of small (SKCa) and large (BKCa) conductance calcium‐sensitive potassium (K)‐channels, respectively, but not by each toxin alone. The selective BKCa inhibitor iberiotoxin cannot replace charybdotoxin in this combination. Since delayed rectifier K‐channels (KV) represent another target for charybdotoxin, we explored the possible involvement of KV in EDHF‐mediated relaxation in this artery.
The KV inhibitors, agitoxin‐2 (0.3 μM), kaliotoxin (0.3 μM), β‐dendrotoxin (0.3 μM), dofetilide (1 μM) and terikalant (10 μM), each in combination with apamin (0.3 μM) had no effect on the EDHF‐mediated relaxation induced by acetylcholine in the presence of Nω‐nitro‐L‐arginine (0.3 mM) and indomethacin (10 μM), inhibitors of nitric oxide (NO) synthase and cyclo‐oxygenase, respectively (n=2–3). Although the KV inhibitor margatoxin (0.3 μM) was also without effect (n=5), the combination of margatoxin and apamin produced a small inhibition of the response (pEC50 and Emax values were 7.5±0.0 and 95±1% in the absence and 7.0±0.1 and 81±6% in the presence of margatoxin plus apamin, respectively; n=6; P<0.05).
Ciclazindol (10 μM) partially inhibited the EDHF‐mediated relaxation by shifting the acetylcholine‐concentration‐response curve 12 fold to the right (n=6; P<0.05) and abolished the response when combined with apamin (0.3 μM; n=6). This combination did not inhibit acetylcholine‐induced relaxations mediated by endothelium‐derived NO (n=5).
A 4‐aminopyridine‐sensitive delayed rectifier current (IK(V)) was identified in freshly‐isolated single smooth muscle cells from rat hepatic artery. None of the cells displayed a rapidly‐activating and ‐inactivating A‐type current. Neither charybdotoxin (0.3 μM; n=3) nor ciclazindol (10 μM; n=5), alone or in combination with apamin (0.3 μM; n=4–5), had an effect on IK(V). A tenfold higher concentration of ciclazindol (0.1 mM, n=4) markedly inhibited IK(V), but this effect was not increased in the additional presence of apamin (0.3 μM; n=2).
By use of membranes prepared from rat brain cortex, [125I]‐charybdotoxin binding was consistent with an interaction at a single site with a KD of approximately 25 pM. [125I]‐charybdotoxin binding was unaffected by iberiotoxin (0.1 μM, n=6), but was increased by apamin in a concentration‐dependent manner (Emax 43±10%, P<0.05 and pEC50 7.1±0.2; n=7–8). Agitoxin‐2 (10 nM) displaced [125I]‐charybdotoxin binding by 91±3% (n=6) and prevented the effect of apamin (1 μM; n=6).
It is concluded that the EDHF‐mediated relaxation in the rat hepatic artery is not mediated by the opening of either KV or BKCa. Instead, the target K‐channels for EDHF seem to be structurally related to both KV and BKCa. The possibility that a subtype of SKCa may be the target for EDHF is discussed.

British Journal of Pharmacology (1997) 121, 141–149; doi:10.1038/sj.bjp.0701108