PNU-89692

ATP‐sensitive K+ (KATP) channel activation is implicated in the vascular hyporeactivity occurring in septic shock. However, channel inhibition with the sulphonylurea receptor (SUR) antagonist, glibenclamide (Glib) fails to reverse lipopolysaccharide (LPS)‐induced vascular hyporeactivity in vitro. We investigated whether inhibitors that act by binding to the KATP channel pore could be effective.Ring segments of endothelium‐intact rat mesenteric artery were incubated with LPS in culture media for either 6 or 20 h before contractile responses to phenylephrine were assessed in the absence or presence of KATP channel inhibitors.The pore‐forming subunit inhibitors barium chloride (BaCl2; 300 μM) and PNU‐37883A (1 μM) significantly reversed hyporeactivity at both time points, although less so at 20 h. In contrast, the SUR inhibitors, Glib (10 μM), tolbutamide (Tolb) (1 mM) and PNU‐99963 (1 μM) were ineffective. In LPS‐incubated tissues, Glib and Tolb antagonised contractions to the thromboxane A2 mimetic, U46619 (9,11‐dideoxy‐9α, 11α‐methanoepoxy prostaglandin F2α) (10−7 M), whereas the pinacidil‐derived inhibitor, PNU‐99963, did not.Contractions to 60 mM KCl were unaffected by LPS at 6 h, but were significantly depressed by LPS at 20 h, suggesting that K+‐channel‐independent pathways contribute to hyporeactivity at the later time point.The inducible nitric oxide synthase (iNOS) inhibitor, 1400 W (10 μM) and Tolb inhibited the production of nitrite induced by LPS, whereas BaCl2 and PNU‐37883A had no effect.In conclusion, KATP channels contribute to LPS‐induced vascular hyporeactivity via the iNOS pathway in rat mesenteric artery. The effectiveness of pore inhibitors over SUR inhibitors of the KATP channel suggests altered SUR function following LPS administration, which cannot be explained by thromboxane receptor inhibition.British Journal of Pharmacology (2005) 144, 367–375. doi:10.1038/sj.bjp.0706065

We investigated the mechanism of action of two novel nonsulphonylurea ATP‐sensitive potassium channel (KATP) inhibitors, PNU‐99963 and PNU‐37883A, on four types of cloned KATP channels.Whole‐cell currents were recorded in a symmetrical potassium (140 mM) gradient in HEK‐293 cells stably expressing Kir6.2/SUR1, Kir6.2/SUR2A, Kir6.2/SUR2B or Kir6.1/SUR2B.PNU‐99963 potently inhibited the four KATP channel clones. The concentration at which half‐maximum current was inhibited (IC50) was 66, 41, 43 and 11 nM for Kir6.2/SUR1, Kir6.2/SUR2A, Kir6.2/SUR2B and Kir6.1/SUR2B, respectively. In contrast, PNU‐99963 up to a concentration of 3 μM had no significant effect on current generated in HEK‐293 cells by transiently expressing Kir6.2Δ26, a C‐terminal truncated pore‐forming subunit of Kir6.2.PNU‐37883A inhibited four types of KATP channels, but to different extents. Inhibition of the putative smooth muscle KATP channel types, Kir6.2/SUR2B (IC50; 15 μM) and Kir6.1/SUR2B (IC50; 6 μM), was significantly greater than inhibition of either the pancreatic β cell or cardiac KATP channel clones. Moreover, PNU‐37883A significantly inhibited currents generated by expressing Kir6.2Δ26 alone, with an IC50 of 5 μM, which was significantly increased to 38 μM when Kir6.2Δ26 was expressed with SUR2B.In conclusion, two structurally different nonsulphonylurea compounds, PNU‐99963 and PNU‐37883A, inhibit KATP channels via different mechanisms, namely through the sulphonylurea receptor (SUR) and the pore‐forming subunits, respectively, although SUR2B reduced the inhibitory effect of PNU‐37883A. While PNU‐99963 potently inhibits all the four cloned KATP channels, PNU‐37883A has a degree of selectivity towards both smooth muscle KATP channels, but could not discriminate between them.British Journal of Pharmacology (2003) 139, 122–128. doi:10.1038/sj.bjp.0705228