Ion channel modulators mediated alterations in NO-induced free radical generation and neutrophil membrane potential

Abstract

The present study investigated the effect of various ion (H⁺ and K⁺) channel modulators on nitric oxide (NO) donors (SNP and SNAP) induced free radical generation and on neutrophil membrane potential. Free radical generation was assessed by DCDHF-DA, using flow cytometry, while membrane potential was measured by a fluorescent dye, DiO-C5-(3). Neutrophil suspension in high potassium containing medium or following addition of NO donors (SNP, SNAP) to the neutrophil suspension led to free radical generation and membrane depolarization. DPI (a dual inhibitor of NADPH-oxidase and NOS), ABAH (MPO inhibitor) and BAPTA-AM (calcium chelator) significantly reduced 80 mM KCl or NO mediated free radical generation. Modulators of large (NS1619), intermediate (Chlorzoxazone) and small conductance (Apamin, chlorzoxazone) calcium activated K⁺ channels (TBA), voltage activated K⁺ channels (Kv) (4AP, 8Br-cGMP), ATP sensitive K⁺ channels (K_ATP) (Glybenclamide, pinacidil), Na⁺,K⁺-ATPase (Ouabain) and Na⁺/H⁺ exchanger (NHE, Amiloride) altered NO-induced neutrophil free radical generation response and membrane polarity. The results obtained thus suggest an association between rat neutrophil membrane depolarization and NO-dependent free radical generation.