Activation of Glucose‐6‐Phosphate Dehydrogenase by Depolarization of Membrane Potential Mediates Vascular Smooth Muscle Contraction

Abstract

We have previously demonstrated that inhibition of glucose‐6‐phosphate dehydrogenase (G6PD) evokes relaxation of vascular smooth muscle (VSMC). So, to elucidate the mechanism of relaxation, we examinedwhether G6PD activation affects contraction of VSMC. We found,using ultracentrifugation, two isoforms (62 & 58 KD) of G6PD were present in bovine coronary artery (BCA); the longer (62 KD)isoform was localized in caveolae fraction and the shorter(58 KD) isoform was detected in cytoplasmic fraction. Both isoforms were functionally active; depolarization of the membrane potential to ‐9mV by KCl (30 mM; n=5) increased G6PD activity in the membrane fraction by ~2‐fold. Mechanistic studies revealed that PKCδ mediated activation of G6PD evoked by depolarization in a time‐dependent manner.Silencing of PKCδ in VSMC and HEK293 decreased (P<0.05; n=5) basal activity (0.25 nmol/min/mg protein) of endogenous G6PD and ectopically expressed G6PD in HEK293 (PKCδ−/G6PD+) cells by 50% as compared to cells transfected with non‐targeting siRNA, and in these PKCδ−cells 30mMKClwas unable to activate G6PD. Interestingly, inhibition of G6PD expression in BCA by siRNAand in aorta of G6PD mutated mice, decreased KCl‐induced contractions. In conclusion, activation of G6PD by depolarization of membrane potential plays a pivotal role in mediatingVSMC contraction.This study is supported by AHA and NIH grant# 0435070N and R01HL85352