Dileep K. Rohra, T Yamakuni, Y Ohizumi Acidosis-induced depolarization is due to activation of chloride channels in SHR and WKY rat aorta Dileep K. Rohra, T Yamakuni, Y Ohizumi

Introduction The contractile state of vascular smooth muscle can be regulated by many factors including changes in pH. Although the pH of blood and extracellular fluids is generally maintained at around 7.4, there are various pathophysiological conditions such as diabetes mellitus, renal dysfunction, and pulmonary edema, in which a significant decrease in pH occurs

Introduction Acidic pH induces a contraction (APIC) in aortas from SHR rats. A significant component of the APIC is associated with depolarization and Ca2+ entry through voltage-dependent Ca2+ channels.

Aim of study The aim of this study was to investigate the mechanism underlying the acidosis-induced depolarization and contraction in vascular tissue from SHR and WKY rats.

Representative tracings showing the effects of acidosis on SHR and WKY rat aortas 5 min // 64.8 mM KCl // pH 7.4 pH 6.5 pH 7.4 WKY // 64.8 mM KCl // pH 7.4 pH 6.5 pH 7.4

pH-contraction relationship in SHR aorta 125 ** 100 ** 75 Contraction (%) n = 8 50 * * 25 7.4 7.2 7.0 6.8 6.6 6.4 6.2 6.0 pHo

Effect of verapamil on acidosis-induced contraction of SHR and WKY rat aortas 0.5 g 5 min 64.8 mM KCl 1 mM verapamil pH 7.4 pH 6.5 pH 7.4 WKY 64.8 mM KCl 1 mM verapamil pH 7.4 pH 6.5 pH 7.4

**p < 0.01 versus control Comparative analysis of the effects of Ca2+ channel inhibitors on APIC in SHR aorta 100 **p < 0.01 versus control n = 7- 8 75 Contraction (%) ** ** 50 25 Control Verapamil (1 mM) Nifedipine (0.1 mM)

**p < 0.01 versus control Comparative effects of Cl- channel inhibitors and verapamil on acidic pH-induced contraction in SHR and WKY aortas SHR WKY 100 **p < 0.01 versus control n = 8 100 ** 75 ** 75 ** ** ** ** ** ** Contraction (%) 50 50 ** ** 25 25 Control Control 0.5 mM DIDS 0.5 mM 9-AC 30 mM IAA-94 0.5 mM DIDS 1 mM Verapamill 0.5 mM 9-AC 30 mM IAA-94 1 mM Verapamill 3 mM Niflumic acid 3 mM Niflumic acid

Effects of niflumic acid on acidic pH-induced Ca2+ mobilization and contraction in SHR aorta 3 mM niflumic acid 1.2 pH 7.4 R340/380 0.8 0.25 g 2 m

Effect of niflumic acid on acidic pH-induced depolarization in cultured vascular smooth muscle cells from SHR aorta pH 6.5 3 mM niflumic acid 5 Membrane potential 3 (arbitrary units) 1 1 min

Conclusion Acidic pH activates Cl- channels resulting in membrane depolarization of smooth muscle cells of both SHR and WKY aortas. Consequent to depolarization, opening of voltage-dependent Ca2+ channels occurs resulting in Ca2+ influx and contraction.

H+ Outside Inside H+ Contraction Scheme of events occurring under acidic pH conditions leading to contraction in SHR and WKY aortas Cl- Ca2+ Outside H+ Depolarization VDCC Inside Cl- Ca2+ H+ Tyrosine phosphorylation (activation) of PI3-kinase Via Akt, Rho kinase (?) Contraction