We assessed whether hypoosmotic swelling of cardiac myocytes activates volume-sensitive Cl⁻ current (I_Cl,swell) via the angiotensin II (AngII)-reactive oxygen species (ROS) signalling cascade. The AngII-ROS pathway previously was shown to elicit I_Cl,swell upon mechanical stretch of β_1D integrin. Integrin stretch and osmotic swelling are, however, distinct stimuli. For example, blocking Src kinases stimulates swelling-induced but inhibits stretch-induced I_Cl,swell.

I_Cl,swell was measured in rabbit ventricular myocytes by whole-cell voltage clamp. Swelling-induced I_Cl,swell was completely blocked by losartan and eprosartan, AngII type I receptor (AT₁) antagonists. AT₁ stimulation transactivates epidermal growth factor receptor (EGFR) kinase. Blockade of EGFR kinase with AG1478 abolished both I_Cl,swell and AngII-induced Cl⁻ current, whereas exogenous EGF evoked a Cl⁻ current that was suppressed by osmotic shrinkage. Phosphatidylinositol 3-kinase (PI-3K) is downstream of EGFR kinase, and PI-3K inhibitors LY294002 and wortmannin blocked I_Cl,swell. Ultimately, AngII signals via NADPH oxidase (NOX) and superoxide anion, •O₂⁻. NOX inhibitors, diphenyleneiodonium, apocynin and gp91ds-tat, eliminated I_Cl,swell, whereas scramb-tat, an inactive gp91ds-tat analogue, was ineffective. •O₂⁻ rapidly dismutates to H₂O₂. Consistent with H₂O₂ being a downstream effector, catalase inhibited I_Cl,swell, and exogenous H₂O₂ overcame suppression of I_Cl,swell by AT₁ receptor, EGFR kinase, and PI-3K blockers. H₂O₂-induced current was not blocked by osmotic shrinkage, however.

Activation of I_Cl,swell by osmotic swelling is controlled by the AngII-ROS cascade, the same pathway previously implicated in I_Cl,swell activation by integrin stretch. This in part explains why I_Cl,swell is persistently activated in several models of cardiac disease.