Chloride Channel Inhibition Blocks the Protection of Ischemic Preconditioning and Hypo-Osmotic Stress in Rabbit Ventricular Myocardium

Abstract

Abstract-The objective of this study was to examine the role of chloride (Cl -) channels in the myocardial protection of ischemic preconditioning (IP). Isolated rabbit ventricular myocytes were preconditioned with 10-minute simulated ischemia (SI) and 20-minute simulated reperfusion (SR) or not preconditioned (control). The myocytes then received 180-minute SI or 45-minute SI/120-minute SR. Indanyloxyacetic acid 94 (IAA-94, 10 mol/L) or 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB, 1 mol/L) was administered before IP or before SI or SI/SR to inhibit Cl -channels. Electrophysiological studies indicate that these drugs, at the concentrations used, selectively abolished Cl -currents activated under hypo-osmotic conditions (215 versus 290 mOsm). IP significantly (PϽ0.001) reduced the percentage of dead myocytes after 60-minute (30.8Ϯ1.3%, meanϮSEM), 90-minute (35.3Ϯ1.3%), and 120-minute (39.2Ϯ1.7%) SI compared with controls (44.7Ϯ1.6%, 54.5Ϯ1.3%, and 58.9Ϯ1.8%, respectively) and after 45-minute SI/120-minute SR (36.3Ϯ0.6%) compared with control (56.6Ϯ2.2%). Hypo-osmotic stress also produced protection similar to IP. IAA-94 or NPPB abolished the protection of both IP and hypo-osmotic stress. In buffer-perfused rabbit hearts preconditioned with three 5-minute ischemia/10-minute reperfusion cycles given before the 40-minute long ischemia and 60-minute reperfusion, IP

The volume-regulated anion channel (LRRC8) in nodose neurons is sensitive to acidic pH

“…It has been shown that enhanced cell volume regulation is a key protective mechanism of ischemic preconditioning (42) and that Cl -channel inhibition blocks the protection of ischemic preconditioning and hypoosmotic stress in ventricular myocardium (43). Therefore, we tested the hypothesis that the acid pH o -conditioned Cl -current is similarly protective in nodose neurons.…”

“…Studies in rabbit ventricular myocyte demonstrated that a Cl -channel that enhances cell volume regulation provides a key protective mechanism of ischemic preconditioning (42). Conversely, the same authors showed that Cl -channel inhibition blocks the protection of ischemic preconditioning and hypoosmotic stress in rabbit ventricular myocardium (43). It is also known that ROS generation is required for ischemic cardiac preconditioning (56).…”

Molecular Basis of Cardioprotection

“…407 The role of chloride channel activation in such attenuation of intracellular edema is contentious. 408,409 Potassium channels may also be involved in cell volume control during IPC to maintain electroneutrality along with chloride efflux. 410 Similar findings on chloride and potassium channels were also reported in cardiomyocytes subjected to plasma dialysate from a RIPC protocol.…”

Remote Ischemic Preconditioning Cardio-protection via Enhanced Cell Volume Regulation Requires Activation of Swelling-Activated Chloride (Icl- Swell) Channels

“…We have demonstrated that pan chloride channel inhibition blocked the protection of IPC against I/R necrosis in both Langendorff perfused hearts and isolated ventricular myocytes [11]. Furthermore, we have determined that the osmotic equivalent for IPC protection in cultured cardiomyocytes is 50-60 mOsm [15] and by quantitative fluorescence imaging of Cl-in the same model observed that IPC causes a net Cl -efflux of ~30 mOsm [10].…”

“…New Zealand White rabbits (weight range 3.0 to 3.5 kg) were used as heart donors to obtain ventricular cardiomyocytes using an enzymatic digestion method we have previously described [9][10][11].…”