Adenosine A1Receptor Activation Reduces Reactive Oxygen Species and Attenuates Stunning in Ventricular Myocytes

Background Myocardial protection by volatile anesthetics involves activation of cardiac adenosine triphosphate-sensitive potassium (K(ATP)) channels. The authors have previously shown that isoflurane enhances sensitivity of the sarcolemmal K(ATP) channel to the opener, pinacidil. Because reactive oxygen species seem to be mediators in anesthetic preconditioning, the authors investigated whether they contribute to the mechanism of the sensitization effect by isoflurane. Methods Ventricular myocytes were isolated from guinea pig hearts for the whole cell patch clamp recordings of the sarcolemmal K(ATP) channel current (I(KAPT)). Free radical scavengers N-acetyl-L-cysteine, carnosine, superoxide dismutase, and catalase were used to investigate whether reactive oxygen species mediate isoflurane facilitation of the channel opening by pinacidil. A possible role of the mitochondrial K(ATP) channels was tested using a blocker of these channels, 5-hydroxydecanoate. Results The mean density (+/- SEM) of I(KAPT) elicited by pinacidil (20 microM) was 18.9 +/- 1.8 pA/pF (n = 11). In the presence of isoflurane (0.55 mM), the density of pinacidil-activated I(KAPT) increased to 38.5 +/- 2.4 pA/pF (n = 9). Concurrent application of isoflurane and N-acetyl-L-cysteine decreased the sensitization effect by isoflurane in a concentration-dependent manner, whereby the densities of I(KAPT) were 32.6 +/- 1.4 (n = 6), 26.2 +/- 2.3 (n = 6), and 19.4 +/- 2.1 pA/pF (n = 8) at 100, 250, and 500 microM N-acetyl-L-cysteine, respectively. Concurrent application of isoflurane and carnosine (100 microM), superoxide dismutase (100 U/ml), or catalase (100 U/ml) attenuated the densities of I(KAPT) to 27.9 +/- 2.6, 27.2 +/- 2.9, and 25.9 +/- 2.2 pA/pF, respectively. None of the scavengers affected activation of I(KAPT) by pinacidil alone. 5-Hydroxydecanoate (100 microM) did not alter the sensitization effect by isoflurane, and the density of I(KAPT) in this group was 37.1 +/- 3.8 pA/pF (n= 6). Conclusion These results suggest that reactive oxygen species contribute to the mechanism by which isoflurane sensitizes the cardiac sarcolemmal K(ATP) channel to the opener, pinacidil.

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Mechano-arrhythmogenicity is enhanced during late repolarisation in ischemia and driven by a TRPA1-, calcium-, and reactive oxygen species-dependent mechanism

10.1101/2021.08.20.457138 ◽

2021 ◽

Author(s):

Breanne Ashleigh Cameron ◽

T Alexander Quinn

Keyword(s):

Reactive Oxygen Species ◽

Ventricular Myocytes ◽

Reactive Oxygen ◽

Oxygen Species ◽

Cellular Mechanisms ◽

Sustained Activity ◽

Intracellular Ca ◽

Rabbit Ventricular Myocytes ◽

Induced Changes ◽

Dependent Mechanism

Background: Cardiac dyskinesis in regional ischemia results in arrhythmias through mechanically-induced changes in electrophysiology ('mechano-arrhythmogenicity') that involve ischemic alterations in voltage-calcium (Ca2+) dynamics, creating a vulnerable period (VP) in late repolarisation. Objective: To determine cellular mechanisms of mechano-arrhythmogenicity in ischemia and define the importance of the VP. Methods and Results: Voltage-Ca2+ dynamics were simultaneously monitored in rabbit ventricular myocytes by dual-fluorescence imaging to assess the VP in control and simulated ischemia (SI). The VP was longer in SI than in control (146±7 vs 54±8 ms; p<0.0001) and was reduced by blocking KATP channels with glibenclamide (109±6 ms; p<0.0001). Cells were rapidly stretched (10-18% increase in sarcomere length over 110-170 ms) with carbon fibres during diastole or the VP. Mechano-arrhythmogenicity, associated with stretch and release in the VP, was greater in SI than control (7 vs 1% of stretches induced arrhythmias; p<0.005) but was similar in diastole. Arrhythmias during the VP were more complex than in diastole (100 vs 69% had sustained activity; p<0.05). In the VP, incidence was reduced with glibenclamide (2%; p<0.05), by chelating intracellular Ca2+ (BAPTA; 2%; p<0.05), blocking mechano-sensitive TRPA1 (HC-030031; 1%; p<0.005), or by scavenging (NAC; 1%; p<0.005) or blocking reactive oxygen species (ROS) production (DPI; 2%; p<0.05). Ratiometric Ca2+ imaging revealed that SI increased diastolic Ca2+ (+9±1%, p<0.0001), which was not prevented by HC-030031 or NAC. Conclusion: In ischemia, mechano-arrhythmogenicity is enhanced specifically during the VP and is mediated by ROS, TRPA1, and Ca2+.

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