Abstract 15020: Activation of Mitochondrial Bk Ca Channels Improves Hemodynamics and Normalizes Phosphoproteomic Changes in Pigs Following Cardioplegic Arrest and Cardiopulmonary Bypass

Abstract

Introduction: BKCa channels are large conductance Ca++-activated K+ channels, which reside in the inner mitochondrial membrane in cardiomyocytes. We have demonstrated that the BKCa channel activator rottlerin (a.k.a mallotoxin) is cardioprotective during cardioplegic arrest in isolated rodent hearts and in vitro models. Here we used a clinically relevant in vivo swine model of cardioplegic arrest and cardiopulmonary bypass (CP/CPB) with and without rottlerin supplemented CP solution. Mitochondrial signaling alterations were identified by TMT-MS analysis of phosphorylation enriched mitochondrial protein fractions. Methods: Yorkshire swine (50 Kg) of both sexes were subjected to hypothermic CP/CPB (1hour) followed by 1 hour reperfusion. Groups included sham (n = 5), CP/CPB alone (n = 8) and CP/CPB with rottlerin supplemented cardioplegia (1uM) (CP/CPB+R) (n = 7). Mitochondria-enriched fractions were isolated from the left ventricular free wall and subjected to Tandem Mass Tag (TMT) labeling coupled mass spectrometry analysis. Results: CP/CPB pigs had reduced myocardial function indicative of myocardial stunning as shown by decreased LVDevP, and ±dP/dt vs sham (P<.05). BKCa activation significantly restored left ventricular function compared to CP/CPB alone (LVDevP: 74.0 ± 2.8 vs . 52.5±5.0 mmHg, P < .01, +dP/dt: 1482±116 vs . 923±126 mmHg/s, P < .01, and - dP/dt: 1120±116 vs . 629±82 mmHg/s, P < .01). TMT-MS analysis identified 3363 proteins. 410 unique as well as numerous novel phosphorylation sites were significantly regulated in CP/CPB vs sham. Rottlerin treatment generally normalized phosphorylation changes with 175 and 63 different vs CP/CPB and sham, respectively. Enrichment analysis demonstrated significant changes in pathways associated with adrenergic and PKA signaling, calcium handling, mitochondrial dynamics, cristae organization, and oxidative phosphorylation. Conclusions: BKCa-activator supplemented cardioplegia significantly improves myocardial functional recovery following CP/CPB in a clinically relevant large animal model. Furthermore, mitochondrial BKCa-channel activation during cardioplegic arrest normalizes numerous signaling alterations associated with myocardial stunning and cardiac surgery.