Abstract 264: Therapeutic Hypothermia Cardioprotection Is Associated with Enhanced pAMPK but Not ATP Preservation
Introduction: Therapeutic Hypothermia (TH) is cardioprotective following ischemia/reperfusion (I/R) injury and improves survival following cardiac arrest in an Akt dependent manner. AMP activated protein kinase (AMPK) is a highly conserved, cardioprotective, sensor of cellular energetic stress and ATP depletion that is activated by ischemia. Phosphorylated AMPK (pAMPK) is also a significant inhibitor of the mammalian target of rapamycin complex 1 (mTORC1) that is activated by Akt but inhibited by TH. In this study we hypothesized that TH enhances pAMPK independent of cellular ATP while increasing cell survival. We further hypothesized that strategies to enhance pAMPK would mimic TH cardioprotection by inhibiting mTORC1.
Methods: Cardiomyocytes isolated from 1-2-day old C57BL6/J mice, were exposed to simulated Ischemia (90 min) followed by reperfusion. For TH, cells were cooled from 37C to 32C during ischemia and the first hour of reperfusion. Protein lysates were collected at serial time points for changes in pAMPK by western blot analyses. ATP measurements were made using a standard bioluminescence assay.
Results: pAMPK was significantly increased during ischemia as compared to controls. During reperfusion, pAMPK decreased but was maintained by TH (n=5, p<.05). Total AMPK was unchanged. Total cellular ATP decreased significantly with ischemia and TH to 6% and 13% of controls (n=5, p<.05). Following 30 minutes of reperfusion, ATP increased to 9% (IR) and 26% (TH) of controls. Although total cellular ATP increased with TH, these changes were not significant and remained substantially reduced from controls. Cardiomyocyte cell death during ischemia was minimal but increased during reperfusion (4% vs. 46%, p.05, n=5). This cell death was attenuated by TH (24%, p<.05, n=5). The pAMPK activator metformin (100um) similarly attenuated reperfusion associated cell death (22%, p<.05, n=4) while inhibiting mTORC1.
Conclusions: TH is associated with enhanced cardiomyocyte survival and pAMPK following IR. These events were not associated with enhanced ATP preservation during ischemia. Pharmacologic strategies to mimic TH effects on AMPK may be beneficial to enhance survival following cardiac arrest.
- © 2011 by American Heart Association, Inc.