Calcium Influx Pathways in the Calcium Paradox
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Project Summary:
The Ca paradox is the observation that massive cell death occurs immediately on restoration of Ca to the perfusate of hearts perfused for a brief period with Ca-free medium. It is important to understand the Ca paradox because it reveals mechanisms of injury relevant to heart preservation for transplantation. It is known that cell lysis on reperfusion with Ca results from increased membrane fragility and is accompanied by enzyme release and massive Ca uptake. It is unclear, however, what molecular mechanisms underlie the Ca paradox, or what causes the massive Ca uptake. We have found that the Ca paradox is inhibited by oxidized ATP, an inhibitor of P2X7 receptors. This is an extremely significant result, because previous work on the Ca paradox in 40 years has not been able to relate the phenomenon to a specific pathway. The objective of this application is to elucidate the mechanism of action of oxidized ATP by which it inhibits the Ca paradox, including its role in preventing abnormal Na and Ca uptake.
Our hypothesis is as follows: The Ca paradox is mediated by myocyte P2X7 or P2X4 receptors, which become activated during Ca-free perfusion via NAD and ATP released via connexin-43 hemichannels. To test our hypothesis we aim to: 1) Define the site of action of oxidized ATP by which it inhibits the Ca paradox. This will utilize three approaches: a proteomic approach using affinity labeling with oATP, P2X7 receptor knockout mice, and pharmacological studies with rat hearts. 2) Determine whether connexin-43 hemichannels mediate the Ca paradox by the release of ATP and NAD, during Ca-free perfusion of rat hearts.
As a result of this project we expect to significantly advance understanding of the mechanism of the Ca paradox, and by extension the pathophysiology of heart preservation injury.
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Cardiothoracic Surgery
- University of Wisconsin Department of Surgery
First published: 07/15/02 Last updated: 08/19/08
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