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Protocol A was used for all inotropy and signaling studies. Ouabain was perfused at 100 or 500 nM for 20 min. Protocol B was used for preconditioning studies. Ouabain was given for 4 min at the dose of 100 nM, as indicated. After ischemia, all hearts were reperfused with standard Krebs-Henseleit buffer for 2 hours.
