Subcellular distribution of energy metabolites in the pre-ischaemic and post-ischaemic perfused working rat heart

Isolated working rat hearts were subjected to 20 min of global ischaemia and either 5 min or 15 min of reperfusion. The subcellular distribution of ATP, ADP, AMP, phosphocreatine and Pi were determined before and after ischaemia by the method of non-aqueous tissue fractionation. Ventricular function and the cytosolic, mitochondrial and ATPase-associated compartmentation of metabolites were measured. After 5 min of reperfusion, only 13 +/- 9% of the pre-ischaemic contractile function was restored compared to 67 +/- 8% after 15 min reperfusion. ATP was reduced in all cellular compartments after 5 min of reperfusion but was only decreased from pre-ischaemic values in the cytosolic compartment after 15 min of reperfusion (17.1 +/- 3.9 nmol/mg vs. 4.3 +/- 1.5 nmol/mg total protein; P less than 0.05). The mitochondrial ATP]/ADP] was reduced from a normal value of 4.36 to 1.79 after 5 min but recovered to 4.62 after 15 min of reperfusion. Most of the Pi was located in the mitochondria or with the ATPase fraction of the cell, with only 16% of the total Pi free in the cytosol. This study indicates that the capacity of the heart to recover function may be compromised during early reperfusion by a 59% increase in mitochondrial phosphate content and during late reperfusion by a reduced cytosolic/mitochondrial concentration ratio of both ATP (from 0.85 to 0.19) and phosphocreatine (from 3.9 to 1.24).