Subcellular heterogeneity in mitochondrial red-ox responses to KATP channel agonists in freshly isolated rabbit cardiomyocytes.

We have used the technique of fluorescent microscopy imaging supplemented with the refined analysis of temporal cartography of the cell fluorescence to investigate the mechanisms of regulation of mitochondrial function and its red-ox state in cardiac cells in vivo. Autofluorescence of flavoproteins of the respiratory chain in the isolated rabbit cardiomyocytes was registered before and after application of mitochondrial KATP channel opener diazoxide (100 and 400 microM). Diazoxide addition resulted in oxidation of flavoproteins. Detailed analysis of these responses showed that they were heterogeneous over space and time. The local responses show rapid jumps. In a few cells, metabolic oscillations developed and could be recorded for tens of minutes. Under these conditions the cells appeared divided into a small number of regions in which mitochondria function synchronously. Local pattern of oxidation switches again and again from a reduced state to the same level of oxidation. All these phenomena where absent when the cells were permeabilized by saponin giving a direct access to mitochondrial KATP channel opener. Cross-correlation analysis revealed a high degree of homogeneity for cells presenting metabolic oscillations, contrarily to those displaying a smooth increase in fluorescence in response to diazoxide. The results are consistent with the view that mitochondria form independent functional units whose behaviour can be synchronised by some unknown cellular factors or metabolites.