SMALL-SCALE PREPARATION OF SKELETAL-MUSCLE MITOCHONDRIA, CRITERIA OF INTEGRITY, AND ASSAYS WITH REFERENCE TO TISSUE FUNCTION

Mitochondria prepared in small scale from skeletal muscle were studied with respiration measurements and low temperature spectroscopy. The m ethod of preparation was developed for 25-100 mg tissue with pigeon br east muscle as model organ. The yield was 40%. Data collected during t he developmental work were used to evaluate criteria of mitochondrial quality. The cytochrome c conservation, i.e. cytochrome c per mitochon drial quantity in the preparation relative to that in the tissue, is a most useful test parameter. It is bounded between 0-100%. Proportiona lity between the state 3 rate and the cytochrome c conservation was no t rejected by statistical tests. The respiratory control ratio (RCR) w as also highly correlated to the cytochrome c conservation. These corr elations might be extrapolated to 100% conservation to give hypothetic al tissue values. The cause for the correlations is discussed. The P/O ratio showed only weak dependence on the cytochrome c conservation an d the state 4 rate showed no dependence. Other, rather insensitive tes t parameters are also discussed. The pigeon breast muscle mitochondria isolated by the final method showed cytochrome c conservation of 73 /- 9% (n = 16). They are compared with pig biceps femoris mitochondria prepared by the same method. The two types of mitochondria show many similarities. Some differences may be explained by a different amount of inner mitochondrial membrane relative to mitochondrial protein. The pig tissue contains ten times less mitochondrial protein than the pig eon tissue does.