CAFFEINE AND CA2-PHOSPHORYLATION IN SAPONIN-SKINNED HUMAN SKELETAL-MUSCLE FIBERS DUE TO ACTIVATION OF ACTOMYOSIN ATPASE( STIMULATE MITOCHONDRIAL OXIDATIVE)

The rate of mitochondrial oxidative phosphorylation of saponin-skinned human muscle fibers from m. vastus lateralis in the presence of gluta mate, malate and ATP is reported to be sensitive to caffeine and to ch anges of free calcium ion concentration. An approximately twofold incr ease in respiration was observed by the addition of 15 mM caffeine, be cause of the efflux of calcium from sarcoplasmic reticulum. Direct add ition of a Ca2+/CaEGTA buffer, containing 1.5 mu M free calcium ions h ad a similar effect. The ATP-splitting activity of skinned fibers was also stimulated by caffeine or calcium. These observations can be expl ained exclusively by the calcium-induced activation of actomyosin ATPa se. (i) Thapsigargin, an inhibitor of the sarcoplasmic reticulum Ca2+- ATPase, had no influence. (ii) In myosin-extracted 'ghost' fibers cont aining intact mitochondria and an intact sarcoplasmic reticulum caffei ne had a negligible effect on oxidative phosphorylation. (iii) The caf feine-induced increase in rate of fiber respiration was concomitant wi th a decrease in mitochondrial membrane potential and a decrease in th e redox state of the mitochondrial NAD system. (iv) The calcium ionoph ore A 23187 caused a stimulation of respiration and ATP-splitting acti vity, similar to caffeine. (v) The calcium dependencies of respiration and ATP splitting activity of saponin-skinned human muscle fibers wer e in experimental error identical. Therefore it is concluded that calc ium efflux from sarcoplasmic reticulum affects oxidative phosphorylati on in skeletal muscle mostly via the stimulation of actomyosin ATPase.