Uptake of creatine phosphate into heart mitochondria: a leak in the creatine shuttle

CrP uptake into isolated rat heart mitochondria was studied using silicone oil centrifugation. Further, the involvement of the mitochondrial adenine nucleotide translocase was examined by measuring CrP accumulation in mitochondria in the presence of substrates and inhibitors of the ATP/ADP-carrier and by investigating uptake kinetics in liposomes reconstituted with purified bovine heart adenine nucleotide translocase protein. CrP is accumulated in the matrix space of isolated rat heart mitochondria and mitoplasts. The uptake is inhibited by carboxyatractyloside, a specific inhibitor of the mitochondrial adenine nucleotide translocase, and by ADP, phosphoenolpyruvate, 3-phosphoglycerate and pyrophosphate, compounds which are able to bind to the carrier. It is not inhibited when the mitochondrial membrane potential is decreased. CrP is transported into reconstituted liposomes at a rate which is about 3 orders of magnitude lower than the rate for ATP uptake. The transport is sensitive to temperature change and to carboxyatractyloside. It is concluded that CrP is specifically taken up by heart mitochondria via the mitochondrial adenine nucleotide translocase. The transport in mitochondria in situ is facilitated by the close local and functional interaction of the mitochondrial creatine kinase and the adenine nucleotide translocase within contact sites between inner and outer mitochondrial membrane. A certain amount of CrP synthesized by the mitochondrial creatine kinase thus escapes its usage at cytosolic energy consuming processes.