Regulation of respiration in brain mitochondria and synaptosomes: restrictions of ADP diffusion in situ, roles of tubulin, and mitochondrial creatine kinase.
The role of ubiquitous mitochondrial creatine kinase (uMtCK) reaction in regulation of mitochondrial respiration was studied in purified preparations of rat brain synaptosomes and mitochondria. In permeabilized synaptosomes, apparent Km for exogenous ADP, Km (ADP), in regulation of respiration in situ was rather high (110 +/- 11 microM) in comparison with isolated brain mitochondria (9 +/- 1 microM). This apparent Km for ADP observed in isolated mitochondria in vitro dramatically increased to 169 +/- 52 microM after their incubation with 1 muM of dimeric tubulin showing that in rat brain, particularly in synaptosomes, mitochondrial outer membrane permeability for ADP, and ATP may be restricted by tubulin binding to voltage dependent anion channel (VDAC). On the other hand, in synaptosomes apparent Km (ADP) decreased to 25 +/- 1 microM in the presence of 20 mM creatine. To fully understand this effect of creatine on kinetics of respiration regulation, complete kinetic analysis of uMtCK reaction in isolated brain mitochondria was carried out. This showed that oxidative phosphorylation specifically altered only the dissociation constants for MgATP, by decreasing that from ternary complex MtCK.Cr.MgATP (K (a)) from 0.13 +/- 0.02 to 0.018 +/- 0.007 mM and that from binary complex MtCK.MgATP (K (ia)) from 1.1 +/- 0.29 mM to 0.17 +/- 0.07 mM. Apparent decrease of dissociation constants for MgATP reflects effective cycling of ATP and ADP between uMtCK and adenine nucleotide translocase (ANT). These results emphasize important role and various pathophysiological implications of the phosphocreatine-creatine kinase system in energy transfer in brain cells, including synaptosomes.
Monge
Claire
C
Laboratory of Fundamental and Applied Bioenergetics, Joseph Fourier University, Grenoble, France.
Beraud
Nathalie
N
Kuznetsov
Andrey V
AV
Rostovtseva
Tatiana
T
Sackett
Dan
D
Schlattner
Uwe
U
Vendelin
Marko
M
Saks
Valdur A
VA
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000201:enzymology*; D019069:Cell Respiration / Q000187:drug effects; D003401:Creatine / Q000494:pharmacology; D052281:Creatine Kinase, Mitochondrial Form / Q000378:metabolism*; D004058:Diffusion / Q000187:drug effects; D004579:Electron Transport / Q000187:drug effects; D004734:Energy Metabolism; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D008297:Male; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology*; D008929:Mitochondria, Heart / Q000187:drug effects / Q000201:enzymology; D000226:Mitochondrial ADP, ATP Translocases / Q000378:metabolism; D010539:Permeability / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D013574:Synaptosomes / Q000187:drug effects / Q000201:enzymology*; D014020:Tissue Extracts; D014404:Tubulin / Q000378:metabolism*
D007527:Isoenzymes; D014020:Tissue Extracts; D014404:Tubulin; D000244:Adenosine Diphosphate; D000226:Mitochondrial ADP, ATP Translocases; D052281:Creatine Kinase, Mitochondrial Form; D003401:Creatine