Effects of spermine on mitochondrial Ca2+ transport and the ranges of extramitochondrial Ca2+ to which the matrix Ca2+-sensitive dehydrogenases respond.
1. Spermine has previously been reported to be an activator of mitochondrial Ca2+ uptake [Nicchitta & Williamson (1984) J. Biol. Chem. 259, 12978-12983]. This is confirmed in the present studies on rat heart, liver and kidney mitochondria by using the activities of the Ca2+-sensitive intramitochondrial dehydrogenases (pyruvate, NAD+-isocitrate and 2-oxoglutarate dehydrogenases) as probes for matrix Ca2+, and also, for the heart mitochondria, by using entrapped fura-2. 2. As also found previously [Damuni, Humphreys & Reed (1984) Biochem. Biophys. Res. Commun. 124, 95-99], spermine activated extracted pyruvate dehydrogenase phosphate phosphatase. However, it was found to have no effects at all on the extracted NAD+-isocitrate or 2-oxoglutarate dehydrogenases. It also had no effects on activities of the enzymes in mitochondria incubated in the absence of Ca2+, or on the Ca2+-sensitivity of the enzymes in uncoupled mitochondria. 3. Spermine clearly activated 45Ca uptake by coupled mitochondria, but had no effect on Ca2+ egress from mitochondria previously loaded with 45Ca. 4. Spermine (with effective Km values of around 0.2-0.4 mM) caused an approx. 2-3-fold decrease in the effective ranges of extramitochondrial Ca2+ in the activation of the Ca2+-sensitive matrix enzymes in coupled mitochondria from all of the tissues. The effects of spermine appeared to be largely independent of the other effectors of mitochondrial Ca2+ transport, such as Mg2+ (inhibitor of uptake) and Na+ (promoter of egrees). 5. In the most physiological circumstance, coupled mitochondria incubated with Na+ and Mg2+, the presence of saturating spermine (2 mM) resulted in an effective extramitochondrial Ca2+ range for matrix enzyme activation of from about 30-50 nM up to about 800-1200 nM, with half-maximal effects around 250-400 nM-Ca2+. The implications of these findings for the regulation of matrix and extramitochondrial Ca2+ are discussed.
McCormack
J G
JG
Department of Biochemistry, University of Leeds, U.K.
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002451:Cell Compartmentation; D004305:Dose-Response Relationship, Drug; D007521:Isocitrate Dehydrogenase / Q000378:metabolism; D007655:Ketoglutarate Dehydrogenase Complex / Q000378:metabolism; D007668:Kidney / Q000201:enzymology; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008928:Mitochondria / Q000378:metabolism*; D008929:Mitochondria, Heart / Q000201:enzymology; D008930:Mitochondria, Liver / Q000201:enzymology; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism; D051381:Rats; D012964:Sodium / Q000494:pharmacology; D013096:Spermine / Q000494:pharmacology*
D011768:Pyruvate Dehydrogenase Complex; D013096:Spermine; D012964:Sodium; D007521:Isocitrate Dehydrogenase; D007655:Ketoglutarate Dehydrogenase Complex; D008274:Magnesium; D002118:Calcium