title: Effects of cytosolic NADH/NAD(+) levels on sarcoplasmic reticulum Ca(2+) release in permeabilized rat ventricular myocytes.
abstract: In the heart ischaemic conditions induce metabolic changes known to have profound effects on Ca(2+) signalling during excitation-contraction coupling. Ischaemia also affects the redox state of the cell. However, the role of cytosolic redox couples, such as the NADH/NAD(+) redox system, for the regulation of Ca(2+) homeostasis has remained elusive. We studied the effects of NADH and NAD(+) on sarcoplasmic reticulum (SR) Ca(2+) release in permeabilized rat ventricular myocytes as well as on Ca(2+) uptake by SR microsomes and ryanodine receptor (RyR) single channel activity. Exposure of permeabilized myocytes to NADH (2 mm; [Ca(2+)](cyt)= 100nm) decreased the frequency and the amplitude of spontaneous Ca(2+) sparks by 62% and 24%, respectively. This inhibitory effect was reversed by NAD(+) (2 mm) and did not depend on mitochondrial function. The inhibition of Ca(2+) sparks by NADH was associated with a 52% decrease in SR Ca(2+) load. Some of the effects observed with NADH may involve the generation of superoxide anion (O(2)(-).) as they were attenuated to just a transient decrease of Ca(2+) spark frequency by superoxide dismutase (SOD). O(2)(-). generated in situ from the xanthine/xanthine oxidase reaction caused a slowly developing decrease of Ca(2+) spark frequency and SR Ca(2+) load by 44% and 32%, respectively. Furthermore, in studies with cardiac SR microsomes NADH slowed the rate of ATP-dependent Ca(2+) uptake by 39%. This effect also appeared to depend on O(2)(-). formation. Single channel recordings from RyRs incorporated into lipid bilayers revealed that NADH (2 mm) inhibited the activity of RyR channels by 84%. However, NADH inhibition of RyR activity was O(2)(-).-independent. In summary, an increase of the cytoplasmic NADH/NAD(+) ratio depresses SR Ca(2+) release in ventricular cardiomyocytes. The effect appears to be mediated by direct NADH inhibition of RyR channel activity and by indirect NADH inhibition (O(2)(-). mediated) of SR Ca(2+)-ATPase activity with a subsequent decrease in SR Ca(2+) content.
authors.affiliation: Department of Physiology, Stritch School of Medicine, Loyola University Chicago, 2160 South First Avenue, Maywood, IL 60153, USA.
authors.forename: Aleksey V
authors.initials: AV
authors.lastname: Zima
authors.forename: Julio A
authors.initials: JA
authors.lastname: Copello
authors.forename: Lothar A
authors.initials: LA
authors.lastname: Blatter
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002463:Cell Membrane Permeability; D003600:Cytosol / Q000378:metabolism*; D004553:Electric Conductivity; D006352:Heart Ventricles; D008297:Male; D008929:Mitochondria, Heart / Q000502:physiology; D032383:Myocytes, Cardiac / Q000378:metabolism*; D009243:NAD / Q000378:metabolism* / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D019837:Ryanodine Receptor Calcium Release Channel / Q000502:physiology; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism
chemical_list: D019837:Ryanodine Receptor Calcium Release Channel; D009243:NAD; D013481:Superoxides; D002118:Calcium