Assessment of sarcoplasmic reticulum Ca2+ flux pathways in cardiomyocytes from rabbits with infarct-induced left-ventricular dysfunction

Abstract

The aim of the study was to correlate intracellular Ca2+ transients with Ca2+ uptake and efflux characteristics of the sarcoplasmic reticulum (SR) in ventricular myocytes isolated from rabbits with left-ventricular dysfunction (LVD). Chronic (8 weeks) ligation of a coronary artery caused marked LVD in rabbits. Measurements of intracellular [Ca2+] were made using Fura-2 on intact, single, left-ventricular myocytes. SR Ca2+ flux rates associated with sarco-endoplasmic reticulum Ca2+ ATPase type 2 (SERCA2)-mediated uptake, ryanodine receptor type 2 (RyR2)-mediated Ca2+ efflux and background SR Ca2+ leak were measured in suspensions of permeabilised myocytes. Measurements on single, permeabilised myocytes were used to assess the steady-state Ca2+ content of the SR and the characteristics of spontaneous SR Ca2+ release. Peak systolic [Ca2+] was significantly lower; time-to-peak and Ca2+ transient duration were significantly longer in LVD myocytes. SERCA2-mediated Ca2+ uptake was reduced to ∼50% in myocytes from the LVD group. Ruthenium red (RuR)-sensitive Ca2+ efflux (mediated by the RyR2) was also reduced in the LVD group by ∼50%, as was the remaining (RuR-insensitive) background Ca2+ leak. Measurements from single, permeabilised myocytes showed a lower steady-state SR Ca2+ content. The frequency and amplitude of spontaneous SR Ca2+ release from LVD hearts was also reduced. Partial inhibition of SERCA2 by thapsigargin depressed both the amplitude and the frequency of spontaneous release. Partial inhibition of RyR2-mediated-Ca2+ efflux with tetracaine enhanced spontaneous Ca2+ release amplitude and decreased frequency. Increased background Ca2+ leak with ionomycin decreased the frequency of spontaneous release. It is concluded that partial inhibition of SERCA2 mimics some aspects of altered SR function in LVD, but reduced RyR2 function cannot explain the other functional alterations observed. Reduced background Ca2+ leak from the SR may compensate partly for the reduced Ca2+ uptake capacity of the SR in the LVD group.