Proceedings of The Physiological Society
University of Oxford (2011) Proc Physiol Soc 23, PC255
Is SERCA overexpression anti- or pro-arrhythmic?
T. P. Collins1, A. R. Lyon1, M. B. Sikkel1, S. E. Harding1, K. T. MacLeod1
1. National Heart and Lung Institute, Imperial College, London, United Kingdom.
Sarcoplasmic reticulum calcium ATPase 2a (SERCA2a) gene therapy improves mechanical function in heart failure and is under evaluation in a clinical trial. Recent work has shown that SERCA2a gene delivery to rats in a chronic heart failure model reduced both spontaneous and catecholamine-induced arrhythmias in vivo, as well as reducing sarcoplasmic reticulum (SR) Ca2+ leak in failing myocytes. For the overexpression of SERCA, rats (male, 200-250g) received a 300μL tail vein injection of AAV9.SERCA2a (2x1011 drp) into their right internal jugular vein (performed under general anaesthesia with 2% isoflurane). Six to eight weeks later, hearts from control (n = 6) and normal+SERCA (n = 5) were harvested and myocytes were enzymatically isolated from the left ventricles. Intracellular Ca2+ transients and SR Ca2+ leak were measured using Fluo-4 fluorescence (and expressed in terms of F/F0) and cellular contraction was detected using an edge tracking system. Values are mean ± S.E.M., compared with either Student’s T-test or ANOVA with Tukey’s post-test comparison as appropriate. The overexpression of SERCA did not cause any significant changes in electrically stimulated contractions or Ca2+ transients (amplitude, rise time, 50 % decay). There was an increase in the SR load measured as peak fluorescence following 20 mM caffeine application (from 9.9 ± 0.6 in control myocytes, n = 30, to 13.2 ± 0.9 in normal+SERCA myocytes, n = 11, P < 0.01). In addition, there was a reduction in the tetracaine dependent SR leak (from 3.0 ± 0.2 in control myocytes, n = 30, to 2.3 ± 0.2 in normal+SERCA myocytes, n = 11, P < 0.05). In control myocytes, 30 nM isoprenaline increased cell shortening by 129 ± 24 % (n = 6, P < 0.001) and Ca2+ transient amplitude by 20.1 ± 3.3 % (n = 11, P < 0.001). In normal+SERCA cells, 30 nM isoprenaline increased cell shortening by 177 ± 45 % (n = 6, P < 0.001) and Ca2+ transient amplitude by 26.3 ± 9.5 % (n = 6, P < 0.05). However, there was a significant increase in the diastolic Ca2+ in normal+SERCA cells (25.4 ± 6.1 %, n = 11, P < 0.05), which means that the increase in Ca2+ transient amplitude is underestimated by the F/F0 value. The increase in diastolic Ca2+ in normal+SERCA cells is due to a large increase in SR leak in the presence of isoprenaline (from 2.3 ± 0.2 to 4.1 ± 0.6, n = 11 for both, P < 0.05). In addition to the increased leak, there was a greater frequency of Ca2+ waves following isoprenaline in normal+SERCA myocytes (from 0.018 ± 0.018 to 0.303 ± 0.054 waves per second, n = 11 for both, P < 0.001). There was no increase in either SR leak or Ca2+ wave frequency in control myocytes treated with isoprenaline. In conclusion, SERCA overexpression can be both pro- and anti-arrhythmic, depending on the type of myocyte that it is administered to (healthy vs. failing) as well as on the inotropic state of the myocyte.
Where applicable, experiments conform with Society ethical requirements