Effects of calsequestrin over-expression on excitation-contraction coupling in isolated rabbit cardiomyocytes

Objective: This study investigated the role of calsequestrin (CSQ) in the control of excitation-contraction (E-C) coupling in the heart. Methods: CSQ over-expression was induced in isolated rabbit ventricular cardiomyocytes using an adenovirus coding for rabbit CSQ (Ad-CSQ). After 24 h of culture, CSQ protein expression was increased by 58 +/- 18% (n = 10). An adenovirus coding for beta-galactosidase (Ad-LacZ) was used as a control. Results: In voltage-clamped, Fura-2-loaded cardiomyocytes, L-type Ca2+ current (I-Ca,I-L) and Ca2+ transient arriplitude were both increased in the Ad-CSQ group by similar to 78%. Doubling the external Ca2+ concentration in the control group (Ad-LacZ) increased the LTCC amplitude to a similar degree (85 +/- 6%), but increased the Ca2+ transient amplitude by 149 +/- 13%. This suggests that SR Ca2+ release may be inhibited upon CSQ over-expression. Alternatively, nifedipine (0.5 mu M) was used to reduce I-Ca,I-L in Ad-CSQ-transfected cells to values comparable to control (Ad-LacZ). Under these conditions, Ca2+ transient amplitude was not different from Ad-LacZ, but the SR Ca2+ content was similar to 60% higher as assessed by both the caffeine-induced Ca2+ release and the accompanying Na+/Ca2+ exchanger current (I-NCX). The cause of the increased I-Ca,I-L is unknown. No change in the expression level of the at-subunit of the L-type Ca channel was observed. beta-Escin-permeabilized cardiomyocytes were used to study Ca2+ sparks imaged with Fluo-3 at 145-155 nmol/L [Ca2+]. Spontaneous Ca2+ spark frequency, duration, width, and amplitude were unchanged in the Ad-CSQ group, but SR Ca2+ content was 48% higher than Ad-LacZ. Conclusions: CSQ over-expression increased SR Ca2+ content but reduced the gain of E-C coupling in rabbit cardiomyocytes. (c) 2005 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.