Copyright © 2005, European Society of Cardiology
Effects of calsequestrin over-expression on excitation–contraction coupling in isolated rabbit cardiomyocytes
aInstitute of Biomedical and Life Sciences, University of Glasgow, Glasgow G12 8QQ, UK
bDepartment of Cardiology and Pneumology, Georg-August-University Goettingen, D-37075 Goettingen, Germany
* Corresponding author. Tel.: +44 141 3305963; fax: +44 141 3304612. Email address: g.smith{at}bio.gla.ac.uk
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 β-galactosidase (Ad-LacZ) was used as a control.
Results: In voltage-clamped, Fura-2-loaded cardiomyocytes, L-type Ca2+ current (ICa,L) and Ca2+ transient amplitude were both increased in the Ad-CSQ group by 
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 µM) was used to reduce ICa,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 60% higher as assessed by both the caffeine-induced Ca2+ release and the accompanying Na+/Ca2+ exchanger current (INCX). The cause of the increased ICa,L is unknown. No change in the expression level of the 
1-subunit of the L-type Ca channel was observed. β-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.
KEYWORDS E–C coupling; SR (function); Calcium (cellular); Ion channels
Time for primary review 21 days
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