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1 Cardiac Membrane Res. Lab., Simon Fraser University, Burnaby, BC, Canada; Cardiovascular Sciences, Child and Family Research Institute, Vancouver, BC, Canada
2 Cardiac Membrane Res. Lab., Simon Fraser University, Burnaby, BC, Canada
3 Cardiology, Hospital de Santa Creus y Sant Pau, Barcelona, Spain
* To whom correspondence should be addressed. E-mail: tibbits{at}sfu.ca.
L-type Ca2+ channel-mediated Ca2+-induced Ca2+ release (CICR) is the dominant mode of excitation-contraction (E-C) coupling in the mature mammalian myocardium but is thought to be absent in the fetal and newborn mammalian myocardium. Furthermore, the characteristics and contributors of E-C coupling at the earliest developmental stages are poorly understood. In this study, we measured [3H](+)PN200-110 (PN) dihydropyridine (DHP) binding capacity, functionality and expression of the L-type Ca2+ channel and cytosolic [Ca2+] ([Ca]i) at various developmental stages (3, 6, 10, 20 and 56 days old) in order to characterize ontogenetic changes in E-C coupling. We found that: 1) the whole cell L-type Ca2+ channel peak current (ICa) density increased slightly in parallel with cell growth, but the current-voltage (I-V) relationship, the steady-state activation and the maximum DHP binding (Bmax) and binding affinity (Kd) did not exhibit significant developmental changes; 2) sarcoplasmic reticulum (SR) Ca2+-dependence of inactivation rates of L-type Ca2+ channel and peak of ICa density were only observed after 10 days of age which temporally coincides with transverse (T)-tubule formation; 3) the relationship between [Ca2+]i and voltage changed from a linear relationship at the earliest developmental stages to a "bell-shaped" relationship at the later developmental stages; presumably corresponding to a switch from reverse mode NCX-dependent to ICa-dependent E-C coupling; and 4) the expression of two different splice variants of CaV1.2, IVS3A and IVS3B, switched from predominantly IVS3A at the earliest stages to IVS3B at the later developmental stages. Our data suggest that while the density of functional DHP receptors (DHPRs) increases only slightly during ontogeny, the enhancement of functional coupling between DHPR and ryanodine receptor (RyR) is dramatic between the second and third weeks after birth. Furthermore, we found that the differential expression of splice variants during development temporally correlated with the appearance of ICa-dependent E-C coupling and T-tubule formation.
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