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Three-Dimensional Distribution of Cardiac Na⁺-Ca²⁺ Exchanger and Ryanodine Receptor during Development

Pauline Dan ^* , Eric Lin ^* , Jingbo Huang ^* , Perveen Biln ^*  and Glen F. Tibbits ^* 

^* Cardiac Membrane Research Laboratory, Simon Fraser University, Burnaby, British Columbia, Canada; and Cardiovascular Sciences, Child and Family Research Institute, Vancouver, British Columbia, Canada

Correspondence: Address reprint requests to Glen F. Tibbits, PhD, Cardiac Membrane Research Laboratory, Kinesiology, Simon Fraser University, Burnaby, BC, Canada V5A 1S6. Tel.: 604-291-3658; Fax: 604-291-3040; E-mail: tibbits{at}sfu.ca.

Mechanisms of cardiac excitation-contraction coupling in neonates are still not clearly defined. Previous work in neonates shows reverse-mode Na⁺-Ca²⁺ exchange to be the primary route of Ca²⁺ entry during systole and the neonatal sarcoplasmic reticulum to have similar capability as that of adult in storing and releasing Ca²⁺. We investigated Na⁺-Ca²⁺ exchanger (NCX) and ryanodine receptor (RyR) distribution in developing ventricular myocytes using immunofluorescence, confocal microscopy, and digital image analysis. In neonates, both NCX and RyR clusters on the surface of the cell displayed a short longitudinal periodicity of 0.7 µm. However, by adulthood, both proteins were also found in the interior. In the adult, clusters of NCX on the surface of the cell retained the 0.7-µm periodicity whereas clusters of RyR adopted a longer longitudinal periodicity of 2.0 µm. This suggests that neonatal myocytes also have a peri-M-line RyR distribution that is absent in adult myocytes. NCX and RyR colocalized voxel density was maximal in neonates and declined significantly with ontogeny. We conclude in newborns, Ca²⁺ influx via NCX could potentially activate the dense network of peripheral Ca²⁺ stores via peripheral couplings, evoking Ca²⁺-induced Ca²⁺ release.

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