Sarcoplasmic Reticulum and Nuclear Envelope Are One Highly Interconnected Ca2+ Store Throughout Cardiac Myocyte

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Abstract

Abstract—

Previous ventricular myocyte studies indicated that ryanodine receptors (RyRs) are in the sarcoplasmic reticulum (SR) and are critical in excitation–contraction coupling, whereas the inositol trisphosphate (InsP3) receptors are separately localized on the nuclear envelope (NucEn) and involved in nuclear Ca2+ signaling. Here, we find that both caffeine and InsP3 receptor agonists deplete free [Ca2+] inside both SR and NucEn. Fluorescence recovery after photobleach (FRAP) was measured using the low-affinity Ca2+ indicator Fluo-5N trapped inside the SR and NucEn (where its fluorescence is high because [Ca2+] is ≈1 mmol/L). After Fluo-5N photobleach in one end of the cell, FRAP occurred, accompanied by fluorescence decline in the unbleached end with similar time constants (τ≈2 minutes) until fluorescence regained spatial uniformity. Notably, SR and NucEn fluorescence recovered simultaneously in the bleached end. Ca2+ diffusion inside the SR-NucEn was also measured. SR Ca2+-ATPase was completely blocked but without acute SR Ca2+ depletion. Then caffeine was applied locally to one end of the myocyte. In the caffeine-exposed end, free SR [Ca2+] ([Ca2+]SR) declined abruptly and recovered partially (τ=20 to 30 seconds). In the noncaffeine end, [Ca2+]SR gradually declined with a similar τ, until [Ca2+]SR throughout the cell equalized. We conclude that the SR and NucEn lumen are extensively interconnected throughout the myocyte. Apparent intrastore diffusion coefficients of Fluo-5N and Ca2+ were estimated (≈8 μm2 sec−1 and 60 μm2 sec−1). This rapid luminal communication may maintain homogeneously high luminal [Ca2+], ensuring a robust and uniform driving force for local Ca2+ release events from either SR or NucEn.

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