Sarcoplasmic reticulum and nuclear envelope are one highly interconnected Ca2+ store throughout cardiac myocyte.
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 (InsP(3)) receptors are separately localized on the nuclear envelope (NucEn) and involved in nuclear Ca(2+) signaling. Here, we find that both caffeine and InsP(3) receptor agonists deplete free [Ca(2+)] inside both SR and NucEn. Fluorescence recovery after photobleach (FRAP) was measured using the low-affinity Ca(2+) indicator Fluo-5N trapped inside the SR and NucEn (where its fluorescence is high because [Ca(2+)] 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 (tau&2 minutes) until fluorescence regained spatial uniformity. Notably, SR and NucEn fluorescence recovered simultaneously in the bleached end. Ca(2+) diffusion inside the SR-NucEn was also measured. SR Ca(2+)-ATPase was completely blocked but without acute SR Ca(2+) depletion. Then caffeine was applied locally to one end of the myocyte. In the caffeine-exposed end, free SR [Ca(2+)] ([Ca(2+)](SR)) declined abruptly and recovered partially (tau=20 to 30 seconds). In the noncaffeine end, [Ca(2+)](SR) gradually declined with a similar tau, until [Ca(2+)](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 Ca(2+) were estimated (&8 microm(2) sec(-1) and 60 microm(2) sec(-1)). This rapid luminal communication may maintain homogeneously high luminal [Ca(2+)], ensuring a robust and uniform driving force for local Ca(2+) release events from either SR or NucEn.
Wu
Xu
X
Department of Physiology, Loyola University Chicago, 2160 S First Ave, Maywood, IL 60153, USA.
Bers
Donald M
DM
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D004058:Diffusion; D007425:Intracellular Membranes / Q000648:ultrastructure; D008856:Microscopy, Fluorescence; D032383:Myocytes, Cardiac / Q000648:ultrastructure*; D009685:Nuclear Envelope / Q000378:metabolism*; D038761:Photobleaching; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
D002110:Caffeine; D002118:Calcium