Ca(2+) homeostasis in sealed t-tubules of mouse ventricular myocytes.
We have recently shown that in mouse ventricular myocytes, t-tubules can be quickly and tightly sealed during the resolution of hyposmotic shock of physiologically relevant magnitude. Sealing of t-tubules is associated with trapping extracellular solution inside the myocytes but the ionic homeostasis of sealed t-tubules and the consequences of potential transtubular ion fluxes remain unknown. In this study we investigated the dynamics of Ca(2+) movements associated with sealing of t-tubules. The data show that under normal conditions sealed t-tubules contain Ca(2+) at concentrations below 100μM. However, blockade of voltage-dependent Ca(2+) channels with 10μM nicardipine, or increasing extracellular concentration of K(+) from 5.4mM to 20mM led to several fold increase in concentration of t-tubular Ca(2+). Alternatively, the release of Ca(2+) from sarcoplasmic reticulum using 10mM caffeine led to the restoration of t-tubular Ca(2+) towards extracellular levels within few seconds. Sealing of t-tubules in the presence of extracellular 1.5mM Ca(2+) and 5.4mM extracellular K(+) led to occasional and sporadic intracellular Ca(2+) transients. In contrast, sealing of t-tubules in the presence of 10mM caffeine was characterized by a significant long lasting increase in intracellular Ca(2+). The effect was completely abolished in the absence of extracellular Ca(2+) and significantly reduced in pre-detubulated myocytes but was essentially preserved in the presence of mitochondrial decoupler dinitrophenol. This study shows that sealed t-tubules are capable of highly regulated transport of Ca(2+) and present a major route for Ca(2+) influx into the cytosol during sealing process.
Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, MI, USA.
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Moench
Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, MI, USA. Electronic address: alopatin@umich.edu.
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Lopatin
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000378:metabolism; D020013:Calcium Signaling; D002478:Cells, Cultured; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D004140:Dinitrophenols / Q000494:pharmacology; D006352:Heart Ventricles / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006706:Homeostasis; D017136:Ion Transport; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D009529:Nicardipine / Q000494:pharmacology; D011188:Potassium / Q000378:metabolism; D012508:Sarcolemma / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D014475:Uncoupling Agents / Q000494:pharmacology
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004140:Dinitrophenols; D014475:Uncoupling Agents; D002110:Caffeine; D009529:Nicardipine; D011188:Potassium; D002118:Calcium
Ca(2+); Calcium; Hyposmotic stress; T-tubules; Ventricular myocytes