Ca(2+) homeostasis in sealed t-tubules of mouse ventricular myocytes.

Journal of molecular and cellular cardiology (2014-05-03)
I Moench, A N Lopatin
ABSTRACT

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.

MATERIALS
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