Cardiomyocytes generating spontaneous Ca2+-transients as tools for precise estimation of sarcoplasmic reticulum Ca2+ transport

Abstract

Spontaneous Ca²⁺-transient (wave) generation in isolated cardiomyocytes is well established phenomenon which poses a lot of questions about myocardial excitability. Current studies of spontaneous Ca²⁺-activity in cardiac cells mainly relate to the kinetic characteristics, classification and simulation of Ca²⁺-events through ryanodine receptor (RyR) activity modeling. Here, for the first time we pay attention to the Ca²⁺-transients having stationary kinetics for correct estimation of the sarcoplasmic reticulum Ca²⁺ transport. In cardiomyocytes generating such type of Ca²⁺-transients, the averaged intracellular calcium ([Ca²⁺]_in) fluorescence practically does not change in time. Stationary Ca²⁺-transients are observed in different animal models (Wistar, SHR, ground squirrels) revealing a common cardiomyocyte phenomenon. They somewhat depend on external Ca²⁺ ([Ca²⁺]_ex) because the [Ca²⁺]_ex lowering to 1 μM in the presence of EGTA disrupts Ca²⁺-wave propagation. At the same time, spontaneous Ca²⁺-transients do not associated with the forward or reverse mode of Na⁺/Ca²⁺ exchanger (NCX), but partially modulated by the L-type Ca²⁺-channels. Among the sarcoplasmic reticulum targets, RyR and SERCA are crucial for Ca²⁺-wave generation and sustained self-oscillation activity. Analysis of the spontaneous wave kinetics reveals that both slopes of the rising wave front and the wave front decline are gradually changed during propagation, which well correlates with the RyR and SERCA activity, respectively. On the contrary, in the electrical field-stimulated myocytes, both slope factors are sharply changed corresponding to 'all-or-nothing' rule, which is fundamental principle for action potential in cardiomyocytes. Furthermore, stimulation of single cardiomyocyte using local electrode appears the deterioration in the [Ca²⁺]_in utilization from the cytosol, which limits the time of observation during the protocol. Obtained data suggest that stationary spontaneous Ca²⁺-transients occurring without actual myocellular excitation represent useful and precise tools for estimation of the sarcoplasmic reticulum Ca²⁺-transport.

Keywords: Ca(2+)-transients; Cardiomyocyte; Ryanodine receptor; SERCA; Spontaneous waves; Voltage-gated calcium channels.