Microdomain [Ca²⁺] near ryanodine receptors as reported by L-type Ca²⁺ and Na+/Ca²⁺ exchange currents

During Ca²⁺ release from the sarcoplasmic reticulum triggered by Ca²⁺ influx through L-type Ca²⁺ channels (LTCCs), [Ca²⁺] near release sites ([Ca²⁺]nrs) temporarily exceeds global cytosolic [Ca²⁺]. [Ca²⁺]nrs can at present not be measured directly but the Na+/Ca2+ exchanger (NCX) near release sites and LTCCs also experience [Ca²⁺]nrs. We have tested the hypothesis that ICaL and INCX could be calibrated to report [Ca²⁺]nrs and would report different time course and values for local [Ca²⁺]. Experiments were performed in pig ventricular myocytes (whole-cell voltage-clamp, Fluo-3 to monitor global cytosolic [Ca²⁺], 37◦C). [Ca²⁺]nrs-dependent inactivation of ICaL during a step to +10 mV peaked around 10 ms. For INCX we computationally isolateda current fraction activated by [Ca²⁺]nrs; values were maximal at 10 ms into depolarization. The recovery of [Ca²⁺]nrs was comparable with both reporters (>90% within 50 ms). Calibration yielded maximal values for [Ca²⁺]nrs between 10 and 15 μmol l⁻¹ with both methods. When applied to a step to less positive potentials (-30 to -20 mV), the time course of [Ca²⁺]nrs was slower but peak values were not very different. In conclusion, both ICaL inactivation and INCX activation, using a subcomponent analysis, can be used to report dynamic changes of [Ca²⁺]nrs. Absolute values obtained by these different methods are within the same range, suggesting that they are reporting on a similar functional compartment near ryanodine receptors. Comparable [Ca²⁺]nrs at +10 mV and -20 mV suggests that, although the number of activated release sites differs at these potentials, local gradients at release sites can reach similar values.