Microdomain Ca 2+ near ryanodine receptors as reported by L‐type Ca 2+ and Na + /Ca 2+ 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.