Sustained subthreshold-for-twitch depolarization in rat single ventricular myocytes causes sustained calcium channel activation and sarcoplasmic reticulum calcium release

Single rat ventricular myocytes, voltage-clamped at -50 to -40 mV, were depolarized in small steps in order to define the mechanisms that govern the increase in cytosolic [Ca²⁺] (Ca_i) and contraction, measured as a reduction in myocyte length. Small (3-5 mV), sustained (seconds) depolarizations that caused a small inward or no detectable change in current were followed after a delay by small (<2% of the resting length), steady reductions in cell length measured via a photodiode array, and small, steady increases in Ca_i measured by changes in Indo-1 fluorescence. Larger (greater than - 30 and less than - 20 mV), sustained depolarizations produced phasic Ca²⁺ currents, Ca_i transients, and twitch contractions, followed by a steady current and a steady increase in Ca_i and contraction. Nitrendipine (or Cd, verapamil, or Ni) abolished the steady contraction and always produced an outward shift in steady current. The steady, nitrendipine-sensitive current and sustained increase in Ca_i and contraction exhibited a similar voltage dependence over the voltage range between - 40 and -20 mV. 2 µM ryanodine in the presence of intact Ca²⁺ channel activity also abolished the steady increase in Ca_i and contraction over this voltage range. We conclude that when a sustained depolarization does not exceed about -20 mV, the resultant steady, graded contraction is due to SR Ca²⁺ release graded by a steady ("window") Ca²⁺ current. The existence of appreciable, sustained, graded Ca²⁺ release in response to Ca²⁺ current generated by arbitrarily small depolarizations is not compatible with any model of Ca²⁺-induced Ca²⁺ release in which the releasing effect of the Ca²⁺ channel current is mediated solely by Ca²⁺ entry into a common cytosolic pool. Our results therefore imply a distinction between the triggering and released Ca²⁺ pools.