A repetitive mode of activation of discrete Ca2+ release events (Ca2+ sparks) in frog skeletal muscle fibres

1. Ca2+ release events (Ca2+ 'sparks'), which are believed to arise from th e opening of a sarcoplasmic reticulum (SR) Ca2+ release channel or a small cluster of such channels that act as a release unit, have been measured in single, frog (Rana pipiens) skeletal muscle fibres. 2. Under conditions of extremely low rates of occurrence of Ca2+ sparks we observed, within individual identified triads, repetitive Ca2+ release even ts which occurred at a frequency more than 100-fold greater than the prevai ling average event rate. Repetitive sparks were recorded during voltage-cla mp test depolarizations after a brief (0.3-2 s) repriming interval in fibre s held at 0 mV and in chronically depolarized, 'notched' fibres. 3. These repetitive events are likely to arise from the re-opening of the s ame SR Ca2+ release channel or release unit operating in a repetitive gatin g mode ('rep-mode') rather than from the random activation of multiple, ind ependent channels or release units within a triad. A train of rep-mode even ts thus represents a series of Ca2+ sparks arising from a single location w ithin the fibre. Rep-mode events are activated among different triads in a random manner after brief repriming. The frequency of repetitive events amo ng all identified events during voltage-clamp depolarization to 0 mV after brief repriming was 3.9 +/- 1.3%. The occurrence of repetitive events was n ot related to exposure of the fibre to laser illumination. 4. The events observed within a rep-mode train exhibited a relatively unifo rm amplitude. Analysis of intervals between identified events in triads exh ibiting rep-mode trains indicated similar variations of fluorescence as in neighbouring, quiescent triads, suggesting there was not a significant numb er of small, unidentified events at the triads exhibiting rep-mode activity . 5. The distribution of rep-mode interspark intervals exhibited a paucity of events at short intervals, consistent with the need for recovery from inac tivation before activation of the next event in a repetitive train. The mea n interspark interval of repetitive sparks during voltage-clamp depolarizat ions was 88 +/- 5 ms, and was independent of membrane potential. 6. The individual Ca2+ sparks within a rep-mode train were similar in avera ge amplitude and spatiotemporal extent to singly occurring sparks, suggesti ng a common mechanism for termination of the channel opening(s) underlying both types of events. The average properties of the sparks did not vary dur ing a train. The relative amplitude of a spark within a rep-mode was not co rrelated with its rise time. 7. Repetitive Ca2+ release events represent a mode of gating of SR Ca2+ rel ease channels which may be significant during long depolarizations and whic h may be influenced by the biochemical state of the SR ryanodine receptor C a2+ release channels.