Elementary Ca2+ release events in mammalian skeletal muscle: effects of the anaesthetic drug thiopental

Abstract

We examined the effect of clinically relevant doses of thiopental (10-100 microM) on Ca2+ release from the sarcoplasmic reticulum of chemically skinned skeletal muscle fibres of the mouse. Elementary Ca2+ release events (ECRE) were recorded with confocal microscopy and were detected and analysed by an automated algorithm. Thiopental at 25 microM evoked a marked increase in ECRE frequency (events/100 microm/s) from 0.64 +/- 0.32 to 1.56 +/- 0.38 (P < 0.001). Incubation with 5 microM ryanodine significantly reduced spontaneous and evoked ECRE frequencies to 0.08 +/- 0.08 (P < 0.001) and 0.39 +/- 0.25 (25 microM thiopental, P < 0.001) respectively. Thiopental-evoked ECRE show different morphological characteristics compared to spontaneous events. Maximum relative amplitudes (DeltaF/F0)max and spatial width (full width at half maximum) of the events were substantially increased. Full duration at half maximum was increased and some very long events (200 ms compared to approximately 30 ms standard) were produced. The rise times as an indicator of the channel open time were slightly increased. Furthermore, the occurrence of repetitive ECRE was observed. These events, in contrast to previous observations in amphibian skeletal muscle fibres, displayed a multitude of different release patterns. In particular, a repetitive ECRE mode with successively decaying amplitudes was identified and the inter-event intervals were analysed. Estimation of the underlying Ca2+ release current suggests that during repetitive events with a decaying amplitude a decreasing amount of Ca2+ was released within the individual release event. Possible underlying mechanisms are discussed. In summary, thiopental seems to be a potent RyR1 agonist and substantially alters the gating mechanisms of RyR Ca2+ release channel clusters already in clinically relevant doses, i.e. doses administered during general anaesthesia.