The free intracellular calcium concentration of suspensions of isolated rat heart cells was monitored during sequential exposures to halothane and caffeine to evaluate cellular mechanisms of the negative inotropic effect of halothane. The calcium-sensitive, fluorescent dye quin2 was used as the indicator of free intracellular calcium. The acute addition of halothane in concentrations greater than or equal to 0.062 mM (0.19 vol%) to suspensions of quiescent rat heart cells at 37 degrees C caused a transient (approximately 1.5 min) increase in free intracellular calcium concentration. The intracellular calcium concentration after the decay of this transient was not detectably different from that prior to the addition of halothane. Neither the reduction of extracellular calcium from 1 mM to 100 nM, nor the prior addition of verapamil (5 microM) decreased this halothane-induced calcium transient. The transient was completely blocked by the prior addition of 10 mM caffeine, which depletes the sarcoplasmic reticulum of calcium. Also, the prior addition of halothane caused a reduction in the calcium transient due to caffeine. The depression of the caffeine-induced calcium transient by halothane was independent of the time interval (up to 4 min) between the additions of halothane and caffeine. These results indicate that halothane causes a net loss of calcium from the sarcoplasmic reticulum of quiescent rat heart cells. Thus, halothane has a direct effect at the sarcoplasmic reticulum, probably an enhancement of calcium release, which may explain its depression of myocardial contractility.