The goal was to test whether isoflurane exerts its depressant effect on the heart by mainly affecting the intracellular Ca2+ transient [Ca2+]i. Intact rat ventricular trabeculae, paced at 0.5 Hz and 30 degreesC with extracellular [Ca2+] ([Ca2+]o) of 2 mM, were used. The [Ca2+]i was monitored using fura 2 injected into the myoplasm. The sarcoplasmic reticulum (SR) Ca2+ content was estimated using rapid cooling with or without caffeine to induce Ca2+ release and contracture. A plot of peak twitch force versus peak [Ca2+]i transient with increasing isoflurane concentration declines linearly so that a 56% reduction in the peak [Ca2+]i transient would abolish twitch force. This relationship is intermediate between those obtained with lowering [Ca2+]o, which depresses twitch force through a reduction of the [Ca2+]i transient, and adding 2,3-butanedione monoxime, which reduces the responsiveness of the contractile system to [Ca2+]i. The isoflurane effect is different from that of halothane with respect to both the above relationship and the rapid-cooling response. Isoflurane abolishes the ability of rapid cooling to liberate Ca2+ from the SR.