The molecular mechanism of the Ca2(+)-induced permeabilization of rat liver mitochondria was evaluated by studying a new effect of the commonly used general anaesthetic Propofol (2,6-diisopropylphenol). The compound was found to induce an apparent uptake of Ca2+ at steady-state in the Ca2(+)-distribution between the medium and the mitochondria, and to inhibit swelling and release of accumulated Ca2+ induced by inorganic phosphate, t-butyl hydroperoxide, diamide or FCCP plus Ruthenium red. The compound did not stimulate the activity of the Ca2(+)-uniporter and it is concluded that the effects seen are due to the inhibition of the Ca2(+)-dependent, unspecific permeability increase. The results suggest two mechanisms whereby Propofol stabilizes the mitochondrial membrane in the presence of Ca2+: (i) by interaction with the putative pore, thus causing its closure; and (ii) by scavenging of free radicals thus inhibiting its opening during oxidative stress.