The role of the adenine nucleotide translocase on Ca2+ homeostasis in mitochondria from brown adipose tissue was examined. It was found that in mitochondria incubated with 50 microM Ca2+, ADP was not needed to retain the cation, but it was required for strengthening the inhibitory effect of cyclosporin on membrane permeability transition as induced by menadione. In addition, carboxyatractyloside was unable to promote matrix Ca2+ release, even though it inhibits the ADP exchange reaction. However, when the Ca2+ concentration was increased to 150 microM carboxyatractyloside did induce Ca2+ release, and ADP favored Ca2+ retention. Determination of cardiolipin content in the inner membrane vesicles showed a greater concentration in brown adipose tissue mitochondria than that found in kidney mitochondria. It suggested that the failure of the adenine nucleotide translocase to influence membrane permeability transition depends on the lipid composition of the inner membrane.