The administration of glucagon or N6O2'-dibutyryl cyclic AMP to fed rats by intraperitoneal injection was associated with a 2-fold increase in the amounts of endogenous Pi and ATP, and an increase in the rate and extent of transport of exogenous Pi (measured in either the presence or the absence of Ca2+) in mitochondria subsequently isolated from the liver. No change was observed in either the maximum rate of transport of exogenous Pi or in the rate of 32Pi exchange. The changes induced by glucagon and dibutyryl cyclic AMP were markedly decreased by the co-administration of cycloheximide. The administration of insulin to rats resulted in an increase of about 1.3-fold in the concentration of endogenous mitochondrial Pi The amounts of endogenous Pi in mitochondria isolated from the livers of starved rats were 3 times those in mitochondria isolated from fed animals. It is concluded that the liver mitochondrial phosphate-transport system may be an important site of hormone action. In the course of these experiments, it was shown that Ca2+ markedly stimulates mitochondrial phosphate transport.