The transport of L-carnitine by lactating rat mammary tissue has been examined. L-carnitine uptake by rat mammary tissue explants isolated from lactating rats, 3-4 days post partum, was via both Na+-dependent and Na+-independent pathways. The Na+-dependent pathway, the predominant route for L-carnitine uptake, was a saturable process: the Km and Vmax were, respectively, 132 microM and 201 pmol/2 h/mg of intracellular water. The Na+-independent pathway, which was non-saturable, had a coefficient of 0.26 microl/mg of intracellular water/2 h. The Na+-dependent component of L-carnitine uptake by mammary tissue explants was cis-inhibited by D-carnitine and acetyl-L-carnitine, but not by choline or taurine. In contrast, the Na+-independent component of L-carnitine uptake was not affected by any of these compounds. The uptake of L-carnitine by mammary tissue isolated from lactating rats, 10-12 days post partum, was qualitatively similar to that by mammary tissue taken from rats during the early stage of lactation. However, L-carnitine uptake was quantitatively lower: this was attributable to a reduction in the Na+-dependent component of L-carnitine uptake. L-Carnitine efflux from rat mammary tissue taken from animals 3-4 days post partum, consisted of at least two components; a fast extracellular component and a slow membrane-limited component. Reversing the trans-membrane Na+-gradient did not stimulate L-carnitine efflux suggesting that the Na+-dependent L-carnitine carrier operates with asymmetrical kinetics. A hyposmotic shock, hence cell-swelling, increased L-carnitine efflux from mammary tissue explants.