Sodium- and potassium-activated adenosine triphosphatase (Na,K-ATPase) activity and [3H]-ouabain binding were examined in homogenates of cerebral cortex, striatum, and hypothalamus of 3-, 8-, and 26-month-old rats to determine if aging-related alterations in energy utilization demonstrated in brain slices and homogenates are potentially associated with alterations in Na,K-ATPase. There were no consistent age-related changes seen in Na,K-ATPase activity, the number of [3H]-ouabain binding sites, or their affinity for ouabain. Moreover, enzyme activities of the two molecular forms of Na,K-ATPase and their inhibition by strophanthidin did not appear to be different in partially purified enzyme preparations obtained from whole brain of 3- and 26-month-old rats. In contrast, the concentration of [3H]-ouabain binding sites was lower in cardiac muscle of senescent rats indicating a reduction in the number of active Na,K-ATPase units. It appears unlikely that aging-related central nervous system changes are associated with alterations in the Na,K-ATPase enzyme system.