Insulin transported from plasma into the central nervous system (CNS) is hypothesized to contribute to the negative feedback regulation of body adiposity. Because CNS insulin uptake is likely mediated by insulin receptors, physiological interventions that impair insulin action in the periphery might also reduce the efficiency of CNS insulin uptake and predispose to weight gain. We hypothesized that high-fat feeding, which both reduces insulin sensitivity in peripheral tissues and favors weight gain, reduces the efficiency of insulin uptake from plasma into the CNS. To test this hypothesis, we estimated parameters for cerebrospinal fluid (CSF) insulin uptake and clearance during an intravenous insulin infusion using compartmental modeling in 10 dogs before and after 7 weeks of high-fat feeding. These parameters, together with 24-h plasma insulin levels measured during ad libitum feeding, also permitted estimates of relative CNS insulin concentrations. The percent changes of adiposity, body weight, and food intake after high-fat feeding were each inversely associated with the percent changes of the parameter k1k2, which reflects the efficiency of CNS insulin uptake from plasma (r = -0.74, -0.69, -0.63; P = 0.015, 0.03, and 0.05, respectively). These findings were supported by a non-model-based calculation of CNS insulin uptake: the CSF-to-plasma insulin ratio during the insulin infusion. This ratio changed in association with changes of k1k2 (r = 0.84, P = 0.002), body weight (r = -0.66, P = 0.04), and relative adiposity (r = -0.72, P = 0.02). By comparison, changes in insulin sensitivity, according to minimal model analysis, were not associated with changes in k1k2, suggesting that these parameters are not regulated in parallel. During high-fat feeding, there was a 60% reduction of the estimated CNS insulin level (P = 0.04), and this estimate was inversely associated with percent changes in body weight (r = -0.71, P = 0.03). These results demonstrate that increased food intake and weight gain during high-fat feeding are associated with and may be causally related to reduced insulin delivery into the CNS.