The present study investigated the inheritance of dietary fat, carbohydrate, and kilocalorie intake traits in an F(2) population derived from an intercross between C57BL/6J (fat-preferring) and CAST/EiJ (carbohydrate-preferring) mice. Mice were phenotyped for self-selected food intake in a paradigm which provided for 10 days a choice between two macronutrient diets containing 78/22% of energy as a composite of either fat/protein or carbohydrate/protein. Quantitative trait locus (QTL) analysis identified six significant loci for macronutrient intake: three for fat intake on chromosomes (Chrs) 8 (Mnif1), 18 (Mnif2), and X (Mnif3), and three for carbohydrate intake on Chrs 17 (Mnic1), 6 (Mnic2), and X (Mnic3). An absence of interactions among these QTL suggests the existence of separate mechanisms controlling the intake of fat and carbohydrate. Two significant QTL for cumulative kilocalorie intake, adjusted for baseline body weight, were found on Chrs 17 (Kcal1) and 18 (Kcal2). Without body weight adjustment, another significant kcal locus appeared on distal Chr 2 (Kcal3). These macronutrient and kilocalorie QTL, with the exception of loci on Chrs 8 and X, encompassed chromosomal regions influencing body weight gain and adiposity in this F2 population. These results provide new insight into the genetic basis of naturally occurring variation in nutrient intake phenotypes.