The hypothesis that vitamin content of the diet during gestation alters macronutrient choice, food intake and the expression of serotonin receptors and proopiomelanocortin (POMC) in the hypothalamus of the offspring was investigated. Pregnant Wistar rats (n = 10/group) were fed the AIN-93G diet containing a multivitamin mix at the recommended (RV) content or10-fold higher (high vitamin, HV) content. Male offspring were weaned to a choice of 10% and 60% casein diets. Intake regulation by the serotonergic system was determined by measuring food choice daily for 7 weeks, and following tryptophan (TRP) or mCPP (a serotonin receptor agonist) injections at 4 and 6 weeks post-weaning. mRNA expressions of hypothalamic serotonin receptor and POMC were measured at birth, weaning and sacrifice (7 weeks post-weaning). No differences were found in body weight at birth or weaning. HV offspring had lower food intake for the duration of the study (P < 0.001), and 11% lower body weight (P < 0.05) and 23% lower fat pad mass (P < 0.05) at 7 weeks post-weaning. They selected less protein following 12 h of food deprivation (P < 0.05) and were less responsive to TRP (P = 0.05) and mCPP (P < 0.05) injections at 6 weeks post-weaning. Expressions of mRNA for serotonin receptors 5-HT1A/2A/2C at weaning (P < 0.01) and of POMC at weaning and 7 weeks post-weaning (P < 0.05) were lower. In conclusion, intake of multivitamins above the requirements during pregnancy affected macronutrient choice, food intake and the expression of serotonin receptors and POMC in the hypothalamus.