Reported here are the effects of a high-fat diet (HFD) fed to dams during pregnancy on the weight, beta- and alpha-cell development, and beta-cell function of their weanling offspring.Offspring were obtained from dams maintained on an HFD for the first, second, or third week of gestation or throughout gestation and then on a standard laboratory diet for the duration of lactation. Weanling weights and circulating glucose and insulin concentrations were measured on postnatal day 21, after which pancreata were excised and snap-frozen for quantitative polymerase chain reaction of glucokinase (GK) or processed for immunohistochemical examination and image analysis (beta- and alpha-cell volume, number, and size, and GK immunoreactivity).All of the weanlings had low body weights and were hypoinsulinemic. In weanlings maintained on an HFD for either the first, second, or third week of gestation, hyperglycemia and a reduction in beta-cell volume and number, in beta- and alpha-cell size, and in both GK messenger RNA expression and immunoreactivity were observed. The development of beta and alpha cells was normal in weanlings maintained on an HFD throughout gestation.Maintenance of dams on an HFD for any single week of gestation results in weanling offspring with an impairment in beta-cell development and function.