An intrauterine hyperglycemic environment has long-lasting effects on the offspring. Recent studies focused on fetal tissues, whereas we studied the development and molecular alteration of the placenta. By intercrossing male and female adult control (C) and first-generation offspring mice with gestational diabetes mellitus (F1-GDM), we obtained four groups of second generation (F2) offspring: 1) C♂-C♀, 2) C♂-GDM♀, 3) GDM♂-C♀, 4) GDM♂- GDM♀. Placental weights in F1-GDM offspring were lower than in the control group. Placental weights in F2-offspring decreased through the paternal line. Placental RNA was extracted and analyzed using microarrays on day18.5 of pregnancy. This revealed 35 upregulated imprinted genes and 10 down-regulated imprinted genes. Dlk1and Gtl2 were especially down-regulated and up-regulated, respectively, due to their abnormal methylation status. These findings suggest that intrauterine hyperglycemia decreased placental weight in the first generation, and this was transmitted paternally to the second generation in mice. They also suggest intrauterine hyperglycemia leads to abnormal placental Dlk1-Gtl2 expression due to DNA methylation in first and second generation mice.