Muscle growth of low birth weight (LBW) piglets may be improved with adapted nutrition. This study elucidated effects of glutamine (Gln) supplementation on the cellular muscle development of LBW and normal birth weight (NBW) piglets. Male piglets (n = 144) were either supplemented with 1 g Gln/kg body weight or an isonitrogeneous amount of alanine (Ala) between postnatal day 1 and 12 (dpn). Twelve piglets per group were slaughtered at 5, 12 and 26 dpn, one hour after injection with Bromodeoxyuridine (BrdU, 12 mg/kg). Muscle samples were collected and myogenic cells were isolated and cultivated. Expression of muscle growth related genes was quantified with qPCR. Proliferating, BrdU-positive cells in muscle sections were detected with immunohistochemistry indicating different cell types and decreasing proliferation with age. More proliferation was observed in muscle tissue of LBW-GLN than LBW-ALA piglets at 5 dpn, but there was no clear effect of supplementation on related gene expression. Cell culture experiments indicated that Gln could promote cell proliferation in a dose dependent manner, but expression of myogenesis regulatory genes was not altered. Overall, Gln supplementation stimulated cell proliferation in muscle tissue and in vitro in myogenic cell culture, whereas muscle growth regulatory genes were barely altered.