Gestational diabetes mellitus (GDM) associates with increased L-arginine transport and extracellular concentration of adenosine in human umbilical vein endothelial cells (HUVECs). In this study we aim to determine whether insulin reverses GDM-increased L-arginine transport requiring adenosine receptors expression in HUVECs. Primary cultured HUVECs from full-term normal (n = 38) and diet-treated GDM (n = 38) pregnancies were used. Insulin effect was assayed on human cationic amino acid transporter 1 (hCAT1) expression (protein, mRNA, SLC7A1 promoter activity) and activity (initial rates of L-arginine transport) in the absence or presence of adenosine receptors agonists or antagonists. A 1 adenosine receptors (A 1 AR) and A 2A AR expression (Western blot, quantitative PCR) was determined. Experiments were done in cells expressing or siRNA-suppressed expression of A 1 AR or A 2A AR. HUVECs from GDM exhibit higher maximal transport capacity (maximal velocity (V max )/ apparent Michaelis Menten constant (K m ), V max /K m ), which is blocked by insulin by reducing the V max to values in cells from normal pregnancies. Insulin also reversed the GDMassociated increase in hCAT-1 protein abundance and mRNA expression, and SLC7A1 promoter activity for the fragment −606 bp from the transcription start point. Insulin effects required A 1 AR, but not A 2A AR expression and activity in this cell type. In the absence of insulin, GDM-increased hCAT-1 expression and activity required A 2A AR expression and activity. HUVECs from GDM pregnancies exhibit a differential requirement of A 1 AR or A 2A AR depending on the level of insulin, a phenomenon that represent a condition where adenosine or analogues of this nucleoside could be acting as helpers of insulin biological effects in GDM.