Gestational diabetes coincides with elevated circulating progesterone levels. We show that progesterone accelerates the progression of diabetes in female db/db mice. In contrast, RU486, an antagonist of the progesterone receptor (PR), reduces blood glucose levels in both female WT and db/db mice. Furthermore, female, but not male, PR−/− mice had lower fasting glycemia than PR+/+ mice and showed higher insulin levels on glucose injection. Pancreatic islets from female PR−/− mice were larger and secreted more insulin consequent to an increase in β-cell mass due to an increase in β-cell proliferation. These findings demonstrate an important role of progesterone signaling in insulin release and pancreatic function and suggest that it affects the susceptibility to diabetes.