The purpose of this study was to determine the in vitro effect of ovine PRL (oPRL) on the dynamics of insulin secretion and dye coupling among islet B cells. The effect of oPRL (2 Î¼g/ml) on insulin secretion was time dependent and reached a maximum on day 4 when there was a 2.4-fold increase in insulin secretion from cultured neonatal rat islets (n = 6, p ≪ 0.001). When islets cultured in the presence of oPRL for 4 days were perifused, 300 mg/dl glucose stimulation resulted in insulin release of 131 Â± 20 Î¼U/ml Â· 100 Î¼g islet tissue as compared to control islets 94 Â± 20 Î¼U/ml-100 Î¼g islet tissue (n = 7, p ≪ 0.02). Stimulation of the islets with a linear 30â€ 250 mg/dl glucose gradient resulted in a threshold for glucose-stimulated insulin secretion of 73 Â± 6 mg/dl glucose for the oPRL treated islets (n = 7) as compared to a threshold of 123 Â± 6 mg/dl glucose for control islets (n = 7, p ≪ 0.001). Mean islet volume was unchanged after 4 days of oPRL treatment but was 34% greater after 8 days (n = 6, p ≪ 0.001). Dye coupling among central islet B cells was also increased after in vitro treatment with oPRL for 4 days. The mean projected area of dye spread was 2-fold greater in the oPRL treated islets (n = 33) in comparison to the control islets (n = 33, p ≪ 0.05). These results indicate that in vitro lactogen treatment, in the form of oPRL, alters insulin secretory behavior and B cell junctional communication and supports our hypothesis that lactogen, insulin secretion, and junctional communication among B cells are related.