ABSTRACT
AIMS/HYPOTHESIS: Cell-cell coupling mediated by gap junctions formed from connexin (CX) contributes to the control of insulin secretion in the endocrine pancreas. We investigated the cellular production and localisation of CX36 and CX43, and gap junction-mediated beta cell coupling in pancreatic islets from rats of different ages, displaying different degrees of maturation of insulin secretion. METHODS: The presence and distribution of islet connexins were assessed by immunoblotting and immunofluorescence. The expression of connexin genes was evaluated by RT-PCR and quantitative real-time PCR. The ultrastructure of gap junctions and the function of connexin channels were assessed by freeze-fracture electron microscopy and tracer microinjection, respectively. RESULTS: Young and adult beta cells, which respond to glucose, expressed significantly higher levels of Cx36 (also known as Gjd2) than fetal and newborn beta cells, which respond poorly to the sugar. Accordingly, adult beta cells also showed a significantly higher membrane density of gap junctions and greater intercellular exchange of ethidium bromide than newborn beta cells. Cx43 (also known as Gja1) was not expressed by beta cells, but was located in various cell types at the periphery of fetal and newborn islets. CONCLUSIONS/INTERPRETATION: These findings show that the pattern of connexins, gap junction membrane density and coupling changes in islets during the functional maturation of beta cells.
Subject(s)
Connexins/metabolism , Insulin-Secreting Cells/metabolism , Islets of Langerhans/metabolism , Animals , Animals, Newborn , Connexin 43/genetics , Connexin 43/metabolism , Connexins/genetics , Female , Fluorescent Antibody Technique , Gap Junctions/metabolism , Immunoblotting , Insulin-Secreting Cells/ultrastructure , Islets of Langerhans/growth & development , Islets of Langerhans/ultrastructure , Male , Microscopy, Electron , Rats , Rats, Wistar , Reverse Transcriptase Polymerase Chain Reaction , Gap Junction delta-2 ProteinABSTRACT
Fetal and neonatal pancreatic islets present a lower insulin secretory response as compared with adult islets. Prolonged culturing leads to an improvement of the glucose-induced insulin secretion response in neonatal pancreatic islets that may involve regulation of gap junction mediated cell communication. In this study, we investigated the effect of culturing neonatal islet cells for varying periods of time and with different glucose medium concentrations on the cellular expression of the endocrine pancreatic gap junction associated connexin (Cx) 36 and Cx43. We report here that the 7-d culture induced upregulation of the expression of these junctional proteins in neonatal islets in a time-dependent manner. A correlation was observed between the increased mRNA and protein expression of Cx36 and Cx43 and the increased insulin secretion following islet culturing. In addition, increasing glucose concentration within the culture medium induced a concentration-dependent enhancement of Cx36 islet expression, but not of Cx43 expression in cultured neonatal islets. In conclusion, we suggest that the regulation of gap junctional proteins by culture medium containing factors and glucose may be an important event for the maturation process of beta cells observed at in vitro conditions.
