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Expression of insulin receptor on clonal pancreatic alpha cells and its possible role for insulin-stimulated negative regulation of glucagon secretion


In pancreatic alpha cells, the existence and function of the insulin receptor has not yet been fully established. In this study, to confirm the expression of functional insulin receptors in pancreatic alpha cells, we performed: 1) insulin receptor binding assay, 2) Northern blot analysis and RT-PCR (reverse transcription-polymerase chain reaction) amplification of insulin receptor mRNA, 3) immunocytochemical staining, 4) biosynthetic labelling of insulin receptor protein using [35S]methionine, 5) analysis of insulin-stimulated autophosphorylation of the insulin receptor in glucagon secreting cell lines, In-R1-G9 and αTC clone 6 cells. Glucagon secretion decreased with the addition of insulin in both cells. The receptor binding studies using [125I-Tyr-A14] insulin revealed that both cells possessed a significant number of insulin receptors (In-R1-G9: K1=2.1×109mol/l−1, K2=6.2×107 mol/l−1, R1=0.2×104, R2=1.86×104 sites/cell; αTC clone 6: K1=2.1×109 mol/l−1, K2=7.3×107 mol/l−1, R1=0.27×104, R2=1.95×104 sites/cell). Northern blot analysis as well as RT-PCR amplification showed the mRNA specific for insulin receptor in both cells. By immunocytochemical staining using anti-insulin receptor α-subunit antibody, positive immunostaining for insulin receptor was observed in both cells. [35S]Methionine labelling of both cells followed by immunoprecipitation using anti-insulin receptor antibody showed the correct size of the insulin receptor protein. The insulin receptor expressed in these cells underwent autophosphorylation by insulin stimulation. It is concluded that functional insulin receptors are properly expressed in In-R1-G9 and αTC clone 6 cells.



Reverse transcriptase-polymerase chain reaction


fetal bovine serum


Dulbecco's modified Eagle's medium


bovine serum albumin


phosphate buffered saline


sodium dodecyl sulphate


saline sodium citrate


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Correspondence to Dr. K. Kisanuki.

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Kisanuki, K., Kishikawa, H., Araki, E. et al. Expression of insulin receptor on clonal pancreatic alpha cells and its possible role for insulin-stimulated negative regulation of glucagon secretion. Diabetologia 38, 422–429 (1995). https://doi.org/10.1007/BF00410279

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Key words

  • Pancreatic alpha cell
  • In-R1-G9
  • αTC clone 6
  • insulin receptor
  • glucagon secretion