Molecular and functional characterization of insulin receptors present on hamster XXXX cells.

@entity1 @entity2755 @entity8277 @entity12983
Studies using pancreas perfusion techniques point to a physiological inhibition of glucagon release by insulin which should be mediated by A cell-residing insulin receptors. In this study, we have characterized the insulin receptors expressed in a hamster @entity12983 A cell line (INR1G9 cells) which is an accepted tool for A cell studies. In receptor binding assays 125I-insulin was displaced with a Kd of 3 nmol/l. Binding was also dependent upon time, temperature and cell number. Insulin concentration-dependently inhibited glucagon secretion (1 mumol: 59%, 100 nmol/l: 71%, 10 nmol/l: 86% of controls). In transient transfection experiments insulin inhibited proglucagon gene transcription (controls: 100%, 100 nmol/l: 54%, 10 nmol/l: 57%, 1 nmol/l: 72%, 100 pmol/l: 96%). Treatment of INR1G9 cells with insulin for 20 h induced a strong downregulation of insulin receptors (controls: 100%, 100 nmol/l: 30%, 10 nmol/l: 70%, 1 nmol/l: 73%, 100 pmol/l: 75%) and of @entity8277 mRNA levels (controls: 100%, 100 nmol/l: 42%, 10 nmol/l: 82%, 1 nmol/l: 84%, 100 pmol/l: 90%). When INR1G9 cells were transiently transfected with a hybrid gene containing the promotor/enhancer region of the @entity1 @entity8277 promotor (1,462 bp) linked to the transcriptional reporter gene @entity2755 acetyltransferase and were treated with insulin it was demonstrated that insulin did not affect the @entity8277 gene transcription. In conclusion, INR1G9 cells express specific receptors for insulin. Insulin inhibits glucagon secretion and proglucagon gene expression via an inhibition of proglucagon gene transcription. Ligand-induced downregulation of the @entity8277 is not mediated by changes of @entity8277 gene transcription and is most likely regulated by posttranscriptional mechanisms, e.g. destabilization of @entity8277 mRNA.
