Radioimmunoassay for Rat Pancreatic a-Amylase and the Effect of Phe-Met-Arg-Phe-Amide on Amylase Secretion in the Isolated Perfused Rat Pancreas

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Abstract
In this study a radioimmunoassay was developed to measure secreted amylase from the isolated perfused rat pancreas. Using Sephadex G-75 gel chromatography, rat pancreatic amylase was purified to a single migrating protein band as determined by SDS polyacrylamide gel electrophoresis. Specificity of a rat pancreatic amylase antiserum, raised in rabbits, was determined using immunodiffusion, immunoelectrophoresis, and immunoblotting techniques. Secreted amylase concentrations, obtained using the radioimmunoassay, were not significantly different than those measured with the amylase enzyme assay. The rat pancreatic amylase radiommunoassay was used to measure the amylase secretion in the isolated perfused rat pancreas. PheMet-Arg-amide (FMRF-NH2) immunoreactivity has been shown to be colocalized with pancreatic polypeptide in the rat pancreatic islet, and evidence suggest that islet peptides modulate amylase secretion from the exocrine pancreas. In the present study, FMRF-NH2 significantly (p < 0.05) suppressed cholecystokinin (CCK)-stimulated amylase secretion by 55%. The average pancreatic amylase secretion in response to CCK was 10.89 .+-. 2.0 .mu.g/ml/min (n = 6); with the addition of FMRF-NH2, CCK-stimulated amylase secretion was reduced to 4.79 .+-. 1.6 .mu.g/ml/min (n = 6). These results are consistent with the insuloacinar hypothesis in that an FMRF-NH2-like substance in the islet may act to modulate the exocrine pancreas.

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