Environmental Circadian Modulators Target Melatonin Receptors in Pancreatic β‐Cells

Abstract

In silico 3D modelling and 2-[125I]-iodomelatonin binding to human MT1 and MT2 melatonin receptors identified the carbamate insecticides, carbaryl and carbofuran, as potential melatonin receptor circadian modulators (M P-G et al., EB Abstract, 2014). In humans, signaling through MT1 and MT2 melatonin receptors modulates insulin secretion from pancreatic β-cells (Int J Mol Sci 14: 6981, 2013). This study determined the pharmacological profile of melatonin receptors and the effect of carbamate insecticides in rat insulinoma cells (INS-1), a model for pancreatic β-cells. Saturation binding with 2-[125I]-Iodomelatonin (1 to 1000 pM) to INS-1 cell membranes yielded a KD of 50.2 ± 13.2 pM and a Bmax of 10.8 ± 2.8 fmol/mg protein (n=5). Competition binding of melatonin and the competitive melatonin receptor antagonist luzindole for 2-[125I]-iodomelatonin binding to INS-1 cell membranes yielded Ki of 83 ± 18 pM (n=9) and 0.50 ± 0.14 µM (n=4), respectively. The affinity of luzindole and the reported low to undetectable levels of MT2 receptor expression in INS-1 cells, suggest that 2-[125I]-iodomelatonin binds primarily to MT1 receptors. The Ki for carbaryl was 2.59 ± 0.54 µM (n=7) while concentrations as high as 1 mM of carbofuran did not compete for 2-[125I]-iodomelatonin binding to INS-1 cell membranes. The presence of GTP (100 µM) did not alter the competition of carbaryl for 2-[125I]-iodomelatonin binding (Ki = 5.39 ± 0.19 µM, n=4). These results suggest that carbaryl may potentially block melatonin receptors in INS-1 cells. Supported by ES 023684 to MLD and RVR.