Characterisation of the effects of SR146131, a novel non-peptide CCK1 receptor agonist, on IMR-32 human neuroblastoma cells

Abstract The effect of {2-[4-(4-chloro-2,5-dimethoxy-phenyl)-5-[2-cyclohexyl-ethyl)-thiazol-2-ylcarbamoyl]-5,7-dimethyl-indol-1-yl}-acetic acid (SR146131), a novel non-peptide agonist of cholecystokinin (CCK) CCK 1 receptors, was compared to the effect of sulphated cholecystokinin octapeptide (CCK-8-S) on CCK 1 receptors of the human neuroblastoma cell line IMR-32. SR146131 inhibited [ 125 I]CCK-8-S binding to IMR-32 cells at nanomolar concentrations. SR146131 and CCK-8-S increased intracellular free Ca 2+ levels ([Ca 2+ ] i ) in the same concentration range (EC 50 =6±2.3 and 1.3±0.14 nM, respectively). Although the shape of the [Ca 2+ ] i increase induced by CCK-8-S and SR146131 was slightly different, extracellular Ca 2+ removal affected the response of both compounds to a similar degree, and the response of both compounds was essentially due to Ca 2+ release from intracellular stores. This was also confirmed by measuring the [Ca 2+ ] i response of single cells: both compounds induced [Ca 2+ ] i oscillations at subnanomolar concentrations and elicited a large peak increase in [Ca 2+ ] i at higher concentrations (EC 50 =0.5±0.04 and 5.7±1.9 nM for CCK-8-S and SR146131, respectively). Both CCK-8-S and SR146131 induced a sustained increase of phosphoinositide turnover in these cells, and acted at similar concentrations (EC 50 =2.7±0.7 and 6±3.1 nM, respectively), although the maximal effect of SR146131 was somewhat lower than the effect of CCK-8-S. These data show that SR146131 activates human CCK 1 receptors on IMR-32 cells in a manner and with a potency similar to that of CCK-8-S.