Vol. 130, No. 1, 1985 BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNKATIONS Julv 16, 1985 Pages 9-15 INHIBITION OF INSULIN RECEPTOR BINDING BY A23187: SYNERGY WITH PHORBOL ESTERS Mustapha ROUIS', Pierre THOMOPOULOS', Catherine Cherier' and ugo TESTA' 'INSERM U-282 and 'U-91, Haoital Henri Mondor, 94010 Cr@teil, France Received May 6, 1985 The ionophore A23187 inhibits the binding of insulin on U-937 monocyte-like cells which had been induced to diffe- rentiate by la, 25-(OH)2 cholecalciferol, while it remains inactive on the undifferentiated cells. A 50% reduction of the specific binding of '251-insulin is observed within 20 to 30 min at 37°C. The effect is obtained at 10-'M to 10s6M A23187. It is reversible in 60 min at 37°C. A suboptimal con- centration of the ionophore potentiates the inhibitory action of phorbol esters on insulin binding. 1965 Ac.idtmic Prvc,i. Inc. The function of the insulin receptor is regulated by various hormones or cellular signalling mechanisms. Insulin itself is a major and permanent controlling factor, through the well-known phenomenon of down-regulation of the receptors (1). In addition, phorbol esters (2,3) probably through the stimulation of protein kinase C (4) and fi-adrenergic agonists, through a cyc ic AMP-dependent process (5-7), induce a rapid inhibition of the binding ability of insulin receptors. In the present work, we investigated the role of cyto- solic calcium in regulating the binding of insulin. Our study has been con d ucted in the human monocyte-like cell line U-937 (8). These cells possess insulin receptors which are sensitive to the inhibitory effect of phorbol esters (2,9). We took advantage of the recently described property of ICC, 25-(OH)2 cholecalciferol to induce the differentiation of U-937 cells to monocytes, while it increases the binding of insulin (10). Our data demonstrate that cytosolic calcium is able to regulate the function of the hormone receptors in the differentiated U-937 cells, while it remains without effect in the uninduced cells. This phenomenon is synergistic with the action of phorbol esters on the binding of insulin.