Liver cells respond to changes in extracellular calcium (Ca2+o). The hepatic functions affected include bile secretion, metabolic activity, liver regeneration, and the response to xenobiotics. In the present study, we demonstrate the presence, in the liver, of the extracellular calcium-sensing receptor (CASR), previously described in the parathyroid and thyroid glands and kidney. CASR mRNA was specifically expressed in hepatocytes and was absent in non-parenchymal liver cells (stellate, endothelial and Kupffer cells). Western blot analysis using a specific CASR antibody showed staining in both whole liver and hepatocyte extracts. Immunohistochemistry and in situ hybridization of rat liver sections showed expression of CASR protein and mRNA by a subset of hepatocytes. The known agonists of the CASR, gadolinium (Gd3+; 0.5-3.0 mM) and spermine (1.25-20 mM), in the absence of Ca2+o, elicited dose-related increases in intracellular calcium (Ca2+ i) in isolated rat hepatocytes loaded with Fura 2-AM. There was a greatly attenuated response to a second challenge with either agonist. The response was also abrogated when IP3-sensitive calcium pools had been depleted by pretreatment with either thapsigargin or phenylephrine, an a 1-adrenergic receptor agonist known to mobilize Ca2+i from IP3-sensitive pools. Addition of the deschloro-phenylalkylamine compound, NPS-R467, but not the S enantiomer, NPS-S467, increased the sensitivity of the Ca2+i mobilization response to 1.25 mM spermine. Bile flow ceased after Ca2+o withdrawal and its recovery was enhanced by spermine in isolated perfused liver preparations. The CASR agonists Ca2+ and Gd3+ increased bile flow and the response to a submaximal Ca2+ concentration was enhanced by NPS-R467 but not the S compound. Thus, the data indicate that rat hepatocytes harbor a CASR capable of mobilizing Ca2+i from IP3- sensitive stores and that activation of the CASR stimulates bile flow.