Calcium (Ca2+) ions serve multiple roles both intra- and extracellularly. We recently cloned a cell surface, Cao(2+)-sensing receptor (CaR) that plays a central role in Cao2+ homeostasis by enabling direct regulation by Cao2+ of parathyroid hormone (PTH) secretion and the function of other tissues involved in mineral ion homeostasis. In parathyroid cells, the CaR activates phospholipase C, thereby raising the levels of inositol trisphosphate (IP3) and releasing Ca2+ from intracellular stores. High Cao2+ also activates Ca2+ influx into parathyroid cells through poorly defined mechanisms that may involve Ca(2+)-permeable, nonselective cation channels (NCC). We now show that human embryonic kidney (HEK293) cells also have NCC and, furthermore, that these channels are regulated by the CaR. We have utilized the cell-attached configuration of the patch clamp technique to characterize the properties of these channels as well as their regulation by various CaR agonists added to the external bath solution. The polycationic CaR agonist, neomycin (100 microM), as well as an elevated concentration of Cao2+ (3 mM), both of which activate the cloned CaR, significantly increased the probability of channel opening (Po) in HEK cells stably transfected with the CaR but not in nontransfected HEK cells which do not contain the receptor. Thus, the activation of the CaR enhances the activity of Ca(2+)-permeable NCC in these cells, which could contribute to the sustained increase in Cai2+ in parathyroid cells which is observed in response to elevated Cao2+. The CaR may also regulate the membrane functions of other CaR-expressing cells (e.g., those in the brain), at least in part, by modulating similar channels.