CALCIUM-PERMEABLE CHANNELS IN RAT HEPATOMA-CELLS ARE ACTIVATED BY EXTRACELLULAR NUCLEOTIDES

Extracellular ATP is known to cause uptake of Ca2+ by rat liver cells. The specific pathway permitting influx of Ca2+ has not yet been identified. In the present investigations, we studied the properties of ATP-evoked Ca-45(2+) uptake in rat hepatoma cell monolayers and then used patch-clamp electrophysiology to identify the channel that may account for this uptake. The results suggest that ATP-stimulated Ca-45(2+) uptake occurs as a result of P2-purinergic receptor interaction because uptake was inhibited by Reactive Blue (100-mu-M), a blocker of this type of receptor. Furthermore, the ability of other adenine nucleotides to stimulate Ca-45(2+) uptake was related to the selectivity sequence for binding to the P2-purinergic receptor. ATP-stimulated Ca-45(2+) uptake occurs primarily through a conductance pore since it was inhibited by 70% upon dissipation of the membrane potential using the K+ ionophore valinomycin. The calcium channel blockers nifedipine and verapamil failed to inhibit Ca-45(2+) uptake, but gadolinium (GdCl3) was an effective blocker. In cell-attached patch-clamp experiments, a single type of channel was activated with ATP (100-mu-M) addition to the bath in 18 of 32 trials. The current-voltage relationship of the ATP-activated channel is identical to that of the stretch-activated channel previously characterized in this laboratory as a calcium-permeable cation-nonselective channel [Am. J. Physiol. 258 (Cell Physiol. 27): C421-C428, 1990]. There are several lines of evidence which suggest that this cation-nonselective channel may account for ATP-stimulated Ca-45(2+) uptake. Both ATP-stimulated Ca-45(2+) uptake and cation channel activation are transient in nature, commencing immediately after ATP addition and reaching a peak at 1 min or less. As in the case of ATP-evoked Ca-45(2+) uptake, the Ca2+-channel blocker GdCl3 inhibited ATP-induced channel activation. ATP-activated channel opening was inhibited in presence of Reactive Blue, suggesting that channel activation was dependent on purinergic receptor interaction. In conclusion, we suggest that a nonselective cation channel may provide the route for ATP-evoked Ca2+ influx in rat hepatoma cells.