AGONIST-SPECIFIC CALCIUM SIGNALING AND PHOSPHOINOSITIDE HYDROLYSIS INHUMAN SK-N-MCIXC NEUROEPITHELIOMA CELLS

Fura-2 digital imaging microfluorimetry was used to evaluate the Ca2signals generated in single clonal human neuroepithelioma cells (SK-N- MCIXC) in response to agonists that stimulate phosphoinositide hydroly sis. Addition of optimal concentrations of either endothelin-1 (ET-1), ATP, oxotremorine-M (Oxo-M), or norepinephrine (NE) all resulted in a n increase in the concentration of cytosolic calcium (Ca-i(2+)) but of different magnitudes (ET-1 = ATP > Oxo-M > NE). The Ca2+ signals elic ited by the individual agonists also differed from each other in terms of their latency of onset, rate of rise and decay, and prevalence of a sustained phase of Ca2+ influx. The Ca2+ signals that occurred in re sponse to ATP had a shorter latency and more rapid rates of rise and d ecay than those observed for the other three agonists. Furthermore, a sustained plateau phase of the Ca2+ signal, which was characteristic o f the response to Oxo-M, was observed in <40% of cells stimulated with ET-1 and absent from Ca2+ signals elicited after NE addition. Removal of extracellular Ca2+ enhanced the rate of decay of Ca2+ signals gene rated by ATP, ET-1, or Oxo-M and, when evident, abolished the sustaine d phase of Ca2+ influx. In the absence of extracellular Ca2+, NE elici ted asynchronous multiple Ca2+ transients. In either the absence or pr esence of extracellular Ca2+, >94% of cells responded to ET-1 or ATP, whereas corresponding values for Oxo-M and NE were similar to 74 and s imilar to 48%. Sequential addition of agonists to cells maintained in a Ca2+-free buffer indicated that each ligand mobilized Ca2+ from a co mmon intracellular pool. When monitored as a release of a total inosit ol phosphate fraction, all four agonists elicited similar (four- to si xfold) increases in phosphoinositide hydrolysis. However, the addition of ET-1 or ATP resulted in larger increases in the net formation of i nositol 1,4,5-trisphosphate than did either Oxo-M or NE. These results indicate that, in SK-N-MCIXC cells, the characteristics of both Ca2signaling and inositol phosphate production are agonist specific.