Measurement of cytosolic calcium (Cai) in the parotid acinar cells is very important in understanding the cause of cystic fibrosis since the parotid cells of cystic fibrosis patients have 60% more calcium and respond to neurostimulants abnormally. The calcium sensitive aequorin seems to be the best indicator for measurement of Cai because it does not have the untoward effect (i.e. buffering Cai, altering the kinetics of Cai transporters, self-quenching etc.) of Quin 2 and Fura 2. Since aequorin must be bulk loaded into the cell, however, it is necessary to confirm that the loading procedure does not alter the normal parotid cell function. This grant application is to determine whether parotid cells can withstand the loading procedure. This will be done by comparing the cGMP, and ATP of the aequorin has altered the amount of amylase released by beta adrenergic or cholinergic stimulation. Finally, we shall determine if there are changes in the evoked enhancement of Ca-45 influx, Ca-45 efflux, cyclic AMP and cyclic GMP due to the presence of aequorin.
