DETECTION OF INTRACELLULAR CALCIUM ELEVATIONS IN XENOPUS-LAEVIS OOCYTES - AEQUORIN LUMINESCENCE VERSUS ELECTROPHYSIOLOGY

Detection of receptor expression in Xenopus oocytes often relies upon functional coupling to second messengers such as Ca2+ or cyclic adenos ine monophosphate. To detect intracellular Ca2+, electrophysiological measurement of the endogenous Ca2+-activated chloride current (I-Cl(Ca )) is often used (Dascal, 1987). An alternative utilizes the Ca2+ sens ing, bioluminescent protein aequorin (Parker and Miledi (1986) Proc. R . Sec. Lend. B, 228: 307-315; Giladi and Spindel (1991) Bio Techniques , 10: 744-747). In the present study the sensitivities of aequorin and electrophysiology for detecting receptor-mediated Ca2+ transients wer e compared. Assays were performed on the same batches of oocytes using either animal serum or ligands of exogenous receptors to generate ino sitol 1,4,5-trisphosphate (InsP(3)) and ultimately elevate intracellul ar Ca2+. Signal amplitudes were controlled by titrating the concentrat ion of animal serum, or titrating the amount of receptor mRNA injected . Both assays detected signals with high concentrations of animal seru m, or with high receptor density. However, aequorin signals were not d etected in experiments with average I-Cl(Ca) current amplitudes below 200 nA. To further evaluate the differences between these two techniqu es, membrane current and bioluminescence were measured simultaneously. Results of these studies suggest that the signals differ due to the s patial distribution of aequorin, the chloride channels, and the calciu m release sites.