Fluorometric measurement of the intracellular free Ca(2+)-concentration in the ciliate Didinium nasutum using Fura-2.
We developed an experimental approach to measure somatic and ciliary Ca(2+)-signals in the ciliate Didinium under voltage clamp conditions using the dye Fura-2. Intracellular pressure injection of Fura-2 molecules did not alter electrophysiological membrane properties besides an expected buffering effect. The intracellular free Ca(2+)-concentration was determined at 2 x 10(-7) M. During membrane excitation, this resting value increased in the cilia; a quantification was not feasible. Within the cell soma, however, the Ca(2+)-level was unchanged within the physiological range of the membrane potential (-70 mV to 0 mV). Increasing the driving force for Ca(2+)-ions via strong hyperpolarization (potentials negative to -200 mV) a centripetal increase in the somatic Ca2+ concentration was found. Our results support the hypothesis that Ca2+ is the intracellular messenger in rapid electromotor coupling in ciliates.
Pernberg
J
J
Arbeitsgruppe Zelluläre Erregungsphysiologie, Fakultät für Biologie, Ruhr-Universität Bochum, Germany.
Machemer
H
H
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D016798:Ciliophora / Q000737:chemistry*; D003593:Cytoplasm / Q000378:metabolism; D005456:Fluorescent Dyes; D016257:Fura-2; D008027:Light; D008433:Mathematics; D008564:Membrane Potentials / Q000502:physiology; D008845:Microinjections; D018408:Patch-Clamp Techniques
D005456:Fluorescent Dyes; D002118:Calcium; D016257:Fura-2