AJP - Cell Physiology, Vol 263, Issue 6 C1302-C1309, Copyright © 1992 by American Physiological Society
Simultaneous fluorescence measurement of calcium and membrane potential responses to endothelin
S. G. Kremer, W. Zeng and K. L. Skorecki
Membrane Biology Group, University of Toronto, Ontario, Canada.
Vasopressin stimulates calcium signaling and chloride-dependent
depolarization in glomerular mesangial cells. We describe a technique
whereby both calcium and membrane potential changes can be simultaneously
monitored with fluorescent probes. This technique was validated by
comparison with single parameter measurements in basal and
vasopressin-stimulated mesangial cells. It was shown that the calibration
for calcium is unaffected by that for membrane potential, whereas the
calibration for membrane potential is affected by prior calcium
calibration. Accordingly, it was necessary to calibrate for the former
first. The technique was then applied to investigate the effects of
endothelin, which was found to elicit a concentration-dependent calcium
release response and a chloride-dependent depolarization of mesangial
cells. The interaction between the calcium signaling response to
vasopressin and endothelin was also investigated. When vasopressin
stimulation occurred subsequent to endothelin stimulation, and vice versa,
a calcium response was still evident. However, these agonists displayed
partial heterologous desensitization in that prior stimulation with
vasopressin attenuated the subsequent response to endothelin, and vice
versa. This suggests the presence of functionally distinct
hormone-responsive calcium pools. The technique of double-parameter
fluorescent measurement outlined could potentially be applied to other
cellular signaling parameters by the use of the appropriate probes.
