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Indo-1 can simultaneously detect Ba2+ entry and Ca2+ blockade at a plasma membrane calcium channel
1Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, PA 19107, USA.
Molecular and Cellular Biochemistry
|October 18, 1995
Summary
This study shows how Indo-1 fluorescence can measure multiple intracellular ions, like calcium and barium, simultaneously. It demonstrates barium
Area of Science:
- Cellular Physiology
- Ion Channel Research
- Biophysical Chemistry
Background:
- Indo-1 is a fluorescent chelator capable of measuring intracellular ion concentrations.
- Simultaneous measurement of multiple ions, such as calcium and barium, presents analytical challenges.
- Understanding ion channel function and regulation is crucial in cellular physiology.
Purpose of the Study:
- To demonstrate the simultaneous determination of intracellular calcium ([Ca2+]) and barium ([Ba2+]) concentrations using Indo-1.
- To investigate the effects of extracellular barium on calcium influx through receptor-operated calcium channels.
- To explore the impact of intracellular barium accumulation on cellular calcium homeostasis.
Main Methods:
- Utilized the fluorescent chelator Indo-1 for simultaneous intracellular ion concentration measurements.
- Employed spectral analysis of Indo-1 fluorescence, considering spectral shape and total signal intensity.
- Applied mathematical analysis to whole spectra for maximum accuracy in ion quantification.
- Stimulated B cell lines with anti-immunoglobulin to activate receptor-operated calcium channels.
Main Results:
- Addition of extracellular barium ([Ba2+]) caused a rapid decrease in intracellular calcium ([Ca2+]) in stimulated B cells.
- This decrease in [Ca2+] was attributed to the blockage of receptor-operated calcium channels by extracellular barium.
- Barium conductance was observed as a slow increase in total Indo-1 fluorescence, indicating barium influx.
- A subsequent slow rise in intracellular calcium was observed, potentially due to barium inhibition of the plasma membrane calcium pump.
Conclusions:
- Indo-1 fluorescence analysis allows for simultaneous measurement of intracellular calcium and barium concentrations.
- Extracellular barium effectively blocks receptor-operated calcium channels, reducing calcium influx.
- Intracellular barium accumulation may impair calcium extrusion mechanisms, affecting cellular calcium homeostasis.