The Effect of pH and Temperature on the Dissociation Constant for Fura-2 and Their Effects on [Ca2+]i in Enterocytes from a Poikilothermic Animal, Atlantic Cod (Gadus morhua)
✍ Scribed by Dennis Larsson; Birgitta Larsson; Ted Lundgren; Kristina Sundell
- Publisher
- Elsevier Science
- Year
- 1999
- Tongue
- English
- Weight
- 96 KB
- Volume
- 273
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
In this study, we investigated the validity of the fluorescent probe fura-2 in determinations of intracellular Ca(2+) concentrations (Ca(2+)), at physiological temperatures, in poikilothermic animals living at low temperatures. The K(d) for fura-2 was found to decrease with increasing temperature (5-37 degrees C) and DeltaH, in the Van't Hoff isochore equation, was determined to be 11.03 kJ/mol, when pH was corrected to 7.2 for all temperatures tested. The absorption maxima (340 nm) and isobestic point (360 nm) for the UV spectra of fura-2 were not affected by temperature. Thus, if pH- and temperature-dependent changes in K(d) are corrected for, fura-2 is a suitable tool for measurements of Ca(2+) at temperatures of 5-37 degrees C. The present study demonstrates that Atlantic cod enterocytes, acclimated to 37 degrees C, show a lower basal Ca(2+) (65 +/- 8 nM) compared to enterocytes acclimated to 10 degrees C (161 +/- 6 nM). Furthermore, addition of 10 mM Ca(2+) increases the Ca(2+) by 526%, when compared to basal Ca(2+), in cells at 37 degrees C but only by 36%, in cells kept at 10 degrees C. Thus, performing experiments at unphysiological temperatures results in cellular responses that would not be observed under physiological conditions.