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Paltauf-Doburzynska, J; Graier, WF.
Temperature dependence of agonist-stimulated Ca2+ signaling in cultured endothelial cells.
Cell Calcium. 1997; 21(1):43-51
Doi: 10.1016%2FS0143-4160%2897%2990095-6
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- Graier Wolfgang
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- Abstract:
- In cultured endothelial cells, the temperature dependence of bradykinin-initiated Ca2+ signaling was studied using Fura-2 technique. Initially, the temperature dependence of the dissociation constant of Fura-2 for Ca2+ was investigated. Temperature-initiated changes in the apparent dissociation constant (K'D) using the ratio (F340/F380) were due to a hypsochromic shift in excitation wavelengths and changes in the effective dissociation constant of Fura-2 for Ca2+ (K"D). Equations were provided to correct the dissociation constant for Fura-2, either for using the common ratio (F340/F380) or the shift corrected ratio (F340-delta lambda/F380-delta lambda). In a simple experimental protocol, the temperature dependence of the transient increase in free intracellular Ca2+ to bradykinin (i.e. Ca2+ release, sequestration and extrusion) and Ca2+/Mn2+ entry through a Ca2+ store-operated Ca2+ entry pathway (SOCP) were determined. While the temperature dependence of intracellular Ca2+ release, sequestration and extrusion (i.e. enzymatically controlled phenomena) were found to follow the same exponential function [t = A x e(-B x T); t, reaction time; A, B, constants; T, experimental temperature in K; K = degree C + 273], Ca2+/Mn2+ entry upon ion application to pre-stimulated cells strictly followed Fick's law of diffusion [t = A x (1/T) x e(B/T); t, reaction time; A, B, constants; T, experimental temperature in K]. In contrast to the temperature dependence of bradykinin-stimulated Ca2+/Mn2+ entry, the temperature dependence of Mn2+ entry on addition of agonist did not correlate with Fick's law of diffusion, but followed the same exponential function obtained for Ca2+ release, sequestration and extrusion. In conclusion, these data suggest that activation of SOCP by autacoid is due to enzymatic mechanism(s), while Ca2+ entry through SOCP, once activated, is due to a diffusion-like phenomenon.
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