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Kinetics of Ca 2+ binding to parvalbumin in bovine chromaffin cells: implications for [Ca 2+ ] transients of neuronal dendrites
Author(s) -
Lee SukHo,
Schwaller Beat,
Neher Erwin
Publication year - 2000
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.2000.t01-2-00419.x
Subject(s) - depolarization , chemistry , time constant , parvalbumin , chromaffin cell , fura 2 , biophysics , ionomycin , calcium , phase (matter) , cytosol , analytical chemistry (journal) , adrenal medulla , endocrinology , biology , biochemistry , catecholamine , neuroscience , chromatography , organic chemistry , electrical engineering , enzyme , engineering
1 κ S 1. The effect of parvalbumin (PV) on [Ca 2+ ] transients was investigated by perfusing adrenal chromaffin cells with fura‐2 and fluorescein isothiocyanate (FITC)‐labelled PV. As PV diffused into cells, the decay of [Ca 2+ ] transients was transformed from monophasic into biphasic. The proportion of the initial fast decay phase increased in parallel with the fluorescence intensity of FITC, indicating that PV is responsible for the initial fast decay phase. 2 The relationship between the fast decay phase and the [Ca 2+ ] level was investigated using depolarizing trains of stimuli. Within a train the relative amplitude of the fast decay phase was inversely dependent on the [Ca 2+ ] level preceding a given stimulus. 3 Based on these observations, we estimated the Ca 2+ binding ratio of PV (κ P ), the apparent dissociation constant of PV for Ca 2+ ( K dc,app ), and the unbinding rate constant of Ca 2+ from PV ( k c‐ ) in the cytosol of chromaffin cells. Assuming free [Mg 2+ ] to be 0.14 m m , we obtained values of 51.4 ± 2.0 n m ( n = 3 ) and 0.95 ± 0.026 s −1 ( n = 3 ), for K dc,app and k c‐ , respectively. 4 With the parameters obtained in the perfusion study, we simulated [Ca 2+ ] transients, using two different Ca 2+ extrusion rates (γ) – 20 and 300 s −1 – which represent typical values for chromaffin cells and neuronal dendrites, respectively. The simulation indicated that Ca 2+ is pumped out before it is equilibrated with PV, when γ is comparable to the equilibration rates between PV and Ca 2+ , resulting in the fast decay phase of a biexponential [Ca 2+ ] transient. 5 From these results we conclude that Ca 2+ buffers with slow kinetics, such as PV, may cause biexponential decays in [Ca 2+ ] transients, thereby complicating the analysis of endogenous Ca 2+ binding ratios (κ S ) based on time constants. Nevertheless, estimates of κ S based on Ca 2+ increments provide reasonable estimates for Ca 2+ binding ratios before equilibration with PV.