Impact of Cytoplasmic Calcium Buffering on the Spatial and Temporal Characteristics of Intercellular Calcium Signals in Astrocytes

Abstract

The impact of calcium buffering on the initiation and propagation of mechanically elicited intercellular Ca2+waves was studied using astrocytes loaded with different exogenous, cell membrane-permeant Ca2+chelators and a laser scanning confocal or video fluorescence microscope. Using an ELISA with a novel antibody to BAPTA, we showed that different cell-permeant chelators, when applied at the same concentrations, accumulate to the same degree inside the cells. Loading cultures with BAPTA, a high Ca2+affinity chelator, almost completely blocked calcium wave occurrence. Chelators having lower Ca2+affinities had lesser affects, as shown in their attenuation of both the radius of spread and propagation velocity of the Ca2+wave. The chelators blocked the process of wave propagation, not initiation, because large [Ca2+]iincreases elicited in the mechanically stimulated cell were insufficient to trigger the wave in the presence of high Ca2+affinity buffers. Wave attenuation was a function of cytoplasmic Ca2+buffering capacity; i.e., loading increasing concentrations of low Ca2+affinity buffers mimicked the effects of lesser quantities of high-affinity chelators. In chelator-treated astrocytes, changes in calcium wave properties were independent of the Ca2+-binding rate constants of the chelators, of chelation of other ions such as Zn2+, and of effects on gap junction function. Slowing of the wave could be completely accounted for by the slowing of Ca2+ion diffusion within the cytoplasm of individual astrocytes. The data obtained suggest that alterations in Ca2+buffering may provide a potent mechanism by which the localized spread of astrocytic Ca2+signals is controlled.

High-ResolutionIn VivoImaging of the Neurovascular Unit during Spreading Depression

“…It thus differs substantially from what one would expect from a phenomenon that is an integral part of SD. We regard these as astrocytic [Ca 2ϩ ] waves, as seen in vitro (Cornell-Bell et al, 1990;Charles et al, 1991;Wang et al, 1997), expressed as a collateral phenomenon of SD but whose origin and significance are unclear.…”

Connexins regulate calcium signaling by controlling ATP release

“…Experiments were carried out at room temperature in a 1:1 mixture of DMEM and F12 (Ham). Single cells were mechanically stimulated with a glass micropipette as earlier described (21). The wave radius was measured as the farthest distance traveled by the wave in any direction from the stimulated cell.…”

“…Wild-type or null-transfectant C6 cells were minimally coupled (4 Ϯ 2% refill of CDCF after photobleach) and were unable to propagate Ca 2ϩ waves (mean wave radius Ͻ50 m). As in astrocytes, calcium signaling in Cx43 ϩ or Cx32 ϩ transfectants was also insensitive to uncoupling agents but inhibited by thapsigargin (5 Ϯ 6% of control) or preloading with the calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetate acetoxymethyl ester (BAPTA; 9 Ϯ 7% of control) (21). Their similar pharmacological characteristics suggest that calcium signaling in astrocytes and Cx-transfected C6 cells are mediated by identical signaling cascades.…”

Two dimensional finite element estimation of calcium ions in presence fo NCX and Buffers in Astrocytes

“…Previous researcher have studied the effect of rapid and excess buffers on cytosolic calcium concentration in nerve cells like neuron, astrocytes, fibrocytes etc [3,4,5,9,10,12,15,17,19,21]. Calcium kinetics in nerve cells is governed by a set of reaction-diffusion equations which can be framed assuming the following bimolecular reaction between [Ca 2+ ] and buffer species [11,12]:…”