Activity-Dependent Calcium Sequestration in Dendrites of Hippocampal Neurons in Brain Slices

Abstract

Synaptic activity-dependent changes in the spatio-temporal distribution of calcium ions regulate important neuronal functions such as dendritic integration and synaptic plasticity, but the processes that terminate the free Ca 2ϩ transients associated with these changes remain unclear. We have characterized at the electron microscopic level the intracellular compartments involved in buffering free Ca 2ϩ transients in dendritic cytoplasm of CA3 neurons by measuring the larger changes in the concentrations of total Ca that persist for several minutes after neuronal activity. Quantitative energy-dispersive x-ray microanalysis of cryosections from hippocampal slice cultures rapidly frozen 3 min after afferent synaptic activity identified a subset of dendritic endoplasmic reticulum (ER) as a highcapacity Ca 2ϩ buffer. Calcium sequestration by cisterns of this subset of ER was graded, reversible, and dependent on a thapsigargin-sensitive Ca 2ϩ -ATPase. Sequestration was so robust that after repetitive high-frequency stimulation the Ca content of responsive ER cisterns increased as much as 20-fold. These results demonstrate that a subpopulation of ER is the major dendritic Ca sequestration compartment in the minutes after neuronal activity.

Excitotoxic Calcium Overload in a Subpopulation of Mitochondria Triggers Delayed Death in Hippocampal Neurons

“…Difference by t test, p Ͻ 0.0001. B, C, In control neurons, total Na, K, and Ca concentrations are consistent with known values for resting pyramidal neurons (Pozzo-Miller et al, 1997). Immediately after NMDA exposure, cytosolic average concentrations of total Na and Ca increase, whereas the concentration of K falls.…”

Shutdown of Translation: Lethal or Protective? Unfolded Protein Response versus Apoptosis

“…These are strictly calcium-dependent processes that need high calcium activity for correct functioning (Lodish and Kong, 1990;Gosh et al, 1991;Kuznetsov et al, 1992;Lodish et al, 1992). Endoplasmic reticulum calcium activity is therefore several orders of magnitude above that of the cytoplasm (Hofer and Machen, 1993;Chen et al, 1996;Pozzomiller et al, 1997). The need for high ER luminal calcium activity is indicated by the fact that depletion of ER calcium stores is a severe form of stress sufficient to cause cell death (for a comprehensive discussion see Paschen and Doutheil, 1999).…”

Correlated measurements of free and total intracellular calcium concentration in central nervous system neurons

“…If caution is exercised to control some of these experimental drawbacks, i.e., by careful calibration of the experimental setup, rational use of indicator concentrations, and limited UV illumination, optical imaging of Ca 2ϩ concentration using ratiometric fluorescent indicators will provide meaningful quantitative measurements of Ca 2ϩ dynamics in living neurons and their finest processes, such as dendrites and spines. Figure 3, and described in detail elsewhere Buchanan et al, 1993;Leapman and Andrews, 1991;Pozzo-Miller et al, 1997) -was used to obtain direct measurements of persistent changes in the distribution of total Ca within dendritic compartments of pyramidal neurons after synaptic stimulation. We focused on the three subcellular compartments that comprise essentially the entirety of these dendrites, namely, endoplasmic reticulum (ER), mitochondria, and cytoplasm.…”

“…All biologically interesting elements between C (Z ϭ 6) and Ca (Z ϭ 20) produce X-rays at characteristic energies within the energy range 0-10 keV (see labels on F), which can be detected, identified, and quantified. Subsequent processing of spectra, as described (Shuman et al, 1976;Buchanan et al, 1993;Pozzo-Miller et al, 1997), provides compartmental concentrations of elements in units of mmol/kg dry weight. dry weight), so that 5.1 Ϯ 1.1 mmol/kg dry weight for control [Ca] ER becomes ϳ1.3 mmol/l.…”