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The regulation of intracellular calcium in cultured hippocampal neurons and the influence of calcium buffers

University of British Columbia

Excessive activation of the major neurotransmitter glutamate receptors has been blamed as a causative mechanism in a large number of neuro-pathological situations via a process known as excitotoxicity. Excessive influx of calcium (Ca²+, and the consequent loss of Ca²+ homeostasis in neurons subject to this form of excessive activation have been suggested to play a central role in the triggering of a series of events ending in neuronal death. Neuronal populations differ in their vulnerability to excitotoxicity and this difference has, controversially, been attributed to the variability in the calcium-load handling capacity of neurons. The latter may possibly reflect the level of expression of the high-capacity calcium-binding proteins such as calbindin-D28K (CaBP). This work aims at studying the potential influence of CaBP expression in cultured hippocampal neurons on agonist- and depolarization-induced Ca²+ responses, and on the vulnerability of these neurons to excitotoxic stimuli. Manipulation of neuronal Ca²+ buffering capacity could also be achieved by loading neurons with BAPTA-like Ca²+ buffers. We tested the hypothesis that enhancing neuronal Ca²+ buffering capacity, either by the expression of CaBP or by loading neurons with BAPTA-like buffers, will render them more resistant to glutamate-induced neuronal death in vitro. We found that neurons expressing CaBP or loaded with an artificial Ca²+ buffer, contrary to our hypothesis, were more vulnerable to excitotoxicity. The mechanism(s) involved in the enhancement of neuronal loss under conditions of increases Ca²+ buffering capacity are likely to include an increase in the net influx of Ca²+, secondary to the impairment ofCa²+-mediated inhibition of Ca²+ influx, and the prolongation of the recovery phase at the end of the excitotoxic stimulus. Such a prolongation may be a result of the increased Ca²+ influx and competition between the recovery mechanisms on one side, and the large number of high affinity Ca²+ binding sites of the buffer on the other. In neurons that express CaBP, an added vulnerability factor may be the consistently higher peak Ca²+ responses to glutamate receptor agonists and to depolarization observed in these neurons when compared to responses in neurons lacking CaBP. The biological effects of enhanced Ca²+ buffering capacity, particularly using fast and mobile buffers, would be subject to the cell-specific nature of the spatial and temporal pattern of the Ca²+ signal. Thus, the influence of Ca²+ buffering on neuronal vulnerability may be difficult to predict on a theoretical basis particularly in view of our results indicating the large impact of many factors in the culture environment on the vital properties of neurons in vitro. [Scientific formulae used in this abstract could not be reproduced.]

Thesis/Dissertation

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2009-06-04

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http://hdl.handle.net/2429/8736

Physiology

University of British Columbia

UBCV

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