The Journal of Neurophysiology Vol. 80 No. 5 November 1998, pp. 2688-2698
Copyright ©1998 The American Physiological Society

Neurotoxicity Mediated by Aberrant Patterns of Synaptic Activity Between Rat Hippocampal Neurons in Culture

John R. McLeod Jr., Maoxing Shen, Daniel J. Kim, and Stanley A. Thayer

Department of Pharmacology, University of Minnesota Medical School, Minneapolis, Minnesota 55455

McLeod, John R., Jr., Maoxing Shen, Daniel J. Kim, and Stanley A. Thayer. Neurotoxicity mediated by aberrant patterns of synaptic activity between rat hippocampal neurons in culture. J. Neurophysiol. 80: 2688-2698, 1998. Reducing the extracellular Mg²⁺ concentration ([Mg²⁺]_o) to 0.1 mM evoked an aberrant pattern of glutamatergic activity in the synaptic network formed by rat hippocampal neurons grown in primary culture. This treatment resulted in a significant increase in neuronal death when maintained for 20-24 h; 0.1 mM [Mg²⁺]_o elicited a stable and repetitive series of intracellular Ca²⁺ concentration ([Ca²⁺]_i) spikes as indicated by indo-1-based microfluorimetry. Fura-2-based digital imaging experiments found that the [Ca²⁺]_i spikes were synchronized for all the neurons in a given field. Thus electrophysiological recordings from individual cells were reasonable representations of the field as a whole, enabling correlation of electrical activity to viability. Underlying each [Ca²⁺]_i spike was an intense burst of action potentials. Whole cell voltage-clamp experiments showed that a burst was composed of fast action currents superimposed on a slow inward current. The N-methyl-D-aspartate (NMDA) receptor antagonist CGS19755 (10 µM) blocked [Ca²⁺]_i spiking, the slow inward current, and the cell death induced by low [Mg²⁺]_o. The L-type Ca²⁺ channel antagonist nimodipine (10 µM) blocked [Ca²⁺]_i spiking, all synaptic activity, and the cell death induced by low [Mg²⁺]_o. The non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 µM) exerted variable effects on [Ca²⁺]_i spiking and blocked the slow inward current only when the cells were held at a relatively negative holding potential. CNQX did not afford any protection from 0.1 mM [Mg²⁺]_o-induced neurotoxicity. [Ca²⁺]_i imaging experiments showed that CNQX inhibited [Ca²⁺]_i spiking in a subset of neurons within an active network. Thus, the neurons that were insensitive to CNQX appear to be those that were destined to die. We characterized an in vitro model that allowed us to correlate specific electrophysiological components of glutamatergic synaptic activity to the subsequent viability of the network. A slow NMDA receptor-mediated inward current was required to elicit [Ca²⁺]_i spiking and neurotoxicity. Non-NMDA receptors did not contribute to synaptically mediated cell death in this model. An L-type Ca²⁺ channel antagonist was neuroprotective when used at concentrations that blocked synaptic activity, suggesting that dendritic L-type Ca²⁺ channels present a useful target for neuroprotective drugs.

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