NMDA Receptor Activation Contributes to a Portion of the Decreased Mitochondrial Membrane Potential and Elevated Intracellular Free Calcium in Strain-Injured Neurons

Abstract

In our previous studies, we have shown that in vitro biaxial strain (stretch) injury of neurons in neuronal plus glial cultures increases intracellular free calcium ([Ca²⁺]_i) and decreases mitochondrial membrane potential (Δψ_m). The goal of this study was to determine whether strain injury, without the addition of exogenous agents, causes glutamate release, and whether NMDA receptor antagonists affect the post-strain injury rise in [Ca²⁺]_i and decrease in Δψ_m. [Ca²⁺]_i and Δψ_m were measured using the fluorescent indicators fura-2 AM and rhodamine-1,2,3 (rh123). Strain injury of neuronal plus glial cultures caused an immediate 100-200 nM elevation in neuronal [Ca²⁺]i and a decline in neuronal Δψ_m by 15 min post-injury. Pretreatment with the NMDA receptor antagonist MK-801 (10 μM) attenuated the [Ca²⁺]_i elevation after mild, but not moderate and severe injury. MK-801 pretreatment reduced the decline in Δψ_m after mild and moderate, but not after severe injury. The NMDA receptor antagonist D-2-amino-5-phosphonopentanoic acid (APV; 100 μM) had effects similar to MK-801. Simultaneous measurement of [Ca²⁺]_i and Δψ_m demonstrated a significant correlation and a temporal relationship between [Ca²⁺]_i elevation and depression of Δψ_m. We conclude that NMDA receptor stimulation contributes to some of the changes in [Ca²⁺]_i and Δψ_m after less severe strain injury. However, after more pronounced injury other mechanisms appear to be more involved.