NMDA Receptor Activation Inhibits Neuronal Volume Regulation after Swelling Induced by Veratridine-Stimulated Na⁺Influx in Rat Cortical Cultures

Abstract

Neurons and glia experience rapid fluctuations in transmembrane solute and water fluxes during normal brain activity. Cell volume must be regulated under these conditions to maintain optimal neural function. Almost nothing is known, however, about how brain cells respond to volume challenges induced by changes in transmembrane solute flux. As such, we characterized the volume-regulatory mechanisms of cultured cortical neurons swollen by veratridine-stimulated Na+ influx. Exposure of cortical neurons to 100 microM veratridine for 10-15 min caused a 1.8- to 2-fold increase in cell volume that persisted for at least 90 min. This volume increase was blocked by extracellular Na+ removal or by exposure to 5 microM tetrodotoxin, indicating that swelling is a result of Na+ entry via Na+ channels. Treatment of cells with veratridine together with various NMDA receptor antagonists had no effect on the magnitude of swelling. NMDA receptor antagonist-treated cells, however, underwent nearly complete volume recovery within 50-70 min after veratridine exposure. This recovery suggests that NMDA receptor activation disrupts neuronal osmoregulatory pathways. Volume regulation was blocked by Ba2+, quinidine, or 5-nitro-2-(3-phenylpropylamino) benzoic acid, indicating that swelling activates volume regulatory K+ and Cl- channels. Veratridine also caused a rapid, transient increase in intracellular Ca2+. Extracellular Ca2+ removal or intracellular Ca2+ chelation prevented or dramatically reduced veratridine-induced increases in intracellular Ca2+ and completely blocked volume recovery. These findings indicate that increases in Ca2+ during cell swelling induced by Na+ influx are required for activation of neuronal volume-regulatory pathways.

Impaired Volume Regulation is the Mechanism of Excitotoxic Sensitization to Complement

“…3 A, C,E). These results confirm and extend previous findings that established the presence of RVD in mouse cortical neurons challenged with hypotonic solution (Inoue et al, 2005) or the sodium channel activator veratridine (Churchwell et al, 1996). Notably, the homeostatic mechanisms are powerful because the neurons reestablished their original cell volume despite persistence of the osmotic stressor.…”

Roles of Volume-Sensitive Chloride Channel in Excitotoxic Neuronal Injury

“…؊ channels activated during excitotoxic stimulation are involved in varicosity formation Although it was found that some volume-regulatory pathway is disrupted during activation of NMDA receptors in cortical neurons (Churchwell et al, 1996), we were able to consistently observe robust activity of the VSOR Cl Ϫ channel, which plays a volume-regulatory role in nonexcitotoxic swollen neurons (Inoue et al, 2005), on varicosities during NMDA receptor stimulation (Fig. 7).…”

“…Each symbol represents the mean current Ϯ SEM (error bars). *At given voltages, data points designated with circles that were significantly different from those designated with squares at p Ͻ 0.05. toxic conditions, volume regulation was inhibited in cortical neurons (Churchwell et al, 1996). Thus, it is important to investigate the activity and roles of volume-regulatory Cl Ϫ channels in cortical neurons during and after an excitotoxic insult.…”

“…Under marked depolarization induced by GluR, the VSOR Cl Ϫ channel would serve as a pathway for swelling-aggravating Cl Ϫ influx (process 1 to 2). This Cl Ϫ inflowmediating activity of VSOR Cl Ϫ channel may explain why volume regulation is impaired in cortical neurons under excitotoxic conditions (Churchwell et al, 1996). However, when neurons repolarize after washout of NMDA, VSOR Cl Ϫ channel activity may be responsible for the volume-regulatory Cl Ϫ efflux (process 4).…”

Disruption of ionic and cell volume homeostasis in cerebral ischemia: The perfect storm

“…Under physiological conditions these two independent permeability pathways mediate electrically coupled loss of K + and Cl − , which is accompanied by an efflux of osmotically obligated water, and REGULATORY VOLUME DECREASE, or RVD [30,33,34]. However, in pathology, anoxic and/or glutamate-driven depolarization disrupts cell volume regulation due to an inhibition of the Na + ,K + -pump, dissipation of K + and Cl − electrochemical gradients, and blockade of volume regulatory channels [13,21,25,35]. Na + begins to accumulate uncontrollably in the cells via leak mechanisms, voltage-gated Na + channels, and other Na + -transporting pathways, which are considered in following sections.…”