Acute Alterations in [Ca²⁺]_iin NG108-15 Cells Subjected to High Strain Rate Deformation and Chemical Hypoxia: Anin VitroModel for Neural Trauma

Abstract

The short-term (less than 2 min) alterations in the intracellular free calcium concentration in differentiated NG108-15 (neuroblastoma cross glioma) cells exposed to dynamic mechanical deformation with and without superimposed chemical hypoxia were determined. A previously developed device, modified for these studies, was used to apply deformations at a magnitude and rate representative of those experienced by neural tissue in Traumatic Brain Injury. Chemical hypoxia was imposed using a combination of 2-deoxy-D-glucose and salicylate, anaerobic and aerobic metabolic blockers, respectively. Real time measurement of intracellular free calcium concentration using Fura-2 and a custom epifluorescence microscopy system provided a quantitative index of cell response. At high rates of deformation (approximately 10 sec-1), increases in intracellular free calcium concentration were exponentially related to the magnitude of the applied deformation. Chemical hypoxia had no effect on this acute response. At low rates of deformation, small increases in intracellular free calcium concentration were independent of the magnitude of the deformation. These findings indicate that strategies for reducing severity of TBI should focus on minimizing the rate of deformation of neural cells. Together with data from animal, physical, and finite element models, these data can be employed in the development of physiologic injury tolerance criteria for the whole head.

The Effect of Poloxamer-188 on Neuronal Cell Recovery from Mechanical Injury

“…The loading parameters used in this study were developed based on our understanding of the magnitude and rate dependence of cell injury. Previous studies have used a variety of techniques for applying mechanical loads to cells including simple elongation of axons (Galbraith et al, 1993), biaxial stretch of two-dimensional culture substrates (Cargill and Thibault, 1996) and fluid shear stress , Blackman et al, 2000. A common feature of these studies was the finding that the injury severity depended strongly on the rate at what the mechanical load was applied.…”

“…Most previous in vitro models of TBI (Regan and Choi, 1994;Mukhin et al, 1997;Shepard et al, 1991;Murphy and Horrocks, 1993;Lucas and Wolf, 199;Cargill and Thibault, 1996;Ellis et al, 1995; have some limitations. The most severe limitations of these models are the non-uniformity of the applied mechanical stimuli and the difficulty in controlling strain and strain rate independently.…”

“…Previous studies have shown that cell injury depends on both the magnitude and rate of the applied load (Blackman et al, 2000;Cargill et al, 1996;Geddes et al, 2003;. To investigate the mechanism of loading rate dependent responses in cultured cells, we used the controlled cell shearing device (CCSD) described by Blackman et al (2000), which has been used previously to demonstrate loading-rate dependent injury of the plasma membrane of endothelial cells.…”

Modelos experimentales de traumatismo craneoencefálico

“…El modelo inicial consistía en el estiramiento bidireccional de dos membranas elásticas entre las que se disponían células en cultivo. Inicialmente este modelo se utilizó para la investigación del daño de cultivos de astrocitos pero rápidamente se aplicó a culti- vos de neuronas y a cultivos mixtos neurogliales 19,83 . Una variación posterior del modelo consistió en el estiramiento de las células en cultivo en una sola dirección 87,71 (Figura 6-B).…”

Traumatic injury induces differential expression of cell death genes in organotypic brain slice cultures determined by complementary DNA array hybridization

“…39,47 Although several in vivo models of TBI appear well suited for the study of the clinically relevant response of the whole animal, 53 the complexity of the in vivo situation prevents free access to the tissue, thereby interfering with spatial or realtime measurements of mechanical or biological parameters. Several in vitro models of mechanical trauma have recently been developed 5,17,48 and possess specific advantages over in vivo models including the precise control of the extracellular environment, easy access to the tissue, and the isolation of specific cellular and molecular events from potentially confounding factors found in vivo.…”