Studies on the Mechanisms Responsible for the Formation of Focal Swellings on Neuronal Processes Using a NovelIn VitroModel of Axonal Injury

Abstract

A novel in vitro model of axonal injury using PC12 cells was designed to introduce traumatic alterations on neuronal processes and to identify mechanisms responsible for the formation of focal swellings by observation with phase-contrast and transmission electron microscopes. The injury on the processes was produced by one-dimensional, horizontal oscillation. The fluid shear stress applied by the oscillation did not exceed 380 dyne/cm2. The injured processes showed two forms. One involved an increase in the terminal diameter of the processes and the other entailed beading along the injured portions. Long-term observation of cellular responses to the mechanical insult disclosed that the terminal swelling coincided with the detachment of growth cones from the culture plate. The finding suggests that the detachment of the growth cone destroys the cytoskeletal network, which determines and maintains the cell shape, resulting in spherical deformation of the processes. When the cytoskeletal destruction occurred at non-terminal sites along the processes, spherical deformations developed slowly, and these appeared as beads. The beading also caused the detachment of the growth cones. As the most proximal bead grew, they absorbed the distal segment and their growth cones were pulled proximally with the spreading cytoskeletal destruction. The processes with terminal swellings as well as the bead segments showed regeneration with time evidence of and growth cone formation.

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

“…These cells have been well characterized and have become a model for many studies related to identification and characterization of neuronal signaling pathways (Heneka et al, 1998), to study cell death (Dubreuil et al, 2003;Kitazawa et al, 2004;Ulloth et al, 2003;Yang et al, 2004), especially apoptosis, in neuronal cells and cytoskeletal structure and function (Kobayashi and Mundel, 1998;Nakayama et al, 2001;Okabe and Hirokawa, 1988;Tahir et al, 1992). Although no other types of cells were examined in the present study, cell viability studies and onset rate dependency of traumatic injury produced on cells with CCSD were similar to previous studies conducted with human derived neuronal cells (Ellis et al, 1995;LaPlaca and Thibault, 1998).…”

Collagen-Dependent Neurite Outgrowth and Response to Dynamic Deformation in Three-Dimensional Neuronal Cultures

“…Enhanced survival and neurite outgrowth was observed in collagen (0.4-0.5 mg/mL) as compared to hydrogels lacking ECM ligands, which produced varying degrees of cell viability and a paucity of neurite outgrowth, together indicating that growth and survival of primary cortical neurons are improved by specific cell-matrix interactions. 46,47,60 Most in vitro models developed to study the response of neural cells to mechanical deformation utilize planar cell culture, and include mechanical stretch 19,23,54 and hydrodynamic shear stress 40,45 systems. Differences in cell morphology and the spatial distribution of cell-cell/cell-ECM interactions in planar vs. 3-D cultures may be important, especially in models of traumatic cellular injury as bulk deformation is translated to cells through physical coupling, which may be unrealistically represented in 2-D culture.…”

Acute and Chronic Neural Stimulation via Mechano-Sensitive Ion Channels

“…cells, although a poor model for neurons, were also subjected to horizontal oscillation to induce shear stress injuries. 135 It was found that injury induced swelling in the neurite terminals and the detachment of growth cones. This resulted in disruption to the cytoskeleton and inability to maintain cell shape.…”