Traumatic brain injury (TBI) occurs when brain tissue is subjected to stresses and strains at high rates and magnitudes, yet the mechanisms of injury and cellular thresholds are not well understood. The events that occur at the time of and immediately after an insult are hypothesized to initiate cell dysfunction or death following a critical cell strain and strain rate. We analyzed neuronal plasma membrane disruption in two in vitro injury models-fluid shear stress delivered to planar cultures and shear strain induction of 3-D neural cultures. We found that insult severity positively correlated with the degree of membrane disruptions in a heterogeneous fashion in both cell configurations. Furthermore, increased membrane permeability led to increases in electrophysiological disturbance. Specifically, cells that exhibited increased membrane permeability did not fire random action potentials, in contrast to neighboring cells that had intact plasma membranes. This approach provides an experimental framework to investigate injury tolerance criteria as well as mechanistically driven therapeutic strategies.