Monitoring Stroke Progression: In Vivo Imaging of Cortical Perfusion, Blood—Brain Barrier Permeability and Cellular Damage in the Rat Photothrombosis Model

Abstract

Focal cerebral ischemia is among the main causes of death and disability worldwide. The ischemic core often progresses, invading the peri-ischemic brain; however, assessing the propensity of the peri-ischemic brain to undergo secondary damage, understanding the underlying mechanisms, and adjusting treatment accordingly remain clinically unmet challenges. A significant hallmark of the peri-ischemic brain is dysfunction of the blood-brain barrier (BBB), yet the role of disturbed vascular permeability in stroke progression is unclear. Here we describe a longitudinal in vivo fluorescence imaging approach for the evaluation of cortical perfusion, BBB dysfunction, free radical formation and cellular injury using the photothrombosis vascular occlusion model in male Sprague Dawley rats. Blood-brain barrier dysfunction propagated within the peri-ischemic brain in the first hours after photothrombosis and was associated with free radical formation and cellular injury. Inhibiting free radical signaling significantly reduced progressive cellular damage after photothrombosis, with no significant effect on blood flow and BBB permeability. Our approach allows a dynamic follow-up of cellular events and their response to therapeutics in the acutely injured cerebral cortex.

Glutamate-Mediated Blood-Brain Barrier Opening: Implications for Neuroprotection and Drug Delivery

“…BBB dysfunction was not associated with a significant decrease in tissue oxygen levels, and, because of neurovascular coupling, tissue oxygen levels rapidly increased above baseline, ruling out tissue hypoxia as a cause for the rapid increase in endothelial permeability during seizures. Prolonged or frequently recurring seizures, as well as ischemic stroke and traumatic brain injury, are associated with increased concentrations of extracellular glutamate (Ͼ50-fold increase and in the 0.1-1 mM range; Benveniste et al, 1984;Rothman and Olney, 1986;Bradford, 1995;Nishizawa, 2001;Parfenova et al, 2006). Our data, showing that a greater TMS-mediated increase in permeability is prevented when neuronal activity is blocked, but not when glutamate is applied, suggests that neuronal release of glutamate is targeting a nonneuronal target.…”

“…Brain insults, including hypoxic-ischemic or traumatic injuries, were shown to be associated with synchronous neuronal hyperexcitability and elevated extracellular glutamate in both animal models and humans (Benveniste et al, 1984;Rothman and Olney, 1986;Nishizawa, 2001). Interestingly, although NMDA-R antagonists were consistently shown to be neuroprotective in animal models of brain injuries (Rao et al, 2001;Miguel-Hidalgo et al, 2002;Huang et al, 2015), their effects on brain vasculature have never been carefully tested.…”

“…Because excess glutamate release is a hallmark of hypoxic-ischemic injuries (Benveniste et al, 1984;Rothman and Olney, 1986;Nishizawa, 2001) and because the dynamic progression of BBB dysfunction has been characterized in the rat cerebral cortex stroke photothrombosis model (Schoknecht et al, 2014), we tested the hypothesis that blocking NMDA-R activation could reduce BBB breakdown in the peri-ischemic brain. A focal ischemic lesion was induced under ketamine/xylazine anesthesia by photothrombosis after an injection of RB (Watson et al, 1987;Prager et al, 2010;Fig.…”

Assessment of Blood–Brain Barrier Disruption in Stroke

“…26 A novel extension of assessing BBB disruption by fluorescence microscopy is the use of an implantable closed cranial window ( Figure 2D). 28 After surgical implantation of the closed cranial window, rats are given 1 to 2 weeks to recover and can then undergo experimental focal ischemia and reperfusion. Implantation of a closed cranial window permits longterm (≤4 weeks) evaluation of microcirculatory parameters such as BBB disruption, cerebral blood flow, and leukocyte behavior.…”

Functional implications of axon initial segment cytoskeletal disruption in stroke

“…Spreading depolarization, on one hand increases the tissue's metabolic demand and on the other hand could decrease the blood supply due to "inverse" neurovascular coupling [103] . The mismatch between the metabolic demand and supply could contribute to the expansion of the damage to the tissues that did not suffer from direct ischemia and thus cause the stroke to grow in the first hour following the cerebrovascular event [104] . The damage to penumbra could be reversible (in the first six hour) and the long-term outcome of the stroke can be dramatically improved with the fast revascularization of penumbra [105] .…”