During stroke, failure of the sodium-potassium ion pump caused by a depletion of ATP in the ischemic core results in anoxic depolarization (AD) propagating through the stroke focus followed by recurring AD-like peri- infarct depolarizations (PIDs) in the penumbra. The functional collapse of plasma membrane ion selectivity associated with propagating AD, causes dramatic neuronal swelling, dendritic beading and spine loss within seconds with associated glial swelling. In essence, swelling is the initial response and a sign of the acute neuronal damage that follows if ischemia is maintained. Recurring PIDs migrate through regions of compromised blood flow, consuming precious energy supplies and expanding the initial site of infarct during subsequent days. This represents a window of opportunity for therapeutic inhibition of these propagating depolarizations. Reduction of the excitotoxic effects of glutamate release caused by these depolarizations has been a traditional approach to prevent ischemic damage. However, an equally viable rationale representing a different and potentially powerful neuroprotective strategy is to target these upstream rapid depolarizations. Morphological and functional differences exist between the brain of humans and animals. Lower mammals appear to be more amenable to neuroprotection than humans. This may suggest that there may be essential differences between human and lower mammals'brain tissue in response to ischemic injury. Therefore, human brain slices as a model system can provide a missing link between animal models and patients and offer a unique chance to identify and study potentially useful therapeutics before advancing to clinical trials. Here, using live human cortical slices prepared from brain tissue resected during surgeries, we propose to inhibit these propagating depolarizations and associated neuronal and glial swelling to better understand and treat their clinical consequences.
The specific aims of this project are: 1) Test candidate therapeutics for efficacy of inhibiting AD in human neocortical slices. 2) Evaluate at the cellular level the neuroprotective effect of candidate therapeutics during onset and recovery from AD-mediated injury in human neocortical slices. AD will be induced by oxygen-glucose deprivation, an in vitro ischemia model, or by application of ouabain, a sodium- potassium pump blocker.
In aim 1, changes in light transmittance (LT) will track AD and associated slice injury in time and space.
In aim 2, cellular dynamics of neurons and astrocytes will be imaged in real time with 2- photon laser scanning microscopy (2PLSM). We will directly correlate structural data from 2PLSM and changes in LT with functional data from field recordings with and without a drug treatment. As a result, we anticipate developing a valuable model for reliable testing and the preliminary screening of candidate therapeutic drugs.
Project Narrative Stroke is the third leading cause of death in industrialized countries and the main source of adult disability. Recent experimental and clinical data imply that the spread of a propagating wave of anoxic depolarization defines the ischemic core and recurring waves of peri-infarct depolarizations expand a cerebral infarct contributing to secondary injury in patients with stroke and traumatic brain injury. Choosing these depolarizing events as targets for therapeutic intervention, this exploratory project aims to develop an assay using human neocortical slices for the preliminary screening of candidate stroke therapeutics before advancing to clinical trial.
| Douglas, Heather A; Callaway, Jennifer K; Sword, Jeremy et al. (2011) Potent inhibition of anoxic depolarization by the sodium channel blocker dibucaine. J Neurophysiol 105:1482-94 |
| Risher, W Christopher; Lee, Mark R; Fomitcheva, Ioulia V et al. (2011) Dibucaine mitigates spreading depolarization in human neocortical slices and prevents acute dendritic injury in the ischemic rodent neocortex. PLoS One 6:e22351 |
| Witcher, Mark R; Park, Yong D; Lee, Mark R et al. (2010) Three-dimensional relationships between perisynaptic astroglia and human hippocampal synapses. Glia 58:572-87 |
