Rodent Stroke Models to Study Functional Recovery and Neural Repair.

Rodent ischemic stroke models are essential research tools for studying this highly prevalent disease and represent a critical element in the translational pipeline for development of new therapies. The majority of ischemic stroke models have been developed to study the acute phase of the disease and neuroprotective strategies, but a subset of models is better suited for studying stroke recovery. Each model therefore has characteristics that lend itself to certain types of investigations and outcome measures, and it is important to consider both explicit and implicit details when designing experiments that utilize each model. The following chapter briefly summarizes the known aspects of the main rodent stroke models with emphasis on their clinical relevance and suitability for studying recovery and neural repair following stroke.

The Finer Aspects of Grid-Walking and Cylinder Tests for Experimental Stroke Recovery Studies in Mice.

The choice of behavioral tests and their proper execution is critically important for experimental and preclinical therapeutic stroke recovery studies, where improvement of impaired neurological function(s) is the main outcome measure. Two tests that focus on spontaneous motor behaviors of the forelimb during gait and exploratory rearing and are expert recommended for stroke recovery studies in mice are grid-walking and cylinder tasks. Both tests have been widely used in various experimental stroke studies to evaluate acute and chronic motor impairment. To facilitate adoption of these tests and consistency of use between different research laboratories, this chapter describes a simple and rigorous protocol and our schemes to successfully perform both tasks in mice and evaluate motor dysfunction and recovery after stroke. In addition, we provide practical tips to minimize experimental bias and acquire data for analyses.

Delayed atomoxetine or fluoxetine treatment coupled with limited voluntary running promotes motor recovery in mice after ischemic stroke.

Currently, there is an unmet need for treatments promoting post-stroke functional recovery. The aim of this study was to evaluate and compare the dose-dependent effect of delayed atomoxetine or fluoxetine therapy (starting on post-stroke day 5), coupled with limited physical exercise (2 hours daily voluntary wheel running; post-stroke days 9 to 42), on motor recovery of adult male mice after photothrombotic stroke. These drugs are selective norepinephrine or serotonin reuptake inhibitors indicated for disorders unrelated to stroke. The predetermined primary end-point for this study was motor function measured in two tasks of spontaneous motor behaviors in grid-walking and cylinder tests. Additionally, we quantified the running distance and speed throughout the study, the number of parvalbumin-positive neurons in the medial agranular cortex and infarct volumes. Both sensorimotor tests revealed that neither limited physical exercise nor a drug treatment alone significantly facilitated motor recovery in mice after stroke. However, combination of physical exercise with either of the drugs promoted restoration of motor function by day 42 post-stroke, with atomoxetine being a more potent drug. This was accompanied by a significant decrease in parvalbumin-positive inhibitory interneurons in the ipsilateral medial agranular cortex of mice with recovering motor function, while infarct volumes were comparable among experimental groups. If further validated in larger studies, our observations suggest that add-on atomoxetine or fluoxetine therapy coupled with limited, structured physical rehabilitation could offer therapeutic modality for stroke survivors who have difficulty to engage in early, high-intensity physiotherapy. Furthermore, in light of the recently completed Assessment oF FluoxetINe In sTroke recoverY (AFFINITY) and Efficacy oF Fluoxetine-a randomisEd Controlled Trial in Stroke (EFFECTS) trials, our observations call for newly designed studies where fluoxetine or atomoxetine pharmacotherapy is evaluated in combination with structured physical rehabilitation rather than alone. This study was approved by the Texas Tech University Health Sciences Center Institutional Animal Care and Use Committee (protocol # 16019).

The Effect of Histone Deacetylase Inhibitors Panobinostat or Entinostat on Motor Recovery in Mice After Ischemic Stroke.

Using rigorous and clinically relevant experimental design and analysis standards, in this study, we investigated the potential of histone deacetylase (HDAC) inhibitors panobinostat and entinostat to enhance recovery of motor function after photothrombotic stroke in male mice. Panobinostat, a pan-HDAC inhibitor, is a FDA-approved drug for certain cancers, whereas entinostat is a class-I HDAC inhibitor in late stage of clinical investigation. The drugs were administered every other day (panobinostat-3 or 10 mg/kg; entinostat-1.7 or 5 mg/kg) starting from day 5 to 15 after stroke. To imitate the current standard of care in stroke survivors, i.e., physical rehabilitation, the animals run on wheels (2 h daily) from post-stroke day 9 to 41. The predetermined primary end point was motor recovery measured in two tasks of spontaneous motor behaviors in grid-walking and cylinder tests. In addition, we evaluated the running distance and speed throughout the study, and the number of parvalbumin-positive neurons in medial agranular cortex (AGm) and infarct volumes at the end of the study. Both sensorimotor tests revealed that combination of physical exercise with either drug did not substantially affect motor recovery in mice after stroke. This was accompanied by negligible changes of parvalbumin-positive neurons recorded in AGm and comparable infarct volumes among experimental groups, while dose-dependent increase in acetylated histone 3 was observed in peri-infarct cortex of drug-treated animals. Our observations suggest that add-on panobinostat or entinostat therapy coupled with limited physical rehabilitation is unlikely to offer therapeutic modality for stroke survivors who have motor dysfunction.

Motor deficit in the mouse ferric chloride-induced distal middle cerebral artery occlusion model of stroke.

Ferric chloride-induced distal middle cerebral artery occlusion (MCAO) model of stroke was described in mice several years ago, however it lacked in-depth evaluation of the post-stroke functional outcomes in the animals. In this study, we reproduced the recently developed model and expanded its characterization by thorough evaluation of blood supply, cerebral infarction, and motor function in adult male and female mice up to 14 days after stroke. Our observations indicate near complete interruption of blood flow in the distal MCA shortly after application of 20 % ferric chloride over the artery through a cranial window, which remained occluded for at least 4 h. As expected, infarction of the brain tissue, documented by TTC and hematoxylin stains, was restricted to the cerebral cortex. We also systematically evaluated motor impairment of the animals in this model. For this, a series of studies were carried out in male and female mice up to 14 days after stroke, and motor function was assessed in cylinder and grid-walking tests in blinded manner. Contrary to our expectations, the results of both motor tests indicated minor, transient motor deficit in mice after stroke. Based on these observations, we conclude that the mouse ferric chloride-induced distal MCAO model is likely not suitable for proof-of-concept and preclinical studies where motor function is an important outcome measure.