Gauging Recovery after Hemorrhagic Stroke in Rats: Implications for Cytoprotection Studies

Abstract

Successful clinical translation of prospective cytoprotectants will likely occur only with treatments that improve functional recovery in preclinical (rodent) studies. Despite this assumption, many rely solely on histopathologic end points or the use of one or two simple behavioral tests. Presently, we used a battery of tests to gauge recovery after a unilateral intracerebral hemorrhagic stroke (ICH) targeting the striatum. In total, 60 rats (N = 15 per group) were stereotaxically infused with 0 (SHAM), 0.06 (MILD lesion), 0.12 (MODERATE lesion), or 0.18 U (SEVERE lesion) of bacterial collagenase. This created a range of injury akin to moderate (from SEVERE to MODERATE or MODERATE to MILD lesion size B30% reduction) and substantial cytoprotection (SEVERE to MILD lesion size-51% reduction). Post-ICH functional testing occurred over 30 days. Tests included the horizontal ladder and elevated beam tests, swimming, limb-use asymmetry (cylinder) test, a Neurologic Deficit Scale, an adhesive tape removal test of sensory neglect, and the staircase and single pellet tests of skilled reaching. Most tests detected significant impairments (versus SHAM), but only a few (e.g., staircase) frequently distinguished among ICH groups and none consistently differentiated among all ICH groups. However, by using a battery of tests we could behaviorally distinguish groups. Thus, preclinical testing would benefit from using a battery of behavioral tests as anything less may miss treatment effects. Such testing must be based on factors including the type of lesion, the postoperative delay and the time required to complete testing.

Skilled reaching impairments follow intrastriatal hemorrhagic stroke in rats

“…Although each of their tests (corner turn test, forelimb use asymmetry (cylinder) test and forelimb placing test) detected ICH injury, marked recovery to pre-surgical levels occurred by 1 month in two of the tests. Similarly, our recent study [24] showed ICH rats near completely recover in behavioral tests (e.g., cylinder, ladder and staircase tasks) otherwise sensitive to focal ischemia [27,29,37] and collagenase-induced ICH [22]. These findings highlight the need to identify sensitive tests for this model of ICH, especially over the long term.…”

“…Pellets on the left stairs may be retrieved only using the left paw, whereas pellets on the right stairs must be obtained using the right paw. Rats were trained over 4 weeks (two 15 min sessions/day and 5 days/week) and were excluded from this test if they did not retrieve at least 9 pellets/side (out of a possible 21) on 3 consecutive days [5,22]. The last 5 days of training were used to calculate baseline performance.…”

“…The first three successful reaches were each analyzed qualitatively [22,28]. We anticipated that only a small proportion of reaches would be successful after ICH, and therefore also analyzed movement components for reaches considered hits and failures.…”

“…Recently, we [22] characterized neurological deficits after ICH induced by striatal infusion of bacterial collagenase, which causes bleeding by disrupting cerebral microvasculature [35]. Although the majority of the tests (e.g., skilled reaching, walking and neglect) detected ICH injury, few distinguished among gradations in injury [22]. Interestingly, the staircase task of skilled reaching was one of the most useful tests for detecting ICH injury and moderate differences in lesion size.…”

“…An ideal test should not only detect the presence of a lesion, but must also detect therapeutic effects. Recently, we [22] characterized neurological deficits after ICH induced by striatal infusion of bacterial collagenase, which causes bleeding by disrupting cerebral microvasculature [35]. Although the majority of the tests (e.g., skilled reaching, walking and neglect) detected ICH injury, few distinguished among gradations in injury [22].…”