Enhanced Neocortical Neural Sprouting, Synaptogenesis, and Behavioral Recovery With d-Amphetamine Therapy After Neocortical Infarction in Rats

Abstract

Background and Purpose—d-Amphetamine administration increases behavioral recovery after various cortical lesions including cortical ablations, contusions, and focal ischemia in animals and after stroke in humans. The purpose of the present study was to test the enhanced behavioral recovery and increased expression of proteins involved in neurite growth and synaptogenesis in d-amphetamine–treated rats compared with vehicle-treated controls after a focal neocortical infarct. Methods—Unilateral neocortical ischemia was induced in male spontaneously hypertensive Wistar rats (n=8 per time point per group) by permanently occluding the distal middle cerebral artery and ipsilateral common carotid artery in 2 groups of rats:d-amphetamine treated (2 mg/kg IP injections) and vehicle treated (saline IP injections). To determine the spatial and temporal distribution of neurite growth and/or synaptogenesis, growth-associated protein (GAP-43), a protein expressed on axonal growth cones, and synaptophysin, a calcium-binding protein found on synaptic vesicles, were examined by immunohistochemical techniques, and both density and distribution of reaction product were measured. Since the resulting infarction included a portion of the forelimb neocortex, behavioral assessments of forelimb function using the foot-fault test of Hernandez and Schallert were performed on the same rats used for immunohistochemical studies during the period of drug action and 24 hours later. A Morris water maze and other indices of behavioral assays were also measured similarly. Recovery times were 3, 7, 14, 30, and 60 days postoperatively. Results—Both GAP-43 and synaptophysin proteins demonstrated statistically significant increases in density and distribution of immunoreaction product as determined by optical density measurements in the neocortex of the infarcted group treated with d-amphetamines compared with vehicle-treated infarcted controls. The GAP-43 was elevated to statistically significant levels in forelimb, hindlimb, and parietal neocortical regions ipsilateral to the infarction only at days 3, 7, and 14. By contrast, the synaptophysin demonstrated no statistically significant changes in expression at 3 or 7 days but demonstrated statistically significant increases at 14, 30, and 60 days in the forelimb, hindlimb, and parietal neocortical regions ipsilateral to the infarction as well as increased distribution in the contralateral parietal neocortex. Behavioral assessment of forelimb function indicated that improved recovery of forelimb placement on the side contralateral to the infarction was statistically significant in the d-amphetamine–treated group compared with the vehicle-treated group (P<0.025). Spatial memory, as measured with the Morris water maze, worsened in the vehicle-treated group compared with the d-amphetamine–treated group at 60 days (P<0.025). Conclusions—These data support the occurrence of neurite growth followed by synaptogenesis in the neocortex in a pattern that corresponds both spatially and temporally with behavioral recovery that is accelerated by d-amphetamine treatment. While the specific mechanisms responsible for d-amphetamine–promoted expression of proteins involved in neurite growth and synaptogenesis and of enhanced behavioral recovery are not known, it is suggested that protein upregulation occurs as a result of functional activation of pathways able to remodel in response to active behavioral performance.