Abstract

Introduction

Materials & Methods

Results

Discussion & Conclusion

References

Discussion
Board

Experiment 1

Subjects: The subjects were 50 male Sprague-Dawley rats, 60 days of age, randomly assigned to one of five conditions:

              Prazosin    Propranolol     Saline
Contusion       n=13         n=7           n=12
Sham            n=12         n=6	

Surgery: Under Halothane anesthesia the traumatic brain injury (TBI) was centered over the hindlimb area of sensorimotor cortex using the weight-drop method [2,7]. Sham operates were anesthetized, the scalp shaved, incised and sutured closed without craniotomy.

Beam Walk Task (BW):

Animals were trained to traverse a narrow, elevated beam using white noise and a bright light as aversive stimuli. BW performance was scored for both ipsi and contralateral hindlimbs using a 7-point rating scale, 7=maximum of 2 footslips – 1=unable to traverse the beam [3,7,], based on their ability to use each hindlimb. This bilateral scoring was done to assess any sedative drug effects which would be manifest in disruption of performance of both limbs.

Tactile Placing:

Rats were held suspended by the abdomen at a 90 degree angle from a piece of Plexiglas [11]. Either the vibrissae or the lateral surface of the forelimb were brought into contact with the Plexiglas. A successful placement was counted if the animal placed the ipsilateral forelimb on the flat surface within 5 seconds of stimulation. Animals were given 10 trials per side for both vibrissae and forelimb stimuli. Normal animals score 10 of 10 for each forelimb.

Drug Doses:

        Prazosin       Propranolol          Saline
Low     2 mg/kg         5 mg/kg         equivalent volume
Medium  4 mg/kg         10 mg/kg        equivalent volume
High    8 mg/kg         20 mg/kg        equivalent volume

Pre-Injury:

Animals were given an acclimation period, then were trained to traverse the beam until they received scores of 7 on three consecutive days. A baseline score was taken for both vibrissae-evoked and forelimb-evoked tactile placing. Animals then underwent surgery per their assigned group.

Initial Deficit and Recovery:

Beginning 24 hours after surgery, rats were assessed for their initial BW deficit and subsequent recovery on all tasks.

Reinstatement Evaluation:

At 1 month after surgery, animals were tested on both tasks, then given a single injection of the low dose of their assigned drug. BW tests were performed at 1, 3 & 6 hours then 1, 2, & 3 days after injection. Tactile placing was assessed between hours 1 & 3, then at 1, 2, & 3 days after injection. The regimen was repeated for the medium and high doses of drug.

At 3 & 6 months after surgery, the entire regimen was repeated; however, the tactile placing tasks were omitted. At 12 months after TBI or sham surgery, 15 remaining rats received the high dose of either PRAZ or SAL. Comparisons of BW scores for the ipsilateral BW scores were compared to the contralateral BW scores to evaluate lateralized effects of treatment ,considered a reinstatement of hemiplegia, versus bilateral effects, considered a result of sedation.

Evaluation of Cumulative Drug Effects:

At the completion of the 6 or 12 month dosing regimen, a subgroup of animals, controls never given PRAZ, were given a medium dose of PRAZ. These rats were compared to PRAZ-treated rats to assess drug tolerance or carryover effects on BW performance.

Histology:

Frozen 40 mm sections were stained using cresyl violet, a Nissl stain. Cysts volumes and extents were measured using a computer-assisted imaging system. The following structures were rated using a histopathology scale 1. This evaluation of subcortical pathology in hippocampal CA1, CA3 and Hillar regions, thalamic VPL, Nucleus Reticularis, Medial Geniculate nuclei, and dorsolateral caudate was done by uninformed raters using the following categories used by others (1):

0=no detectable pathology 1=mild pathology
2=moderate pathology
3=severe pathology.

Statistical Analyses:

BW data was analyzed by calculating the area under the curve over 6 hours using the trapezoidal method of approximation. Repeated measures ANOVA and contrasts using the Bonferroni adjustment were analyzed for all measures.

Experiment 2:

Subjects:

The subjects were 16 male Sprague-Dawley rats, 90 days of age, randomly assigned to one of four conditions:

 Group            Drug       Number of Animals
 Surgery        Prazosin         n=4
 Sham           Prazosin         n=4
 Surgery        Clonidine        n=4
 Sham           Clonidine        n=4

Surgery: Same as Experiment 1 (above).

Beam Walk Task (BW): Same as Experiment 1 (above).

Drug Doses:

Prazosin        Clonidine
0.4 mg/kg       4 mg/kg

Procedures

Pre-Injury:

Animals were given an acclimation period, then were trained to traverse the beam until they received scores of 7 on three consecutive days. Animals then underwent TBI or sham surgery per their assigned group.

Initial Deficit and Recovery:

Beginning 24 hours after surgery, rats were assessed for their initial BW deficit.

Reinstatement Evaluation:

At 1 month after surgery, recovered animals were tested on the beam walk task then given three single injections spaced three hours apart of their assigned drug. BW tests were performed at 1, 2 & 3 hours after injection.

One week after the first set of injections, a cross-over design was used so in the second phase all rats received the other drug from the one given previously and the testing regimen was repeated.

Statistical Analyses:

BW data was analyzed using t- tests and correlations were analyzed using pearson r.

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