Effects of early rolipram treatment on histopathological outcome after controlled cortical impact injury in mice

Abstract

Traumatic brain injury (TBI) pathology includes contusions, cavitation, cell death, all of which can be exacerbated by inflammation. We hypothesized that an anti-inflammatory drug, rolipram, may reduce pathology after TBI, since in several CNS injury models rolipram reduces inflammation and improves cell survival and functional recovery. Adult male C57BL/6 mice received a craniotomy over the right parietotemporal cortex. Vertically directed controlled cortical impact (CCI) injury was delivered. Naïve controls were used for comparison. At 30 min post-surgery, animals were treated with vehicle or rolipram (1 mg/kg), and then once per day for 3 days. On day 3, the brains were systematically sectioned and stained to visualize the resulting pathology using hematoxylin and eosin (H&E) staining and NeuN immunocytochemistry. Total parietotemporal cortical contusion and cavity volume were significantly increased in rolipram-treated as compared to vehicle-treated CCI animals. Contusion areas at specific bregma levels indicated a significant effect of drug across bregma levels. Neuronal cell loss in the dentate hilus and area CA3 of the hippocampus were similar between vehicle and rolipram-treated animals. Although rolipram is well known to reduce pathology and inflammation in several other CNS injury models, the pathology resulting from CCI was worsened with rolipram at this particular dose and administration schedule. These studies suggest that consideration of the unique characteristics of TBI pathology is important in the extrapolation of promising therapeutic interventions from other CNS injury models.

Fig. 1

Cortical contusions after CCI injury in mice. Representative images of H&E stained sections at −2.4 mm posterior from bregma for vehicle-treated (A) and rolipram-treated (B) mice. Scale bar 500 µm. Magnification of the contused cortex for vehicle (C) and rolipram (D) treatment. The contoured contusion and cavity areas are denoted by the dashed lines. Scale bar 250 µm. Total contusion volume quantification (E) and contusion areas across bregma levels (F). Mean±SEM, n=6/group, **p<0.01 for CCI+vehicle versus CCI+rolipram.

Fig. 2

Rolipram did not affect dentate hilar cell loss. Representative images of the dentate hilus immunostained with NeuN at bregma level −2.1 mm for vehicle-treated (A, B), and rolipram-treated (C, D) mice. Scale bar 100 µm. Estimated numbers of dentate hilus NeuN-positive cells between bregma levels −1.6 to −2.7 mm (E). Mean±SEM, n=6/group.

Fig. 3

Neuronal loss in area CA3 after CCI injury is not affected by rolipram treatment. Images taken at bregma level −1.9 mm for the ipsilateral side of vehicle-treated (A) and rolipram-treated animals (B) indicate small areas of neuronal loss (arrows). No significant cell loss was observed on the contralateral side with either vehicle (C) or rolipram treatment (D). Scale bar 100 µm. Quantification of CA3 cell loss between bregma levels −1.9 to −2.1 mm indicated a significant loss of neurons in both vehicle- and rolipram-treated animals on the ipsilateral side (E). Mean±SEM, n=6/group.