Lateral fluid percussion injury

All experimental procedures were approved by the Institutional Animal Care and Use Committee and were conducted in accord with the recommendations provided in the Guide for the Care and Use of Laboratory Animals. Protocols were designed to minimize pain and discomfort during the injury procedure and recovery. Lateral FPI was carried out similar to that described previously.^17–19 Briefly, rats were initially anesthetized using 5% isoflurane with a 1:1 N₂O/O₂ mixture and then maintained with a 2.5% isoflurane with 1:1 air/O₂ mixture by a face mask. Animals were mounted on the stereotaxic frame; a 4.8-mm-diameter craniotomy was carefully made midway between bregma and lambda. The craniotomy used for hub placement was centered at 4.4 mm from midline, placing the epicenter of the injury over the right parietal cortex. A hub (modified from a 20-gauge needle) was implanted into the burr hole and affixed to the skull by contact adhesive and dental cement. Once the assembly was secured, the rat was removed from the anesthesia and allowed to regain its toe pinch reflex. Immediately upon regaining this reflex, the rat was injured using an FPI device and a pressure of 1.5 atmosphere (atm) over base room pressure. Previous studies have shown that this injury magnitude does not cause visible brain damage, nor overt hippocampal damage, but does cause axonal injury as detected by diffusion tensor imaging.²⁰ Acute neurological responses were recorded post-injury. On average, injured animals regain their response to toe pinch at 72.4 ± 5.6 sec, tail pain reflexes by 100.1 ± 5.3 sec, and their righting response (ability to right itself three consecutive times after being placed on its back) by 387 ± 11.2 sec. By comparison, sham-operated animals require only 36.3 ± 8.4 sec to regain their righting response (after subtraction of the tail pinch reflex). Post-injury, the hub and surrounding dental cement were immediately removed and the incision closed by wound clips. Sham-operated animals received all the aforementioned surgical procedures except hub implantation and the injury. Animals' body temperature was maintained at 37°C during the surgery using a rectal thermometer coupled to a heating pad.