Midline (central) fluid percussion model of traumatic brain injury in pediatric and adolescent rats

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OBJECTIVE

Experimental traumatic brain injury (TBI) models hold significant validity to the human condition, with each model replicating a subset of clinical features and symptoms. TBI is the leading cause of mortality and morbidity in children and teenagers; thus, it is critical to develop preclinical models of these ages to test emerging treatments. Midline fluid percussion injury (FPI) might best represent mild and diffuse clinical brain injury because of the acute behavioral deficits, the late onset of behavioral morbidities, and the absence of gross histopathology. In this study, the authors sought to adapt a midline FPI to postnatal day (PND) 17 and 35 rats. The authors hypothesized that scaling the craniectomy size based on skull dimensions would result in a reproducible injury comparable to the standard midline FPI in adult rats.

METHODS

PND17 and PND35 rat skulls were measured, and trephines were scaled based on skull size. Custom trephines were made. Rats arrived on PND10 and were randomly assigned to one of 3 cohorts: PND17, PND35, and 2 months old. Rats were subjected to midline FPI, and the acute injury was characterized. The right reflex was recorded, injury-induced apnea was measured, injury-induced seizure was noted, and the brains were immediately examined for hematoma.

RESULTS

The authors’ hypothesis was supported; scaling the trephines based on skull size led to a reproducible injury in the PND17 and PND35 rats that was comparable to the injury in a standard 2-month-old adult rat. The midline FPI suppressed the righting reflex in both the PND17 and PND35 rats. The injury induced apnea in PND17 rats that lasted significantly longer than that in PND35 and 2-month-old rats. The injury also induced seizures in 73% of PND17 rats compared with 9% of PND35 rats and 0% of 2-month-old rats. There was also a significant relationship between the righting reflex time and presence of seizure. Both PND17 and PND35 rats had visible hematomas with an intact dura, indicative of diffuse injury comparable to the injury observed in 2-month-old rats.

CONCLUSIONS

With these procedures, it becomes possible to generate brain-injured juvenile rats (pediatric [PND17] and adolescent [PND35]) for studies of injury-induced pathophysiology and behavioral deficits, for which rational therapeutic interventions can be implemented.

ABBREVIATIONS FPI = fluid percussion injury; PND = postnatal day; TBI = traumatic brain injury.

Article Information

Correspondence Rachel K. Rowe: Barrow Neurological Institute, Phoenix Children’s Hospital, Phoenix, AZ. rkro222@email.arizona.edu.

INCLUDE WHEN CITING Published online April 20, 2018; DOI: 10.3171/2018.1.PEDS17449.

Disclosures The authors report no conflict of interest concerning the materials or methods used in this study or the findings specified in this paper.

© AANS, except where prohibited by US copyright law.

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Figures

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    Experimental timeline and comparison of developmental stages in rats and humans. The rats were subjected to TBI at PND17 before the onset of puberty, at PND35 during puberty, or at 2 months, which is after full sexual maturation and onset of adulthood in the rat. Puberty in humans occurs with an average age range of 10–14 years old (yo). Full sexual maturation and adulthood in humans occurs at approximately 18–24 years of age.

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    Trephines adapted for the different stages of rat development. Custom-designed and machined 3.0- and 4.0-mm-diameter trephines (left and center, respectively) are displayed with the 4.7-mm trephine used in adult rats (right).

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    Cranial surgery for hub placement in the pediatric (PND17) rat. A: The head of the rat is swabbed with alcohol and Betadine and secured in a stereotactic frame with a continuous flow of inhaled isoflurane via a nosecone. B: A midline scalp incision is made to expose the skull, and the overlying fascia is removed. C: Vetbond (3M) tissue adhesive is applied to secure a thin (translucent) cross-section of weed trimmer line directly on the midline between the bregma and lambda sutures. D: Using the weed trimmer line as an anchor, a 3.0-mm-diameter trephine is used to perform a craniectomy to expose the underlying dura. E: Small drops of cyanoacrylate gel are placed on the outside of the constructed injury hub, and the hub is centered atop the craniectomy. F and G: After the cyanoacrylate gel dries, the injury hub is covered in methyl methacrylate cement, and the injury hub is filled with saline.

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    Cranial surgery for hub placement in the adolescent (PND35) rat. A: The head of the rat is shaved, swabbed with alcohol and Betadine, and secured in a stereotactic frame with a continuous flow of inhaled isoflurane via a nosecone. B: A midline scalp incision is made to expose the skull, and the overlying fascia is removed. C: A Dremel tool with a burr bit is used to make a pilot hole to anchor the trephine via a centering pin. D: A 4.0-mm-diameter trephine is used to perform the craniectomy to expose the underlying dura. E: Small drops of cyanoacrylate gel are placed on the outside of the constructed injury hub, and the hub is centered atop the craniectomy. F: After the cyanoacrylate gel dries, the injury hub is covered in methyl methacrylate cement, and the injury hub is filled with saline.

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    Midline FPI in the PND17 and PND35 rats. A and B: The injury hub is filled with sterile saline and connected to the male fitting of the injury device. PND17 rats are connected to the device via an extension tube (A), and PND35 rats are connected directly to the device (B). C and D: The PND17 (C) or PND35 (D) rat is positioned and held parallel to the ground on its right side. The pendulum is released once a positive toe-pinch response is achieved. E: The rats are placed supine on a heating pad and monitored until restoration of the righting reflex. Of note, in this adaptation we observed a high incidence of acute postinjury seizure in PND17 rats. F: The condition/appearance of the surgical site and brain tissue beneath the injury site is observed for herniation and hematoma (left, uninjured sham; right, brain-injured rat).

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    Acute physiological characterization of the adapted midline FPI. A: Midline FPI suppressed the righting reflex response in all rats. PND17 rats had a significantly shorter righting reflex time than did PND35 rats. However, there was no difference between PND17 (n = 11) and PND35 (n = 11) rats and the 2-month-old adult rats (n = 11). B: Midline FPI induced apnea in juvenile but not adult rats. PND17 rats had significantly longer apnea durations than those for PND35 and 2-month-old adult rats. C: Midline FPI induced seizures in PND17 but not adult rats; 73% of PND17 rats experienced seizures compared with 9% of PND35 rats and 0% of adults. D: There was a significant relationship between righting reflex times and the presence of seizure in which righting reflex times decreased by 92.26 seconds on average if the animal had a seizure. Data are presented as the mean ± SEM; *p < 0.05. Sham values were not included in the analyses.

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