A Comparative Study of Two Blast-Induced Traumatic Brain Injury Models: Changes in Monoamine and Galanin Systems Following Single and Repeated Exposure

Abstract

Repeated mild blast-induced traumatic brain injury (rmbTBI), caused by recurrent exposure to low levels of explosive blast, is a significant concern for military health systems. However, the pathobiology of rmbTBI is currently poorly understood. Animal models are important tools to identify the molecular changes of rmbTBI, but comparisons across different models can present their own challenges. In this study, we compared two well-established rodent models of mbTBI, the "KI model" and the "USU/WRAIR model." These two models create different pulse forms, in terms of peak pressure and duration. Following single and double exposures to mild levels of blast, we used in situ hybridization (ISH) to assess changes in mRNA levels of tyrosine hydroxylase (TH), tryptophan hydroxylase (TPH2), and galanin in the locus coeruleus (LC) and dorsal raphe nucleus (DRN). These systems and their transmitters are known to mediate responses to stress and anxiety. We found increased mRNA levels of TH, TPH2 and galanin in the LC and DRN of single-exposed rats relative to sham rats in the KI but not the USU/WRAIR model. Sham mRNA values measured in the USU/WRAIR model were substantially higher than their KI counterparts. Double exposure caused similarly significant increases in mRNA values in the KI model but not the USU/WRAIR model, except TPH2 and galanin levels in the DRN. We detected no cumulative effect of injury in either model at the used inter-injury interval (30 min), and there were no detectable neuropathological changes in any experimental group at 1 day post-injury. The apparent lack of early response to injury as compared to sham in the USU/WRAIR model is likely caused by stressors (e.g., transportation and noise), associated with the experimental execution, that were absent in the KI model. This study is the first to directly compare two established rodent models of rmbTBI, and to highlight the challenges of comparing findings from different animal models. Additional studies are needed to understand the role of stress, dissect the effects of psychological and physical injuries and to identify the window of increased cerebral vulnerability, i.e., the inter-injury interval that results in a cumulative effect following repeated blast exposure.

Keywords: animal models; anxiety; catecholamines; dorsal raphe nucleus; locus coeruleus; neuropeptide; post-traumatic stress disorder; transmitter coexistence.

Figure 1

A schematic representation of the KI model (A) and USU/WRAIR model (B). The positive phase durations and peak pressures are measured at the animal level in both models.

Figure 2

ISH analysis of transcript levels for TH and galanin in the LC following exposure to single or double mbTBI, using two different models of TBI, in two different laboratories. Quantification of transcript levels of TH (a) and galanin (b) revealed that both were significantly increased bilaterally at 1 day post-exposure in the single and double exposed groups, relative to their respective shams using the KI model. While the same trend was seen in exposed groups vs. shams in the USU/WRAIR model, the elevations were not statistically significant in any of the transcripts. There did not appear to be a cumulative effect of repeated exposure in either model. (c–r) Representative dark field ISH photomicrographs of emulsion-dipped sections show the distribution and levels of TH (c–j) and galanin (k–r) transcripts levels. TH mRNA levels in the single (c), and double (d) exposed groups in the LC, relative to sham single (e), and double (f) groups using the KI model. (g–j) Show TH mRNA distribution and levels in the single (g), and double (h) exposed groups using the USU/WRAIR mbTBI model, and their respective sham groups; single (i), and double (j). Photomicrographs (k–r) show galanin transcript levels: the single (k) and double (l) exposed groups using the KI model, and their respective shams, single (m) and double (n); the single (o), and double exposed (p), and single (q), and double (r) sham groups, using the USU/WRAIR model. Data are presented as mean ± SEM (^**p < 0.01, ^****p < 0.0001). ISH, in situ hybridization; LC, locus coeruleus; TH, tyrosine hydroxylase.

Figure 3

ISH analysis of transcript levels for TPH2 and galanin in the mid/caudal DRN following exposure to single or double mbTBI, using two different models of TBI, in two different laboratories. Quantification of transcript levels of TPH2 (a) and galanin (b) mRNA levels showed a significant increase at 1 day post-exposure in the single and double exposed groups relative to their respective shams using the KI model. While the same trend of an elevation was also observed in exposed groups vs. shams using the USU/WRAIR model, this was only statistically significant in the double exposure group. There did not appear to be a cumulative effect of repeated exposure in either model. (c–r) Representative dark field ISH photomicrographs of emulsion-dipped sections show distribution and levels of TPH2 (c–j) and galanin (k–r) transcripts. TPH2 mRNA levels in the single (c), and double (d) exposed groups, relative to sham single (e), and double (f) groups using the KI model. (g–j) Show single (g), and double (h) exposed groups using the USU/WRAIR model, and their respective single (i), and double (j) sham groups. Photomicrographs (k-r) show galanin transcript levels: the single (k) and double (l) exposed groups using the KI model, and their respective shams, single (m) and double (n); the single (o), and double exposed (p), and single (q), and double (r) sham groups, using the USU/WRAIR model. Data are presented as mean ± SEM (^*p < 0.05, ^**p < 0.01, ^****p < 0.0001). ISH, in situ hybridization; DRN, dorsal raphe nucleus; TPH2, tryptophan hydroxylase 2.