Traumatic brain injury induces an adaptive immune response in the meningeal transcriptome that is amplified by aging.

Traumatic Brain Injury (TBI) is a major cause of disability and mortality, particularly among the elderly, yet our mechanistic understanding of how age renders the post-traumatic brain vulnerable to poor clinical outcomes and susceptible to neurological disease remains poorly understood. It is well established that dysregulated and sustained immune responses contribute to negative outcomes after TBI, however our understanding of the interactions between central and peripheral immune reservoirs is still unclear. The meninges serve as the interface between the brain and the immune system, facilitating important bi-directional roles in healthy and disease settings. It has been previously shown that disruption of this system exacerbates inflammation in age related neurodegenerative disorders such as Alzheimer's disease, however we have an incomplete understanding of how the meningeal compartment influences immune responses after TBI. Here, we examine the meningeal tissue and its response to brain injury in young (3-months) and aged (18-months) mice. Utilizing a bioinformatic approach, high-throughput RNA sequencing demonstrates alterations in the meningeal transcriptome at sub-acute (7-days) and chronic (1 month) timepoints after injury. We find that age alone chronically exacerbates immunoglobulin production and B cell responses. After TBI, adaptive immune response genes are up-regulated in a temporal manner, with genes involved in T cell responses elevated sub-acutely, followed by increases in B cell related genes at chronic time points after injury. Pro-inflammatory cytokines are also implicated as contributing to the immune response in the meninges, with ingenuity pathway analysis identifying interferons as master regulators in aged mice compared to young mice following TBI. Collectively these data demonstrate the temporal series of meningeal specific signatures, providing insights into how age leads to worse neuroinflammatory outcomes in TBI.

Delayed treatment with ceftriaxone reverses the enhanced sensitivity of TBI mice to chemically-induced seizures.

The pathophysiological changes that occur after traumatic brain injury (TBI) can lead to the development of post-traumatic epilepsy, a life-long complication of brain trauma. The etiology of post-traumatic epilepsy remains unknown, but TBI brains exhibit an abnormal excitatory / inhibitory balance. In this study, we examine how brain injury alters susceptibility to chemically-induced seizures in C57Bl/6J mice, and if pharmacological enhancement of glutamate transporters can reduce chronic post-traumatic seizures. We found that controlled cortical impact (CCI) mice display delayed susceptibility to pentylenetetrazol (PTZ)-induced seizures. While CCI mice have no change in seizure susceptibility at 7d post-injury (dpi), at 70dpi they have reduced latency to PTZ-induced seizure onset, higher seizure frequency and longer seizure duration. Quantification of glutamate transporter mRNA showed that levels of Scl1a2 and Scl1a3 mRNA were increased at 7dpi, but significantly decreased at 70dpi. To test if increased levels of glutamate transporters can ameliorate delayed-onset seizure susceptibility in TBI mice, we exposed a new cohort of mice to CCI and administered ceftriaxone (200mg/kg/day) for 14d from 55-70dpi. We found that ceftriaxone significantly increased Scl1a2 and Scl1a3 in CCI mouse brain at 70dpi, and prevented the susceptibility of CCI mice to PTZ-induced seizures. This study demonstrates cortical impact can induce a delayed-onset seizure phenotype in mice. Delayed (55dpi) ceftriaxone treatment enhances glutamate transporter mRNA in the CCI brain, and reduces PTZ-induced seizures in CCI mice.

Sequential Isolation of Microglia and Astrocytes from Young and Aged Adult Mouse Brains for Downstream Transcriptomic Analysis.

In aging, the brain is more vulnerable to injury and neurodegenerative disease, but the mechanisms responsible are largely unknown. Evidence now suggests that neuroinflammation, mediated by resident brain astrocyte and microglia populations, are key players in the generation of inflammatory responses and may influence both age related processes and the initiation/progression of neurodegeneration. Consequently, targeting these cell types individually and collectively may aid in the development of novel disease-modifying therapies. We have optimized and characterized a protocol for the effective sequential isolation of both microglia and astrocytes from the adult mouse brain in young and aged mice. We demonstrate a technique for the sequential isolation of these immune cells by using magnetic beads technology, optimized to increase yield and limit potential artifacts in downstream transcriptomic applications, including RNA-sequencing pipelines. This technique is versatile, cost-effective, and reliable for the study of responses within the same biological context, simultaneously being advantageous in reducing mice numbers required to assess cellular responses in normal and age-related pathological conditions.

The Effect of Traumatic Brain Injury on Sleep Architecture and Circadian Rhythms in Mice-A Comparison of High-Frequency Head Impact and Controlled Cortical Injury.

Traumatic brain injury (TBI) is a significant risk factor for the development of sleep and circadian rhythm impairments. In this study we compare the circadian rhythms and sleep patterns in the high-frequency head impact (HFHI) and controlled cortical impact (CCI) mouse models of TBI. These mouse models have different injury mechanisms key differences of pathology in brain regions controlling circadian rhythms and EEG wave generation. We found that both HFHI and CCI caused dysregulation in the diurnal expression of core circadian genes (Bmal1, Clock, Per1,2, Cry1,2) at 24 h post-TBI. CCI mice had reduced locomotor activity on running wheels in the first 7 d post-TBI; however, both CCI and HFHI mice were able to maintain circadian behavior cycles even in the absence of light cues. We used implantable EEG to measure sleep cycles and brain activity and found that there were no differences in the time spent awake, in NREM or REM sleep in either TBI model. However, in the sleep states, CCI mice have reduced delta power in NREM sleep and reduced theta power in REM sleep at 7 d post-TBI. Our data reveal that different types of brain trauma can result in distinct patterns of circadian and sleep disruptions and can be used to better understand the etiology of sleep disorders after TBI.

High-frequency head impact causes chronic synaptic adaptation and long-term cognitive impairment in mice.

Repeated head impact exposure can cause memory and behavioral impairments. Here, we report that exposure to non-damaging, but high frequency, head impacts can alter brain function in mice through synaptic adaptation. High frequency head impact mice develop chronic cognitive impairments in the absence of traditional brain trauma pathology, and transcriptomic profiling of mouse and human chronic traumatic encephalopathy brain reveal that synapses are strongly affected by head impact. Electrophysiological analysis shows that high frequency head impacts cause chronic modification of the AMPA/NMDA ratio in neurons that underlie the changes to cognition. To demonstrate that synaptic adaptation is caused by head impact-induced glutamate release, we pretreated mice with memantine prior to head impact. Memantine prevents the development of the key transcriptomic and electrophysiological signatures of high frequency head impact, and averts cognitive dysfunction. These data reveal synapses as a target of high frequency head impact in human and mouse brain, and that this physiological adaptation in response to head impact is sufficient to induce chronic cognitive impairment in mice.