Blast waves from detonated military explosive reduce GluR1 and synaptophysin levels in hippocampal slice cultures

Abstract

Explosives create shockwaves that cause blast-induced neurotrauma, one of the most common types of traumatic brain injury (TBI) linked to military service. Blast-induced TBIs are often associated with reduced cognitive and behavioral functions due to a variety of factors. To study the direct effects of military explosive blasts on brain tissue, we removed systemic factors by utilizing rat hippocampal slice cultures. The long-term slice cultures were briefly sealed air-tight in serum-free medium, lowered into a 37 °C water-filled tank, and small 1.7-gram assemblies of cyclotrimethylene trinitramine (RDX) were detonated 15 cm outside the tank, creating a distinct shockwave recorded at the culture plate position. Compared to control mock-treated groups of slices that received equal submerge time, 1–3 blast impacts caused a dose-dependent reduction in the AMPA receptor subunit GluR1. While only a small reduction was found in hippocampal slices exposed to a single RDX blast and harvested 1–2 days later, slices that received two consecutive RDX blasts 4 min apart exhibited a 26–40% reduction in GluR1, and the receptor subunit was further reduced by 64–72% after three consecutive blasts. Such loss correlated with increased levels of HDAC2, a histone deacetylase implicated in stress-induced reduction of glutamatergic transmission. No evidence of synaptic marker recovery was found at 72 h post-blast. The presynaptic marker synaptophysin was found to have similar susceptibility as GluR1 to the multiple explosive detonations. In contrast to the synaptic protein reductions, actin levels were unchanged, spectrin breakdown was not detected, and Fluoro-Jade B staining found no indication of degenerating neurons in slices exposed to three RDX blasts, suggesting that small, sub-lethal explosives are capable of producing selective alterations to synaptic integrity. Together, these results indicate that blast waves from military explosive cause signs of synaptic compromise without producing severe neurodegeneration, perhaps explaining the cognitive and behavioral changes in those blast-induced TBI sufferers that have no detectable neuropathology.

Neuronal susceptibility to beta‐amyloid toxicity and ischemic injury involves histone deacetylase‐2 regulation of endophilin‐B1

“…Since HDAC2 suppresses Endo-B1b/c expression (see Figure 2) while mediating Aβ toxicity (see Figure 4), it prompted us to determine if HDAC2 also influences Aβ -induced changes in mitochondrial membrane potential. Consistent with the observation that HDAC2 knockdown sustained Endo-B1b/c expression and cell viability despite Aβ 25-35 treatment (Figure 4), HDAC2 knockdown also maintained mitochondrial membrane potential, as measured by JC-1 fluorescence, in the presence of Aβ [25][26][27][28][29][30][31][32][33][34][35] ( Figure 5A). These findings collectively suggest that HDAC2 suppression of Endo-B1b/c mediates a decline in mitochondrial function caused by Aβ toxicity.…”

“…full-length oligomerized Aβ 1-42 ( Figure 3A), thereby rationalizing the use of Aβ [25][26][27][28][29][30][31][32][33][34][35] to emulate Aβ toxicity in subsequent experiments.…”

“…Treatment with Aβ [25][26][27][28][29][30][31][32][33][34][35] resulted in significant activation of caspase-3 and neuronal cell death ( Figure 3B,C). Aβ [25][26][27][28][29][30][31][32][33][34][35] treatment of cultured neurons also significantly increased HDAC2 protein and decreased levels of N-terminally acetylated histone H3 ( Figure 3D,E). Infection with lentivirus expressing shRNA against HDAC2 prior to Aβ [25][26][27][28][29][30][31][32][33][34][35] treatment strongly mitigated Aβ [25][26][27][28][29][30][31][32][33][34][35] toxicity and histone deacetylation ( Figure 3B,C,E).…”

“…To create a more stable reagent to study Aβ toxicity in vitro, a smaller peptide A β [25][26][27][28][29][30][31][32][33][34][35] has been utilized (1,21). We previously reported that Aβ [25][26][27][28][29][30][31][32][33][34][35] treatment induced neuronal caspase-3 activation that was attenuated by Endo-B1c overexpression (37). We replicated these findings using The levels of the Endo-B1a and Endo-B1b/c bands in wild-type and HDAC2KO neurons in culture were determined for individual blots, as represented in D, by densitometry.…”

“…Excitotoxicity is a common factor in models of brain injury including traumatic and epileptic insults. These paradigms also show HDAC2 upregulation and benefit from the application of HDAC inhibitors (13,18,33). Excitotoxicity may be mediated through HDAC2-dependent epigenetic changes impinging on mitochondria, which play a major role in dictating cellular susceptibility to excitotoxicity.…”