title: Acute death of astrocytes in blast-exposed rat organotypic hippocampal slice cultures.
abstract: Blast traumatic brain injury (bTBI) affects civilians, soldiers, and veterans worldwide and presents significant health concerns. The mechanisms of neurodegeneration following bTBI remain elusive and current therapies are largely ineffective. It is important to better characterize blast-evoked cellular changes and underlying mechanisms in order to develop more effective therapies. In the present study, our group utilized rat organotypic hippocampal slice cultures (OHCs) as an in vitro system to model bTBI. OHCs were exposed to either 138 ± 22 kPa (low) or 273 ± 23 kPa (high) overpressures using an open-ended helium-driven shock tube, or were assigned to sham control group. At 2 hours (h) following injury, we have characterized the astrocytic response to a blast overpressure. Immunostaining against the astrocytic marker glial fibrillary acidic protein (GFAP) revealed acute shearing and morphological changes in astrocytes, including clasmatodendrosis. Moreover, overlap of GFAP immunostaining and propidium iodide (PI) indicated astrocytic death. Quantification of the number of dead astrocytes per counting area in the hippocampal cornu Ammonis 1 region (CA1), demonstrated a significant increase in dead astrocytes in the low- and high-blast, compared to sham control OHCs. However only a small number of GFAP-expressing astrocytes were co-labeled with the apoptotic marker Annexin V, suggesting necrosis as the primary type of cell death in the acute phase following blast exposure. Moreover, western blot analyses revealed calpain mediated breakdown of GFAP. The dextran exclusion additionally indicated membrane disruption as a potential mechanism of acute astrocytic death. Furthermore, although blast exposure did not evoke significant changes in glutamate transporter 1 (GLT-1) expression, loss of GLT-1-expressing astrocytes suggests dysregulation of glutamate uptake following injury. Our data illustrate the profound effect of blast overpressure on astrocytes in OHCs at 2 h following injury and suggest increased calpain activity and membrane disruption as potential underlying mechanisms.
authors.affiliation: Department of Neurosurgery, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Department of Cell Biology, Neurobiology & Anatomy, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Clement J. Zablocki Veterans Affairs Medical Center, Milwaukee, Wisconsin, United States of America.
authors.forename: Anna P
authors.initials: AP
authors.lastname: Miller
authors.affiliation: Department of Neurosurgery, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Clement J. Zablocki Veterans Affairs Medical Center, Milwaukee, Wisconsin, United States of America.
authors.forename: Alok S
authors.initials: AS
authors.lastname: Shah
authors.affiliation: Department of Neurosurgery, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Clement J. Zablocki Veterans Affairs Medical Center, Milwaukee, Wisconsin, United States of America.
authors.forename: Brandy V
authors.initials: BV
authors.lastname: Aperi
authors.affiliation: Department of Neurosurgery, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Department of Cell Biology, Neurobiology & Anatomy, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Clement J. Zablocki Veterans Affairs Medical Center, Milwaukee, Wisconsin, United States of America.
authors.forename: Shekar N
authors.initials: SN
authors.lastname: Kurpad
authors.affiliation: Department of Neurosurgery, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Clement J. Zablocki Veterans Affairs Medical Center, Milwaukee, Wisconsin, United States of America.
authors.forename: Brian D
authors.initials: BD
authors.lastname: Stemper
authors.affiliation: Department of Neurosurgery, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Department of Cell Biology, Neurobiology & Anatomy, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America.|Clement J. Zablocki Veterans Affairs Medical Center, Milwaukee, Wisconsin, United States of America.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Glavaski-Joksimovic
mesh_terms: D000818:Animals; D017209:Apoptosis; D001253:Astrocytes / Q000378:metabolism / Q000473:pathology; D001753:Blast Injuries / Q000209:etiology / Q000378:metabolism / Q000473:pathology; D001930:Brain Injuries / Q000209:etiology / Q000378:metabolism / Q000473:pathology; D016923:Cell Death; D002463:Cell Membrane Permeability; D004195:Disease Models, Animal; D027342:Excitatory Amino Acid Transporter 2 / Q000235:genetics / Q000378:metabolism; D005107:Explosions; D015870:Gene Expression; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D006624:Hippocampus; D059748:Proteolysis; D051381:Rats; D046509:Tissue Culture Techniques
chemical_list: D027342:Excitatory Amino Acid Transporter 2; D005904:Glial Fibrillary Acidic Protein; C493395:Slc1a2 protein, rat