Advanced and High-Throughput Method for Mitochondrial Bioenergetics Evaluation in Neurotrauma.
Mitochondrial dysfunction is one of the key posttraumatic neuropathological events observed in various experimental models of traumatic brain injury (TBI). The extent of mitochondrial dysfunction has been associated with the severity and time course of secondary injury following brain trauma. Critically, several mitochondrial targeting preclinical drugs used in experimental TBI models have shown improved mitochondrial bioenergetics, together with cortical tissue sparing and cognitive behavioral outcome. Mitochondria, being a central regulator of cellular metabolic pathways and energy producer of cells, are of a great interest for researchers aiming to adopt cutting-edge methodology for mitochondrial bioenergetics assessment. The traditional way of mitochondrial bioenergetics analysis utilizing a Clark-type oxygen electrode (aka. oxytherm) is time-consuming and labor-intensive. In the present chapter, we describe an advanced and high-throughput method for mitochondrial bioenergetics assessments utilizing the Seahorse Biosciences XF(e)24 Flux Analyzer. This allows for simultaneous measurement of multiple samples with higher efficiency than the oxytherm procedure. This chapter provides helpful guidelines for conducting mitochondrial isolation and studying mitochondrial bioenergetics in brain tissue homogenates following experimental TBI.
Brain Trauma Neuroprotection and Neurorestoration Branch, Center for Military Psychiatry and Neuroscience, Walter Reed Army Institute of Research, Silver Spring, MD, USA. jignesh.d.pandya.civ@mail.mil.
Jignesh D
JD
Pandya
Spinal Cord and Brain Injury Research Center, University of Kentucky, Lexington, KY, USA.
Patrick G
PG
Sullivan
Brain Trauma Neuroprotection and Neurorestoration Branch, Center for Military Psychiatry and Neuroscience, Walter Reed Army Institute of Research, Silver Spring, MD, USA.
Lai Yee
LY
Leung
Brain Trauma Neuroprotection and Neurorestoration Branch, Center for Military Psychiatry and Neuroscience, Walter Reed Army Institute of Research, Silver Spring, MD, USA.
Frank C
FC
Tortella
Brain Trauma Neuroprotection and Neurorestoration Branch, Center for Military Psychiatry and Neuroscience, Walter Reed Army Institute of Research, Silver Spring, MD, USA.
Deborah A
DA
Shear
Brain Trauma Neuroprotection and Neurorestoration Branch, Center for Military Psychiatry and Neuroscience, Walter Reed Army Institute of Research, Silver Spring, MD, USA.
Ying
Y
Deng-Bryant
D000818:Animals; D019069:Cell Respiration / Q000187:drug effects; D055808:Drug Discovery; D004353:Drug Evaluation, Preclinical; D004734:Energy Metabolism; D057166:High-Throughput Screening Assays; D008297:Male; D055432:Metabolomics / Q000379:methods; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D010085:Oxidative Phosphorylation; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D020196:Trauma, Nervous System / Q000188:drug therapy / Q000209:etiology* / Q000378:metabolism*
D017382:Reactive Oxygen Species
Brain metabolism; Mitochondria; Mitochondrial bioenergetics; Neurotrauma; Oxidative phosphorylation; Therapeutic drugs preclinical screening; Traumatic brain injury