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 XFe24 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.
Key wordsNeurotrauma Traumatic brain injury Mitochondria Mitochondrial bioenergetics Brain metabolism Oxidative phosphorylation Therapeutic drugs preclinical screening
The views of the authors do not purport or reflect the position of the Department of the Army or the Department of Defense (para 4-3, AR 360-5). The authors declare that there are no conflicts of interest regarding this protocol. This research is funded by Combat Casualty Care Research Program (H_026_2014_WRAIR), and NIH/NINDS grant NS 48191 (PGS).
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