Measurement of Mitochondrial Toxicity Parameters in Embryonic Hippocampus

Abstract

Recent discoveries have focused on mitochondria functions in the neuroscience research for approaches to study mitochondria dysfunction in neurodegenerative diseases. Mitochondrion is one of the organelles that is possibly worst affected in cognitive impairments. These are known as “powerhouse” of the cell as they are the main source of generation of ATP through aerobic respiration. They have role in oxidative phosphorylation and metabolism, they play central role in cell differentiation, apoptosis, oxygen sensing and detoxification of reactive oxygen species, innate immunity, mitochondrial matrix calcium, and maintenance of cell quality and regulation of cytoplasmic. There is a relationship between mitochondrial dysfunction and cognitive disorder that may be related to certain neurotoxins or mutations in mitochondrial DNA as well as the nuclear. Evaluating compounds for mitochondrial toxicity is an important capability for evaluation of cognitive effects by drugs. Studying mitochondria isolated from individual mouse brain regions is a challenge because of small amount of the available brain tissue. There are conventional techniques for isolation and purification of mitochondria from ventral midbrain, hippocampus, or striatum. The utilization of alcohol within pregnancy impairs the development of the unborn offspring and can lead to a plethora of anatomical, behavioral, and cognitive abnormalities.

In here, a method for isolation of brain mitochondria from mouse is described. The method utilizes a refrigerated tabletop microtube centrifuge, and produces research grade quality mitochondria in amounts sufficient for performing multiple enzymatic and functional assays, thereby eliminating the necessity for pooling mouse brain tissue. A method for measuring ROS measurement, mitochondrial membrane potential, mitochondrial swelling, cytochrome c release, and mtDNA alterations after exposure to drugs is also included.