Chapter 8 Measurement of membrane permeability and permeability transition of mitochondria

Abstract

This chapter presents experimental procedures designed to measure apoptosis associated mitochondrial membrane permeabilization, both in intact cells and in isolated mitochondria. Mitochondria are essential for the evolution of complex animals in aerobic conditions. They carry out most cellular oxidations and produce the bulk of an animal cell's ATP. The permeability transition (PT) pore, also called the mitochondrial megachannel or multiple conductance channel, is a dynamic multiprotein complex located at the contact site between the inner and the outer mitochondrial membranes, one of the critical sites of metabolic coordination among the cytosol and the mitochondrial intermembrane and matrix spaces. In isolated mitochondria, opening of the PT pore causes matrix swelling with consequent distension and local disruption of the mitochondrial outer membrane, release of soluble products from the intermembrane space, dissipation of the mitochondrial inner transmembrane potential, and release of small molecules up to 1500 Da from the matrix, through the inner membrane. In several models of apoptosis, pharmacological inhibition of the PT pore is cytoprotective, suggesting that opening of the PT pore can be rate limiting for the death process. Irrespective of this possibility, mitochondrial membrane permeabilization is a general feature of apoptosis.