Permeabilized muscle fibre
Historical Perspective
The skinned muscle fibre technique was originally invented by Endo (Endo 1967), in dependence on the mechanically skinned skeletal muscle fiber method, first introduced by Natori (Natori 1954). It was further adapted and modified by Veksler (Veksler et al. 1987) and Saks (Saks et al. 1998) for cardiac muscle fibres and also for skeletal muscle fibres. Then as now, it was of great importance βto have the characters closely resembling those of the intact fibreβ (Natori 1954). The idea was: βfrom a physiological point of view, it is desirable to study the contractile system in a state as close to the living state as possibleβ (Saida and Nonomura 1978).
Method
After removal of the tissue form the species, the procedure allots mechanical separation of muscle tissue in a relaxing solution. Thereafter, muscle tissue is incubated in saponin by gentle agitation. Saponin, a chemical permeabilization agent of plant origin, selectively attacks cholesterol rich (plasma) membranes of cells or muscle ο¬bers, leaving intracellular membrane structures (mitochondria and ER) as well as myoο¬laments and the cytoskeleton intact. Consequently, membrane barrier integrity is lost and the cytosol and all solutes are washed out and the composition of the intracellular milieu is quickly equilibrated with the incubation medium. The mitochondria (in situ) are then able to use various substrates within a given protocol.
References
1.Endo M (1967) Regulation of contraction relaxation of cycle (in japanese). . Gen Assoc Jpn Med Congr Proc. XVII. (1):193-197 2.Natori H (1954) The property and contraction process of isolated myofibrils. Jikei Med J, vol 1. First published 3.Saida K, Nonomura Y (1978) Characteristics of ca2+- and mg2+-induced tension development in chemically skinned smooth muscle fibers. J Gen Physiol, vol 72. First published 4.Saks VA, Veksler VI, Kuznetsov AV, Kay L, Sikk P, Tiivel T, Tranqui L, Olivares J, Winkler K, Wiedemann F, Kunz WS (1998) Permeabilized cell and skinned fiber techniques in studies of mitochondrial function in vivo. Mol Cell Biochem 184 (1-2):81--100 5.Veksler VI, Kuznetsov AV, Sharov VG, Kapelko VI, Saks VA (1987) Mitochondrial respiratory parameters in cardiac tissue: A novel method of assessment by using saponin-skinned fibers. Biochim Biophys Acta 892 (2):191--196
