GENERAL METABOLISM in vitro 75 



phorylation. With such preparations it is possible to study the 

 details of metabolism either in terms of the individual, or as a 

 sequence of, enzymic reactions. Dispersions can be prepared in 

 distilled water, various buffers, salines or in isotonic sucrose. 

 Generally the maintenance of metabolism in suspensions requires 

 more exacting conditions than in the slice, since disintegration of 

 cellular structures permits many enzymes to assume degradative 

 rules. Co-factors usually added include magnesium, calcium and 

 potassium salts, inorganic phosphate, nucleotides such as adeno- 

 sine triphosphate, components of the cytochrome system such as 

 cytochrome-c and diphosphopyridine nucleotide, and substrates 

 such as pyruvate together with fumarate or malate. Respiration in 

 such preparations is not maintained as in slices, oxygen uptake 

 decreasing markedly after 45 min at 37 °C. 



Particulate Preparations 



Dispersions of brain, containing many systems actively degrad- 

 ing phosphates, are not ideally suited to the detailed study of 

 oxidative phosphorylation. For this purpose particulate fractions 

 can be isolated which contain essentially the total oxidative capacity 

 of the tissue. In comparison with other tissues, principally liver 

 (see Lindberg and Ernster, 1954), relatively little work has been 

 carried out with brain particles. After the disintegration of the 

 tissue in a suitable medium, usually 0-25 M sucrose containing 

 ethylenediaminetetra-acetate, fractions containing nuclei, mito- 

 chondria and microsomes can be obtained by differential centri- 

 fugation. The mxitochondria are usually obtained free from major 

 cellular structures (Brody and Bain, 1951; Abood et al., 1952; 

 Christie et al, 1953; Hesselbach and Dubuy, 1953; Brody and 

 Bain, 1954; Narayanaswami and Mcllwain, 1954; Jordan and 

 March, 1956; Aldridge, 1957; Dubuy and Hesselbach, 1958). The 

 scheme of fractionation described by Brody and Bain (1952) has 

 been almost universally accepted as providing a suitable separation 

 of the main groups of particles in dispersions of whole brain and is 

 illustrated in Fig. 10. The groups of particles obtained are far 

 from homogeneous and also suffer from a degree of overlap with 

 the preceding and following groups. The nuclear fraction has been 

 subdivided into 5-7 distinct sub-fractions (Heald, 1959) and the 

 mitochondrial fraction contains at least four distinct groups of 

 particles (Hebb and Whittaker, 1958). Similar subdivisions were 



