TRICARBOXYLATE CYCLE 97 



than pyruvate oxidation by iodoacetate, so one can visualize a reasonably 

 selective action over 10-30 min if the proper concentration is chosen, with 

 the selectivity decreasing with time. There is one fact which is strongly 

 established: when iodoacetate or iodoacetamide is used at 1 mM or above, 

 there can be no selective block of the EM pathway. Indeed, in most cases 

 concentrations well below 1 mM are by no means selective. These concen- 

 trations are those in the regions of 3-PGDH and the cycle. In cellular prep- 

 arations the external concentration in the medium may be higher than 

 within the cell, and this will vary greatly with the permeability properties 

 of the cells and the pH, but most of the work discussed in this section in- 

 dicates that iodoacetate inhibits to some extent the oxidation of pyruvate 

 in cells when present in concentrations between 0.1 and 1 mikf in the me- 

 dium. If a selective EM pathway block in a particular tissue is desired, it 

 is necessary to examine the effects of different concentrations of iodoacetate 

 on glycolysis and pyruvate oxidation so as to obtain evidence that under 

 the particular conditions such a block can be achieved. Lower concentra- 

 tions than are usually used would be expected to be more effective. I believe 

 that it is possible in most cells to obtain an inhibition of the EM pathway 

 which is selective enough for relating this pathway to some cell activity, 

 but to do this requires the proper preliminary experiments to establish the 

 optimal conditions. This matter will be taken up again when the effects of 

 iodoacetate on cellular function are discussed. 



General Effects on the Cycle and Mitochondria 



The data presented in Tables 1-13 and 1-14 suggest that several steps in 

 the cycle, in addition to the initial reaction of pyruvate, are sensitive to 

 iodoacetate, but the results are often so inconsistent that it is difficult to 

 evaluate the importance of these inhibitions. a-Ketoglutarate oxidase is 

 perhaps as sensitive as pyruvate oxidase, while isocitrate dehydrogenase, 

 succinate oxidase, and malate dehydrogenase are moderately inhibited. The 

 over-all effect of these simultaneous inhibitions on the cycle may be greater 

 than any single inhibition, but there have been no studies of these over-all 

 effects in isolated mitochondria. Several reports on citrate levels as influenc- 

 ed by iodoacetate have been made but are difficult to interpret. Iodoace- 

 tate does not act like fluoroacetate as far as is known and does not form an 

 inhibitory iodocitrate. Kalnitsky (1948) showed in kidney homogenates that 

 only fluoroacetate of all the haloacetates leads to citrate accumulation. One 

 would indeed expect iodoacetate to decrease citrate levels, due to either a 

 reduction in the formation of pyruvate or the utilization of pyruvate, and 

 such has been found in mammary gland by Terner (1955). However, Gal et 

 al. (1956) reported some citrate accumulation in the kidneys of rats given 

 1.8 mg/kg iodoacetate ( + 65% ) while citrate decreases in the liver ( — 37% ), 

 but only two animals were used. Citrate fermentation in Aspergillus niger 



