VARIATION OF INTRACELLULAR INHIBITION WITH pH 725 



possess a rather fixed internal anion concentration and for an anion to 

 enter the cell, either a cation must be taken in simultaneously or an internal 

 anion released from the cell in exchange. The amount of exchangeable 

 anion in many cells is very low; for example, the chloride concentration in 

 nerve and muscle is usually less than 10 .mM. Of course, some cells have 

 mechanisms for the active transport of organic anions and the entrance of 

 a possible inhibitor anion would dei3end on how readily it would react with 

 this mechanism. 



It is also possible for the the ionic form to be active without penetrating 

 into the cell through an action in some manner on the cell surface. The 

 anions of the tricarboxylic acid cycle substrates exert small but definite 

 actions on the membrane properties of cardiac cells and, through these 

 changes, on the contractility (Webb and Hollander, 1956). These actions are 

 presumably unrelated to their metabolic role and in a similar manner the 

 anions of inhibitors could alter tissue function independently of any en- 

 zyme action. 



The plotting of equiactive concentrations of inhibitors, or their un- 

 dissociated forms, against pH provides a means of testing the activity of 

 the ionic forms and it was concluded by Simon and Beevers (1952), from the 

 results on many types of cells and substances, that the ionic form can be 

 of importance, at least over a certain range of pH. The ion would not have 

 to penetrate well to have a very significant effect on the inhibition in long- 

 term experiments, such as in studies on growth. The penetration of the 

 ion should actually reduce the inhibition produced by the uncharged form 

 in many cases, because the ion, instead of accumulating in the cell as it 

 is formed from the un-ionized inhibitor, will leave the cell. The results will 

 often depend on how long the cells are incubated with the inhibitor and 

 particularly on whether equilibrium has been reached or not. Even if only 

 the un-ionized form penetrates, in a i^H range much above the p^^, the 

 rate or entrance of HI may be very slow due to the low concentration gra- 

 dient and equilibrium will be achieved only slowly. If HI penetrates more 

 rapidly than I~, HI may enter initially and the concentration of I"" build 

 up in the cell, later to be reduced as I~ diffuses out of the cell. The best 

 conclusion is probably never to ignore the ionic form until it has been ex- 

 perimentally proved to be impermeant or inactive. 



Some confusion has occurred in the past from the use of the term active 

 form when applied to the effects of weak acids or bases on cells. The un- 

 charged form has frequently been called active when it has been found that 

 the cellular action is dependent on its concentration. However, within the 

 cell it may well not be the active form on the enzyme. It would be better 

 to distinguish between the permeant form, that which enters the cell pre- 

 dominantly, and the active form, the form that combines with and inhibits 

 the enzyme. These two forms are often different. Usually it is the ionized 



