ASER ROTHSTEIN 



75 



were not only taking up K+, but they were also exchanging cellular K+ for 

 K*^ in the medium. 



The leakage of K+ associated with active metabolism has been investigated 

 recently at very low K+ concentrations (47). Typical data are presented in 

 figure 5. If cells are suspended in 3 X io~* m/1. KCl plus glucose, at pH 4.5, 

 the K+ concentration drops rapidly to about 5 X io~* m/1. but no further. If 

 the initial concentration is 5 X io~* m/1., then no changes in concentration occur. 

 However, if the cells are suspended in distilled water, K+ leaks out of the cells 

 rapidly until its concentration in the medium builds up to 5 X io~^ m/1. Thus 

 at 5 X io~^ m/1. a steady state condition exists in which the rate of leakage and 

 uptake are exactly equal. In the absence of substrate, there is no steady state 

 condition, but K+ continues to leak out at a slow rate for long periods of time. 



Fig. 5. Gain or loss of K+ by 

 media of different initial K""" con- 

 centrations containing actively 

 metabolizing yeast cells. 



The kinetics of K+ leakage can be readily characterized if the K+ concentra- 

 tion can be maintained at levels below the steady state concentration, 5 X lo"^ 

 m/1. in this case. Ordinarily, it is difficult to study K+ leakage and uptake at 

 low K+ concentrations because the cells establish the steady state in a few 

 minutes. The problem has been resolved by passing the medium continuously 

 through a column of packed cells at a rate such that the medium is in contact 

 with the cells for only a short period of time. Samples of the medium are taken 

 continuously by a standard fraction collector and analyzed for ionic content. 

 By this technique K+ leakage or uptake can be studied in a system in which 

 the K+ concentration can be controlled at very low values. At pn 4.0, there is a 

 constant leakage of K+ from the cells at a rate of 3.4 mu/kg of cells each hour. 

 After glucose is added to the system, the rate of leakage increases about four 

 fold to 12.5 mm/kg hr. (fig. 6). At pH 2.0, the rate of leakage is markedly in- 

 creased, 16 mM/kg hr. without substrate, and 46 mM/kg hr. with substrate. 

 At the latter rate, one third of the cellular K+ leaks out in an hour. The de- 

 creased rate of leakage observed after 165 minutes in figure 6 at pn 2 is probably 



