FATIGUE-PRODUCTS OF NERVE-CENTERS 525 



find that acids are set free in the central nervous system as a result 

 of its activity or fatigue. The actual demonstration of an increase in 

 acidity of the brain-substance as a result of prolonged excitation has 

 proved difficult on account of the slightness of the change of hydrogen 

 ion concentration which is involved, owing to the buffer-action of the 

 tissues and tissue-fluids, and the technical difficulties, almost insuper- 

 able it would appear, which attend the utilization of adequate electro- 

 chemical methods of estimating the hydrogen ion changes in nervous 

 tissues. We can, however, perceive the changed reaction of the brain 

 after excessive stimulation by the employment of a simple indicator, 

 provided, however, that instead of employing the change of color of 

 the indicator as a sign or measure of acidity, we employ the change 

 in its solubility in a solvent which is immiscible in water. 



If to ten cubic centimeters of a concentrated (two per cent.) and 

 very faintly acid solution of Neutral Red in water we add a single drop 

 of tenth-normal potassium hydroxide the color of the solution does not 

 perceptibly change, but nevertheless a great change is seen in respect 

 to the lipoid-solubility of the neutral red if we shake up the original 

 and the faintly alkaline solutions with Ethyl Acetate, from which any 

 admixture of acetic acid has been previously carefully removed. On 

 shaking up with the faintly acid solution of neutral red the ethyl 

 acetate remains absolutely colorless, while on shaking it up with the 

 faintly alkaline solution the ethyl acetate layer is stained deep yellow. 

 In two ways the indicator is rendered moire sensitive by this method; 

 in the first place a trace of the yellow modification of neutral red, 

 which would be invisible in watery solution owing to the great excess 

 of the red modification, is removed by the ethyl acetate and thereby 

 rendered visible. In the second place, let us suppose that the Coeffi- 

 cient of Distribution: 



concentration in lipoid layer 

 concentration in aqueous layer 



is 100 : 1 for the yellow modification of neutral red, and zero for the red 

 modification. Then at any given concentration "b" of hydroxyl ions, 

 if "y" be the concentration of the red modification and "x" that of the 

 yellow modification : 



x = kf(b)y 



where "k" is a constant and f (b) is some function of the alkalinity 

 not necessarily known or defined. Now let this solution be shaken up 

 with ethyl acetate, and let the concentration of the yellow modi- 

 fication in the watery layer now be "x," while that of the red modifi- 

 cation is "y," and that of the yellow modification in the ethyl acetate 

 layer is "x?," then we have: 



xi = kf(b)yi 



x 2 = lOOxi 



x? = 100kf(b)yi 



