FREE AMINO ACIDS IN ANIMAL TISSUE 347 
scopically observable intracellular structures, since tumor cells which were affected 
severely by cytotoxic agents were still found capable of maintaining high concen- 
trations of the ninhydrin-reactive substances. Particularly striking was the failure 
to produce any change, whatsoever, in the amino acid pattern of Yoshida sarcoma 
cells after treatment with podophyllin at a time when severe damage to individual 
cells was evident, as reflected in abnormalities at the cell surface as well in mito- 
chondrial and nuclear structures. Even sudden and complete cessation of the cir- 
culation to a portion of the left ventricle of the dog achieved by ligation of the 
descending coronary artery resulted in only a relatively slow loss of the high con- 
centrations of some of the free amino acids found in the normal tissue, the decrease 
becoming considerable between 8 and 16 h after ligation when the histologically 
observed damage to the myocardium was of such an extent that many of the cellular 
constituents of both high and low molecular weight undoubtedly were lost from the 
injured tissue. 
The data presented suggest that the steady-state concentrations of the various 
detectable constituents are regulated largely by their own separate metabolic servo- 
mechanisms. Although the mechanisms for the regulation of amounts of the indivi- 
dual substances may possibly interact with one another at one point or another, 
there are some examples of changes produced in the concentration of one constituent 
without marked effects occurring in any of the others, even though known metabolic 
relations exist between the constituents which change and those that remain essen- 
tially constant. In a following communication (BAXTER AND Roserts, this Sym- 
posium) will be discussed the specific elevations of GABA in brain and f-alanine in 
other tissues by administration of hydroxylamine or aminooxyacetic acid to rats. 
Administration of lethal doses of ammonium acetate to rats produced large in- 
creases only in free aspartic acid and alanine levels in liver and in glutamine in 
muscle, testes and brain. Remarkable increases in glutamine content, and no other 
consistent changes, were found in the brains of dogs after the bilateral intracarotid 
infusion of lactate-Ringer’s solution containing ammonium hydroxide. Early in 
regression of tumors changes in glutamine content were found and not in other 
amino acids. A number of experimental procedures discussed in previous sections 
were found to result in changes in only a few of the large number of the detectable 
constituents, e.g. the changes found in muscle of potassium-deficient rats and serum 
and tissues of vitamin A-deficient rats. The above observations point to the relatively 
independent regulation of the amounts of a number of the substances which can be 
shown in other types of experiments to be closely related to each other metabolically. 
Thus, the quantity of glutamic acid, the precursor of GABA in brain, remains 
essentially unchanged whether the amount of GABA is increased by blocking its 
utilization with hydroxylamine or aminooxyacetic acid or decreased by inhibiting 
its formation with thiosemicarbazide (see BAXTER AND ROBERTS, this Symposium). 
Since the glutamine synthetase activity in brain is much greater than glutamic 
dehydrogenase it might have been expected that the great accumulation of glutamine 
occurring in brain during rapid infusion of ammonium salts would have been accom- 
panied by a decrease in the level of glutamic acid, the precursor of glutamine, or at 
least by a decrease in aspartic acid or some other amino acid which can transaminate 
with a-ketoglutarate to form glutamic acid. Such changes were not found. The 
above considerations emphasize that although our knowledge of possible metabolic 
References p. 348/349 
