The Chemical Labeling of Glutamate Decarboxylase in

Mouse brain glutamate decarboxylase(s) was specifically titrated in vivo and in crude brain homogenates by a combination of gabaculine and [a-'H]acetylenic yaminobutyric acid. This specific titration is based on the differential spectra of action of these two mechanism-based enzyme inactivators. The specificity of the titration in vitro was demonstrated by showing that the time course of radioactivity incorporation exactly paralleled the time course for glutamate decarboxylase inactivation. Furthermore, pretreatment of the crude homogenate with aminooxyacetic acid and a-methyltrans-3-dehydroglutamate, two inactivators of glutamate decarboxylase which function by entirely different mechanisms, decreased count i n c o ~ o r a t ~ o n ([n3H]acetylenic y-aminobutyric acid) to the same extent as the activity was decreased. Injection of [a-3H]acetylenic y-aminobutyric acid intraperitoneally after gabaculine injection led to incorporation of 0.46 nmol of inactivatorfmouse brain, when approximately 70% of the enzyme was inactivated. This means that there is approximately 0.66 nmol of glutamate decarboxylase/ 0.5 g of mouse brain, assuming the stoichiometry of inactivator bound to enzyme is one. This value is similar to the one (0.646 nmol) obtained from a calculation based on the enz-yme purification data (Wu, J.-Y. (1974) in y-Arninobutyric Acid in Neroous System Function (Roberts, E., Chase, E. N., and Tower, D. B., eds) pp. 755, Raven Press, New York).