MECHANISM OF PRODUCING EXCITATORY TRANSMITTER 387 



AN EXPERIMENTAL METHOD CONCERNING THE 

 GENESIS OF AN EXCITATORY TRANSMITTER 



The method we used was to introduce a substance into the cerebrospinal 

 fluid of a dog and, at the same time, to introduce a coenzyme in order to 

 transform the substance into the excitatory transmitter. As in the experiment 

 described in Table 2, we took folic acid instead of vitamin B12 in a concentra- 

 tion which by itself caused no motor effects. When combined with GABA 

 and ATP it produced strong seizures. Vitamin Bi could be used instead of 

 ATP in this case and the result indicated that it acts as energy liberator in the 

 reaction. 



The method hitherto used to detect enzymatic actions in tissues consisted 

 in the artificial addition of substrate to homogenates or slices of brain in vitro 

 and one obtained the expected substances from the incubation fluid or used 

 the output of CO2 as an indicator. This was essentially the procedure in 

 biochemistry. But here we found a method of neurochemistry whereby the 

 substrate is introduced in vivo into the cerebrospinal fluid of a dog so that it 

 can contact the cells of the brain that contain the enzymatic systems whose 

 substrate was increased by the artificial addition. It was assumed that if this 

 excess of substrate drives the reaction in the direction of synthesis of a new 

 substance at a rate which is greater than that of the normal state, this resulting 

 substance should be detectable by any indicator, in our case, the production of 

 a seizure. From the experiments just mentioned we could formulate the reac- 

 tion that produces the excitatory chemical transmitter in central nervous 

 system (at least in the motor system). 



GABA plus vitamin B12 plus Bi — cc* (1) 



GABA plus folic acid plus Bi ^* cc* 



Instead of vitamin B12 we tested vitamin B2, biotin, lipoic acid and vitamin 

 C, but these substances did not accelerate the reaction. Pantothenic acid was, 

 however, found to produce clonic convulsions according to the following 

 reaction : 



GABA plus pantothenic acid plus Bi -> cc (2) 



It was now a question of whether the reaction of formulae (1) and (2) repre- 

 sented the same sequence (that is, whether pantothenic acid had the same 

 action as B12 or folic acid), or if there were quite different reactions leading to 

 two different substances. 



When GABA, vitamin B12, pantothenic acid and vitamin Bi were combined, 

 the seizure was delayed or the concentration of each substance needed to be 

 increased in order to produce a seizure. In other words, the reactions of 

 vitamin B12 on the one hand and the reaction of pantothenic acid on the other 



* cc = clonic convulsions. 



