EXTRACTION OF EXCITATORY SUBSTANCE 377 



syringe was filled with 10 ml of absolute methyl alcohol so as to mix with 

 5 ml of the cerebrospinal fluid collected during the seizure. After centrifuga- 

 tion, the fluid was reduced by evaporation on a 50°C water bath to 48% of 

 the original volume. This fluid produced a seizure when it was introduced 

 into the ventricles of the receiver dog. The extracted fluid was dried to a 

 yellow-brown mass and kept for 2 weeks at room temperature of 15°C. After 

 this period the fluid was taken up to the original volume with water. Two 

 millihtres were introduced into the receiver dog's ventricle. Violent seizures, 

 especially clonic convulsions, were produced with a latent period of 5-10 sec 

 and they continued for 160-180 sec. Thus we obtained a crude excitatory 

 substance from central nervous system wliich is active at least in the motor 

 system of dogs. 



ANALYSIS OF THE CRUDE EXCITATORY SUBSTANCE 



For the sake of convenience we named the effective component of the 

 extract "excitine". This excitine was found to be rather unstable at tempera- 

 tures above 40 "C, that is its activity was reduced during the process of 30- 

 60 sec of evaporation. 



The methanol-extracted fluid lost 50 % of its eff'ectiveness during 2 weeks of 

 storage at room temperature (15'C during the day and 3-7 C at night). When 

 it was kept in a dry state, however, its eff'ectiveness was not absolutely lost. 

 If the extracted fluid was kept at a pH of 2-3 for 6-10 hr, its effectiveness 

 was reduced by one-half. The active agent was highly unstable in an oxygen 

 atmosphere. If one bubbled pure oxygen through the fluid it became quite 

 ineff"ective within 60 sec. When the fluid was kept in open air the excitatory 

 action was abohshed during 1 or 2 weeks. On the other hand, when the fluid 

 was kept in contact with nitrogen the activity was maintained for over 3 weeks. 



Further analysis is now going on in our laboratory which makes us believe 

 that it seems difficult but not hopeless to obtain purification and identification 

 of the substance. 



REFERENCE 



Hayashi, T. (1959) Neurophysiology and Neurochemistry of Convulsion. Dainihon-Tosho, 

 Tokyo. 



