326 DEVELOPMENT OF ASCIDIANS 



divide. The same concentration used on the eggs at 2-, 4-, and 

 8-cell stages produces a similar result; that is, the eggs are 

 blocked. Solutions at O.OIM inhibit the enzyme partially; at this 

 concentration development is also impossible. Only at O.OOIM 

 or 2 X 10~^ and 3 X 10~'^ M is some development possible. 

 Stronger solutions (0.2 or O.IM) can be used only with the un- 

 fertilized egg without any consequent damage to development. 

 After treatment, even prolonged for 12 hours, it can be fertilized 

 in normal sea water and start development. Under such condi- 

 tions the CO. is inhibited, as shown by the Nadi, and the inhibi- 

 tion lasts longer ( Reverberi, 1956b ) . 



Sodium malonate and selenite are considered good inhibitors 

 of succinodehydrogenase. The inhibition can be checked histo- 

 chemically by tetrazolium salts. In our experiments with ascidian 

 eggs solutions 5 X 10- to 1 X 10"^ M and 6 X 10"^ to 1 X 10"^ 

 M, respectively for malonate and for selenite, are effective. At 

 these concentrations, however, the inhibition of the enzyme is 

 only partial, as revealed by the positive histochemical reaction. 

 Malonate and selenite do not inhibit the activity of the CO. 



Development of the Ascidian Egg with Blocked Enzymes 



The central problem of our research was to establish whether 

 the ascidian egg with one or more blocked enzymes develops 

 into a normal larva or into a larva with specific and constant ab- 

 normalities. Eggs at different stages of development were sub- 

 mitted to the action of sodium azide, malonate, or selenite, or 

 sodium azide plus malonate ( or selenite ) . 



Treatment ivith Sodium Azide. UnfeHilized eggs can be 

 treated at length and at high concentrations (0.1 to O.OIM). In 

 such solutions the CO. is completely inhibited, fertilization and 

 development are possible in normal sea water or in diluted azide 

 solutions, and the inhibition of the enzyme persists. The segmen- 

 tations of the eggs are normal and synchronous with the controls. 

 Tadpoles develop which have a normal "head" but an abnormal 

 tail which is distorted and physiologically impotent. The tadpoles 

 move poorly and verv irregularly, and they are easily exhausted 

 (Fig. 2^7). 



