CHEMICAL ALTERATION OF GROWTH 

 AND DIFFERENTIATION 



FUBPCSE : The study of the effect of a specific chemical, Lithium Chloride, on early 

 amphibian development. 



MATEEIALS : 



Biological : Early gaatrula stages of Anura (stage #9) and Urodela (stage #10). 



Technical : Various solutions as follows:* 



1. Balanced lithium solution (Hall, 1914-2): 



Lithium chloride 1.12 gr. 



Lithium sulphate 1.5 gr. 



Calcium chloride 0.I5 gr. 



Potassium bl phosphate .... O.Qlt- gr. 



Sodium bicarbonate 0.5 gr. 



Glass dist. water 1000.0 cc. 



2. Unbalanced lithium solution (Hall, 1942): 



Lithium chloride I.09 gr. 



Lithium sulphate 1.5 gr. 



Sodium bicarbonate 0.11 gr. 



Glass dist. wBter 1000.0 cc. 



3. Unbalanced sodium solution (Hall, 19'+2): 



Sodium chloride 1.5 gr. 



Sodium sulphate 1-97 gr. 



Sodium bicarbonate 0.25 gr. 



Glass dist. water 1000.0 cc. 



h. Pure lithium chloride solutions: 



(a) 1.9 grams LlCl in 1,000 cc. of glass distilled water. 



(b) 1.9 grams of LiCl in 250 cc. of glass distilled water. 



5. Control solutions: 



For Anura: Standard ( Holtfreter' s) solution 

 For Urodela: Urodele Growing Solution 



METHOD: 



Precautions : 



1. Glass distilled water should be usedj if possible, to make up tne experimental 

 solutions to avoid even the minutest traces of metallic or other ions. 



2. In returning the exposed (experimental) gastrulae to the nonnal medium, it is 

 wise to pass them through several changes to remove all adherent Lithium. 



Control : This consists of similar gastrula, similarly denuded of their membranes, but 

 kept in control media. Conditions of temperature, light, etc. must be identiceil. 



Procedure : 



1. Prepare 12 finger bowls, each with 50 cc. of solution as follows: Two finger 

 bowls (a and b) each, of solutions 1, 2, 5, U, and tne appropriate control 5. 

 Cover them and iceep them at a cool temperature. (About 18°C. for Anura or lit-°C. 

 for Urodela. ) 



2. Strip eggs (Anura or Urodela) of their Jelly membranes. Do not attempt to re- 

 move the vitelline membranes, particularly of the Anura. The stages should be the 

 same and in early gastrula (Anura #9 and Urodela #10). A total of at least 500 

 undamaged eggs should be quickly prepared. (Since it takes 5 hours to reach 

 stage #10 in the Anura, at least this much time should be allowed for the denud- 

 ing.) 



Note: Effects similar to those produced by Lithium Chloride may be produced by un- 

 balanced solutions of NaCl, NaOH, ethyl alcohol, NHj^OH, MgCl^, Mg(N0j)2, ether, 

 chloroform, and by x-raye. 



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