THYROIDECTOMY AND 

 EARLY AMPHIBIAN DEVELOPMENT 



PUBPOSE : By meana of surgical extirpation to determine the functional relationship of the 

 thyroid anlage in the early larra to met amorphic changes in the later tadpole. 



MATERIALS : 



Biological : Anura (stage #1?) or Urodele (stage #51) larvae. 



Technical : Standard Equipment. 



METHOD: 



Precautions : 



a. Avoid Injury or removal of heart anlage which Is Just posterior to the thyroid 

 anlage . 



b. Avoid post-operative bacterial infection. If necessary, use 0.1^ sodium sulfa- 

 diazine in operating and in culture media. 



Control : The control consists of embryos of similar age and stage in which similar 

 surgical incisions are made but without the removal of any tissue. Such animals 

 should be given Identical treatment as the experimentals. 



Procedure : 



1. Bemove the embryos from their Jelly capsules and fertilization membranes. If 

 there is any muscular activity, anesthetize them ini/5,000 MS 222 (freshly made 

 up). Ciliary movement cannot be reduced by narcosis. 



2. Make a shallow depression in the Permoplast (or paraffin) of the operating dish. 

 Use Urodele Operating Medium for Urodeles and 2X Standard Solution for Anura, 

 adding I'jt sodium sulfadiazine if there is difficulty with infection. Place the 

 embryo on its left side, head away from the operator. 



5, Insert the point of a double-edged leincet (or operating glass needle) between the 

 position of the thyroid and the heart anlage, (see diagrams) and make an outward 

 cut through the throat ectoderm. Bemove a wedge of tissue, the apex of which 

 reaches the floor of the pharynx Just at the point of the slightly pigmented 

 thyroid evagination. Carefully excavate the cells with the hair loop, avoiding 

 particularly the heart mesoderm. Part of the pharyngael floor will, of necessity, 

 be removed with the thyroid. (If the student finds this operation difficult, re- 

 fresh his memory of the position of the thyroid anlage by studying both trans- 

 verse and sagittal sections of tall-bud stages. A complete dissection study of 

 the living tall-bud stage prior to thyroid extirpation is definitely recommended, 

 for this is a delicate operation.) 



h. Transfer the operated embryo to an agar- base in a #2 Stender filled with operat- 

 ing medium for about 50 minutes during which the wound will heal. Then transfer 

 the embryo to Urodele Growing Medium or Standard Solution (depending upon the 

 genus) for further development, preferably at a temperature slightly below that 

 of the laboratory. Begin feeding at appropriate stage of development. 



OBSERVATION AMD TABULATION OF DATA : 



Operated and control embryos must be given identical treatment with respect to volume, 

 medium, light, food, etc. 



1. Make sketches at 15 minute intervals of the wound healing of the operated animals. 



2. At weekly intervals following the operation, make drawings (or photographs) of 

 thyroidectomlzed and control embryos. 



5. Select some embryos that show definite effect of thyroidectony at a time when 

 metamorphlc changes are beginning to appear in the controls, and treat them with 

 1/1,000,000 thyroxin to determine whether It is possible to compensate for the 

 loss of the thyroid gland in bringing the larvee through the critical stage of 

 metamorphosis. (Consult exercise on "Thyroid and Amphibian Metamorphosis".) 



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