42. HYPOPHYSECTOMY AND 

 EARLY AMPHIBIAN DEVELOPMENT 



PURPOSE: By means of surgical extirpation to determine the relation ol the epithelial 

 hypophysis to early amphibian development. 



MATERIALS: 



Biological: Early tail-bud stages of any amphibian {Anura stage #18, Urodela stage 

 #29). 



Technical : Standard equipment. 



METHOD: 



Precautions: 



1. Use only embryos that are not injured during removal from their jelly cap- 

 sules. 



2. Avoid bacterial contamination following the operation. If this becomes a 

 factor, operate in 0. 1% sodium sulfadiazine in Standard Solution (or Oper- 

 ating Medium for Urodeles). 



3. Endeavor to remove the hypophysis completely, but no other tissue. (The 

 success of this operation can be tested only by subsequent histological exam- 

 ination. ) 



Control: The control for this experiment consists of surgical cutting in the vicinity 

 of the hypophysis, but without removal of any cells. 



Procedure : 



1. Remove all coverings, including the vitelline (fertilization) membrane, from 

 a group of embryos at the appropriate stage (see above). Place them in 2X 

 Standard Solution or in Spring Water (Anura) or in Urodele Operating Me- 

 dium (Urodeles) over a base of agar. The agar will prevent adhesion of the 

 epidermis to the glass bottom of the dish. 



2. Prepare an operating dish with base of soft paraffin or of Permoplast, and, 

 with a ball tip, mould a depression so that the embryo can be held securely 

 with its face looking upward toward the operator. 



3. Locate the hypophyseal groove (from stomodeum to hypophysis) and its dor- 

 sal hypophyseal pit. This is the region of ingrowing of the pigmented, ecto- 

 dermal cells which constitute the hypophysis. Use a double-edged lancet, 

 or micro- (glass) needles to remove the wedge of pigmented hypophysis. 

 This anlage' grows inward between the roof of the pharynx and the floor of 

 the brain (infundibulum) but neither of these other tissue areas should be 

 disturbed, if possible. With small hair loop excavate all pigmented cells 

 from the hypophyseal pit. * 



4. Leave the embryo in operating medium until the wound heals, about 30 min- 

 utes, then return it to the normal culture medium (i. e. , Standard for Anura 

 and Growing Medium for Urodeles). Keep operated embryos separately in 

 #2 Slenders, with agar bases, preferably at temperatures slightly below 

 that of the laboratory. 



OBSERVATIONS AND TABULATION OF DATA: 



1. Make sketches immediately after the operation and at 1 5 minute intervals during the 

 healing process of the hypophyseal area. Healing should be complete within 1 hour. 



* Sec section on "Thyroidectomy and Early Development" for photograph of sagittal section of Anuran embryo showing hypophysis, 

 page 321. 



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