VITAL STAINING AND MOR PHOGENET I C MOVEMENTS 



225 



Presumptive medullary plate : Locate a region atout 2/5 of the distance from 

 the dorsal lip of the early "blastopore to the center of the animal pole and 

 mark with Neutral Eed. This region should not invaginate and may be followed 

 until it "becomes enclosed in the neural folds. 



5- The Somite Area of the Blastula : "Using the map of "Vogt (derived hy the vital dye 

 method) locate the presumptive somite area ( dorso-lateral to the crecentric blasto- 

 pore) and stain (on either side) with Neutral Bed. When the embryo reaches the 

 tail-bud stage (stage #17 or #l8) dissect it with needles and a hair loop to ex- 

 pose the somites beneath the dorso-lateral ectoderm. 



Presumptive Ejye- forming Areas : This can be accomplished best by using stage #15 

 of Amblystoma and consulting the exercise on Eye Field Operations to determine the 

 presumptive area to be stained. With "Urodele material either Nile blue sulphate 

 or Neutral Eed may be used, since there is less pigment than in the Anuran egg. 



The presumptive eye-forming areas will be incorporated within the brain to 

 give rise to the optic vesicles. The stain must be applied anteriorly over the 

 transverse neural folds. Embryos thus stained should be dissected at stages #20, 

 #26, and #50 to locate the position of the previously stained areas. 



Lateral Line Organs : Select specimens of Amblystoma punctatum at stage #50, when 

 the otic (auditoiy) vesicles begin to form. There may be some muscular activity 

 making it necessary to confine the embryo more rigidly, or even to cover it with 

 a piece of cover glass during the staining. Locate the otic vesicles, Just above 

 the second visceral arch. Apply the dye to the otocyst and to the epidermis Just 

 posterior to it for about 30 minutes. 



The lateral line organs may be followed superficially from their point of 

 origin into the tail, hence this part of the exercise requires a series of draw- 

 ings to indicate the path of the dye as it moves posteriorly with the development 

 of the lateral line system. The dye should pass posteriorly across the somites, 

 and subsequently a smaller line may be seen passing dorsal to the somites, both 

 reaching the tail (see Stone, I955). 



Specific Organ Anlagen : Consult the accompanying figures and photographs for the 

 presumptive areas for the nasal placodes, balancers, lens, gills, etc. (also see 

 Carpenter, 1957) and stain any specific area to check the "fate-mapa" that have 

 derived by this procedure. 



S:JDl\ 



Schematic representation Indicating on the map the posi- 

 tion of the parts of an anterior trunk segment (a) and 

 of a posterior trunk segment (b) . 



(From Pasteels, 19^+2: Jour. Exp, Zool. 89:255) 



