ORGANIZATION AND CELL-LINEAGE OF ASCIDIAN EGG. 11 



vesicle contains an abundant granular precipitate, an enormous nucleolus, and at 

 wide intervals within the vesicle, but chiefly near the unclear membrane, a few 

 deeply staining chromatic granules. These granules arc small at this time and it is 

 difficult to determine their exact shape, though many of them appear to be V- or 

 5T-shaped; they are the bivalent chromosomes of the first maturation division. 

 Close around the germinal vesicle and extending out nearly to the periphery of the 

 egg is the yolk, which exists in the form of spherules, imbedded close together in 

 the granular cytoplasm. Finally there is the peripheral layer of deeply staining 

 protoplasm in which the test cells were formerly imbedded and which contains no 

 yolk, but numerous refractive spherules much smaller than those of the yolk. 



In the living eggs of Cynthia this peripheral layer is clear and transparent and 

 contains uniformly but sparsely distributed yellow pigment, which seems to be asso- 

 ciated with these small refractive spherules. This pigment is soluble in alcohol 

 and hence cannot be observed in fixed and prepared material ; on the other hand, 

 the alcohol in which large numbers of these eggs have been preserved, has the color 

 of a solution of potassium bichromate. The test cells of Cynthia also contain yel- 

 low pigment granules which are gathered close around the nuclei of these cells. It 

 is noticeable that most of the viscera of Cynthia contain this same yellow or orange 

 pigment, the ovaries being especially highly colored. This pigment is much denser 

 in some individuals than in others, and correspondingly one finds some ova in which 

 there is little or none of the pigment, while in others it is very abundant. In gen- 

 eral the animals which have little of the pigment in their viscera are those which 

 produce eggs with little or no pigment, while those in which the viscera are deeply 

 pigmented produce well-pigmented eggs. The central yolk mass of the living egg 

 of Cynthia is of a slaty gray color, while the germinal vesicle is clear and trans- 

 parent. Therefore, in the living egg of this species of ascidian, three areas can be 

 distinguished with great clearness before the maturation divisions begin, the 

 peripheral layer of protoplasm containing the yellow pigment, the central mass 

 of srrav volk and the clear germinal vesicle. 



In Ciona and Molgula also these three areas are distinguishable in the living 

 egg before maturation, but not so clearly as in Cynthia. In Ciona the peripheral 

 layer is nearly transparent, the yolk is a brownish red, while the germinal vesicle 

 is also transparent. In Molgula both the peripheral layer and the germinal vesicle 

 are transparent, while the yolk is gray, with a faint lilac tinge. A brief inspection 

 of the eggs of Boltenia shows that in this genus the yolk is a bright red. 



This peripheral layer of protoplasm, which is present in all the ascidian ova 

 which I have studied, is, both in living and in stained material, the most striking 

 feature of the egg before maturation and fertilization. It is surprising therefore 

 that in spite of this fact it has received so little attention from those who have 

 studied the ovarian history of the ascidian egg: in fact, with a single exception, I 

 cannot be sure that it has ever been mentioned by any previous writer on this sub- 

 ject. In his paper on the origin of the test cells, Morgan (1890) figures and de- 

 scribes this "peripheral /.one of protoplasm" in an unidentified species of Clavellina 



