728 DR. E. B. WILSON ON THE DEVELOPMENT OF RENILLA. 
only those features have been mentioned which appear of interest in connexion with 
the following study of Renilla, we may pass to a description of the observations.* 
SEGMENTATION OF THE EGG AND FORMATION OF THE GERMINAL LAYERS. 
$1. External features of segmentation. 
Renilla, like most other Aleyonaria, is dicecious, and on account of the rather 
marked difference in colour between the ova and spermatic capsules, the sexes may 
usually be distinguished by external examination. During the months of May, June, 
and July many Renillas were found with the cavities of the polyps packed with the 
lead-coloured ovaries or the whitish spermatic capsules. The egg or mass of sperm- 
cells is enclosed in a very distinct follicle of ciliated entoderm cells which is ruptured 
at the time of spawning, the eggs being thus discharged into the gastric cavity and 
thence passed out to the exterior. 
The eggs make their exit through the mouths of the sexual polyps, and the time 
occupied in spawning is very short. They are vomited forth in great masses, 
together with a considerable quantity of mucus, by a reversed peristaltic movement of 
the cesophagus, the entire colony being usually in a state of complete expansion. 
The mass of eggs is often held for some time clasped in the tentacles before being 
thrown off into the water. All of the polyps in the central part of the disc spawn 
simultaneously ; those near the edge of the disc often do not spawn with the others, 
* It is perhaps worth while to describe briefly the methods employed in the preparation of the 
embryos. For sections of the early stages the most satisfactory method is that recommended by 
Bosretsky and so successfully employed by Mayer, Harscuex, and others. The eggs were heated in 
sea-water to about 60° C., and maintained at that temperature for two or three minutes in order to 
coagulate thoroughly the protoplasm. They were then hardened for twenty-four hours in potassium 
bichromate, washed two hours in sea-water, and then gradually hardened in alcohol (50 per cent. three 
hours, 75 per cent. three hours, 90 per cent. six hours, and then transferred to absolute alcohol). After 
standing twenty-four hours in picro-carmine, and again soaking a few hours in absolute alcohol, they 
were embedded in paraffin and vaseline and cut with the sledge microtome, 
For sections of later stages the embryos were exposed for a few minutes to very dilute osmic acid 
zo per cent., or less) until a barely perceptible brown tint was produced. After thorough washing they 
were transferred to weak, strong and absolute alcohol, and stained and embedded as before. 
For isolation of the muscle-fibres and other elements of the tissues, the method recommended by the 
Hertwic brothers was employed. The larve were placed for ten or fifteen minutes in a mixture of 
equal parts of ;15 per cent. osmic acid and 2 per cent. acetic acid in sea-water, then thoroughly washed 
and soaked for several days in 1 per cent. acetic acid in sea-water. They were then stained in toto and 
teased in glycerine. 
With other methods of hardening I have had no success. Boprersky’s method is highly to be 
recommended for the early stages, and affords very clear and satisfactory preparations. 
