Adults are abundant on the floats and wharf piles around Woods Hole, Mass. They are far more common than M. citrina, but in the past workers have often confused the two species. Since M. citrina is viviparous and releases larvae instead of eggs, there is little reason for this mistake in identification. The animals are hermaphroditic.

Probably there is no fixed season. Individuals over 12 mm. in length seem to breed continuously (Berrill, 1931).

A. Care of Adults: These animals keep very well when placed in large fingerbowls on the water table. A continuous gentle stream of sea water should be supplied and dead material removed promptly, since the water will foul very rapidly.

B. Procuring Gametes: For experimental work it is usually advisable to obtain naturally-shed eggs. Molgula sheds soon after dawn; however, this process may be delayed by placing several animals in large fingerbowls, and keeping them in the dark until they are needed. They will usually shed about 15 minutes after they are brought into the light.

Eggs and sperm may also be obtained by cutting open the tests of individuals over 12 mm. in length, and pipetting gametes from the genital ducts. This procedure may yield immature as well as mature eggs, since the oviducts in Molgula are short. The eggs should be passed through several changes of sea water, to free them of perivisceral fluid.

C. Preparation of Cultures: Naturally-shed eggs should be pipetted to fingerbowls of fresh sea water. To artificially-obtained eggs, enough sperm should be added to cause a faint milkiness in the water. The water should be replaced after one or two hours. It is probably advisable to use gametes from different individuals.

The bowls of fertilized eggs should be kept on a water table, and the water changed three or four times during subsequent development. As soon as the larvae begin to swim, they should be decanted or pipetted to fingerbowls of fresh sea water or, if a study of metamorphosis is desired, isolated in separate drops of sea water in Syracuse dishes. When they have firmly attached to these dishes, sea water should be added. The dishes with attached larvae can be stored in an inverted position in wooden racks which are submerged in aquaria of running sea water.

D. Removal of the Chorion: The chorion can be digested off with the stomach juice of crabs or with proteolytic enzymes, before or after fertilization (Berrill, 1932, 1937). This technique is useful for experimental purposes.

A. The Unfertilized Egg: The egg is opaque, with colored yolk; measurements of its diameter vary from 100 microns (Conklin, 1905) to 115 microns (Grave, 1926). Outer follicle cells, which are rounded, form a compact layer around the chorion, and a few inner follicle cells are present. A perivitelline space is visible. In the oviduct the germinal vesicle breaks down, and the egg proceeds to the metaphase of the first maturation division. Eggs are shed at this stage and remain in it until fertilization or death. (See the paper by Berrill, 1931, for further details.)

B. Fertilization and Cleavage: The eggs are fertilized as they are shed into the water. Cleavage is equal up to the fourth division, and separates the egg into future right and left halves, as in Styela. Gastrulation occurs between the sixth and seventh cleavages, and is a rather specialized form of true invaginative gastrulation.

C. Rate of Development: Berrill (1931) states that at 19û C. the blastopore is closed and the tail bud visible in four hours; hatching occurs in 8 to 11 hours. Secretion of the test, caudal degeneration, and outgrowth of the ampullae (i.e., metamorphosis) occur 18 to 24 hours after insemination.

D. Later Stages of Development and Metamorphosis: The larvae normally hatch by means of enzymatic digestion of the chorion. The free-swimming urodele-like tadpoles have vertical tail-fins and a large sensory vesicle containing an otolith which is not destroyed during metamorphosis. There are no gill slits or adhesive papillae visible, and the siphons are undeveloped. The alimentary tract is poorly developed and very yolky. For diagrams of larvae, see the papers by Berrill (1931) and Grave (1926).

A few of the early stages of metamorphosis are figured by Berrill and by Grave. As has already been indicated, fixation is followed by tail degeneration and the outgrowth of a long, primary ectodermal ampulla. When this is fully formed, additional ampullae appear, and in the final state there are two present on one side of the body and three on the other. Pulsations appear early in the primary ampulla, which probably has a respiratory function.

REFERENCES

BERRILL, N. J., 1931. Studies in tunicate development. II. Abbreviation of development in the Molgulidae. Phil. Trans. Roy. Soc., London, ser. B, 219: 281-346.

BERRILL, N. J., 1932. The mosaic development of the ascidian egg. Biol. Bull., 63: 381-386.

BERRILL, N. J., 1937. Culture methods for ascidians. In: Culture Methods for Invertebrate

Animals, edit. by Galtsoff et al., Comstock, Ithaca, pp. 564-571

CONKLIN, E. G., 1905. The organization and cell-lineage of the ascidian egg. J. Acad. Nat. Sci, Philadelphia, ser. 2, 13: 1-119.

GRAVE, C., 1926. Molgula citrina (Alder and Hancock). Activities and structure of the free swimming larva. J. Morph., 42: 453-471.