DE. W. H. E AN SOM ON THE OVUM OF OSSEOHS FISHES. 
457 
how impregnation is effected, and afterwards to consider the conditions of the protoplas- 
mic movements. 
1. Earlier sequences of impregnation. 
To trace the changes which follow the action of the spermatczooids npon the egg in 
their earliest stages, it is necessary to fecundate artificially upon the stage of the micro- 
scope. For this purpose I used Powell and Lealakd’s animalcule-cage, a glass ring 
being fastened upon its lower plate, to convert it into a cell, and a portion of the thin 
glass cover being cut away so as to permit the fertilizing agent to be applied at its edge. 
The male and female fishes about to be used may be conveniently manipulated, if the 
spinal cord be first divided just behind the gill-covers, after which they live very well 
in water for forty-eight hours or more. The semen is not easily pressed from the male, 
and hence in these experiments it is convenient to have a number of them ready, and 
to open the abdominal cavity and use a fragment of testis ; this, if quite ripe, will im- 
pregnate the eggs in the cell if pushed just under the cut edge of the glass cover, so as 
to be not too much exposed to the action of water, which soon arrests the movements 
of the spermatozooids. It will not do to put the male fish into water after the abdomen 
has been opened, but if it be kept moist the spermatozooids live some time. 
a. Formation of the breathing-chamber. — The earliest change which occurs after an egg 
is fecundated, is the formation of a space between the yelk-sac and the outer surface of 
the yelk. This space is the breathing-chamber of Newport ; it commences first close 
to the micropyle and gradually extends over the rest of the yelk-ball*, being complete 
in from three to five minutes after the spermatozooids have been applied to the edge of 
the glass cover, in successful experiments. It begins by a withdrawal of the funnel of 
the micropyle from the pit in the discus proligerus, so that as water enters the funnel is 
gradually shortened, and at length may be almost effaced. This withdrawal was seen to 
begin in about fifteen seconds after the first spermatozooid was seen to enter ; but ordi- 
narily it is visible about one minute after the testis has been applied to the eggs. After 
the breathing-chamber has been once formed, it for some minutes longer increases in 
size, by expansion of the yelk-sac, so as to efface the indents on the surface of the 
egg, increase its size, and render it globular, tense, and elastic, remarkably resisting, 
and difficult to injure, the very reverse of its state before it had imbibed water. 
The formation of the breathing-chamber is, I think, not entirely due to the entrance 
of water, but in part to a contraction of the substance of the yelk, which commonly 
produces a flattening of one surface. In the eggs of Gasterosteus it seems certain that 
the water mainly enters to fill the breathing-chamber through the micropyle, and under 
ordinary circumstances, not by imbibition through the yelk-sac. It is somewhat difficult 
to conceive how the passage of such a minute body as a spermatozooid through the tube 
of the micropyle, closed as it seems to be only by the viscid secretion of the oviduct, 
* This is only ascertained to he the fact in Gasterosteus. In most other osseous fishes water enters freely 
through the yelk-sac, and the breathing-chamber may probably commence simultaneously at all parts of the 
surface. In frogs I believe that I have witnessed its commencement, as in Gasterosteus, first near the micropyle, 
3 q 2 
