15. E M B R Y O G E N E S I S in vitYO
575
I I I . IN VITRO
C U L T U R E OF M A M M A L I A N O V A
Many attempts have been made to cultivate in vitro the ova or
embryos of Mammalia, at different stages of their development. They
fall into four groups: (1) fertilization in vitro, followed by development
in vitro; (2) development in vitro, starting at early stages (culture of tubal
egg); (3) development in vitro beginning at the blastocyst stage (culture
of uterine egg); (4) experiments on the egg in vitro, then reimplantation
in the uterus.
A. F E R T I L I Z A T I O N A N D D E V E L O P M E N T IN
VITRO
Experiments with in vitro insemination of mammalian ova, removed
from the ovary, the tubes or the uterus, have been attempted by many
authors and on several different species, e.g. rabbit, guinea-pig, rat
and man.
The recent publication of Thibault and Dauzier (1961), based on
their own work and on the work of Chang (1959) and of Austin and
Bishop (1957), tends to show that most of these attempts were negative;
in the majority of cases, activation of the ovum and experimental
parthenogenesis only was achieved, but not fertilization.
In fact, impregnation in vitro requires that several conditions be
strictly fulfilled: (1) the maintenance of temperature at 37°C during
all the manipulations in order to avoid parthenogenetic development;
(2) the necessity for the activation or "capacitation" of spermatozoa in
the female genital ducts. This activation renders them able to traverse
through the zona pellucida, but, inversely, it also makes them sensitive
to a sperm-repellent substance emitted by the oocytes; (3) this substance must be eliminated in vitro by prolonged washing of the ova.
In vivo, it is probably neutralized by another substance secreted in the
genital passages. These two substances can be considered as similar to
fertilizin and to antifertilizin of invertebrates.
These conditions have been studied in rabbits in particular: they
are not valid for all species.
Whether the eggs be fertilized or parthenogenetic, abortive developments—generally not going beyond the first stages of segmentation—
have been obtained with ova of rabbits (Pincus, 1939) and with the
human ova (Rock and Menkin, 1944; Menkin and Rock, 1948). One
of the most characteristic results was obtained by Shettles (1955) on a
human ovum. Removed in a mature ovarian follicle, the egg was
explanted with the follicular fluid in the presence of mucus from the
Fallopian tubes and of fresh human spermatozoa. The egg divided
and, after 72 h, attained a stage of about thirty-two cells.
575
I I I . IN VITRO
C U L T U R E OF M A M M A L I A N O V A
Many attempts have been made to cultivate in vitro the ova or
embryos of Mammalia, at different stages of their development. They
fall into four groups: (1) fertilization in vitro, followed by development
in vitro; (2) development in vitro, starting at early stages (culture of tubal
egg); (3) development in vitro beginning at the blastocyst stage (culture
of uterine egg); (4) experiments on the egg in vitro, then reimplantation
in the uterus.
A. F E R T I L I Z A T I O N A N D D E V E L O P M E N T IN
VITRO
Experiments with in vitro insemination of mammalian ova, removed
from the ovary, the tubes or the uterus, have been attempted by many
authors and on several different species, e.g. rabbit, guinea-pig, rat
and man.
The recent publication of Thibault and Dauzier (1961), based on
their own work and on the work of Chang (1959) and of Austin and
Bishop (1957), tends to show that most of these attempts were negative;
in the majority of cases, activation of the ovum and experimental
parthenogenesis only was achieved, but not fertilization.
In fact, impregnation in vitro requires that several conditions be
strictly fulfilled: (1) the maintenance of temperature at 37°C during
all the manipulations in order to avoid parthenogenetic development;
(2) the necessity for the activation or "capacitation" of spermatozoa in
the female genital ducts. This activation renders them able to traverse
through the zona pellucida, but, inversely, it also makes them sensitive
to a sperm-repellent substance emitted by the oocytes; (3) this substance must be eliminated in vitro by prolonged washing of the ova.
In vivo, it is probably neutralized by another substance secreted in the
genital passages. These two substances can be considered as similar to
fertilizin and to antifertilizin of invertebrates.
These conditions have been studied in rabbits in particular: they
are not valid for all species.
Whether the eggs be fertilized or parthenogenetic, abortive developments—generally not going beyond the first stages of segmentation—
have been obtained with ova of rabbits (Pincus, 1939) and with the
human ova (Rock and Menkin, 1944; Menkin and Rock, 1948). One
of the most characteristic results was obtained by Shettles (1955) on a
human ovum. Removed in a mature ovarian follicle, the egg was
explanted with the follicular fluid in the presence of mucus from the
Fallopian tubes and of fresh human spermatozoa. The egg divided
and, after 72 h, attained a stage of about thirty-two cells.