Mather, Wharton B. (Wharton Burdett)

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Surgery of the Rabbit Embryo "in utero"
BY
WHARTON B. MATHER, M.Sc.
(The Department of Zoology, University of Queensland, Brisbanp)
VOLUME II
DEPARTMENT OF BIOLOGY
1950
THE UNIVERSITY OF QUEENSLAND PRESS
BRISBANE
DATE OF PUBLICATION:
13th MARCH, 1950
NUMBER 16
SURGERY OF THE RABBIT EMBRYO
"IN UTERO"
By WHAH.TON B. 1VL\THER, M.Sc.*
(The Department of Zoology, University of Queensland, Brisbane).
1. INTRODUCTION.
In 1925, three alltbors~NicllOlas, Bors and S\t,,~nson---published papers
suggesting the feasibility of the direct surgical approach to the late mJ.mmalian
embryo. Nicholas, working with rats, found that it was possihle to excise limbs,
eyes or tails from 12-20 clay embryos "in utero" and shU obtain them viable at
normal term. Bors, working with rabbits, founel that it was possible to excise
limbs from L6-25 day embryos and to obtain them viable at 28 days. However,
he did not let his operated animals go to term because of the danger that they
would be eaten by the mother. Swenson was not interested in the surgery of the
embryo but in foetal movement. Previous work on the foetal movements of
milmmalian embryos had been done without amesthesia which Swenson considered
unsatisfactory. However, it was undesirable to Use an anacsthetic which might
affect embryonic moycment and be describes a technique of ligating the carotid
arteries and jugular veins, thus causing amesthesia by cerebral ana:mia. Using
this method, he opened the uterus and withdrew the embryos into warm saline
during the last third of pregnancy and studied the foetal movements.
In spite of the interesting field that the direct approach to the embryo " in
utero" offered, it was not until Hmo that Hooker and Nicholas made use of the
technique to investigate regeneration after spinal cord section in the last third of
pregnancy of the rat. They reported negative results. Three years later, Larsell
(19:J;3) reported substantial recluction in the size of the scapula after limb excision
ill the 16-19 day embryo.
Quite reccntly, Hall and Schneiderhan (HHi5) have reported spinal ganglia
hypoplasia in the rat after limb excision during the last third of pregnancy, and
Jost (Hl46) , interested in thC' dfect of the embryonic gonads on secondary sex
characteristics, has reported successful embryonic castration of rabbits during the
last third of pregnancy.
Fillillly, Wells (1047), also interested in sex' problems, has found that the late
rat embryo may be brought into the body cavity ancl shU survive provided that
the umbilical corel is left attached to the placenta in the uterus.
In spite of the above, little has been described Oil tile c:daj)s of the technique
of the direct approach to the mammalian embryo" in utero," the stage of pregnancy
when interference can be successfully carried out and the reliability of the method.
With these points in mind, it seemed worth while to investigate further the
possibilities of this method. The rabbit was chosen as the experimental animal
and it was decided to remove a limb or the tail from the foetus at various stages
of development and to observe the result.
IT. TECHNIQUE.
Strict aseptic precal~tions ha ve been observed in all operations as it is considered
that the freedom from infection achieved fully offsets the extra labour involved.
The following procedure has been adopted in making the operations :-
* This work was co,rrif;r1 out while on the sto,ff at the Zoology Depanmegt, The Uu,iversity
of Melbourne.
Dead
Dead
Living
Living
Dead
Living
,:~7 daSi"s.
:31 da3Ts.
:>0 days.
:30 da)Ts.
:31 days.
:11 dilYS.
7
:;
o
+X
1
a -+ X
-I- x
I)
I)
deliverf_,:cl
10
IS ,1"\,,
I d:,·\.;
:?l,I;):,,;
21 da-i/s
:?2 (b)';,;
;\~::(~ at
Clr'i~'rati()n
V
../\
X
8X
2X
Embryos
present
.) + X
++ 2X
-I-
X
indica1cs err.llYO oreratcd on or removed.
SURGERY OF THE RABBIT EMBRYO" IN UTERO"
SURGFhY nF HAIC,BIT EMBHYCS "IN TJTEIW."
lirnb excision
Lirnb e'xcision
CaEsaJe"n section
Limb excisioll
Tail excision
Limb excision
Tatl excision
Caesarean
Caesarean
Type of (?I:eration
I
2
.,
,)
.J.
[)
(I
7
8
\)
number
III. RESULTS.
In all, nine opErations have been can led oul: on rabbit embryos" in utero,"
the data on these operations being set out in Table 1. in the order in which they'
were carded out.
B. Operation (in operat,,:ng
1. Scrub up ,wd put on gown, mask :mel with the aid oJ an assistant.
2. Lay OlIt instruments on a ;:,terile towel. (While this is being done the
assistalltstarts the open etlwr amf:sthc,-ia in the (r,encral laborawrv ,md wheels the
animal into the operating laboratory.) c. J
3. Swab skin with aniiseptic Zepharin).
4. Clip on "fenestrated sheet" with towel clips so that the operative are,"
is exposed.
5. Incise skin (2in. incision) in mid lil~c, I in. in front of pubic synlphysis.
6. Incise muscle layers.
7. Incise peritoneum.
8. Lift Ollt lIteri, count etnbryos present, and kwc 11 umber recorded by assistant,
9. Pack uteri with warm saline pacb.
10. Make iin. incision ill uterus and amnion. on the anti-mesometrial side.
immediately over limb ot tail oJ the embryo.
11. Excise limb or tail and discard.
12. Suture uterus with two fmc interruptell cat gut ,;uture:::.
13. Remove packs and place uterus back in
14. Stlture peritoneum and muscle layers with five Cine interrupted cat sutUrE·S.
15. Dust on slllphacliazine powder.
16. Sutnre skin \vith five tlne interrupted sutures,
A. Preparation (in general laboratory).
1. I.nject 0.04 grains of atropine sulphate intravenously (an ear vein is con-
venient) to prevent excessive muCous Jonnation in the respiL:tory tract.
2. Strap down onopel-ating table.
3. Shave abdomen.
4
SURGERY OF THE RABBIT EMBRYO" IN UTERO"
IV. DISCUSSION.
In operations 3, 8 and 9 it was intended to carry out either a tailor limb
excision, but the embryo protruded through the opening in the uterus and had
to be removed. Reynolds (1947) points out that the tension on the wall of the
uterus is greatest while the conceptus is spheroidal and reaches a maximum at
22 days, just before the conceptus becomes cylindrical. Thus, it is understandable
that the embryos tend to protrude if operated on before 22 days. He also points
out that there is a preponderance of foetal death when there is maximum tension
on the uterus wall, a fact also militating against a high percentage of successful
" in utero" operations hefore this time. Two operations were attempted at 14 days
without success; in the second, an attempt being made to relieve the intra-uterine
pressure with a syringe before opening the uterus. However, this was unsuccessful.
1t remains to be seen if successful operations can be carried out at this stage
with the aid of an assistant operator and whether it is possible to operate between
14 and 18 days. Bors (1925) reports a partially successful operation at 16 days in
which a badly injured embryo reached the size of a typical 22 day embryo at 28 days.
In operation 1, although a limb excision was carried out, the mother aborted
at 27 days, and the operated embryo was apparently absorbed.
In operation 2, although the unoperated embryos were born alive, the operated
embryo was apparently absorbed and similarly in operation 5 all but one unoperated
embryo were absorbed.
In operation 4, the mother was found dead in her pen with the gut herniated
through the body wall incision. The unoperatecl embryo had been born and the
operated embryos were still in the uterus but was of normal 30 day size, indicating
that it had lived up to the time of the death of the mother.
A similar gut herniation occurred during the observed parturition of rabbit 6,
but three unoperated and the operated embryo were born alive. The most successful
experiment was number 7, where all the unoperated and the operated embryos
were obtained alive at parturition. Jost (1946) and his school have used this
technique in their studies on embryonic sex characteristics in the last third of
pregnancy and he gives a brief note on his method not, however, supported by
experimental detail. On the other hand, Bors (1925) reports the results of three
successful operations at 21 days, but his work differs from the present investigation
in that he terminated his experiments at 28 days without letting them run to term.
V. CONCLUSIONS.
It is concluded that it is possible to operate on the rabbit embryo" in utero"
after the critical twenty-second day with a reasonable expectation of success, but
that before this time the operations become progressively more difficult and less
reliable with the technique at present used.
VI. SUMMARY.
The technique of "iJ;! utero" operations on the rabbit embryo is described
with a discussion of the results obtai ned at various stages of gestation.
ACKNOWLEDGMENT.
The author wishes to express his appreciation of the assistance given by
Mr. James Guthrie, Jum., who acted as ana:sthetist.
SURGERY OF THE RABBIT EMBRYO" IN UTERO"
VII. REFEREKCES.
BOHS, E. (1925) : Die Methodik del' intrauterine Operation am uberlebenden SaugetierIoetus.
Arch. Entw. Mech. 115 : fl55.
HALL, E. K. & SCfINEIDERHAN, M. A. (1945) : Spinal ganglia hypoplasia after limb amputation
in the foetal rat, J. Compo N eurol. 82 !J.
HOOKER, D. & NICHOLAS, J. S. (1!J30) : Spinal cord section in rat foetuses. J. Compo Neurol.
50 : 413.
JOST, A. (1!J4fl) : Sur la differenciation sexuelle de l'embryon de lapin. Eemarques au sujet de
certaines operations chirurgicals sur l'em bryon. Camp. Rend. Soc. BioI. 140 : 461.
LAHSELL, O. (1!J:l3) : The d.eyelopment of the scapula following intrauterine fore limb amputation
in the rat. Anat. l'{ec ..'56 : 57.
NICHOLAS, 1. S. (l.925): Notes on the application of experimental method upon mammalian
embryos. Anat. IZec. 31 : :355.
REYNOLDS, S. IL M. (1047) The relatiou of hydrostatic conditions in the uterus to the size
and shape of the conceptus during pregnancy. A concept of uterine accommodation.
Anat. ICZec. 95 : 283.
SWENSON, E. A. (l.!J25): The use of cerebral anaemia in experimental embryological studies
upon mammals. Anat. Eec.:lO 147.
WELLS, L. J. (l94fl) : A method for subjecting foctal. rats to lapamtomy and repeated sub-
cutaneous injection. Anat. Eec. !J4 : 530.


Surgery of the rabbit embryo "in utero"

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Surgery of the rabbit embryo "in utero"

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