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Dominant lethals in young female mice
SHORT COMMUNICATIONS
231
I BATCHELOR, A. L., AND P. W. EDMONDSON, Fast-neutron irradiation of large animals, in : Biological E~ects of Neutron and Proton Irradiations, Vol. I I , International Atomic Energy Agency, Vienna, 1964, pp. 3-13. 2 BATCHELOR, A. L., t~. J. s. PHILLIPS AND A. G. SEARLE, A c o m p a r i s o n of the mutagenic effectiveness of chronic neutron- and F-irradiation of mouse spermatogonia, Mutation Res., 3 (I966) 218-229. 3 CARTER, T. C., M. F. LYON AND R. J. S. PHILLIPS, Genetic h a z a r d of ionizing radiations, Nature, 182 (1958 ) 409 . 4 RUSSELL, W. L., The effect of radiation dose rate and fractionation on mutation in mice, in: F. H. SOBELS (Ed.), Repair from Genetic Radiation Damage, Pergamon, Oxford, 1963, pp. 205217 . 5 RUSSELL, W. g., Studies in mammalian radiation genetics, Nt*cleonics, 23 (1965) 53-62. 6 RUSSELL, W. L., The nature of the dose-rate effect of radiation on mutation in mice, Japan J. Genet., 4 ° , Suppl. (1965) 128-14o. 7 SEARLE, A. G., Progress in mammalian radiation genetics Proc. 3rd Internat. Congr, Radiation Research, Cortina, 1966, in the press. 8 SEARLE, A. G., AND R. j. s. PHILLIPS, Genetic effects of high L E T radiation in mice, Radiation Res., in the press.
Received October 7th, 1966 Mutation Res., 4 (1967) 229-231
Mutation Research Elsevier Publishing Company, Amsterdam Printed in The Netherlands
Dominant lethal$ in young female mice Trenimon is a very effective mutagen in male Drosophila ~ and male C3H mice 3, Female mice, IO-II weeks old, of the same stock, fertilized between the 8th and 9th hours and between the 2nd and the 32nd days after intraperitoneal injection of Trenimon proved to be much less sensitive to the induction of dominant lethal mutations than male mice 4. This is mainly in agreement with the findings of CATTANACH for TEM in adult female mice 1. These results suggested that alkylating agents such as Trenimon or TEM do not constitute a great genetic hazard to oocytes. To be certain we injected Trenimon intraperitoneally in progressingly younger females, and caged each treated mouse at the age of IO weeks for one week with one male. Each female received a single injection of o.125 mg Trenimon/kg in physiological saline solution before mating. The controls received only saline solution. Time of mating was ascertained by the appearance of a vaginal plug. The females were autopsied after 13.5-16.5 days of gestation, and the numbers of dead and living embryos and corpora lutea were recorded. The measure for induced dominant lethal mutations reported here was a significant increase in the rate of dead/total implantations over the control values. To test the sensitivity o f the earliest stages of ovogenesis, too, we treated embryos in utero by intraperitoneal injection of the mother. Females treated in utero were also mated at the age of IO weeks, and examined for the number of dead implantations and so on. Abbreviations : TEM, triethylenenlelanline ; Treninlon, 2,3,5-triethylene-imino-benzoquinone-I,4.
Mutation Res., 4 (1967) 231-233
232
SHORT COMMUNICATIONS
The test for homogeneity (modification of the Z ~ method of BRANDT-SNEDECOR according to WEBER 7) showed that there was no heterogeneity between the various control series (P > o.I). Therefore we combined the control values, and compared each experiment with the combined control series by means of the Z 2 method. Fig. I demonstrates that there exists a sensitive phase to Trenimon in young female mice. The increase in dead implantations over the control is significant (P < 0.0027) in weeks 7, 6, 5, 4, 3, and IO days (after birth), but it is not significant before (I week) and after (8 and 9 weeks) this sensitive period. Whether there are significant differences within this sensitive phase will be discussed after more results have been accumulated 5. $o ~o 0
/ 30
Q.
2O t~
N m
fo~ooo
e oge of treoted ~
; in weeks after birth
dey$ post-coitus
Fig. i. Percentages of dead/total implantations after injection of o.125 mg/kg Trenimon into C3H-~ mice of various ages. The injection of Trenimon to 58 five-week-old females also yielded no dominant lethals (55 dead to 454 total implantations ---- 12.1% ; P = 0.3) when mated 3.5 weeks later at the age of 8.5 weeks. Animals treated in utero on the I4th and I5th days of embryonic development also showed no induction of dominant lethals (P = o.78 ; o.7o ). Results of still earlier treatment (II, and especially 7, days post-coitus of the mother), however, suggest a mutagenic action of Trenimon during this stage of embryonic ovogenesis (P -----o.o5; P < o.oi ; P ~ 0.0o27 for the sum of the both f f values). A possible explanation of the last-mentioned different sensitivity may be that up to the I l t h day post-coitus the gonads of mouse embryos contain mainly so-called primordial or primitive germ cells or oogonia. Afterwards the female germ cells begin to enter the first meiotic prophase e. The teratogenic side-effects on intra-uterinely treated embryos, the cytotoxic effects on oocytes, the pre-implantation egg losses, the influence of the time interval between treatment and mating, the importance of the specific age and some other conditions of the treated animals and other related problems will be discussed in detail elsewhere 6. Mutation Res., 4 (1967) ~31-233
SHORT COMMUNICATIONS
233
Briefly we can say that because of the quick decomposition of Trenimon, and the observation that the shortest intervals between treatment and mating yielded no significant increase in dead implantations, it seems certain that the observed effects in the sensitive phase are not due to the teratogenic action of stored Trenimon. When treating the mothers with chemicals one cannot completely exclude the possibility that lesions of the uterus or adnexa influence the ratios of dead implantations, but the specific sensitivity pattern (especially that of the embryonic stages of ovogenesis) and the negative results after treatment of 5-week-old females and mating them with 8.5-week-old males, makes this improbable. We therefore assume that the observed effects are in fact genetic ones, that means induced dominant lethals.
Institute for Anthropology and Human Genetics, University of Heidelberg (Germany)
GUNTER ROHRBORN HELGA BERRANG
I CATTANACH, ]~[., The effect of triethylene melamine on the fertility of female mice, Intern. J. Radiation Biol., I (1959) 288-299. 2 Lt3ERS, H., AND G. R(JHRBORN, Chelnische Konstitution und mutagene Wirkung, III. "~thylenimine, Mutation Res., 2 (1965) 29-44. 3 R6HRBORN, G., Die mutagene Wirkung yon Trenimon bei der m~innlichen Maus, Humangenetih, i (1965) 576-578. 4 R6HRBORN, G., l~ber einen Geschlechtsunterschied in der mutagenen Wirkung von Trenimon bei der weiblichen Maus, Humangenetik, 2 (1966) 81-82. 5 R6HRBORN, G., in preparation. 6 SNELL, G. D., in G. D, SNELL (Ed.), Biology of the Laboratory Mouse, Dover, New York, 1941, P. 55. 7 WEBER, E., Grundriss der biologischen Statistih, 4th ed., Fischer, Jena, 1961.
Received September 5th, 1966 Revised manuscript received October 28th, 1966 Mutation Res., 4 (1967) 231-233
231
I BATCHELOR, A. L., AND P. W. EDMONDSON, Fast-neutron irradiation of large animals, in : Biological E~ects of Neutron and Proton Irradiations, Vol. I I , International Atomic Energy Agency, Vienna, 1964, pp. 3-13. 2 BATCHELOR, A. L., t~. J. s. PHILLIPS AND A. G. SEARLE, A c o m p a r i s o n of the mutagenic effectiveness of chronic neutron- and F-irradiation of mouse spermatogonia, Mutation Res., 3 (I966) 218-229. 3 CARTER, T. C., M. F. LYON AND R. J. S. PHILLIPS, Genetic h a z a r d of ionizing radiations, Nature, 182 (1958 ) 409 . 4 RUSSELL, W. L., The effect of radiation dose rate and fractionation on mutation in mice, in: F. H. SOBELS (Ed.), Repair from Genetic Radiation Damage, Pergamon, Oxford, 1963, pp. 205217 . 5 RUSSELL, W. g., Studies in mammalian radiation genetics, Nt*cleonics, 23 (1965) 53-62. 6 RUSSELL, W. L., The nature of the dose-rate effect of radiation on mutation in mice, Japan J. Genet., 4 ° , Suppl. (1965) 128-14o. 7 SEARLE, A. G., Progress in mammalian radiation genetics Proc. 3rd Internat. Congr, Radiation Research, Cortina, 1966, in the press. 8 SEARLE, A. G., AND R. j. s. PHILLIPS, Genetic effects of high L E T radiation in mice, Radiation Res., in the press.
Received October 7th, 1966 Mutation Res., 4 (1967) 229-231
Mutation Research Elsevier Publishing Company, Amsterdam Printed in The Netherlands
Dominant lethal$ in young female mice Trenimon is a very effective mutagen in male Drosophila ~ and male C3H mice 3, Female mice, IO-II weeks old, of the same stock, fertilized between the 8th and 9th hours and between the 2nd and the 32nd days after intraperitoneal injection of Trenimon proved to be much less sensitive to the induction of dominant lethal mutations than male mice 4. This is mainly in agreement with the findings of CATTANACH for TEM in adult female mice 1. These results suggested that alkylating agents such as Trenimon or TEM do not constitute a great genetic hazard to oocytes. To be certain we injected Trenimon intraperitoneally in progressingly younger females, and caged each treated mouse at the age of IO weeks for one week with one male. Each female received a single injection of o.125 mg Trenimon/kg in physiological saline solution before mating. The controls received only saline solution. Time of mating was ascertained by the appearance of a vaginal plug. The females were autopsied after 13.5-16.5 days of gestation, and the numbers of dead and living embryos and corpora lutea were recorded. The measure for induced dominant lethal mutations reported here was a significant increase in the rate of dead/total implantations over the control values. To test the sensitivity o f the earliest stages of ovogenesis, too, we treated embryos in utero by intraperitoneal injection of the mother. Females treated in utero were also mated at the age of IO weeks, and examined for the number of dead implantations and so on. Abbreviations : TEM, triethylenenlelanline ; Treninlon, 2,3,5-triethylene-imino-benzoquinone-I,4.
Mutation Res., 4 (1967) 231-233
232
SHORT COMMUNICATIONS
The test for homogeneity (modification of the Z ~ method of BRANDT-SNEDECOR according to WEBER 7) showed that there was no heterogeneity between the various control series (P > o.I). Therefore we combined the control values, and compared each experiment with the combined control series by means of the Z 2 method. Fig. I demonstrates that there exists a sensitive phase to Trenimon in young female mice. The increase in dead implantations over the control is significant (P < 0.0027) in weeks 7, 6, 5, 4, 3, and IO days (after birth), but it is not significant before (I week) and after (8 and 9 weeks) this sensitive period. Whether there are significant differences within this sensitive phase will be discussed after more results have been accumulated 5. $o ~o 0
/ 30
Q.
2O t~
N m
fo~ooo
e oge of treoted ~
; in weeks after birth
dey$ post-coitus
Fig. i. Percentages of dead/total implantations after injection of o.125 mg/kg Trenimon into C3H-~ mice of various ages. The injection of Trenimon to 58 five-week-old females also yielded no dominant lethals (55 dead to 454 total implantations ---- 12.1% ; P = 0.3) when mated 3.5 weeks later at the age of 8.5 weeks. Animals treated in utero on the I4th and I5th days of embryonic development also showed no induction of dominant lethals (P = o.78 ; o.7o ). Results of still earlier treatment (II, and especially 7, days post-coitus of the mother), however, suggest a mutagenic action of Trenimon during this stage of embryonic ovogenesis (P -----o.o5; P < o.oi ; P ~ 0.0o27 for the sum of the both f f values). A possible explanation of the last-mentioned different sensitivity may be that up to the I l t h day post-coitus the gonads of mouse embryos contain mainly so-called primordial or primitive germ cells or oogonia. Afterwards the female germ cells begin to enter the first meiotic prophase e. The teratogenic side-effects on intra-uterinely treated embryos, the cytotoxic effects on oocytes, the pre-implantation egg losses, the influence of the time interval between treatment and mating, the importance of the specific age and some other conditions of the treated animals and other related problems will be discussed in detail elsewhere 6. Mutation Res., 4 (1967) ~31-233
SHORT COMMUNICATIONS
233
Briefly we can say that because of the quick decomposition of Trenimon, and the observation that the shortest intervals between treatment and mating yielded no significant increase in dead implantations, it seems certain that the observed effects in the sensitive phase are not due to the teratogenic action of stored Trenimon. When treating the mothers with chemicals one cannot completely exclude the possibility that lesions of the uterus or adnexa influence the ratios of dead implantations, but the specific sensitivity pattern (especially that of the embryonic stages of ovogenesis) and the negative results after treatment of 5-week-old females and mating them with 8.5-week-old males, makes this improbable. We therefore assume that the observed effects are in fact genetic ones, that means induced dominant lethals.
Institute for Anthropology and Human Genetics, University of Heidelberg (Germany)
GUNTER ROHRBORN HELGA BERRANG
I CATTANACH, ]~[., The effect of triethylene melamine on the fertility of female mice, Intern. J. Radiation Biol., I (1959) 288-299. 2 Lt3ERS, H., AND G. R(JHRBORN, Chelnische Konstitution und mutagene Wirkung, III. "~thylenimine, Mutation Res., 2 (1965) 29-44. 3 R6HRBORN, G., Die mutagene Wirkung yon Trenimon bei der m~innlichen Maus, Humangenetih, i (1965) 576-578. 4 R6HRBORN, G., l~ber einen Geschlechtsunterschied in der mutagenen Wirkung von Trenimon bei der weiblichen Maus, Humangenetik, 2 (1966) 81-82. 5 R6HRBORN, G., in preparation. 6 SNELL, G. D., in G. D, SNELL (Ed.), Biology of the Laboratory Mouse, Dover, New York, 1941, P. 55. 7 WEBER, E., Grundriss der biologischen Statistih, 4th ed., Fischer, Jena, 1961.
Received September 5th, 1966 Revised manuscript received October 28th, 1966 Mutation Res., 4 (1967) 231-233