- Email: [email protected]
Dose-response studies on the induction of dominant lethal mutations in postspermatogonial stages of the mouse treated with triethylenemelamine (TEM)
EUROPEAN ENVIRONMENTAL
MUTAGEN SOCIETY
41
(C3H x IOI), CBA/CaLacY and C57BL/6Y to the mutagenic action of dES at a dose of 150 mg/kg (intrascrotal) were compared. The I& males were more sensitive than CBA (P < 0.01). dES did not influence mortality of F, male descendants. It is necessary to test the ability of some of the drugs, that gave contradictory results in the various dominant lethal experiments, to induce recessive lethals (embryonic mortality in the first back-cross generation). The backcross test was used in an attempt to induce mutations with dES (dose of 6 x 6 = 36 mg/kg). IOI male F, from treated mice and 97 male I;, mice in the control group were estimated on the basis of embryonic lethality in BC, progeny. dES increased pre-implantation mortality only in the EC, generation of the treated group (P < 0.001). In controls the distribution of F, males on the viability (I-E/CL) of its BC, descendants was binomial. The deviation from a binomial curve in the treated group was due to the existence of lethal carriers. 16 F, males from treated spermatogonial cells and 5 males in the control group were probable lethal carriers (P < 0.05). Among 16 lethal carriers from the treated group, two had dominant effects on embryonic lethality and were gonadal mosaics for reciprocal translocation (T4z /Y). Consequently the estimation of induced recessive lethal frequency is difficult on the basis of the difference between embryonic mortality in BC, generations in treated and control groups only, without knowledge on the nature of induced lethals. But for testing of chemicals it is sufficient that lethal carriers could be found. It is, however, necessary to test the male mice before the treatment and to eliminate heterozygotes for spontaneous lethals. Abbreviation:
dES,
diethyl
sulphate
32 MATTER, B. E.,* AND W. M. GEKEROSO, ratory, Oak Ridge, Tenn. (U.S.A.).
Biology
Division,
Oak Ridge
National
Labo-
Dose-response studies on the induction of dominant lethal mutations in postspermatogonial stages of the mouse treated with triethylenemelamine (TEM)** Experiments on dose effects of EMS in the induction of dominant lethal mutations suggest that the shape of the curve departs markedly from linearity (W. M. GENEROSO. Abstract, and Annual EMS Meeting, 1971, Washington, D.C.). Certainly it cannot be expected that the shape of the curve will be the same for all chemicals. The present report is on the effects of TEM dose on the induction of dominant lethal mutations in different postspermatogonial stages. TEM was selected because, in many respects, its effects in the mouse are similar to those of radiation and because there might be a marked difference between TEM and EMS in the ratio of genetically effective dose to lethal dose. *Present address: Medical and Biological Research Division, Sandoz Ltd., Base], Switzerland. **Research jointly sponsored by the National Institute of General Medical Sciences, Food and Drug Administration, and the U.S. Atomic Energy Commission under contract with Union Carbide Corporation. .%bbreviations: EMS, ethyl methanesulfonate; TEM, triethylenemelamine.
2nd ANNUAL MEETING,
42
~INKOVY
CASTLE
rz-week-old (101 x C3H)F, male mice were injected intraperitoneally with various concentrations of TEM. The frequencies of dominant lethal mutations were measured in germ cells treated in the early spermatozoa and mid-spermatid stages (4.5 to 7.5 and 11.5 to 15.5 days post treatment, respectively). These posttreatment intervals were selected on the basis of results of preliminary work with a dose of 0.2 mg/kg. The mid-spermatid stage was found to be most sensitive for dominant lethal induction with TEM. Treated males were mated to (C3H x C57BL)F, virgin females. Between 40 to 60 females per dose were sacrificed for uterine analysis. The frequencies of dominant lethal mutations were calculated as a reciprocal of the ratios between living implants in experimental and control animals. The results indicate that TEM is extremely effective in the induction of dominant lethal mutations in both stages studied. The mutagenic effect was already clearly detectable at the 0.05 mg/kg dose. For early spermatozoa, the frequency increased with dose, being 1.2, 9.7, 20.8, 55.9, 80,7 and 91.9% for 0.035, 0.05, 0.1, 0.2, 0.3 and 0.4 mg/kg doses of TEM, respectively. For spermatids, the frequencies were found to be slightly higher at corresponding doses, namely 4.5, 15.1, 39.5,73.2, 92.9 and 96.80/b. The shape of the TEM dose-response curve differs from that of EMS. Another interesting difference between TEM and EMS that has importance from a practical point of view is revealed when comparison is based on the ratio between the genetically effective dose and the lethal dose. For EMS, the lowest dose that kills IOOO/~ of the animals is approx. 525 mg/kg; for TEM this dose is 5 mg/kg. The lowest genetically effective doses (as measured by dominant lethals) for EMS and TEM are 150 mg/kg and 0.05 mg/kg, respectively, which yielded comparable frequencies of dominant lethals. Thus, the ratio of genetically effective dose to lethal dose for TEM is I/IOO while for EMS it is only 1/3.5 .TEM, therefore, in contrast to EMS, is mutagenic far below the toxic level. Obviously, these results have important implications for the practical problems of evaluating mutagenic effects of chemicals. 33 MOLJTSCHEN, J., AND M. MOUTSCHEK-DAHMEN,
Genetics Department,
University
of
Liege, Liege (Belgium). Hazards
of prostaglandins
for chromosome
damage
Prostaglandins are distributed over a large range of animal tissues. Their physiological properties in animal cells have long been investigated, and the structure of their molecules has been fairly well established. However, nothing is known so far about their possible activity on plant cells, especially on nuclear systems. In this investigation, several plant materials were treated with prostaglandin E,. Root-tips of Nigella damascelza, a material suitable for chromosome breakage studies were treated at concentrations ranging from 2.102 to 5.10~ rig/ml during 15 h. A high cytotoxicity of the substance was observed. A dose of 5.10~ ng/ml/r5 h (or higher) produced complete mitotic inhibition, karyorhexis or pycnosis and special nuclear budding which might be related to some specific modifications of DNA synthesis. At a lower dose of 200 ng/ml/r5 h, chromosomal aberrations were observed, some of which seemed to be more specific than those obtained with classical physical or chemical mutagens. The exact time of action of this substance during the mitotic
MUTAGEN SOCIETY
41
(C3H x IOI), CBA/CaLacY and C57BL/6Y to the mutagenic action of dES at a dose of 150 mg/kg (intrascrotal) were compared. The I& males were more sensitive than CBA (P < 0.01). dES did not influence mortality of F, male descendants. It is necessary to test the ability of some of the drugs, that gave contradictory results in the various dominant lethal experiments, to induce recessive lethals (embryonic mortality in the first back-cross generation). The backcross test was used in an attempt to induce mutations with dES (dose of 6 x 6 = 36 mg/kg). IOI male F, from treated mice and 97 male I;, mice in the control group were estimated on the basis of embryonic lethality in BC, progeny. dES increased pre-implantation mortality only in the EC, generation of the treated group (P < 0.001). In controls the distribution of F, males on the viability (I-E/CL) of its BC, descendants was binomial. The deviation from a binomial curve in the treated group was due to the existence of lethal carriers. 16 F, males from treated spermatogonial cells and 5 males in the control group were probable lethal carriers (P < 0.05). Among 16 lethal carriers from the treated group, two had dominant effects on embryonic lethality and were gonadal mosaics for reciprocal translocation (T4z /Y). Consequently the estimation of induced recessive lethal frequency is difficult on the basis of the difference between embryonic mortality in BC, generations in treated and control groups only, without knowledge on the nature of induced lethals. But for testing of chemicals it is sufficient that lethal carriers could be found. It is, however, necessary to test the male mice before the treatment and to eliminate heterozygotes for spontaneous lethals. Abbreviation:
dES,
diethyl
sulphate
32 MATTER, B. E.,* AND W. M. GEKEROSO, ratory, Oak Ridge, Tenn. (U.S.A.).
Biology
Division,
Oak Ridge
National
Labo-
Dose-response studies on the induction of dominant lethal mutations in postspermatogonial stages of the mouse treated with triethylenemelamine (TEM)** Experiments on dose effects of EMS in the induction of dominant lethal mutations suggest that the shape of the curve departs markedly from linearity (W. M. GENEROSO. Abstract, and Annual EMS Meeting, 1971, Washington, D.C.). Certainly it cannot be expected that the shape of the curve will be the same for all chemicals. The present report is on the effects of TEM dose on the induction of dominant lethal mutations in different postspermatogonial stages. TEM was selected because, in many respects, its effects in the mouse are similar to those of radiation and because there might be a marked difference between TEM and EMS in the ratio of genetically effective dose to lethal dose. *Present address: Medical and Biological Research Division, Sandoz Ltd., Base], Switzerland. **Research jointly sponsored by the National Institute of General Medical Sciences, Food and Drug Administration, and the U.S. Atomic Energy Commission under contract with Union Carbide Corporation. .%bbreviations: EMS, ethyl methanesulfonate; TEM, triethylenemelamine.
2nd ANNUAL MEETING,
42
~INKOVY
CASTLE
rz-week-old (101 x C3H)F, male mice were injected intraperitoneally with various concentrations of TEM. The frequencies of dominant lethal mutations were measured in germ cells treated in the early spermatozoa and mid-spermatid stages (4.5 to 7.5 and 11.5 to 15.5 days post treatment, respectively). These posttreatment intervals were selected on the basis of results of preliminary work with a dose of 0.2 mg/kg. The mid-spermatid stage was found to be most sensitive for dominant lethal induction with TEM. Treated males were mated to (C3H x C57BL)F, virgin females. Between 40 to 60 females per dose were sacrificed for uterine analysis. The frequencies of dominant lethal mutations were calculated as a reciprocal of the ratios between living implants in experimental and control animals. The results indicate that TEM is extremely effective in the induction of dominant lethal mutations in both stages studied. The mutagenic effect was already clearly detectable at the 0.05 mg/kg dose. For early spermatozoa, the frequency increased with dose, being 1.2, 9.7, 20.8, 55.9, 80,7 and 91.9% for 0.035, 0.05, 0.1, 0.2, 0.3 and 0.4 mg/kg doses of TEM, respectively. For spermatids, the frequencies were found to be slightly higher at corresponding doses, namely 4.5, 15.1, 39.5,73.2, 92.9 and 96.80/b. The shape of the TEM dose-response curve differs from that of EMS. Another interesting difference between TEM and EMS that has importance from a practical point of view is revealed when comparison is based on the ratio between the genetically effective dose and the lethal dose. For EMS, the lowest dose that kills IOOO/~ of the animals is approx. 525 mg/kg; for TEM this dose is 5 mg/kg. The lowest genetically effective doses (as measured by dominant lethals) for EMS and TEM are 150 mg/kg and 0.05 mg/kg, respectively, which yielded comparable frequencies of dominant lethals. Thus, the ratio of genetically effective dose to lethal dose for TEM is I/IOO while for EMS it is only 1/3.5 .TEM, therefore, in contrast to EMS, is mutagenic far below the toxic level. Obviously, these results have important implications for the practical problems of evaluating mutagenic effects of chemicals. 33 MOLJTSCHEN, J., AND M. MOUTSCHEK-DAHMEN,
Genetics Department,
University
of
Liege, Liege (Belgium). Hazards
of prostaglandins
for chromosome
damage
Prostaglandins are distributed over a large range of animal tissues. Their physiological properties in animal cells have long been investigated, and the structure of their molecules has been fairly well established. However, nothing is known so far about their possible activity on plant cells, especially on nuclear systems. In this investigation, several plant materials were treated with prostaglandin E,. Root-tips of Nigella damascelza, a material suitable for chromosome breakage studies were treated at concentrations ranging from 2.102 to 5.10~ rig/ml during 15 h. A high cytotoxicity of the substance was observed. A dose of 5.10~ ng/ml/r5 h (or higher) produced complete mitotic inhibition, karyorhexis or pycnosis and special nuclear budding which might be related to some specific modifications of DNA synthesis. At a lower dose of 200 ng/ml/r5 h, chromosomal aberrations were observed, some of which seemed to be more specific than those obtained with classical physical or chemical mutagens. The exact time of action of this substance during the mitotic