[Comparative analysis of ionizing radiation and xenobiotics influence on spermatogenic epithelium and dominant lethal mutations output in laboratory animals].

The study covered state of spermatogenic epithelium and dominant lethal mutations output in mice of BALB/c and CBA lines, subjected to total gamma-irradiation and in Wistar rats after intraperitoneal injection of potassium bichromate (K2Cr2,O7) in small and sublethal doses. The BALB/c line mice under low irradiation dose (0.25 Gy) demonstrated stimulation effect on spermatogenic epithelium, but in the CBA line mice no such effect was seen. Both mice lines under irradiation of 0.25 Gy and 1.0 Gy demonstrated increase in pathologic sperm counts and in percentage ofpreimplantation embryonal death. In rats, injection of potassium bichromate in doses of 0.028 mg/kg and 2.8 mg/kg increased number of micronuclear spermatids, larger pathologic sperm counts and percentage of postimplantation deaths. Thus, lower general embryonal deaths under radiation exposure is due to preimplantation embryonal deaths, under exposure to 6-valent chromium--is due to postimplantation losses.

Tests of induction in mice by acute and chronic ionizing radiation and ethylnitrosourea of dominant mutations that cause the more common skeletal anomalies.

Assessment-of-Dominant-Damage (ADD) experiments explored induction by proven specific-locus mutagens of dominant mutations that cause skeletal anomalies, cataracts, and stunted growth in offspring of mutagenized male mice. The data set reported here includes 6134 offspring. Mutagenic treatments included 600 R (i.e., approximately 6 Gy) of X-rays delivered in about 7 min, 600 R of gamma rays delivered over about 110 days, and four weekly intraperitoneal (i.p.) injections of 77.5 mg/kg of ethylnitrosourea (ENU). The results reported in this paper are restricted to mutations induced in stem-cell spermatogonia and to the 34 more common skeletal anomalies (i.e., those found in 0.5% or more of the control offspring). Mutation induction was demonstrated for eight anomalies in the acute X-ray experiment and for 17 anomalies (including those same eight anomalies) in the ENU experiment. In spite of the surprisingly high mutation rates found for these treatments, there was no hint of any induction of such dominant mutations by 600 R of chronic gamma radiation. Our results suggest that several anomalies related to variation in the sacralization pattern may be particularly useful for revealing induction of dominant mutations.

[The genetic monitoring of the house mouse population from areas contaminated by radionuclides as a result of the accident at the Chernobyl Atomic Electric Power Station]. / Geneticheskii monitoring populiatsii domovykh myshei iz raionov, zagriaznennykh radionuklidami v rezul'tate avarii na Chernobyl'skoi AES.

In 1986-1994, genetic effects of radiation on males of wild mice caught in areas of radioactive contamination after the Chernobyl reactor accident were studied. The gamma-radiation dose rates on the soil surface in the areas varied from 0,02 to 200 mR/h. The frequency of reciprocal translocations in mouse spermatocytes was relatively low and increased linearly with increasing of the dose rate. The levels of recessive lethal mutations in mouse populations did not depend on the radiation dose rate in the areas and decreased with a time.

[ Multigeneration carcinogenesis in the evaluation and prognosis of delayed radiation effects]. / Znachenie multigeneratsionnogo kantserogeneza v otsenke i prognoze otdalënnykh radiatsionnykh éffectov.

Radiation multigeneration carcinogenesis' mechanisms are associated with inheritance of non-specific genome lesions. Evidence indicates that the effect is significantly enhanced by some chemical carcinogens action on the offspring of irradiated parents. Hereditary character of the phenomenon is a prerequisite to accumulation of these genetic determinants in the population. The risk of radiation multigeneration carcinogenesis for man could be as much as 0.4 x 10(-2) Sv-1 over one generation.

[The effect of low and sublethal doses of ionizing radiation on the function of the spermatogenic epithelium and the yield of dominant lethal mutations in different strains of mice]. / Vliianie malykh i subletal'nykh doz ioniziruiushchego izlucheniia na sostoianie spermatogennogo épiteliia i vykhod dominantnykh letal'nykh mutatsii u myshei raznykh linii.

There was a 30% increase in spermatozoids with microheads and a 3-fold increase in "young" spermatozoids at later times after irradiation of BALB/c mice a dose of 1.0 Gy. With radiation doses of 0.1-1.0 Gy, BALB/c and CBA mice (which differ in viability of spermatogenous cells, the number of Sertoli cells and total embryonal death rate) exhibited a significant increase in the preimplantation death at the stage of spermatogonia and stem cells. The results obtained indicate reduction of the fertilizing capacity of sperm which is probably realized as dominant lethal mutations.

[An experimental assessment of the genetic effects of low doses of internal irradiation]. / Eksperimental'naia otsenka geneticheskikh éffektov malykh doz vnutrennego oblucheniia.

Genetic study of rats showed the possibility of registration of the minor doses of the inside and outside irradiation. The postmiosis mutations, number of reciprocal translocations in spermatogonia may be used for indexation of the effects.

Detection of gamma-ray-induced DNA damages in malformed dominant lethal embryos of the Japanese medaka (Oryzias latipes) using AP-PCR fingerprinting.

Adult male fish of the medaka HNI strain exposed to 9.5 Gy or 19 Gy (0.95 Gy/min) of gamma-rays were mated with non-irradiated female fish of the Hd-rR strain. Genomic DNA was prepared from malformed individual embryos which were expected to be dominant lethal and used for AP-PCR fingerprinting. By the use of a part of the T3 promoter sequence (20 mer), which, to our knowledge, is not found in the medaka genome as an arbitrary primer, we found polymorphisms in genomic fingerprints which could distinguish the parental strains. On the other hand, we found that the fingerprints of F1 hybrids were the sum of those of their parents. Based on these findings, we analyzed the fingerprints of genomic DNA of each severely malformed embryo, because we expect that radiation-induced genomic damages resulting in severe malformation and eventually in dominant lethals should be detected as changes in paternal fingerprints of F1 hybrids. Indeed, we succeeded in detecting changes in genomic DNA as loss of some paternal bands in fingerprints of malformed embryos. One of 10 malformed embryos obtained from 9.5 Gy gamma-irradiated males had lost one band of the paternal origin and 4 of 12 malformed embryos obtained from 19 Gy gamma-irradiated males had lost 5 bands. These results indicated a possibility that quantitative as well as qualitative estimation of gamma-ray-induced DNA damages can be made by this method which does not require the functional selection based on a specific target gene.

An enhancement of the yield of X-ray-induced Minute mutations in the c3G female-ywmf-2 male system of Drosophila melanogaster.

The possible enhancement of the yield of X-ray-induced Minute mutations in the c3G female-ywmf-2 male system which is proposed to be responsible for the high production of spontaneous Minute mutations was investigated. To determine and compare the yield of X-ray-induced Minute mutations exactly, four series of crosses were made: (a) ywmf-2 male x Oregon-R (OrR) female crosses, spontaneous Minute mutations were scored; (b) ywmf-2 male x c3G female crosses, spontaneous Minute mutations produced in the c3G female-ywmf-2 male system were evaluated; (c) X-irradiated ywmf-2 male x OrR female crosses, the yield of Minutes induced by X-rays in the different stages of male germ cells was evaluated; and (d) X-irradiated ywmf-2 male x c3G female crosses, the yield of X-ray-induced Minutes in the c3G female-ywmf-2 male system was evaluated. The results show that the yield of X-ray-induced Minutes recorded in the c3G female-ywmf-2 male system is 2.37-16.55 times more than that in the ywmf-2 male x OrR female crosses. This finding clearly indicates that the yield of these mutations is greatly enhanced in the c3G female-ywmf-2 male system. This possibly suggests that the c3G female-ywmf-2 male system may be responsible not only for the high production of spontaneous Minute mutations but also for the high formation of radiation-induced Minutes.

Mechanisms for dominance: Adh heterodimer formation in heterozygotes between ENU or X-ray induced null alleles and normal alleles in Drosophila melanogaster.

To study mechanisms for dominance of phenotype, eight ENU- and four X-ray-induced mutations at the alcohol dehydrogenase (Adh) locus were analyzed for partial dominance in their interaction with normal alleles. All ENU and one of the X-ray mutations were single base substitutions; the other three X-ray mutations were 9-21 base deletions. All but one of the 12 mutant alleles were selected for this study because they produced detectable mutant polypeptides, but seven of the 11 producing a peptide could not form dimers with the normal peptide and the enzyme activity of heterozygotes was about half that of normal homozygotes. Four mutations formed dimers with a decreased catalytic efficiency and two of these were near the limit of detectability; these two also inhibited the formation of normal homodimers. The mutant alleles therefore show multiple mechanisms leading to partial enzyme expression in heterozygotes and a wide range of dominance ranging from almost complete recessive to nearly dominant. All amino acid changes in mutant peptides that form dimers are located between amino acids 182 and 194, so this region is not critical for dimerization. It may, however, be an important surface domain for catalyzation.

Development of a possible nonmammalian test system for radiation-induced germ-cell mutagenesis using a fish, the Japanese medaka (Oryzias latipes).

To develop a specific-locus test (SLT) system for environmental mutagenesis using vertebrate species other than the mouse, we first established a tester stock of the fish medaka (Oryzias latipes) that is homozygous recessive at three loci. The phenotypic expression of these loci can be easily recognized early in embryonic development by observation through the transparent egg membrane. We irradiated wild-type males with 137Cs gamma-rays to determine the dose-response relationships for dominant lethal and specific-locus mutations induced in sperm, spermatids, and spermatogonia. Through observation of 322,666 loci in control offspring and 374,026 loci in offspring obtained from 0.64-, 4.75-, or 9.50-Gy-irradiated gametes, specific-locus mutations were phenotypically detected during early development. These putative mutations, designated "total mutation," can be recognized only in embryos of oviparous animals. The developmental fate of these mutant embryos was precisely followed. During subsequent embryonic development, a large fraction died and thus was unavailable for test-crossing, which was used to identify "viable mutations." Our medaka SLT system demonstrates that the vast majority of total mutations is associated with dominant lethal mutations. Thus far only one spontaneous viable mutation has been observed, so that all doubling calculations involving this endpoint carry a large error. With these reservations, however, we conclude that the quantitative data so far obtained from the medaka SLT are quite comparable to those from the mouse SLT and, hence, indicate the validity of the medaka SLT as a possible nonmammalian test system.

Induction of dominant lethal mutations by combined X-ray-acrylamide treatment in male mice.

The induction of mutations following combined treatment with acrylamide (AA) plus X-rays has been determined using the dominant lethal mutations test in Pzh:SFISS male mice. Combinations of a mutagenic dose of both agents (1.00 Gy, 125 mg/kg b.w.) and a non-mutagenic dose, i.e., a dose that alone does not produce dominant lethals (0.25 Gy, 25 mg/kg b.w.), were used. For the discussion of the effects of combined action of X-rays and acrylamide the term 'enhancement in risk' was used whenever the effects observed after combined exposure significantly exceeded the sum of the effects produced separately by the agents. Such an enhanced risk has been observed in late spermatids after combined action of X-rays and AA at non-mutagenic doses, and in spermatozoa, spermatids and late spermatocytes after exposure to mutagenic doses.

Heterochromatization and euchromatization of whole genomes in scale insects (Coccoidea: Homoptera).

In several families of scale insects (coccids), the sex of an embryo is determined by the number of genetically active genomes present (one = males, two = females). In mealybugs (Pseudococcidae), both males and females develop from fertilized eggs but, in the embryos that develop into males, the set of chromosomes (genome) of paternal origin (PG) becomes heterochromatic (H) and genetically inactive and is not transmitted to the offspring. The mechanism that reduces the number of active genomes in male embryos may vary between families and even between congeneric species. Thus, in male embryos of most armored scale species (Diaspididae), the PG is eliminated, while in a few species it becomes H. In two genera of soft scales (Coccidae), males develop from unfertilized eggs when one of two identical genomes of maternal origin becomes H. In most male tissues, one genome remains H. However, in several tissues that become polyploid by endoreduplication, the PG becomes E and genetically active. The tissues in which the PG becomes E often vary between species and the analysis of hybrid males demonstrated that whether the PG becomes H or remains E is determined by the genome of maternal origin. The euchromatization of the PG in the haploid sector of mosaic male embryos and the presence of spermatocytes with two E genomes (instead of one E and one H), following the irradiation of young mealybug males, strongly suggest that the maintenance of the H state requires the presence of a genetically active genome.

[Selective systems for obtaining recessive ribosomal suppressors in saccharomycete yeasts]. / Selektivnye sistemy dlia polucheniia retsessivnykh ribosomnykh supressorov u drozhzhei sakharomitsetov.

Recessive mutations only occurring in two genes (ribosomal suppressors sup1 and sup2) can be obtained using special selective system. We demonstrate that the absolute selectivity of the system is based on selection for simultaneous reversions to prototrophy in mutants requiring adenine and histidine in haploids marked by two different nonsense mutations--his7-1 (UAA) and ade1-14 (UGA, this being identified in the present study). In support to this conclusion, we developed an analogous system utilising his7-1 (UAA) and lys2-87 (UGA). The selectivity of the system is shown to be influenced both by the choice of nonsense alleles and by genotypic background.

Studies of the induction of dominant lethals and translocations in male mice after chronic exposure to microwave radiation.

Male C3H mice were exposed to 100 W m-2 of 2.45 GHz continuous-wave microwave radiation for 6 h per day for a total of 120 h over an 8-week period. The exposure level was chosen so that the specific energy absorption rate (SAR) would be approximately equal to the level of 4 W kg-1 which is considered by a number of organizations to be a threshold for adverse biological effects. At the end of the treatment period the mice were mated with a different group of (C3H x 101) F1 hybrid females each week for the following 8 weeks. There was no significant reduction in pregnancy rate, preimplantation survival or postimplantation survival in the exposed group compared to sham-exposed controls. At the end of the mating period a cytogenetic analysis was carried out of meiotic chromosome preparations of testicular tissue, thus sampling cells that were stem cell spermatogonia during the treatment regime. The results showed no difference in the frequency of reciprocal translocations between the sham and treated groups, or in the frequency of cells with autosome or sex chromosome univalents. Low levels of fragments and exchanges were found in both groups. It is concluded that there is no evidence in this experiment to show that chronic exposure of male mice to 2.45 GHz microwave radiation induces a mutagenic response in male germ cells. This conclusion is in agreement with the observations of Berman et al. (1980), who reported a lack of male germ cell mutagenesis after repetitive or chronic exposure of rats to 2.45 GHz.

Induction of congenital malformation in mice by parental irradiation: transmission to later generations.

In order to investigate the genetic basis of the increased incidence of congenital malformations in the offspring of irradiated mice, the frequency of malformations among the offspring of individual F1 sons of irradiated females was studied in detail. Among 90 fully tested F1 sons of females which had been mated 15-21 days after receiving 360 cGy X-rays 4 were definite or probable carriers of dominant genes giving a low penetrance of malformations. This confirms that the malformations seen in the first generation are of genetic origin and can be transmitted to later generations. However, the incidence and penetrance of the mutant genes detected were too low to account for all the anomalies found in the first generation. It was concluded that the genetic basis of the original anomalies was heterogeneous, with some due to genetic changes of high penetrance and rapidly eliminated, and others due to genes of low penetrance like those found in this work. Other malformations, in both the irradiated and control series, were probably of non-genetic origin.

X-ray- and chemically induced germ-line mutation causing phenotypical anomalies in mice.

A large and significant increase of phenotypical anomalies was observed in the progeny of ICR parent mice treated before mating with X-rays, urethane, 7,12-dimethylbenz[a]anthracene, ethylnitrosourea (ENU), and 4-nitroquinoline 1-oxide, but the increase was not significant with furylfuramide. Major types of induced anomalies were cleft palate, dwarf, open eyelid, tail anomalies, and exencephalus. Dwarf, open eyelid and tail anomalies were predominant types of viable anomalies and were inherited as if they were dominant mutations with varying expressivity or penetrance. Incidence of prenatal anomalies increased with treated doses of X-rays, urethan, or ENU for both spermatozoa and spermatogonia. Spermatogonia were less sensitive to X-rays and urethane than spermatozoa, while ENU induced a very high incidence of prenatal anomalies by the spermatogonial treatment. In contrast to the previous works with X-rays, there was a clear, almost linear increase of anomalies in the dose range from 0 to 216 rad after spermatogonial exposure. For treatment of oocytes, there was also a clear increase with doses of X-rays and urethane. Doubling doses of X-rays for prenatal anomalies were 12 rad for spermatozoa, 27 rad for spermatogonia, and 19 rad for mature oocytes. These values are similar to those for ordinary mouse mutations. However, the mean rate of prenatal anomalies per rad (1.2 X 10(-4), 6.6 X 10(-5) and 9.1 X 10(-5) for spermatozoa, spermatogonia and mature oocytes, respectively) and that for 1 micrograms/g of ENU (3.4 X 10(-4) for spermatogonia) were 4-40 times higher than that of ordinary mutation in mice, because overall phenotypical abnormalities were scored in this study. Information obtained from the work on phenotypical anomalies is valuable to assess genetic risk of radiation and chemicals, because a majority of human genetic diseases show this kind of irregular and uncertain inheritance and most of the induced anomalies are similar to those found in humans.

X-ray-induced dominant lethal mutations in mouse oocytes detected by an in vitro assay.

Female mice were X-irradiated with 0.5-4.5 Gy 2 h before mating to unirradiated males of the same strain. The dominant lethal frequencies (DLF) were determined by growing the embryos in vitro from the two-cell stage and determining the relative rates of successful embryogenesis to the blastocyst stage and to the trophectoderm outgrowth with proliferated inner cell mass stage. The DLF increased with increasing dose, the two linear aspects having a breakpoint at about 1.5 Gy. The nature of embryo failure was also dose dependent. At doses less than 2.0 Gy embryos failed predominantly after blastocyst formation, but at higher doses the embryos failed both before and after blastocyst formation. Over the dose range tested, the frequency with which lesions leading to dominant lethality were induced [i.e., -ln(1 - DLF)] increased linearly with increasing dose.