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A DOMINANT LETHAL STUDY IN MALE RATS AFTER REPEATED EXPOSURES TO VINYL CHLORIDE OR VINYLIDENE CHLORIDE Robert D. Short, Jan L. Minor, Joseph M. Winston, Cheng-Chun Lee Pharmacology and Toxicology, Midwest Research Institute, Kansas City, Missouri Male CD rats were exposed 6 hr/day for 5 days/wk to 0, 50, 250, or 1,000 ppm of vinyl chloride or 55 ppm of vlnylidene chloride. Starting on week 11 of exposure, these males were mated with untreated females. There was no evidence of either preimplantation loss or postlmplanation loss in pregnant females that resulted from these matings. Consequently, it was concluded that these exposures did not produce germinal mutation, as manifested by a dominant lethal effect, In male rats. INTRODUCTION Vinyl chloride (VC) and vinylidene chloride (VDC) are structurally related monomers used in the synthesis of plastics. VC is carcinogenic in both humans (Creech and Johnson, 1974) and animals (Viola et al., 1971; Maltoni and Lefemine, 1975; Lee et al., 1977). In addition, both VC and VDC are mutagenic in a variety of microbial systems (Loprieno et al., 1976; Bartsch et al., 1975). Epidemiological studies suggest that VC produces germinal mutations, as manifested by increased fetal loss, in humans (Infante et al., 1976). However, there-was no evidence of a dominant lethal mutation in male mice exposed to VC for 5 days and sequentially mated with untreated females (Anderson et al., 1976). The purpose of this study was to determine whether repeated exposures to VC or VDC, for 11 wk, produced germinal mutations of the dominant lethal type in male rats. METHODS CD rats (Charles River Breeding Laboratories, North Wilmington, Massachusetts) were used in this study. Adult males weighing 180-200 g were exposed 6 hr/day for 5 days/wk to 0, 50, 250, or 1,000 ppm of VC or 55 ppm of VDC (Lee et al., 1977). All rats were given free access to The authors gratefully acknowledge the competent technical assistance of Brett Ferguson and Tim Unger. Requests for reprints should be sent to Robert O. Short, Pharmacology and Toxicology, Midwest Research Institute, 425 Volker Boulevard, Kansas City, Missouri 64110. 965 Journal of Toxicology and Environmental Health, 3:965~96S, 1977 Copyright 1977 by Hemisphere Publishing Corporation SL 040588 966 R. D. SHORT ET AL. feed and water except during the daily exposure period, when feed was withheld. Since potential effects of these compounds on specific stages of the spermatogenic cycle were not of immediate concern, we employed a modification of the dominant lethal study described by Green et al. (1977). During week 11 of exposure, each male was housed with two unexposed virgin females for at least seven successive evenings or until the male mated with two females. During the day, the males continued to be exposed to VC or VDC. The females were examined in the morning for evidence of mating as demonstrated by the presence of sperm in vaginal smears or the presence of a vaginal plug. On gestational day 13, females were sacrificed and the number of corpora lutea and implants (viable and dead) was determined. RESULTS Although VC exposure did not significantly reduce the ratio of fertile to exposed males, there was a reduced number of males, in the group exposed to 1,000 ppm of VC, that mated with two females (Table 1). In addition, there was a reduced ratio of pregnant to mated females in the TABLE 1. Number and Classification of Rats in Study with Vinyl Chloride (VC) and Vinylidene Chloride (VDC) No. of rats at chamber concentration (ppm) VC VDC Classification 0 50 250 1,000 55 Exposed male rats Tested 12 12 12 12 11 Mated with two females 12 10 11 8a 9 Mated with one female 021 2 2 Mated with no females 000 2 0 Fertile" 12 11 10 10 10 Unexposed female rats Mated 24 22 23 Pregnant 24 20 16 With one dead implant 757 With two dead implants 3 2 0 With > three dead implants 2 2 1 18 14 5 1 0 20 3 1 2 `'Significantly different from control (Fisher's exact probability test, p < 0.05). ^Fathered at least one litter with one or more viable implants. SL 040589 DOM TABl Corpi Implc Impl; (Viab "Mr grot Of V 8-9 une; and prei lute to t wit! pro< sarc moi wer rela loss or imp fern VC to rea; pre, dor imp was pro * . SHORT ETAL. feed was ;tages of the employed a 3reen et al. >d with two or until the tinued to be morning for ti in vaginal 13, females (viable and io of fertile i the group Table 1). In isles in the W DOMINANT LETHAL STUDY WITH VC AND VDC 967 TABLE 2. Mating Results in Study with Vinyl Chloride (VC) and Vinylidene Chloride (VDC) Chamber concentration (ppm) VC VDC Measurement 0 50 250 1,000 55 Corpora lutea per dam Implants per dam Implants per corpora lutea X 100 (Viable/total implants) X 100 15.8*0.6" 13.7 0.8 86 3 92 2 14.3 0.8 12.0 0.9 83 5 92 3 15.2 0.3 13.4 0.5 88 4 93 3 15.1 0.6 11.9 0.9 81 6 95 2 15.1 0.5 14.2 0.5 94 2 93 3 "Mean SE calculated on a per dam basis for the number of pregnant rats indicated in Table 1. 4 groups mated with males exposed to 250 or 1,000 ppm of VC or 55 ppm of_VDC. The mating period ranged from 7 days for the control group to *8-9 days for the treated groups. Pregnancies that resulted from the mating of exposed males with unexposed females were normal in terms of the number of corpora lutea and implants (Table 2). In these females, there was no evidence of preimplantation loss, as measured by the ratio of implants to corpora lutea, or postimplantation loss, as measured by the ratio of viable implants to total implants. DISCUSSION The present study was incorporated into an ongoing carcinogenic study with VC and VDC to evaluate the potential of these compounds for producing dominant lethal effects. In the carcinogenic study, hemangiosarcoma of the liver and lung occurred in CD rats exposed for 9-12 months to 250 or 1,000 ppm of VC (Lee et al., 1977). In addition, there were no lesions in the testes or accessory organs indicative of a treatment^ related effect on reproductive performance. In the present study, neither preimplantation nor postimplantation losses were observed in pregnancies that'jesuTleJTrom the mating of VCor VDC^eXpoSed~males (Table' 2). However, the occurrence of a preimplantatiorTToS^vasTuggested by the increased number of nonoreenant J females in the groups mated with males exposed to 250 or 1 r000 ppm nT VC or 55""ppm j>f VDC (Table 1). The relevance of this latter observation to the" detection of a dominant lethal effect is questionable for two reasons. First, there was no indication of a preimplantation loss in pregnant rats. Second, a postimplantation loss is more indicative of dominant lethality than a preimplantation loss. Since neither pre-' implantation nor postimplantation losses were observed in pregnant rats, it was concluded that the VC and VDC exposures used in this study did not produce dominant lethal mutations in the germinal cells of male rats. 968 R. D. SHORT ET Al_ REFERENCES Anderson, 0., Hodge, M, C. E. and Purchase, I. F. H. 1976. Vinyl chloride; Dominant lethal studies in male CD-I mice. Mutat. Res. 40:359-370. Bartsch, H., Malaveille, C, Montesano, R. and Tomatis, L 1975. Tissue-mediated mutagenicity of vinylidene chloride and 2-chlorobutadiene in Salmonella typhimurium. Nature (Land.) 255:641-643. Creech, J. L, Jr. and Johnson, M. N. 1974. Angiosarcoma of liver in the manufacture of polyvinyl chloride. J. Occup. Med. 16:150-151. Green, S., Moreland, F. M. and Flamm, W. G. 1977. A new approach to dominant lethal testing. Toxicol. Appl. Pharmacol. 39:549-552. Infante, P. F., Wagoner, J. K. and Waxweiler, R. J. 1976. Carcinogenicity, mutagenicity, and teratogenic risks associated with vinyl chloride. Mutat Res, 41:131-142. Lee, C. C., Bhandari, J. C., Winston, J. M., House, W. B., Peters, P. J., Dixon, R. L. and Woods, J. S. 1977. Inhalation toxicity and carcinogenicity of vinyl chloride and vinylidene chloride. Environ. Health Perspect,, in press. Loprieno, N., Barale, R. and Baroncelli, S. 1976. Evaluation of the genetic effects induced by vinyl chloride monomer (VCM) under metabolic activation: Studies in vitro and in vivo. Mutat, Res, 40:85-95. Maltoni, C. and Lefemine, G. 1975. Carcinogenicity bioassays of vinyl chloride, current results. Ann. N.Y. Acad. Scl. 246:195-218. Viola, P. L., Bigotti, A. and Caputa, A. 1971. Oncogenic response of rat skin, lungs, and bones to vinyl chloride. Cancer Res. 31:516-519. Received July 22, 7977 Accepted September 11, 1977 0^0591 INTR< The h index, 19 of or exp 1974, anc in Canada effervesce 2-yr-old orophary cleanser survey ir populatie The ai also thank and Science The c construed : Reque Commissio Journal of Copyright