34 ° 0th ANSI;AIr MI,IET1NG, MIAMI BI,;A(H, I¢I.A.

24 h, hycanthone or TEM injection at I0 h, sacrificing at 4o h. Pyrogen alone was comparable to a negative saline control in the induction of mutation at all levels tested (e, o.e, o.oz mg/kg). Temperature monitoring showed linear increases in tem- perature with log increases in pyrogen doses. This l<)g increase in pyrogen dosage also yielded a linear decrease in micronuclei when administered in conjunction with hv- canthone (zoo mg/kg). When administered with a variation in hvcanthone dosage (zoo, 33, xo mg/kg), z mg/kg pyrogen yielded a decrease in micronuclei folh)wing a curve parallel to the dose-response curve of hycanthone alone. When administered with a variation in TEM dosage (o.5, o.25, o.I mg/kg), 2 mg/kg pyrogen yielded an

O O/ apparent threshold effect" a 3 <o decrease in micronuclei associated with pvrogen at the o.I and o.e5 mg/kg TEM levels doubled at the 0.5 mg/kg TEM level.

61 MITRA, N., AND H. V. MALL1NG, National Institute of Environmental Health Sciences, P. O. Box 12233, Research Triangle Park, N.C. 277o 9 (U.S.A.).

Induction of dominant lethal mutations in male mice by four individual chemicals

The frequency of dominant lethal nmtations in male mice was used as a tneasure to determine the synergistic effect of four chemicals mitomycin C, 5-bromodeoxy- uridine (BUdR), procarbazine and triethylenemelamine (TEM). Eight-week-old males from inbred DBA/eJ strain were injected in four sets of experiments.

(A) Single injection with each chemical. (B) Injections given once a week with the same chemical for four consecutive

weeks. (C) Single injection of each chemical with four times the concentration used

previously. (D) Chemicals injected once a week for four consecutive weeks in the following

order: mitolnycin C, BUdR, procarbazine and TEM. Control animals received simul- taneous injections with Hank's Balanced salt solution. From the tenth week of the first injection, each male was mated at one-week intervals for 8 weeks to virgin fe- males from inbred C57BL/6 J strain. On the seventeenth day after start of mating, the females were checked for dominant lethals. Control values for intra-uterine death rates throughout the course of several replicas was 1.0c){~. The observed frequency of intra-uterine deaths in the nmltiply treated experiment was approximately 15'}o, even though the proposed additive effect was expected to be less than 4'~,- The percentage of dominant lethals dropped from the fifteenth day of mating.

A comparison of the results of the single doses with the lnultiple treatment will be discussed.

62 GREEN, S. *, I;'. M. M()RELANI)* AND (;. W . tgLAMM * *, *Food and Drug Administration, Washington, D.C. ; **National Cancer Institute, Bethesda. Md. (U.S.A.).

A more refinedapproach to dominant lethal testing

One of the major problems in evaluating dominant lethal data has been the extensive amount of information required by present standards to assess definitively

A M E R I C A N E N V I R O N M E N T A L M U T A G E N SOCIETY 341

a positive or a negative response. In an a t tempt to reduce the voluminous amount of data and retain what is considered pertinent information, a new approach was in- vestigated.

The new approach consisted of t reatment for the entire spermatogenic cycle and subsequent mating for two weeks. Weanling and adult male rats were employed as well as two distinct supplies of triethylenemelamine (TEM). All animals were of the Sprague-Dawley strain. Ten animals were assigned to each group. Weanlings (aver- age weight IOO g) were intubated five days per week for ten weeks with TEM at dos- ages of 0.025, 0.050 or 0.075 mg/kg/day. Adult animals (average ewight 24 ° g) were exposed to the same treatment schedule but at a dosage of 0.050 mg/kg/day. The animals were then mated each of two weeks to two virgin females.

Statistical analyses of the number of dead implants per pregnant female and of the number of females with one or more dead implants or two or more dead implants indicated significant increases in all indexes for the adults and a more variable res- ponse on the part of the weanlings. There were no differences in responses that could be ascribed to the TEM supply.

These observations, when compared to those obtained from a more traditional dominant lethal s tudy of TEM, indicate that there is no loss of pertinent information by employing this approach, except that of stage sensitivity. I t reduces the amount of data required and assesses the more pertinent dominant lethal effects, that of post- implantation losses.

63 GENEROSO, W. M., S. W. HUFF AND K. T. CAIN, Biology Division, Oak Ridge National Laboratory, Oak Ridge, Tenn. 37830 (U.S.A.).

Comparative inducibility by 6-mercaptopurine of dominant lethal mutations and heritable translocations in early meiotic male germ cells and differentiating spermatogonia of mice

Until recently, the following three points appeared to be generally true in male mice. First, that chemicals which clearly produced chromosome aberrations in male mice are either alkylating agents, or are known to be transformed in vivo into alkyla- ting forms. Second, clear-cut chromosome breakage effects induced by chemicals, as detected by dominant lethal mutation and heritable translocation tests, were found only when male post-meiotic germ cells and spermatocytes that were in late meiotic stages were treated. And third, in the postmeiotic stages, heritable translocations are a more sensitive and reliable endpoint than dominant lethal mutations in detecting chromosome breakage.

Recent results with 6-mercaptopurine (6-MP) clearly indicate that these gene- ralizations are no longer valid. This compound is not an alkylating chemical nor is it known to be transformed into one. I t was found that 6-MP, unlike all other known clear-cut chromosome breakers, induced dominant lethal mutations only in late differentiating spermatogonia and early spermatocytes (RAY AND HYNECK, Environ. Health Perspect., 6: 27-36; GENEROSO, PRESTON AND BREWEN, Mutation Res., in press). Cytological analysis of the germ cells affected as they pass through the dia- kinesis-metaphase I stage, revealed that chromatid deletions were the likely cause of