Specific-locus test shows ethylnitrosourea to be the most potent mutagen in the mouse.

Abstract

Use of the specific-locus test to measure the frequency of transmitted gene mutations induced in mouse spermatogonia has shown ethylnitrosourea to be by far the most potent mutagen yet discovered in the mouse. The dose used, 250 mg/kg, gave a mutation rate 5 times as high as had been obtained with 600 R, the most effective acute dose of x-rays. Compared to procarbazine, heretofore the most mutagenic chemical known in the mouse, ethylnitrosourea proved to be 15 times more mutagenic than the peak effect obtained with the most effective dose of procarbazine. Because of its high mutagenicity, ethylnitrosourea can serve as a model compound in exploring the effect of such factors as dose response, dose fractionation, sex, and cell stage on the mutagenic action of a chemical. Ethylnitrosourea is clearly the mutagen of choice for the production of any kind of desired new gene mutations in the mouse. More than a score of chemicals, most of them well-known mutagens in other organisms, have been tested by the mouse specific-locus method. Only three, however, have shown a clear-cut positive mutagenic effect in treated mouse spermatogonial stages (1), and the most effective dose of the most potent of these three-namely, procarbazine (2)-produced only approximately one-third as many mutations as had been obtained with 600 R (1 R = 2.6 X 10-4 coulombs/kg) of acute x-irradiation. The impression was developing that perhaps the mammalian testis was efficiently protected against damage by chemicals. The possibility also existed that the spermatogonia might be effective at repairing mutational damage. It began to appear possible that perhaps no chemical could break through the mammalian body's defense barriers to produce more than a moderate mutagenic effect in spermatogonia.Although repair and other defense mechanisms may still prove to operate against many substances, the present results show that N-ethyl-N-nitrosourea (ENU) provides a dramatic exception.MATERIALS AND METHODS ENU (purchased from Bio-Clinical Laboratories, Bohemia, NY) was dissolved in phosphate buffer (3) adjusted to pH 6. The dose injected intraperitoneally was matched to the body weight of each animal by adjusting the volume of the injected solution, which never exceeded 1 ml. Wild-type (101XC3H)Fl male mice 9-19 weeks old were injected with a single dose (250 mg/kg) of ENU. All animals given this dose survive, whereas a 300-mg/kg dose kills about 40%. All injections were completed within less than 1 hr after the chemical was dissolved. The injected males were mated to females of our standard specific-locus test strain (T) which is homozygous for seven marker genes (4). Each male was mated to a group of either two or four females and moved to a new group of females each week. After each 7-week period the males were rotated back to the original group of females to start the cycle over again. The offspring were scored for mutations at the seven loci. RESULTS AND DISCUSSIONThe 250-mg/kg dose induced a long period of sterility. The males were fertile for a...

Flow cytometric analysis of mouse spermatogenic function following exposure to ethylnitrosourea

“…The effects of ENU, a powerful carcinogen and mutagen (24,36), on mouse testicular function have been measured by classical criteria using light and electron microscopy and compared with flow cytometric evaluation of the same samples. ENU had no significant effect on body weights of the mice, whereas it caused up to a 50% loss of testicular weights, indicating the sensitivity of the testis to the action of this chemical.…”

Assessing human germ‐cell mutagenesis in the Postgenome Era: A celebration of the legacy of William Lawson (Bill) Russell

“…Later in his career, Russell pioneered the application of the specific-locus test to study the induction of germ-cell mutations by chemicals. These studies led to what is perhaps one of the most important legacies of his work in germ-cell mutagenesis-his discovery that ethylnitrosourea (ENU) is a highly potent germ-cell mutagen in the mouse [Russell, et al, 1979]. Because ENU induces primarily single base-pair changes, it has become a highly valuable tool for gene discovery in laboratories around the world [Michaud et al, 2005].…”

Functional annotation of mammalian genomic DNA sequence by chemical mutagenesis: A fine-structure genetic mutation map of a 1- to 2-cM segment of mouse chromosome 7 corresponding to human chromosome 11p14-p15

“…T he supermutagenicity of N-ethyl-N-nitrosourea (ENU) (1) has made it possible to recover heritable mutations at high frequency from mouse spermatogonial stem cells. Consequently, it is possible to use phenotype-driven strategies to recover new ENU-induced mutations in specific, preselected loci for the generation of new alleles (1)(2)(3)(4), in whole-genome screens for genes comprising components of complex pathways or phenotypes (5-10), or, with the use of chromosomal rearrangements, in a wide variety of genes within specific chromosomal regions (11)(12)(13)(14)(15)(16)(17)(18).…”