We used high-density microarrays to evaluate the possible mechanisms by which 2-methoxyacetic acid (MAA) disrupts spermatogenesis. Levels of mRNA transcripts were determined in total RNA isolated from testes of MAA-treated (650 mg/kg i.p.) or concurrent control rats killed 4, 8, 12, or 24 h postexposure (PE). Germ cell death was examined in testis sections using in situ staining for DNA fragmentation. MAA treatment caused increased death of pachytene spermatocytes starting 8 h PE and increasing dramatically at 12 and 24 h PE. Microarray results indicated that at 4 h PE the transcript levels of seven different genes were significantly overrepresented in the testes of MAA-exposed animals. One gene (histone H1 zero [H1f0]) was significantly overrepresented in MAA-treated samples at 4, 8, and 12 h PE. Because expression of this gene has been associated with increased acetylation of core histones, we examined MAA-induced changes in the acetylation of histones H4 (HISTH4) and H3 (HISTH3) in testis nuclear protein. Western blots of acid-extracted testis nuclei indicated that the levels of tetraacetyl histone H4 (4acHIST1H4) and of diacetyl histone H3 (2acHIST1H3) were elevated by MAA treatment at 4, 8, and 12 h PE. Using the same antibodies, 4acHIST1H4 and 2acHIST1H3 were localized primarily to elongating spermatids in testis sections from control animals. At 4 h PE, staining for either histone modification was dramatically increased in spermatogonia and all primary spermatocyte populations except for dividing spermatocytes. MAA treatment of testis nuclear protein extracts from unexposed animals caused both a significant increase in histone acetyltransferase activity and a significant inhibition of histone deacetylase activity, suggesting that increased core histone acetylation results from a combination of these complementary modes of action. Our results indicate that exposure to MAA causes increased acetylation of core histones in several testis germ cell populations, including those in prophase of meiosis, a large proportion of which die rapidly following this treatment.