EVALUATION OF SISTER-CHROMATID EXCHANGES IN MOUSE SPERMATOGONIA - A COMPARISON BETWEEN THE CLASSICAL FLUORESCENCE PLUS GIEMSA STAINING AND AN IMMUNOCYTOCHEMICAL APPROACH

Sister-chromatid exchanges (SCE) were analyzed in mouse spermatogonia using two different protocols for bromodeoxyuridine (BrdU) exposure an d detection. With the classical approach, based on subcutaneous implan tation of agar-coated BrdU tablets and fluorescence plus Giemsa (FPG) staining a satisfactory differentiation of spermatogonial metaphases w as obtained with 50 or 25 mg BrdU per mouse (two or one tablets respec tively). Alternatively, the immunodetection of BrdU was carried out af ter exposure to a very low BrdU concentration (three injections i.p., 3 mg/kg b.w. each, at 5-h intervals), and after exposure to one BrdU t ablet; SCE frequencies evaluated in this way were lower than those fou nd after classical FPG staining, even when the mice were exposed to th e same BrdU concentration (one tablet, 25 mg BrdU). We concluded that the two methodologies may have different sensitivities with respect to SCE detection. In addition, when the effect of a treatment with mitom ycin C was tested (1 mg/kg b.w., at time intervals ranging from 24 h t o 5 days), no sister-chromatid differentiation was obtained with the m ultiple injection protocol, or with one BrdU tablet. By contrast, with two BrdU tablets and FPG, well differentiated metaphases were found a t any time interval tested after MMC treatment, and the peak frequency of SCE (3.4 times the baseline) was observed at 55 h after treatment, as expected on the basis of cell cycle duration in spermatogonia. In summary, even though the use of medium-low concentrations of BrdU was successful in untreated animals, these protocols appeared inadequate t o detect SCE induction by MMC. It is possible that, in the presence of cell cycle delay induced by the treatment, interferences with the rat e of BrdU uptake produce an unsatisfactory differentiation of sister c hromatids.