Cytogenetic and immunofluorescence analysis of benzo[a]pyrene-DNA adduct formation and chromosome damage in larval brain neuroblasts of Drosophila melanogaster.

Recently we have evaluated the relationship between benzo[a]-pyrene(BaP)-DNA adducts, determined by 32P-postlabelling, and clone frequencies in the somatic mutation and recombination test (SMART) in Drosophila melanogaster. Following that study we proceeded to characterise further the mechanism of induction of genetic damage in vivo by BaP in Drosophila by cytogenetic analysis of larval brain neuroblasts. Third stage larvae were treated with 4 and 10 mM BaP for 24, 48 or 72 h. In all cases, the larvae were killed 72 h after the beginning of treatment, entailing 48, 24 or 0 h post-treatment recovery in BaP-free medium, respectively. At the end of the treatment the following data were collected: (i) the types and levels of chromosome aberrations in neuroblast metaphase and anaphase nuclei; (ii) the distribution and level of BaP-DNA adducts, revealed by indirect immunofluorescence in neuroblast nuclei using an anti-(BaP-DNA) antibody. The results indicate that BaP induces chromosome breaks, deletions and exchanges in this system. In particular, chromosome exchanges decrease as the post-treatment recovery time increases, and the dynamics of breaks and deletions appear to be inversely related to those of the exchanges. This suggests that exchanges may require few preconditions to occur and are thus expressed soon after treatment. Chromosome breaks and deletions could require multiple single events before the actual damage is expressed (even some cell divisions away from the end of treatment). The immunofluorescence analysis suggests that BaP-DNA adducts are more abundant in the heterochromatin of the neuroblast nuclei.