Effects of ageing on spermatozoal chromatin and its sensitivity to in vivo and in vitro oxidative challenge in the Brown Norway rat

Abstract

The goals of our study were to examine chromatin packaging and integrity in spermatozoa taken from the caput and cauda epididymides of young (4-month-old) and old (21-month-old) Brown Norway rats and to assess whether spermatozoal sensitivity to oxidative treatments is altered with age. Oxidative treatments consisted of (i) in vivo oxidative challenge by systemic administration of the glutathione-depleting drug l-buthionine-[S,R]-sulphoximine (BSO) and (ii) in vitro oxidative challenge by incubating collected spermatozoa with hydrogen peroxide (H(2)O(2)). Chromatin parameters assessed included quantification of thiols, nuclear chromomycin A3 (CMA3) penetration, DNA breaks by TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling (TUNEL) and ease of DNA dissociation by acridine orange (AO) staining. In spermatozoa from older rats, we found decreases in thiols, CMA3 penetration and the percentage of cells that undergo DNA dissociation. Administration of BSO had oxidizing effects on the thiol groups. It also decreased CMA3 penetration and DNA dissociation and increased TUNEL staining. Furthermore, BSO treatment sensitized cauda epididymidis spermatozoa, from older animals, to H(2)O(2). Overall, we show that spermatozoa from older rats have altered chromatin packaging and integrity and that spermatozoa from the cauda epididymidis are more responsive to combined in vivo and in vitro oxidative challenge than spermatozoa from young rats.