Cell Tissue Res (2004) 318: 353–363 DOI 10.1007/s00441-004-0985-4 REGULAR ARTICLE Apichart Ngernsoungnern . Wattana Weerachatyanukul . Arpornrad Saewu . Siriporn Thitilertdecha . Prasert Sobhon . Prapee Sretarugsa Rat sperm AS-A: subcellular localization in testis and epididymis and surface distribution in epididymal sperm Received: 20 April 2004 / Accepted: 3 August 2004 / Published online: 30 September 2004 # Springer-Verlag 2004 Abstract In this study, we investigated the subcellular compartmentalization of arylsulfatase-A (AS-A) in the testis and epididymis as well as the surface distribution in rat epididymal sperm. Testicular AS-A was compartmen- talized specifically to the area underneath the outer acrosomal membrane of the acrosomal granule and to the dorsal aspect of the sperm acrosome. Epididymal AS- A was synthesized in the endoplasmic reticular (ER) network of principal cells and secreted into epididymal lumen as evident by its reactivity in the apical cytoplasm and vesicles therein underneath stereocilia. In clear cells, AS-A reactivity was found throughout the cytoplasmic machineries involved in endocytosis. Surface distribution of AS-A was initially detectable at the concave ridge as early as in sperm of the initial segment (IS). AS-A was additionally localized to the post-acrosomal region in caput (CP), corpus (CO) and cauda (CD) epididymal sperm. The expression levels of surface AS-A gradually increased during sperm transit from IS to CD epididymi- dis. These results favored the adsorption of AS-A from epididymal fluid onto the sperm surface, rather than shunting from the acrosome as a consequence of capac- itation-associated membrane priming. Keywords Sperm . Arylsulfatase-A . Testis . Epididymis . Sperm maturation . Rat (Wistar) Introduction In mammals, testicular spermatozoa lack progressive mobility and fertilizing capacity. They must transit along the complicated milieu of the epididymis and undergo a number of molecular modifications in order to acquire these physiological abilities (Yanagimachi 1994; Cooper 1998). Epididymal epithelium is highly active in both secretory and absorptive functions under the influence of androgens (Cooper et al. 1987; Hermo et al. 1994). While principal cells, the major cell types of the epididymal epithelium, serve the major task in protein biosynthesis and secretion, clear cells with all endocytotic machineries are responsible mostly for the uptake of luminal contents into the cells (Hermo and Robaire 2002). Moreover, selective protein expression and a differential secretory function of principal cells have also been documented in various parts of epididymis (Andonian and Hermo 1999). A number of intraluminal secretory products of principal cells have been characterized, some of which are hydro- lytic enzymes that play a role in sperm membrane modification (Cooper 1998; Toshimori 1998). With an increasing number of secretory proteins being character- ized in the epididymal lumen, the mechanism of how these proteins are transported onto the sperm surface has recently been a challenged issue for many investigators to elucidate the complicated regulatory process of epidid- ymal epithelium within this single convoluted tubular organ. AS-A (E.C. 3.1.6.8) has long been characterized biochemically in the lysosomes of somatic cells (Kelly et al. 1989; Fujii et al. 1992) and also in the mammalian sperm acrosome (Dudkiewicz 1984; Brandon et al. 1997). AS-A exerts its desulfation activity at the optimal pH of 5.5 on a small synthetic substrate, p-nitrocatecholsulfate (NCS), and on glycolipid substrates that are strongly expressed in mammalian male germ cells, sulfogalacto- sylceramide (SGC) and sulfogalactosylglycerolipid (SGG), in the presence of a detergent or saposin B (Mehl and Jatzkewitz 1968; Gadella et al. 1992). Transcriptional expression of AS-A can be detected This work was supported by Research Initiate Grant funded by Faculty of Science, Mahidol University to W.W. A. Ngernsoungnern . A. Saewu . S. Thitilertdecha Department of Anatomy, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand W. Weerachatyanukul (*) . P. Sobhon . P. Sretarugsa Department of Anatomy, Faculty of Science, Mahidol University, Rama 6 Rd., Payathai, Bangkok, 10400, Thailand e-mail: scwwy@mahidol.ac.th Fax: +66-2-3547168