Characterization of nucleoside transport systems in cultured rat epididymal epithelium

Abstract

The nucleoside transport systems in cultured epididymal epithelium were characterized and found to be similar between the proximal (caput and corpus) and distal (cauda) regions of the epididymis. Functional studies revealed that 70% of the total nucleoside uptake was Na(+) dependent, while 30% was Na(+) independent. The Na(+)-independent nucleoside transport was mediated by both the equilibrative nitrobenzylthioinosine (NBMPR)-sensitive system (40%) and the NBMPR-insensitive system (60%), which was supported by a biphasic dose response to NBMPR inhibition. The Na(+)-dependent [(3)H]uridine uptake was selectively inhibited 80% by purine nucleosides, indicating that the purine nucleoside-selective N1 system is predominant. Since Na(+)-dependent [(3)H]guanosine uptake was inhibited by thymidine by 20% and Na(+)-dependent [(3)H]thymidine uptake was broadly inhibited by purine and pyrimidine nucleosides, this suggested the presence of the broadly selective N3 system accounting for 20% of Na(+)-dependent nucleoside uptake. Results of RT-PCR confirmed the presence of mRNA for equilibrative nucleoside transporter (ENT) 1, ENT2, and concentrative nucleoside transporter (CNT) 2 and the absence of CNT1. It is suggested that the nucleoside transporters in epididymis may be important for sperm maturation by regulating the extracellular concentration of adenosine in epididymal plasma.

Nucleoside Transport at the Blood-Testis Barrier Studied with Primary-Cultured Sertoli Cells

“…Time-and temperature-dependent uridine uptake was observed, and an excess of unlabeled uridine and NBMPR inhibited this uptake (data not shown), suggesting that the nucleoside transporters take up essential nucleosides for spermatogenesis. In primary-cultured rat epididymal epithelial cells, ENT1, ENT2, and CNT2 are expressed and sodium-dependent transporter CNT2 mainly mediates uridine uptake (Leung et al, 2001). Thus, Sertoli cells and epididymal cells appear to have in common CNT2, but the contribution of other nucleoside transporters may be different between these two types of cells, although both of these cells should contribute differentiation and maturation of spermatogenic cells.…”

“…Adenosine is related to the acquisition of motility of sperm (Aitken et al, 1986;Vijayaraghavan and Hoskins, 1986), and nucleoside transport systems were characterized in rat epididymis epithelial cells (Leung et al, 2001). However, there is no information about nucleoside transport systems at the BTB, which is formed mainly by Sertoli cells.…”

Basolateral Uptake of Nucleosides by Sertoli Cells Is Mediated Primarily by Equilibrative Nucleoside Transporter 1

“…Studies investigating epididymal drug transport are limited, but one study revealed that ENT1, ENT2, and CNT2 are present in the epididymis (Leung et al, 2001). Without knowing the localization of these transporters, it is difficult to speculate on their potential impact on drug transport.…”

Carnitine/organic cation transporter OCTN2-mediated transport of carnitine in primary-cultured epididymal epithelial cells

“…Epididymal epithelial cells were isolated from 5-week-old Sprague-Dawley rats (Saitama Experimental Animal Supply Co. Ltd, Saitama, Japan) according to the method reported previously (Kierszenbaum et al 1981, Leung et al 2001. To minimize contamination of non-epithelial cells and inhibition of attachment of epithelial cells to culture dishes by spermatozoa, we used immature rats, which do not contain spermatozoa (Kierszenbaum et al 1981, Leung et al 2001.…”

“…To minimize contamination of non-epithelial cells and inhibition of attachment of epithelial cells to culture dishes by spermatozoa, we used immature rats, which do not contain spermatozoa (Kierszenbaum et al 1981, Leung et al 2001. Briefly, epididymides were dissected out and minced into small fragments.…”