V ATPase is a complex enzyme

V-ATPase is a complex enzyme that is composed of several subunits, which are assembled into two domains: trans-membrane (V0) and cytosolic (V1) domains [3,5]. Epididymal epithelial Griseofulvin synthesis of rats has been reported to contain V-ATPase which is distributed at the apical pole of the narrow and clear cells [2]. These cells are known to be involved in generating low epididymal fluid pH [6]. Some of V-ATPase subunits have more than one isoform [7]. As an example, four “a” isoform and two “b” isoform of V-ATPase subunit are present in the human genome [8,9]. Subunit a1 was expressed ubiquitously while subunit B1 was found in the epididymis and other tissues, while B2 was found to be ubiquitously expressed in the intracellular organelles [9]. The importance of V-ATPase to male reproduction has been demonstrated in mice in which lack of transcription factor, Foxil, a master regulator of V-ATPase in clear cells of epididymis affects male fertility due to inability of the sperm to move up the female genital tract to fertilize the oocyte [[10], [11], [12]]. In epididymal clear cells, V-ATPase function is under hormonal controls with hormone known to influence V-ATPase activity include angiotensin II (AngII). Ang II stimulates V-ATPase function via binding to Ang II type II receptors (AGTR2), which are found in the adjacent basal cells [13]. Once binding to AGTR2, nitric oxide (NO) is released from the basal cells which then diffuses out to stimulate proton secretion in clear cells via activation of cGMP pathway [14]. In vas deferens, the role of hormones in controlling V-ATPase expression and functional activity is largely unknown. Therefore, in this study, we hypothesized that testosterone affects V-ATPase expression and functional activity in vas deferens through which this hormone control fluid pH of this organ. Thus the objectives of this study were to identify the functional activity, expression level and distribution of V-ATPases in vas deferens under testosterone influence. The finding of this study could help to explain the mechanisms underlying testosterone-mediated effect on vas deferens fluid pH where dysregulation could adversely affect male fertility.
Materials and methods
Results
Discussion In this study, orchidectomy was performed in order to minimize the influence of endogenous testosterone on vas deferens morphology and functions. Our findings, which found that plasma testosterone level in rats decreased following orchidectomy, was consistent with the report by previous study [19], which indicate that the main source of testosterone in males is the testes. In males, no other organs were able to compensate for the decrease in testosterone production by the testis, and this was in contrast to the female where elevation in estrogen levels following ovariectomy was due to compensatory hyperplasia of the adrenal gland [24]. Our findings further demonstrated that testosterone was able to induce secretion of fluid in vas deferens, as well as causing the fluid pH and HCO3− content to decrease. A report showed that testosterone was able to induce fluid and electrolyte secretion in androgen-sensitive male reproductive organs including seminal vesicles and prostate [25]. The findings from this study which found that vas deferens fluid secretion rate, pH and HCO3− content could be influence by testosterone further added information with regards to the role of this hormone in controlling vas deferens fluid parameters. In this study, in vivo perfusion technique was used, where changes in the secretion rate, pH and electrolyte content of vas deferens fluid were monitored in life animal model. Therefore, this technique ideally reflects changes in the internal milieu of vas deference fluid environment [20]. The findings from this study have shown that V-ATPase is a channel that is involve in testosterone-induced acidification of vas deferens fluid. A specific V-ATPase inhibitor, bafilomycin A1 was added to confirm the functional activity of V-ATPase where upon its administration, testosterone-induce decrease in the pH and HCO3− content of vas deferens fluid was reversed. The effect of bafilomycin confirms the role of V-ATPase in mediating these changes. Bafilomycin A1 is a macrolide antibiotic isolated from Streptomyces species and has been reported able to inhibit V-ATPase via binding to the V0 sector subunit c of V-ATPase complex which results in inhibition of H+ translocation, thus inducing accumulation of H+ in the cytoplasm of the cells [26]. However, bafilomycin A1 has no effect on vas deferens fluid secretion rate in rats which either possess endogenous testosterone or those treated with exogenous testosterone. These suggested that testosterone-induced fluid secretion in vas deferens could involve other protein transporter such as aquaporins, which is reported to be present in vas deferens [27]. The role of aquaporins in vas deferens fluid secretion need further elucidation. The finding that V-ATPase is involve in luminal fluid acidification is consistent with its role in causing low luminal fluid pH in many organs [28]. This effect is important since maintaining the low pH of vas deferens fluid is crucial for sperm survival as well as help to inhibit sperm activation [4]. Failure of vas deferens fluid acidification could result in poor sperm maturation and this can also cause premature motility of the sperm [6]. It is also reported that in vas deferens, V-ATPase contributes to approximately 80% of proton secretion into the lumen [29], implying that this channel is indeed essential for vas deferns fluid acidification.