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a Dipartimento di Fisiologia Generale ed Ambientale, Università degli Studi di Bari, 70126 Bari, Italy
b Centre for Medically Assisted Procreation, Clinica S. Maria, 70124 Bari, Italy
ABSTRACT
Spermatogenesis and sperm maturation and storage are accompanied by significant movements of water, and multiple aquaporin transmembrane water channels (AQPs) have been recognized in the male reproductive tract. Nevertheless, the involvement of aquaporins in male reproductive physiology is mostly unknown. Here the expression and localization of AQP8 in rat spermatogenesis is defined and compared to that of AQP7, another aquaporin expressed in male germ cells. AQP8 mRNA was found in testis but not in epididymis, whereas the AQP7 transcript was present in both locations. By immunoblotting, the AQP8 protein was detected as a 25-kDa band and a 32- to 40-kDa diffuse component corresponding to the core and glycosylated protein, respectively. Membrane fractionation revealed AQP8 both in microsomal and plasma membrane-enriched fractions of rat testis while no apparent bands were detected in epididymis. AQP7 appeared as a 23- to 24-kDa band and was found both in testis and epididymis. By immunofluorescence, AQP8 labeling was found intracellularly as well as over the plasma membrane of germ cells throughout spermatogenesis. AQP7 was present in spermatids and spermatozoa and was predominant over the plasma membrane. AQP8 may be involved in the cytoplasmic condensation occurring during differentiation of spermatids into spermatozoa and in the generation of seminiferous tubule fluid.
epididymis, gamete biology, sperm, spermatogenesis, testis
INTRODUCTION
Fluid homeostasis is a critical process in the male reproductive tract [1, 2]. Sertoli cells are known to secrete fluid to form a fluid-filled tubular lumen [3] and water loss from germ cells has been suggested to contribute to the seminiferous tubule fluid [4]. Luminal fluid serves as a vehicle for sperm transportation and contributes to sperm maturation [5]. Because the seminiferous fluid of many mammalian species is hyperosmotic [6], it was postulated that osmotically driven cell water extrusion (70% reduction in cell volume) is one of the major mechanisms by which round spermatids progress to elongated spermatids during spermiogenesis [4, 7]. The fluid secreted by seminiferous tubules is significantly rearranged during its passage along the reproductive tract by secretion of nutrients and reabsorption of solutes and water. Efferent ducts are responsible for absorbing 90% of the seminiferous fluid [8]. Additional fluid is reabsorbed along the epididymis leading to sperm concentration and storage [9]. Less known is the mechanism of the fluid transepithelial movement taking place in the vas deferens, although the maintenance of an adequate luminal environment is required for further maturation and survival of the male gamete [10]. Fluids rich in nutrients are secreted by seminal vesicles and prostate and are needed for sperm to survive and fertilize eggs. The water permeability of human spermatozoa is among the highest reported for mammalian cells [11]. Moreover, it has been proven that the osmotic balance regulates cell fusion during mating in Saccharomyces cerevisiae, an observation suggesting that osmotically driven water and/or solute membrane transport through the gamete plasma membrane may be critical for sperm-egg fusion [12].
While understanding of the physiology of spermatogenesis and sperm maturation, storage, and survival is steadily increasing, the mechanisms of fluid secretion and reabsorption in the male reproductive tract remain largely undefined. Recognition of multiple members of the aquaporin family of transmembrane water channels (AQPs) [13] in secretory and absorptive male reproductive segments suggests roles for these proteins in male reproductive physiology. AQP1 is localized to the plasma membrane of nonciliated efferent duct cells and on the basolateral membrane of epithelial cells in the ampulla of the vas deferens, seminal vesicles, and prostate [14]. AQP2 is constitutively expressed in the apical membrane of the ampulla of the vas deferens [15], AQP7 is expressed in spermiogenesis [16, 17], AQP9 is found in the Leydig cells [18] and ciliated epithelial cells of the epididymal duct [19]. The mRNAs of AQP8 and AQP10 have been detected in the seminiferous tubules [20] and in the Sertoli cells [21], respectively. While appealing physiological roles have already been hypothesized for AQP1, AQP2, and AQP7 in the male reproductive tract [14, 15, 17] the functional significance of AQP8, AQP9, and AQP10 remains to be elucidated.
The AQP8 cDNA has been cloned recently from rat testis [20], rat pancreas and liver [22], mouse colon, placenta, liver, heart, submandibular glands [23], and human testis [24]. We recently cloned, characterized the structural organization, and mapped the mouse Aqp8 gene [25]. We also performed the chromosomal mapping of the human AQP8 gene [26]. Marked genomic and evolutionary differences exist between AQP8 and other known mammalian aquaporins [24–26]. This may confer unknown distinctive biophysical and regulatory properties to AQP8. No information is available regarding AQP8 at a protein level.
This study was undertaken to characterize the AQP8 polypeptide and define its localization in the rat male reproductive tract. The expression of AQP8 was compared to that of AQP7, an aquaporin also expressed in male germ cells. Potential physiological roles for AQP8 are hypothesized.
MATERIALS AND METHODS
Reverse Transcription-Polymerase Chain Reactionand RNase Protection Assays
Reverse transcription-polymerase chain reaction (RT-PCR) analysis was carried out using the GeneAmp RNA PCR core kit (Perkin-Elmer, Branchburg, NJ). Briefly, total RNAs of testis, efferent ducts, epididymis, vas deferens, seminal vesicles, prostate, spleen, and liver were isolated from Wistar rats or mouse testis using TRIzol reagent (Life Technologies, Gaithersburg, MD) and reverse transcribed. The resulting first-strand cDNA was employed to amplify a 732-base pair (bp) fragment of the rat and mouse AQP8 coding region using the primers RSA8-start (5'-CGGGATCCATGGCTGACAGTTACCAT-3') and RSA8-stop (5'-CGGAATTCACCTCGACTTTAGAAT-3'), or mAQP8-29 (5'-AGGTGAAGACCAGCATGGCTGG-3') and mAQP8-9 (5'-AGGCGGGTTTTCTCATCTCC-3'), respectively. A similar procedure was used for the RT-PCR analysis of AQP7 using the specific primers RSA7-start (5'-ATGGCCGGTTCTGTGCTG-3') and RSA7-stop (5'-TCTAAGAACCCTGTGGTGG-3'). RT-PCR reactions were normalized using a pair of rat ß-actin primers, ACT-forward (5'-GTTTGAGACCTTCAACACCC-3') and ACT-reverse (5'-CCAATGGTGATGACCTGGCC-3'). For the RNase protection studies, a 199-bp fragment of rat AQP8 cDNA was amplified by PCR using the mAQP8-10 (5'-TGGCTCATGGGCTGGCCTTG-3') and rAQP8-1 (5'-CTTTCCTCTGGACTCACCAC-3') primers and cloned into the EcoRI/EcoRI site of the pCR2.1 vector (Invitrogen, San Diego, CA) by leading to the pRPr8 plasmid. pRPr8 was then linearized with HindIII and used as a template for in vitro transcription of an 
-32P-labeled antisense mRNA probe synthesized from T7 promoter using the MAXIscript kit (Ambion, Austin, TX). The RNase protection assays were performed with 10 µg of rat tissues listed above by using the RPAIII kit (Ambion) as we previously described [25].
Sodium Dodecyl Sulfate-PAGE and Immunoblotting
Homogenates of rat male reproductive segments were prepared by a modification of a protocol previously reported by Brown et al. [14]. Tissues were cut into small pieces that were dispersed in ice-cold PBS and homogenized by 50 strokes in a glass/glass Dounce homogenizer. The homogenate was centrifuged at 800 x g for 15 min to remove nuclei and cell debris. The clear supernatant was then used for the Western blot studies. For the preparation of the plasma membrane- and microsomal membrane-enriched fractions, tissues were suspended in ice-cold homogenizing buffer (0.25 M sucrose, 10 mM Tris-HCl, pH 7.5) with protease inhibitors (1 mM PMSF, 1 µg/ml leupeptin, and 1 µg/ml pepstatin), homogenized by 35 strokes in a Potter A and centrifuged at 800 x g for 10 min at 4°C to remove nuclei and cell debris. The resulting supernatant was recovered and centrifuged at 17 000 x g for 45 min at 4°C (plasma membrane-enriched fraction). For the isolation of the microsomal membrane-enriched fraction (200 000 x g fraction), the 17 000 x g supernatant was centrifuged at 200 000 x g for 1 h at 4°C and the resulting pellet solubilized in Laemmli sample buffer before being submitted to Western blot analyses. For the immunoblotting experiments, samples were denatured at 90°C for 4 min and resolved on a 13% polyacrylamide gel (60 Μg/well). Separated proteins were electrotransferred onto Immobilon-P membranes (Millipore, Bedford, MA). After transfer, the membrane was blocked with Blotto (20 mM Tris-HCl, pH 7.5, 0.15 M NaCl, 1% Triton X-100) containing 5% nonfat dry milk (blocking buffer) for 1 h and then incubated with rabbit anti-ratAQP8 immune serum (Alpha Diagnostic International, San Antonio, TX) diluted 1:5000 in blocking buffer. In some control experiments blots were incubated with a rabbit anti-human AQP1 immune serum (1:2000). After washing (4 x 15 min) in blocking buffer, the membrane was incubated for 1 h with peroxidase-conjugated goat anti-rabbit IgG antibody (Sigma Chemical Co., St. Louis, MO). After washing (2 x 10 min) in blocking buffer and in Blotto (3 x 10 min), blots were revealed for peroxidase activity by enhanced chemiluminescence (ECL Plus kit; Amersham, Buckinghamshire, U.K.).
Deglycosylation
For N-glycosidase F (PNGase F) digestion, 60 µg of the testis plasma membrane-enriched fractions were incubated at room temperature for 6 h in the presence of 2 U ofN-glycosidase F (Boehringer Mannheim, Mannheim, Germany). The enzymatic reaction was stopped by adding one volume of 2x Laemmli buffer and boiling the resulting mixture at 90°C for 4 min. Samples were then analyzed by immunoblotting.
Immunocytochemistry
Adult male Wistar rats were decapitated after ether anesthesia, and the tissues from the reproductive tract were isolated and fixed in P.L.P. fixative (containing 2% paraformaldehyde, 10 mM sodium periodate, and 75 mM lysine, in sodium phosphate buffer, pH 7.4) and then washed three times in PBS for 10 min. The tissues were incubated in PBS containing 30% sucrose. Cryostat sections (5 µm) were prepared and placed on silanized microscope slides. Sections were kept in PBS for 10 min. After blocking in 0.1% gelatin in PBS for 15 min, cells were incubated for 2 h at room temperature with rabbit anti-rat AQP8 affinity-purified antibodies (Alpha Diagnostic International) or with the rabbit anti-ratAQP7 affinity-purified antibodies (kindly provided by Dr. A. Frigeri), both at a concentration of 100 ng/µl. Control experiments were performed using antibodies preadsorbed with an excess of the immunizing peptide. The sections were then washed 3 x 5 min with 0.1% gelatin in PBS and incubated for 60 min with a 1:200 dilution of fluorescein isothiocyanate-coupled goat anti-rabbit antibodies (Sigma), followed by washing once in high salt PBS (PBS supplemented with 2.7% NaCl) and twice in regular PBS for 5 min. The slides were then mounted in mounting medium (50% glycerol in 0.2 M Tris-HCl, pH 8.0, containing 2.5% n-propyl-gallate to retard quenching of the fluorescence). The samples were examined with a Leica DMRXA photomicroscope equipped for epifluorescence, and digital images were obtained with a cooled charge-coupled device camera (Princeton Instruments, Princeton, NJ).
RESULTS
AQP8 and AQP7 mRNA Expression in Rat Seminiferous Tubules and Epididymis
RT-PCR was performed to define the expression of AQP8 and AQP7 mRNA in the rat male reproductive tract. Abundant AQP8 transcript was found in testis (Fig. 1) as expected based on previous reports [20]. In contrast, no AQP8 expression was found in efferent ducts, epididymis, vas deferens, seminal vesicles, and prostate (Fig. 1). Because spleen is known not to express AQP8 [22], total RNA from rat spleen was employed as negative control. By RT-PCR, the AQP8 mRNA was also detected in mouse testis (Fig. 1). The pattern of expression of AQP8 in the rat male reproductive tract was confirmed by RNase protection analysis (data not shown). The AQP7 transcript was strongly detected in testis and epididymis (Fig. 1). Control RT-PCR reactions to discard the possibility of a genomic amplification of AQP8 and AQP7 in the rat and mouse total RNAs were performed (data not shown).
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Western Blotting Characterization of AQP8 and AQP7in Rat Testis and Epididymis
A Western blotting characterization of the AQP8 protein in testis was performed with polyclonal rabbit antibodies developed against a 16-amino acid peptide corresponding to the carboxy terminus of the predicted rat AQP8 polypeptide. By Western blotting of a rat testis homogenate, the AQP8 antiserum recognized a strong 25-kDa discrete band and a diffuse band of 32–40 kDa corresponding to the core AQP8 protein and the N-glycosylated form of AQP8 (glyAQP8), respectively (Fig. 2A). Consistent with a glycosylation of the AQP8 protein, treatment of the testis homogenate with PNGase F resulted in the disappearance of the 32- to 40-kDa component and a relative increase in the intensity of the 25-kDa band (Fig. 2C). The 25-kDa size of AQP8 in testis was confirmed in other immunoblotting experiments including a rat kidney membrane fraction containing the AQP1 core protein whose size is known to be of 28 kDa (Fig. 2E). An equivalent AQP8 pattern was obtained with cell homogenates prepared from mouse testis (data not shown). No immunoreactivity was noted with the antiserum depleted of the anti-AQP8 antibodies by preadsorption with a 20:1 molar excess of the immunizing peptide (Fig. 2D) and the preimmune serum as well (data not shown). Consistent with the results of the RT-PCR and RNase studies, no apparent AQP8 bands were recognized in the homogenates from epididymis (Fig. 2B) and vas deferens, seminal vesicles, and prostate (data not shown).
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Parallel Western blot experiments were carried out to evaluate the expression of the AQP7 protein in the rat male reproductive tract. In line with the above mRNA studies and the results of a previous report [27], both in rat testis and epididymis homogenates, the AQP7 antiserum strongly reacted with a 23- to 24-kDa band corresponding to the AQP7 protein (Fig. 2, F and G). As expected based on the absence of N-glycosylation consensus sequences, no glycosylated forms of AQP7 protein were observed.
The membrane expression of the AQP8 and AQP7 proteins in testis was evaluated by immunoblotting of plasma membrane-enriched (17 000 x g) or microsomal (200 000 x g pellet depleted of the 17 000 x g pellet) fractions obtained by differential centrifugation. Interestingly, AQP8 was detected both in plasma and intracellular membranes (Fig. 3A), suggesting that this aquaporin may have a considerable intracellular expression in addition to its plasma membrane location. A different pattern was seen with anti-AQP7 testis immunoblots because AQP7 was predominant in the plasma membrane fraction while it was significantly less represented in the intracellular membrane fraction (Fig. 3B).
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Immunofluorescent Localization of AQP8 and AQP7in Testis and Epididymis
By immunofluorescence using affinity-purified anti-AQP8 antibodies we found AQP8 strongly expressed in adult rat seminiferous tubules throughout spermatogenesis (Fig. 4, a–c). These data are consistent with previous in situ hybridization studies [20]. In line with the above Western blot studies, staining was seen both within the cytoplasmic compartment and the plasma membrane and was particularly strong in the deep portion of the adluminal compartment. No apparent staining was found over the Sertoli cells as well as the Leydig and the peritubular cells.
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Considerable AQP7 fluorescence was seen throughout spermiogenesis while no labeling of spermatogonia and spermatocytes (Fig. 4, a'–c') occurred. AQP7 staining was restricted to the cytoplasmic mass of differentiating spermatids and became steadily more intense over the plasma membrane of elongated spermatids at the luminal rim of the seminiferous epithelium prior to spermiation (Fig. 4b', inset). This pattern confirmed previous immunohistochemical studies [17]. The plasma membrane and cytoplasmic portion of the residual bodies remaining in the adluminal compartment were clearly immunoreactive (Fig. 4c', arrows). After spermiation, the AQP7 labeling was seen on the plasma membrane of the cytoplasmic droplet (Fig. 4c', upper inset) and anterior tail domain of testicular spermatozoa (Fig. 4c', lower inset).
Anti-AQP8 immunostaining was very weak in the epididymis where it was localized at the level of the cytoplasmic droplet of spermatozoa (Fig. 5, a–c). A very faint intracellular labeling of the epididymal duct epithelial cells was occasionally seen that, however, might not be specific. In contrast to AQP8, an intense AQP7 immunolabeling over the plasma membrane of the cytoplasmic droplet and anterior tail domain of epididymal spermatozoa was observed (Fig. 5, a'–c'). No AQP7 immunoreactivity was noted over the head and posterior tail membranes, an observation consistent with the highly organized membrane domains of mammalian spermatozoa [28].
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No staining of seminiferous tubules, mature spermatozoa, epididymal epithelium, and smooth muscle cells surrounding the epididymal duct was seen with the rabbit preimmune sera or preadsorbed affinity-purified AQP7 antibodies (data not shown).
DISCUSSION
This study provides a first biochemical characterization of the AQP8 protein and establishes the cellular and subcellular localization of AQP8 in rat testis. Moreover, the immunofluorescent localization of AQP7 in epididymal spermatozoa is also defined.
While the existence of a glycosylated form of AQP8 was predicted based on the presence of a typical N-glycosylation consensus sequence located at Asn141 in the extracellular loop C [20, 22], the 25-kDa size of AQP8 observed by Western blot of rat testis does not correspond to the 28.06-kDa molecular mass (263 amino acids) predicted based on the length of the rat AQP8 open reading frame [20, 22]. The reduced size of AQP8 may have three possible explanations: 1) alternate use of translational initiation sites, 2) increased mobility of the AQP8 protein, and 3) posttranslational modification. In the first case, because rat AQP8 cDNA has four potential translation-initiating sites [20], it is possible, at least in testis, that AQP8 mRNA is translated starting from the fourth downstream site by leading to a polypeptide of 241 residues having a molecular mass of 25.59 kDa. This possibility is corroborated by the fact that the fourth potential translation-initiating site is the one with the best Kozak consensus. Thus, like AQP4 [29], AQP8 might exist in multiple forms owing to the alternate use of translational initiation sites depending on the expression of AQP8 in specific tissues. The 25-kDa size was also observed with AQP8 immunoblots of mouse testis (data not shown). AQP8 biosynthesis may therefore be characterized by a complex pattern of regulation, as also suggested by the lack of a TATA box, the presence of three mRNA start sites, and the presence of an unusual splicing site observed in the mouse Aqp8 gene [25]. However, the possibility that an increased mobility of AQP8 during its SDS-PAGE resolution might be possible because this feature is often expressed by hydrophobic integral membrane proteins [30]. Similarly to AQP8, the 23- to 24-kDa molecular mass exhibited by AQP7 resulted considerably smaller than the predicted 28.881 kDa [16]. A posttranslational shortening of the AQP8 protein represents a modification that has never been reported among aquaporins. Future studies aiming to define the N-terminal amino acid sequences of the AQP8 and AQP7 polypeptides should answer these questions.
The subcellular localization of AQP8 resembles rather than that of AQP2 in the renal collecting duct [31] that of AQP6, which has been found to be resident exclusively in intracellular vesicles where its anion/water channel activities are regulated by pH [32]. This suggests the possibility for recycling of AQP8 between intracellular vesicles and the plasma membrane and that this redistribution might be under hormonal control. Aquaporins under hormonal regulation are already known [31, 33, 34]. Hence, a possible regulation of AQP8 in the male reproductive tract by LH (indirectly by testosterone) or FSH, two hormones known for controlling spermatogenesis and its initiation, deserves investigation.
The presence of AQP8 in all differentiating stages of germ cells suggests roles for this aquaporin in spermatogenesis. As elimination of water by germ cells may be one way to make the phagocytosis of a smaller volume of cytoplasm manageable, a role for AQP8 in concert with AQP7 to produce the cytoplasmic condensation associated with the transformation of round spermatids in spermatozoa is conceivable. In rat spermatids, the apparent redundancy of aquaporins may relate to the different functional properties expressed by AQP8 and AQP7. In fact, AQP8 has been shown to transport water [20, 22], whereas rat AQP7 has been found to be permeable to small neutral solutes such as glycerol and urea in addition to water [16]. Moreover, AQP8 might also possess other biophysical features yet unknown as suggested by its distinctive phylogeny when compared to other AQPs [24–26]. Because the seminiferous tubule fluid is thought not to be solely of Sertoli cell origin but also to be generated by water lost from germ cells [2, 35], AQP8 might contribute in generating the fluid serving as a vehicle for sperm transportation and maturation. This possibility is corroborated by the appearance of AQP8 (and not AQP7) in the testis in the first 2 wk of postnatal life (unpublished results), which suggests a possible involvement of AQP8 in the secretion of water to form a fluid-filled tubular lumen, one of the first morphological events announcing the beginning of spermatogenesis.
Absence of AQP8 in male reproductive duct epithelial cells, seminal vesicles, and prostate indicates that AQP8 is not involved in the fluid movement occurring during sperm concentration and maturation. In contrast, the intense expression of AQP7 in epididymal spermatozoa suggests that besides its function in spermiogenesis [17] this mercury-resistant aquaporin may have roles in sperm maturation and storage and be responsible for the high water permeability characterizing mammalian spermatozoa [36].
In summary, this work provides a Western blot characterization of the AQP8 protein and describes its localization in the rat male reproductive tract. The localization of AQP7 in testicular and epididymal spermatozoa is also defined. Although further work will be required to define fully the biophysical, biochemical, and regulatory features of AQP8, this study represents a basis for future studies aimed to define the physiological significance of AQP8. Useful information may also derive from the phenotypic analysis of transgenic Aqp8 knockout mice we are currently generating.
ACKNOWLEDGMENTS
We thank Prof. Soren Nielsen and Dr. Marie Louise Elkjaer for valuable discussions, Dr. Antonio Frigeri for providing the purified anti-AQP7 antibodies and Drs. Patrizia Gena, Chiara Di Nunno, Giuseppe Procino, and Elena Sarcina for their helpful contributions.
FOOTNOTES
First decision: 27 November 2000.
1 Support of the European Community (EU-TMR network grant no. ERB 4061 PL 97-0406) is gratefully acknowledged. 
2 Correspondence: Giuseppe Calamita, Dipartimento di Fisiologia Generale ed Ambientale, Università degli Studi di Bari, via Amendola 165/A, 70126 Bari, Italy. FAX: 39 0805443388; calamita{at}biologia.uniba.it
Accepted: January 16, 2001.
Received: October 18, 2000.
REFERENCES
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|
|
 |
|
  C.-H. Yeung, C. Callies, A. Rojek, S. Nielsen, and T. G. Cooper Aquaporin Isoforms Involved in Physiological Volume Regulation of Murine Spermatozoa Biol Reprod, February 1, 2009; 80(2): 350 - 357. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C Callies, T G Cooper, and C H Yeung Channels for water efflux and influx involved in volume regulation of murine spermatozoa Reproduction, October 1, 2008; 136(4): 401 - 410. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
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|
|
|
|
 |
|
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|
|
|
|
 |
|
 H.-F. Huang, R.-H. He, C.-C. Sun, Y. Zhang, Q.-X. Meng, and Y.-Y. Ma Function of aquaporins in female and male reproductive systems Hum. Reprod. Update, November 1, 2006; 12(6): 785 - 795. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Yang, D. Zhao, E. Solenov, and A. S. Verkman Evidence from knockout mice against physiologically significant aquaporin 8-facilitated ammonia transport Am J Physiol Cell Physiol, September 1, 2006; 291(3): C417 - C423. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Da Silva, C. Silberstein, V. Beaulieu, C. Pietrement, A. N. Van Hoek, D. Brown, and S. Breton Postnatal Expression of Aquaporins in Epithelial Cells of the Rat Epididymis Biol Reprod, February 1, 2006; 74(2): 427 - 438. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 U. Laforenza, E. Cova, G. Gastaldi, S. Tritto, M. Grazioli, N. F. LaRusso, P. L. Splinter, P. D'Adamo, M. Tosco, and U. Ventura Aquaporin-8 Is Involved in Water Transport in Isolated Superficial Colonocytes from Rat Proximal Colon J. Nutr., October 1, 2005; 135(10): 2329 - 2336. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
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|
|
|
|
|
|
M. Albrizio, A. C. Guaricci, F. Maritato, R. L. Sciorsci, G. Mari, G. Calamita, G. M. Lacalandra, G. G. Aiudi, R. Minoia, M. E. Dell'Aquila, et al. Expression and subcellular localization of the {micro}-opioid receptor in equine spermatozoa: evidence for its functional role Reproduction, January 1, 2005; 129(1): 39 - 49. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. O. Hernandez-Gonzalez, D. Mornet, A. Rendon, and D. Martinez-Rojas Absence of Dp71 in mdx3cv mouse spermatozoa alters flagellar morphology and the distribution of ion channels and nNOS J. Cell Sci., January 1, 2005; 118(1): 137 - 145. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Rossato, C. Galeazzi, M. Ferigo, and C. Foresta Antisperm antibodies modify plasma membrane functional integrity and inhibit osmosensitive calcium influx in human sperm Hum. Reprod., August 1, 2004; 19(8): 1816 - 1820. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J.-H. Lignot, C. P. Cutler, N. Hazon, and G. Cramb Immunolocalisation of aquaporin 3 in the gill and the gastrointestinal tract of the European eel Anguilla anguilla (L.) J. Exp. Biol., September 1, 2002; 205(17): 2653 - 2663. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. H. Badran and L. S. Hermo Expression and Regulation of Aquaporins 1, 8, and 9 in the Testis, Efferent Ducts, and Epididymis of Adult Rats and During Postnatal Development J Androl, May 1, 2002; 23(3): 358 - 373. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
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