Water Permeability of an Ovarian Antral Follicle Is Predominantly Transcellular and Mediated by Aquaporins

Abstract

Ovarian folliculogenesis is characterized, in part, by the formation and expansion of the fluid-filled antrum. Development of this cavity requires water influx, which may occur by transcellular or pericellular transport mechanisms. To assess the contribution of these mechanisms to the water permeability of an antral follicle, the rate of (3)H(2)O and (14)C-inulin (a complex sugar restricted to the extracellular compartment) uptake into isolated follicles was determined. The rate of H(2)O movement was 3.5-fold greater than that of inulin, suggesting that water enters a follicle primarily by transcellular pathways. Preincubation of the follicles with 50 microM HgCl(2) [a nonspecific aquaporin (Aqp) inhibitor] decreased H(2)O movement to levels seen with inulin, indicating that transcellular water movement is mediated through Aqp. To demonstrate the functional presence of Aqp in granulosa cells, we show that swelling in response to a hypotonic insult is attenuated by preincubation with 50 microM HgCl(2). Flow cytometry demonstrated the presence of Aqps-7, -8, and -9, thus identifying candidate Aqp potentially mediating water movement into antral follicles. These results suggest that water permeability of antral follicles occurs primarily through transcellular mechanisms, which may be mediated by Aqps -7, -8, and/or -9 in granulosa cells.

The potential role of caveolin‐1 in inhibition of aquaporins during the AVD

“…Thymocytes were harvested as described above and either used immediately or cultured (5×10 6 cells/ml) in serum‐free medium for 24 h (37°C and 5% CO 2 ) to induce apoptosis. Thymocytes were then labelled with anti‐AQP2–AQP7 Abs (Alpha Diagnostic, San Antonio, TX, U.S.A.) or anti‐AQP8 and anti‐AQP9 Abs [produced by our laboratory (McConnell et al, 2002)] using a Cytofix/Cytoperm kit (PharMingen, Franklin Lakes, NJ, U.S.A.). Briefly, cells were fixed and permeabilized in 150 μl of Cytofix/Cytoperm solution for 20 min on ice and washed twice with 200 μl of 1×Perm/Wash solution (proprietary solution provided with the Cytofix/Cytoperm kit).…”

“…Sections were rehydrated in 1×PBS for 10 min and then incubated in blocking solution (10%, v/v, normal goat serum) for 20 min before incubation with primary Abs. Rabbit anti‐rat AQP8 and AQP9 [Abs produced in our laboratory as described previously (McConnell et al, 2002)] and mouse anti‐rat TNF‐R1 (Santa Cruz Biotechnology, Santa Cruz, CA, U.S.A.) were diluted to 1:100 with PBS, applied to slides and allowed to incubate for 12 h at room temperature (25°C) in a humidified chamber. The slides were then washed in PBS (three times for 10 min each) and Texas Red‐conjugated goat anti‐rabbit IgG secondary Ab (1:500 dilution; Sigma—Aldrich, St. Louis, MO, U.S.A.) and FITC‐labelled goat anti‐mouse IgG secondary Ab (1/500 dilution; Sigma—Aldrich) were added to sections and allowed to incubate at room temperature for 2 h. The slides were again washed in PBS (three times for 10 min each), coverslips were mounted and the slides were stored at 4°C until analysis.…”

Increased female fertility in aquaporin 8‐deficient mice

“…A previous study reported expression of AQP7, AQP8, and AQP9 in rat follicular granulosa cells and indicated that water permeability of antral follicles from rat ovary was predominantly transcellular and mediated by AQPs (6). However, the physiological importance of the AQPs in female fertility is unknown.…”

“…Mammalian aquaporins (AQPs) are a family of water‐transporting membrane proteins selectively expressed in cell types of various organs where they may play important physiological functions (1, 2). Among the 13 members of mammalian AQP family, AQP8 has been shown to express extensively in the reproductive system (in testis and sperm in the male reproductive system and in ovary, oviduct, uterus, placenta, amnion, chorion, and cervix in the female reproductive system) (3–10). Yang et al (11) studied the role of AQP8 in male reproductive function in a knockout mouse model.…”

The expression profile of <i>AQP1</i>, <i>AQP5</i> and <i>AQP9</i> in granulosa and theca cells of porcine ovarian follicles during oestrous cycle and early pregnancy

“…The Aqp9 expression was previously found in rat granulosa cells (McConnell et al, 2002). Moreover, the fluctuations of Aqp9 transcripts during growth and oocyte maturation in rats were observed (Ford et al, 2000).…”