Channel-Dependent Permeation of Water and Glycerol in Mouse Morulae1

Abstract

The cryosensitivity of mammalian embryos depends on the stage of development. Because permeability to water and cryoprotectants plays an important role in cryopreservation, it is plausible that the permeability is involved in the difference in the tolerance to cryopreservation among embryos at different developmental stages. In this study, we examined the permeability to water and glycerol of mouse oocytes and embryos, and tried to deduce the pathway for the movement of water and glycerol. The water permeability (L(P), microm min(-1) atm(-1)) of oocytes and four-cell embryos at 25 degrees C was low (0.63-0.70) and its Arrhenius activation energy (E(a), kcal/mol) was high (11.6-12.3), which implies that the water permeates through the plasma membrane by simple diffusion. On the other hand, the L(p) of morulae and blastocysts was quite high (3.6-4.5) and its E(a) was quite low (5.1-6.3), which implies that the water moves through water channels. Aquaporin inhibitors, phloretin and p-(chloromercuri) benzene-sulfonate, reduced the L(p) of morulae significantly but not that of oocytes. By immunocytochemical analysis, aquaporin 3, which transports not only water but also glycerol, was detected in the morulae but not in the oocytes. Accordingly, the glycerol permeability (P(GLY), x 10(-3) cm/min) of oocytes was also low (0.01) and its E(a) was remarkably high (41.6), whereas P(GLY) of morulae was quite high (4.63) and its E(a) was low (10.0). Aquaporin inhibitors reduced the P(GLY) of morulae significantly. In conclusion, water and glycerol appear to move across the plasma membrane mainly by simple diffusion in oocytes but by facilitated diffusion through water channel(s) including aquaporin 3 in morulae.

Developmental Ability of Vitrified Mouse Oocytes Expressing Water Channels

“…b The osmotically inactive volume of oocytes/embryos was assumed to be 0.15 because that of mature oocytes and 4-cell embryos was 0.15 in a previous study [5], and that of 2-cell embryos measured for the present study was 0.15 ± 0.04 (obtained from 9 embryos). into blastocysts.…”

“…[ 5,6]. Since the glycerol-permeability was quite low, mouse oocytes could not survive after vitrification with a glycerol-based solution.…”

“…We also showed in the mouse that permeability to water and cryoprotectants was low in oocytes and early embryos, whereas permeability to water, glycerol and ethylene glycol was high in morulae [4], and that aquaporin 3 (AQP3), an aquaglyceroporin, plays an important role in the high permeability [5,6]. Therefore, the high tolerance of morulae to cryopreservation would be attributable to their high permeability via water/cryoprotectant channels.…”

Contrasting Effects of Glycerol and Urea Transport on Rat Pancreatic β-Cell Function

“…However, we were not able in the present study to investigate the specific role of AQP7 in the β-cell response to urea and glycerol owing to the lack of a suitable AQP blocker. A number of sulphydryl reagents, including silver and mercury salts, have been used to study osmolyte transport via aquaporins [30,31], but these compounds appeared to cause major irreversible cytotoxic effects on β-cells. Thus, the exact role of AQP7 in the β-cell responses to urea, glycerol and related osmolytes awaits the identification of new selective and non-toxic AQP blockers or further studies using transgenic animals lacking the AQP7 gene.…”

Expression of Aquaporins in Human Embryos and Potential Role of AQP3 and AQP7 in Preimplantation Mouse Embryo Development

“…As an aquaglyceroporin that is permeable to water and small neutral solutes including glycerol and urea [1,2,3,4,5], the functions of AQP3 in the transport of water and cryoprotectants in mouse oocytes and embryos have been investigated in several studies [9,10,11]. AQP3 facilitates water and glycerol diffusion in mouse morula [10,12]. Increased expression of AQP3 in mouse oocytes by cRNA injection improves the survival of mouse oocytes after cryopreservation [11].…”