091 Aquaporin 9 plays a significant role in the channel-dependent movement of Me2SO and acetamide in mouse morulae

Abstract

The permeability to water and cryoprotectants of the plasma membrane affects the suitable condition for cryopreservation of the cell. Previously, we showed that water, glycerol, ethylene glycol, Me 2 SO, and acetamide move through mouse morulae by facilitated diffusion. We also showed that the facilitated diffusion of water, glycerol, and ethylene glycol relies on aquaporin 3, an aquaglyceroporin (Edashige et al., 2007), and suggested that AQP9, another aquaglyceroporin, can transport Me 2 SO, acetamide, and other cryoprotectants when AQP9 was exogenously expressed in mouse oocytes (Edashige et al., in Cryo 2011). In the present study, we examined the transporting-specificity of AQP9 endogenously expressed in mouse morulae. In addition, we examined whether AQP9 expressed in morulae was involved in the tolerance to cryopreservation. As shown in previous studies, the suppression of the endogenous expression of AQP9 in morulae decreased the permeability to Me 2 SO and acetamide but not that to other cryoprotectants. And the exogenous expression of AQP9 in mouse oocytes, in which water/cryoprotectant channels are not expressed at the functional level, significantly increased not only the permeability to Me 2 SO and acetamide but also to glycerol and ethylene glycol. Cystic fibrosis transmembrane conductance regulator (CFTR), which has been reported to modulate the transporting-property of AQP9, was markedly expressed in oocytes but not in morulae. The suppression of the endogenous expression of CFTR in oocytes decreased the permeability to glycerol and ethylene glycol of oocytes enhanced by exogenous expression of AQP9 but not that to Me 2 SO and acetamide. These results indicate that the transporting-property of AQP9 is modulated by CFTR, and suggest that AQP9 endogenously expressed in morulae transports only Me 2 SO and acetamide because CFTR is not expressed in morulae. When mouse morulae were vitrified with DFS20, a Me 2 SO -based solution, by a one-step method, most of intact morulae survived and had the ability to develop into blastocysts after warming, whereas only 40% of AQP9-suppressed morulae survived and no embryos had the ability to develop, suggesting that AQP9 plays an important role in the tolerance to vitrification. These results will provide important information for understanding the movement of cryoprotectants in mouse embryos. Source of funding: The Japan Society for the Promotion of Science. Conflict of interest: None declared. keisuke@cc.kochi-u.ac.jp