Evidence that aquaporin-8 is located in the basolateral membrane of rat submandibular gland acinar cells

It is possible that, during primary saliva formation, aquaporins (AQPs) facilitate transcellular water flow across acinar cells to the lumina of salivary glands. In the rat submandibular gland (rSMG) AQP5 is localized in the apical membranes of acinar cells. The pres ence of a basolateral AQP in the same cell type has not been reported. We have therefore used immunofluorescence confocal microscopy to determine the subcellular localization of a newly discovered aquaporin, AQP8, in rSMG epithelial cells. The antibodies we used were made against the amino- or carboxyl-terminus (anti-rAQP8(NT) and anti-rAQP8(CT), respectively) of an AQP8 cloned from rat pancreas and liver (rAQP8). Two lines of evidence suggest that both antibodies are suitable for immunolocalization studies. First, results of immunofluorescence confocal microscopy studies show that both antibodies bind to the plasma membranes of 293 cells infected with an adenovirus encoding rAQP8. Second, results of immunoblots of membranes from infected cells suggest that both antibodies bind to glycosylated and non-glycosylated forms of rAQP8. When tested in frozen sections of rSMG, we could not detect the binding of anti-rAQP8(NT) to any membranes. In contrast, anti-rAQP8(NT) binds to the basolateral membranes of acinar (but not ductal) epithelia, suggesting that rAQP8 resides in the basolateral membranes of acinar cells. Lack of anti-rAQP(NT) binding to basolateral membranes suggests that this epitope is not available in the membranes. Our evidence for the basolateral localization of rAQP8 in acinar cells, coupled with previous findings that AQP5 is localized apically in the same cells, raises the possibility that water crosses the acinar epithelium through these channels during primary saliva formation.