Regulation of the dynamic localization of the rat Bsep gene-encoded bile salt export pump by anisoosmolarity

Canalicular transport via the bile salt export pump (Bsep) represents the rate-controlling step in taurocholate excretion, whose capacity is under osmotic control. The short-term effects of anisoosmolarity and Ca2+-withdrawal on the localization of Bsep and the tight junction proteins Zo-1 and occludin were studied in perfused rat liver by immunohistochemistry, confocal microscopy, and densitometry. Under normoosmotic conditions, Bsep was found in the canalicular membrane and showed a punctate intracellular localization. Hypoosmolarity resulted in the translocation of intracellular Bsep to the canalicular membrane, whereas hyperosmolarity induced a retrieval of Bsep. Following hyperosmolar retrieval of Bsep and multidrug resistance protein 2 (Mrp2) from the canalicular membrane, in the putative intracellular vesicles Bsep and Mrp2 colocalized in 15% of these vesicles, whereas 85% stained either positive for Bsep (61%) or Mrp2 (24%). Anisotonicity had no effect on the linear staining patterns of occludin and Zo-1, indicating no increase in paracellular permeability. Omission of calcium produced cholestasis characterized by a disruption of occludin, whereas the localization of Zo-1, Bsep, and Mrp2 remained unaffected. It is concluded (1) that hyperosmolarity induces retrieval of Bsep from the canalicular membrane, which correlates to cholestasis. Hypoosmolarity leads to choleresis accompanied by a rapid recruitment of intracellular Bsep to the canalicular membrane. (2) Bsep- and Mrp2-specific vesicles participate in the short-term osmoregulation of canalicular secretion, however, a cause-effect relationship between bile salt excretion and transporter localization remains to be established. (3) Ca2+-depletion induces cholestasis by disruption of occludin-determined tight junctional permeability, whereas internalization of canalicular transporters play a minor role. (HEPATOLOGY 2001;33:509-518.)