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Am J Physiol Cell Physiol (December 5, 2001). doi:10.1152/ajpcell.00367.2001
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Articles in PresS, published online ahead of print December 5, 2001
Am J Physiol Cell Physiol, 10.1152/ajpcell.00367.2001
Submitted on August 2, 2001
Accepted on November 28, 2001

Cholangiocytes exhibit dynamic, actin-dependent apical membrane turnover

R Brian Doctor1*, Rolf Dahl1, Laura Fouassier1, Gordan Kilic1, and J Gregory Fitz1

1 Medicine, University of Colorado Health Sciences Center, Denver, CO, USA

* To whom correspondence should be addressed. E-mail: brian.doctor{at}uchsc.edu.

The present studies of cholangiocytes utilized complementary histologic, biochemical and electrophysiologic methods to identify a dense population of sub-apical vesicles, quantify the rates of vesicular trafficking and assess the contribution of the actin cytoskeleton to membrane trafficking. FM(1-43) fluorescence measured significant basal rates of total exocytosis (1.33 ± 0.16 % plasma membrane/min) in isolated cholangiocytes and apical exocytosis in cholangiocyte monolayers. Cell surface area remained unchanged, indicating there was a concurrent, equivalent rate of endocytosis. FM(1-43) washout studies showed 36% of the endocytosed membrane was recycled to the plasma membrane. 8-cpt-cAMP (cAMP analogue) increased exocytosis by 71 ± 31% while Rp-cAMPs (PKA inhibitor) diminished basal exocytosis by 53 ± 11%. A dense population of 140 nm sub-apical vesicles arose, in part, from apical membrane endocytosis. Phalloidin staining showed a sub-population of the endocytosed vesicles was encapsulated by F-actin. Further, membrane trafficking was inhibited by disrupting the actin cytoskeleton with cytochalasin D (51 ± 13 % of control) or jasplakinolide (58 ± 9 % of control). These studies indicate there is a high rate of vesicular trafficking at the apical membrane of cholangiocytes and suggest that both cAMP and the actin cytoskeleton contributes importantly to these events.




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