Cholangiocyte primary cilia are chemosensory organelles that detect biliary nucleotides via P2Y₁₂ purinergic receptors

Cholangiocytes, the epithelial cells lining intrahepatic bile ducts, contain primary cilia, which are mechano- and osmosensory organelles detecting changes in bile flow and osmolality and transducing them into intracellular signals. Here, we asked whether cholangiocyte cilia are chemosensory organelles by testing the expression of P2Y purinergic receptors and components of the cAMP signaling cascade in cilia and their involvement in nucleotide-induced cAMP signaling in the cells. We found that P2Y₁₂ purinergic receptor, adenylyl cyclases (i.e., AC4, AC6, and AC8), and protein kinase A (i.e., PKA RI-β and PKA RII-α regulatory subunits), exchange protein directly activated by cAMP (EPAC) isoform 2, and A-kinase anchoring proteins (i.e., AKAP150) are expressed in cholangiocyte cilia. ADP, an endogenous agonist of P2Y₁₂ receptors, perfused through the lumen of isolated rat intrahepatic bile ducts or applied to the ciliated apical surface of normal rat cholangiocytes (NRCs) in culture induced a 1.9- and 1.5-fold decrease of forskolin-induced cAMP levels, respectively. In NRCs, the forskolin-induced cAMP increase was also lowered by 1.3-fold in response to ATP-γS, a nonhydrolyzed analog of ATP but was not affected by UTP. The ADP-induced changes in cAMP levels in cholangiocytes were abolished by chloral hydrate (a reagent that removes cilia) and by P2Y₁₂ siRNAs, suggesting that cilia and ciliary P2Y₁₂ are involved in nucleotide-induced cAMP signaling. In conclusion, cholangiocyte cilia are chemosensory organelles that detect biliary nucleotides through ciliary P2Y₁₂ receptors and transduce corresponding signals into a cAMP response.