Regulation of Ca²⁺signaling in rat bile duct epithelia by inositol 1,4,5-trisphosphate receptor isoforms

Abstract

Cytosolic Ca 2؉ (Ca i 2؉ ) regulates secretion of bicarbonate and other ions in the cholangiocyte. In other cell types, this second messenger acts through Ca 2؉ waves, Ca 2؉ oscillations, and other subcellular Ca 2؉ signaling patterns, but little is known about the subcellular organization of Ca 2؉ signaling in cholangiocytes. Therefore, we examined Ca 2؉ signaling and the subcellular distribution of Ca 2؉ release channels in cholangiocytes and in a model cholangiocyte cell line. The expression and subcellular distribution of inositol 1,4,5-trisphosphate (InsP 3 ) receptor (InsP 3 R) isoforms and the ryanodine receptor (RyR) were determined in cholangiocytes from normal rat liver and in the normal rat cholangiocyte (NRC) polarized bile duct cell line. Subcellular Ca 2؉ signaling in cholangiocytes was examined by confocal microscopy. All 3 InsP 3 R isoforms were expressed in cholangiocytes, whereas RyR was not expressed. The type III InsP 3 R was the most heavily expressed isoform at the protein level and was concentrated apically, whereas the type I and type II isoforms were expressed more uniformly. The type III InsP 3 R was expressed even more heavily in NRC cells but was concentrated apically in these cells as well. Adenosine triphosphate (ATP), which increases Ca

Cyclic AMP Regulates Bicarbonate Secretion in Cholangiocytes Through Release of ATP Into Bile

“…Forskolin increases cytosolic Ca 2+ in cholangiocytes. (a) ACh (100 μM) induces a rapid, transient increase in Ca 2+ , as has been described previously (9). (b) Forskolin (100 μM) induces a gradual, progressive increase in Ca 2+ , which is blocked by suramin (50 μM).…”

“…To test this, the effects of forskolin on Ca 2+ were measured in intrahepatic bile duct segments isolated from rat livers, and then Ca 2+ _signaling in individual cells within the bile duct segments was examined using confocal microscopy ( Figure 7) (8; 9). Bile ducts were stimulated with forskolin (100 μM), or with ACh (100 μM) as a positive control that is known to cause an InsP3-mediated increase in Ca 2+ in this cell type (9). Both ACh (Figure 7a) and forskolin (Figure 7b) increased Ca 2+ in cholangiocytes, although the forskolin-induced increase was more gradual.…”

“…Ca 2+ signaling in cholangiocytes is mediated entirely by the InsP3R, because this is the only type of intracellular Ca 2+ release channel expressed in these cells (9). Cholangiocytes express all three isoforms of the InsP3R; the types I and II InsP3R are found throughout the cell, whereas the type III isoform is concentrated in the apical region (9).…”

“…Cholangiocytes express all three isoforms of the InsP3R; the types I and II InsP3R are found throughout the cell, whereas the type III isoform is concentrated in the apical region (9). In order to understand the relative role of each isoform in the regulation of bicarbonate secretion, we used isoform-specific siRNA (13) to eliminate them from IBDU.…”

“…However, loss of expression of the inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R) appears to be a general feature of cholestatic conditions (8). Because the InsP3R is the only intracellular Ca 2+ release channel in cholangiocytes (9), this suggests that secretion mediated by cAMP and CFTR must depend in part upon InsP3R expression. The purpose of this study was to determine whether and how each of the three isoforms of the InsP3R affects cAMP-and CFTR-mediated bicarbonate secretion in cholangiocytes.…”