Bile canalicular contraction and dilatation in primary culture of rat hepatocytes - Possible involvement of two different types of plasma membrane Ca2+-Mg2+-ATPase and Ca2+-pump-ATPase

Increasing evidence has indicated that bile canalicular contraction is mediated by the nonmuscular Ca2+-calmodulin-actomyosin system, and the contraction facilitates canalicular bile flow. The aim of the present study was to examine, by electron cytochemistry, how the expression of two types of plasma membrane Ca2+-ATPase, i.e., Ca2+-Mg2+-ATPase and Ca2+ pump-ATPase, is related to the dynamic changes of bile canalicular contraction. Hepatocytes isolated from male Wistar rat liver by collagenase perfusion were cultured to form a primary monolayer. The canalicular dynamics in the couplets and triplets were analyzed by time-lapse cinematography. The Ca2+-Mg2+-ATPase activity was identified by the electron cytochemical method of Ando. Ultrastructural localization of Ca2+ pump-ATPase was examined by immunogold electron microscopy. We found that cytochemical reaction products showing the presence of Ca2+-Mg2+-ATPase activity were localized on the luminal side of the bile canalicular membranes. Immunogold particles, indicating the presence of Ca2+ pump-ATPase; were located mainly on the cytoplasmic side of the bile canalicular membranes. The expression of both Ca2+-ATPases on the canalicular membranes was enhanced during the contracting stage of bile canaliculi, whereas their expression was diminished in the dilating stage. We conclude that two different types of bile canalicular Ca2+-ATPase may be involved in the regulation of canalicular contractility to control the extrusion of intracytoplasmic free calcium ions into the canalicular lumen.