Bile canaliculi contract autonomously by releasing calcium into hepatocytes via mechanosensitive calcium channel.

Drug-induced hepatocellular cholestasis leads to altered bile flow. Bile is propelled along the bile canaliculi (BC) by actomyosin contractility, triggered by increased intracellular calcium (Ca). However, the source of increased intracellular Ca and its relationship to transporter activity remains elusive. We identify the source of the intracellular Ca involved in triggering BC contractions, and we elucidate how biliary pressure regulates Ca homeostasis and associated BC contractions. Primary rat hepatocytes were cultured in collagen sandwich. Intra-canalicular Ca was measured with fluo-8; and intra-cellular Ca was measured with GCaMP. Pharmacological modulators of canonical Ca-channels were used to study the Ca-mediated regulation of BC contraction. BC contraction correlates with cyclic transfer of Ca from BC to adjacent hepatocytes, and not with endoplasmic reticulum Ca. A mechanosensitive Ca channel (MCC), Piezo-1, is preferentially localized at BC membranes. The Piezo-1 inhibitor GsMTx-4 blocks the Ca transfer, resulting in cholestatic generation of BC-derived vesicles whereas Piezo-1 hyper-activation by Yoda1 increases the frequency of Ca transfer and BC contraction cycles. Yoda1 can recover normal BC contractility in drug-induced hepatocellular cholestasis, supporting that Piezo-1 regulates BC contraction cycles. Finally, we show that hyper-activating Piezo-1 can be exploited to normalize bile flow in drug-induced hepatocellular cholestasis.