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

Abstract

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 (Ca²⁺). However, the source of increased intracellular Ca²⁺ and its relationship to transporter activity remains elusive. We identify the source of the intracellular Ca²⁺ involved in triggering BC contractions, and we elucidate how biliary pressure regulates Ca²⁺ homeostasis and associated BC contractions. Primary rat hepatocytes were cultured in collagen sandwich. Intra-canalicular Ca²⁺ was measured with fluo-8; and intra-cellular Ca²⁺ was measured with GCaMP. Pharmacological modulators of canonical Ca²⁺-channels were used to study the Ca²⁺-mediated regulation of BC contraction. BC contraction correlates with cyclic transfer of Ca²⁺ from BC to adjacent hepatocytes, and not with endoplasmic reticulum Ca²⁺. A mechanosensitive Ca²⁺ channel (MCC), Piezo-1, is preferentially localized at BC membranes. The Piezo-1 inhibitor GsMTx-4 blocks the Ca²⁺ transfer, resulting in cholestatic generation of BC-derived vesicles whereas Piezo-1 hyper-activation by Yoda1 increases the frequency of Ca²⁺ 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.