The use of isolated rat hepatocyte couplets, microscopic optical planimetry and quantitative microfluorimetry in hepatobiliary physiology

Abstract

Bile formation requires the vectorial transport of water and solutes into small canaliculi enclosed by two adjacent hepatocytes and further modifications by the biliary epithelium. Direct study of canalicular bile secretion has been hampered by the inacessibility of the tiny canalicular space to micropunture techniques and by the loss of polarity which rapidily occurs when hepatocytes are dissociated. For example, it is still debated whether the bile acid ursodeoxycholate (UDCA) stimulates hypercholeresis biliary HC03 secretion by activating hepatocellular +anq Na /H exchange or as a consequence of its reabsorption by the bile ducts. Part of the methodological limitations can be overcome with the use of isolated rat epatocyte couplets (IRHC). These isolated hepatocytes, which remain undissociated after liver collagenase digestion, retain their bile secretory polarity, and, after short term culture, reform a canalicular space, with competent tight junctions, and polariied expression of the canalicular marker Mg-ATPase. IRHC are also able to transport cholephilic flourescent anions and bile acids into the canalicular space, which rapidly expands following administration of choleretic bile acids. We have applied quantitative microfluorimetric techniques and microscopic opti;z$,;$animetry, to directly assess the effects of UDCA on exchange activity, luminal pH and canalicular expansion rates in these polarized hepatocytes.