Effects of hydrophobic compounds on anion channels expressed in pancreatic duct cells

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Anion conductance in pancreatic duct cells is mainly underlied by Cystic Fibrosis Transmembrane-conductance Regulator (CFTR) anion channels. In this study, we investigated the effects of hydrophobic compounds on CFTR channels using patch-clamp technique.Small negatively-charged hydrophobic aromatic compounds like aromatic carboxylic acids induced both a decrease in single channel conductance and an increase in open probability (Po) simultaneously. On the other hand, small electro-neutral hydrophobic aromatic compounds increased Po without decreasing single channel conductance. These results suggested that the small aromatic compounds bind to a hydrophobic binding site inside the channel pore with a fast kinetics leading to reduce the single channel conductance, and also increase Po by binding to another site located in the intracellular side of CFTR channel.We also investigated mechanisms of ATP-dependent gating of CFTR channel as well as effects of cholesterol on the CFTR gating. Our … More results for the ATP-dependent gating strongly supported the "nucleotide binding domain (NBD) dimmerization hypothesis" that two NBDs make a dimmer with two ATP molecules in their interface, leading to open the channel pore, and the sequential hydrolysis of ATP binding to NBD2 breaks the dimmer, leading to channel closing. We also found that cholesterol content in plasma membrane changed the gating kinetics of CFTR channel significantly. Totally our results suggested that cholesterol affects rate of the ATP hydrolysis at the nucleotide binding domain via changing the mechanical resistance to the conformational change of the membrane spanning domains during channel pore gating.Taken together, the present study showed that effects of hydrophobic compounds on CFTR widely varied dependent on its peculiar characteristics of each compound, not their common feature of ‘hydrophobicity', which suggested us the existence of a complicated feed back system via lipid metabolism. Our novel findings for ATP-dependent gating significantly contribute to understanding of gating mechanisms of CFTR itself and other ABC transporters as well.We also performed some basic studies for epithelial BK_<Ca> channels and GABA_A channels expressed in peripheral tissue, which are important for the next step in this research field. Less