title: Activation of ionic channels by deoxycholate in frog and human cell lines.
abstract: Humans, after extensive ileal resection, frequently suffer from diarrhoea, which may be due to an increased delivery of deoxycholate (DOC) to the large intestine. In the frog skin the addition of DOC (0.5 mM) to the apical side induced the activation of amiloride-sensitive Na+ channels and an increase in the unidirectional Cl- fluxes. Here we used two established cell lines (A6 and Caco2) to study the effect of DOC on ion channels at cell and membrane level using the patch-clamp technique. In A6 cells subcultured directly on Petri dishes and studied in the whole-cell configuration, DOC induced an increase in cell conductance of 110.3 +/- 4 pS pF-1 (N = 8) which was reduced to 89 +/- 14 pS pF-1 (N = 8) by the addition of DIDS (0.5 mM), The absolute values of these two effects were not statistically different (P < 0.2). In Caco2 cells, the addition of DOC (0.5 mM) induced, after 1 min, an increase in cell conductance of 583 +/- 16 pS pF-1 (N = 8) which was reduced to 560.4 +/- 16 pS pF-1 (N = 8) by DIDS (0.5 mM) and N-phenylanthranilic acid (DPC; 0.5 mM). The two values were not statistically different (P < 0.4). In Caco2 cells subcultured under the same conditions, DOC induced an increase in cell conductance of 1710 +/- 64 pS pF-1 (N = 6). Subsequent addition of amiloride (0.1 mM) reduced the cell conductance to 1558 +/- 33 pS pF-1 (N = 6). These two mean values were statistically different allowing for an error of the second kind < 0. 05. In cells in which DOC produced a conductance increase of 1010 +/- 10 pS pF-1, gadolinium (0.5 mM) induced a fall in cell conductance of 1800 +/- 10 pS pF-1. In Caco2 cells, addition of DOC (0.5 mM) to the bath reversibly induced the appearance of or an increase in channel activity in patches studied in cell-attached and excised inside-out configuration. In inside-out experiments (N = 13) DOC (0. 5 mM) induced the appearance of channel activity with conductances and reversal potentials (Er) of 27.7 +/- 1.9 pS and 0.8 +/- 5.7 mV, respectively. In cell-attached patches (N = 13) these values were 24.9 +/- 4.4 pS and -18.1 +/- 6.4 mV. In excised inside-out patches from Caco2 cells, subjected to electrochemical gradients for Na+, K+ and Cl-, (+85, -85 and 0 mV, respectively), addition of DOC also induced an increase in the baseline conductance and a shift in the reversal potential from values around +25 mV to values around 0 mV. Bile salts activated both anionic and cationic channels and did not require the presence of intracellular factors for these effects. We suggest that they act at the membrane level.
authors.affiliation: Instituto de Ciencias Biomedicas de Abel Salazar, University of Oporto and Faculdade de Medicina Veterinária, Technical University of Lisbon, Portugal.
authors.forename: A C
authors.initials: AC
authors.lastname: Maurício
authors.forename: K T
authors.initials: KT
authors.lastname: Ferreira
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D018938:Caco-2 Cells; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002460:Cell Line; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism; D003840:Deoxycholic Acid / Q000494:pharmacology*; D003967:Diarrhea / Q000209:etiology; D006801:Humans; D007082:Ileum / Q000601:surgery; D007473:Ion Channels / Q000187:drug effects* / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D015222:Sodium Channels / Q000187:drug effects / Q000378:metabolism; D014982:Xenopus laevis; D062367:ortho-Aminobenzoates / Q000494:pharmacology
chemical_list: D002121:Calcium Channel Blockers; D018118:Chloride Channels; D007473:Ion Channels; D015222:Sodium Channels; D062367:ortho-Aminobenzoates; D003840:Deoxycholic Acid; D000584:Amiloride; C001209:fenamic acid; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid