PubMed Health⌕ Search

PubMed · 12433664

Characterization of regulatory volume decrease in freshly isolated mouse cholangiocytes.

Abstract

Cell volume regulation plays a vital role in many cell functions. Recent study indicates that both K(+) and Cl(-) channels are important for the regulatory volume decrease (RVD) of cholangiocarcinoma cells, but its physiological significance is unclear due to the tumorous nature of the cells used. This present study reports the RVD of normal mouse cholangiocytes by using freshly isolated bile duct cell clusters (BDCC). A relatively simple and practical method of measuring the cross-sectional area of BDCCs by quantitative videomicroscopy was used to indirectly measure their volumes. Mouse cholangiocytes exhibited RVD, which was inhibited by 5-nitro-2'-(3-phenylpropylamino)-benzoate, DIDS, and glibenclamide, suggesting its dependence on certain chloride channels, such as volume-activated chloride channels. It is also inhibited by barium chloride but not by tetraethylammonium chloride, indicating its dependence on certain potassium channels. However, cAMP agonists had no significant effect on the RVD of BDCCs. This indirect method described can be used to study the RVD of cholangiocytes from normal as well as genetically altered mouse livers.

Explore related subjects

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Won Kyoo Cho. 2002. Characterization of regulatory volume decrease in freshly isolated mouse cholangiocytes.. https://doi.org/10.1152/ajpgi.00256.2002

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related citations

Postmitochondrial regulation of apoptosis by bicarbonate.

Ion homeostasis may play a role in the regulation of apoptosis. The current study has shown such a role for bicarbonate (HCO(3)(-)). In apoptosis triggered by ATP depletion, the proapoptotic molecule Bax translocated from the cytosol to mitochondria, followed by cytochrome c release from the organelle, caspase activation, and development of apoptotic morphology. Apoptosis was significantly ameliorated, when HCO(3)(-) was omitted from the incubation medium. The HCO(3)(-) dependence was also demonstrated for apoptosis induced by staurosporine in HeLa cells. Of significance, when HCO(3)(-) was reintroduced, apoptosis was restored. The Cl(-)/HCO(3)(-) exchanger inhibitor DIDS suppressed apoptosis in HCO(3)(-)-containing medium, further supporting a role for intracellular HCO(3)(-) in apoptosis regulation. We subsequently examined HCO(3)(-)-dependent steps in the apoptotic cascade. Translocation of Bax and cytochrome c was not suppressed by the omission of HCO(3)(-), suggesting HCO(3)(-) regulation at postmitochondrial levels. In vitro reconstitution of caspase activation using exogenous cytochrome c and cytosolic extracts was not HCO(3)(-) dependent. HCO(3)(-) was not required for the enzymatic activity of recombinant caspases either. In conclusion, the results have provided compelling evidence for HCO(3)(-) regulation of apoptosis. Such regulation takes place at postmitochondrial levels, downstream of Bax/cytochrome c translocation.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

NHE3 inhibition activates duodenal bicarbonate secretion in the rat.

We examined the effect of inhibition of Na+/H+ exchange (NHE) on duodenal bicarbonate secretion (DBS) in rats to further understand DBS regulation. DBS was measured by using the pH-stat method and by using CO2-sensitive electrodes. 5-(N,N-dimethyl)-amiloride (50 microM; DMA), a concentration that selectively inhibits the NHE isoforms NHE1 and NHE2, but not NHE3, did not affect DBS. Nevertheless, 3 mM DMA, a higher concentration that inhibits NHE1, NHE2, and NHE3, significantly increased DBS. Moreover, S1611 and S3226, both specific inhibitors of NHE3 only, or perfusion with Na+-free solutions, dose dependently increased DBS, as measured by pH-stat and CO2-sensitive electrode, without affecting intracellular pH. Coperfusion with 0.1 microM indomethacin, 0.5 mM DIDS, or 1 mM methazolamide did not affect S3226-induced DBS. Nevertheless, coperfusion with 0.1 and 0.3 mM 5-nitro-2-(3-phenylpropylamino) benzoic acid, which inhibits the cystic fibrosis transmembrane conductor regulator (CFTR), dose dependently inhibited S3226-induced DBS. In conclusion, only specific apical NHE3 inhibition increased DBS, whereas prostaglandin synthesis, Na+-HCO3- cotransporter activation, or intracellular HCO3- formation by carbonic anhydrase was not involved. Because NHE3 inhibition-increased DBS was inhibited by an anion channel inhibitor and because reciprocal CFTR regulation has been previously shown between NHE3 and apical membrane anion transporters, we speculate that NHE3 inhibition increased DBS by altering anion transporter function.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

Independence of apical Cl-/HCO3- exchange and anion conductance in duodenal HCO3- secretion.

Reduced gastrointestinal HCO3- secretion contributes to malabsorption and obstructive syndromes in cystic fibrosis. The apical HCO3- transport pathways in these organs have not been defined. We therefore assessed the involvement of apical Cl-/HCO3- exchangers and anion conductances in basal and cAMP-stimulated duodenal HCO3- secretion. Muscle-stripped rat and rabbit proximal duodena were mounted in Ussing chambers, and electrical parameters, HCO3- secretion rates, and 36Cl-, 22Na+, and 3H+ mannitol fluxes were assessed. mRNA expression levels were measured by a quantitative PCR technique. Removal of Cl- from or addition of 1 mM DIDS to the luminal perfusate markedly decreased basal HCO3- secretion but did not influence the HCO3- secretory response to 8-bromo-cAMP, which was inhibited by luminal 5-nitro-2-(3-phenylpropylamino)-benzoate. Bidirectional 22Na+ and 36Cl- flux measurements demonstrated an inhibition rather than a stimulation of apical anion exchange during cAMP-stimulated HCO3- secretion. The ratio of Cl- to HCO3- in the anion secretory response was compatible with both Cl- and HCO3- being secreted via the CFTR anion channel. CFTR expression was very high in the duodenal mucosa of both species. We conclude that in rat and rabbit duodena, an apical Cl-/HCO3- exchanger mediates a significant part of basal HCO3- secretion but is not involved in the HCO3- secretory response to cAMP analogs. The inhibitor profile, the strong predominance of Cl- over HCO3- in the anion secretory response, and the high duodenal CFTR expression levels suggest that a major portion of cAMP-stimulated duodenal HCO3- secretion is directly mediated by CFTR.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗