Channels In Epithelial Cells Research Articles

Down-regulation of cystic fibrosis transmembrane conductance regulator gene expression by agents that modulate intracellular divalent cations.

In cystic fibrosis (CF), epithelial cells are unable to normally up-regulate apical membrane Cl- secretion in response to agents which increase cyclic AMP, but they do increase Cl- secretion in response to increases in intracellular Ca2+. Since intracellular divalent cations regulate the expression of many genes, we hypothesized that mobilization of intracellular Ca2+ and/or other divalent cations might modulate not only Ca(2+)-dependent Cl- channels but also cystic fibrosis transmembrane conductance regulator (CFTR) gene expression. To evaluate this concept, HT-29 human colon carcinoma cells were cultured under various conditions designed to manipulate intracellular divalent cation concentrations and CFTR gene expression was quantified at the levels of transcription, mRNA accumulation, mRNA half-life, and protein. Exposure to the divalent cation ionophores A23187 and ionomycin (agents which increase intracellular divalent cation concentrations) caused dose- and time-dependent reductions of CFTR mRNA levels, which could be blocked by the use of Ca(2+)- and Mg(2+)-free media. Ionophore-induced CFTR gene modulation was also observed with T84 human colon carcinoma cells and freshly isolated normal human bronchial epithelial cells. Incubation of HT-29 cells with thapsigargin, an agent that releases Ca2+ from intracellular stores, or in medium containing increased extracellular concentrations of Ca2+ or Mg2+ also caused down-regulation of CFTR mRNA levels. Transcription run-on analysis showed that, parallel with the decrease in CFTR mRNA levels, A23187 reduced the rate of transcription of the CFTR gene, while CFTR mRNA transcript half-life was unaffected. Consistent with the down-regulation of CFTR gene expression, CFTR protein levels also decreased after exposure to A23187. Thus, despite the independence of Ca(2+)-dependent Cl- channels and cyclic AMP-dependent CFTR-related Cl- channels in epithelial cells, increases in intracellular divalent cation concentrations down-regulate the expression of the CFTR gene at the transcriptional level, with consequent decreases in CFTR mRNA and protein.

Ion Channels and Colonic Salt Transport

The focus of this review is the ion transport functions of the vertebrate colon, in particular the role of ion channels in the maintenance and regulation of ion transport processes. A great deal has been learned in the last decade about the properties of ion channels in epithelial cells, and it seems appropriate to review our current understanding of the function of these elements in colonic epithelial transport. I like to think of epithelial ion channels in general, and colonic ion channels in particular, as working class ion channels. While neuronal ion channels are up in the penthouse subserving a variety of cerebral functions, the channels that inhabit colonic epithelial cells are quietly at work doing the tedious, but nevertheless necessary, jobs associated with preserving electrolyte balance . Absorption and Secretion

Blockade of Ca<sup>2+</sup>-Dependent Cl<sup>–</sup> Current in Xenopus Oocytes by C<sup>–</sup> Channel Blockers and Antimalarial Drugs

Effects of Cl^– channel blockers and antimalarial drugs on the Ca²⁺-dependent Cl^– current in <i>Xenopus</i> oocytes were studied using a two-microelectrode voltage-clamp technique. Chloride channel blockers, such as stilbene disulfonic acids and carboxylic acids, inhibit the Cl^- current in oocytes. The calculated half-inhibitory concentrations (IC₅₀) of these blockers on the oocyte Cl^– current demonstrate an order of potency of 4,4’-diisothiocyanatostilbene-2,2’-disulfonic acid > anthracene-9-carboxylic acid > diphenylamine-2-carboxylic acid > indanyloxyacetic acid. This is different from the potency observed in the outwardly rectifying Cl^- channels of epithelial cells. The blocking effect of these Cl^– channel blockers on the Ca²⁺-dependent Cl^– current in oocytes was observed to be stronger at positive membrane potentials. In addition, various antimalarial drugs, such as quinine’s derivatives and artesunate, inhibit the Ca²⁺-dependent Cl^– current in oocytes. These inhibitory effects on cytoplasmic membrane Cl^– channels may be responsible for the pharmacological action and toxic side effects of these compounds.

Polarized Distribution of Cation Channels in Epithelial Cells

Our aim is to establish whether Madin-Darby canine kidney (MDCK) cells have polarized ion channels. We first observe that cation currents are almost exclusively carried by K⁺; Na⁺ and Ca²⁺ currents are instead negligible. Accordingly, we investigate the presence of K⁺ channels, discriminating those on the apical from those on the basolateral side. The apical side has four different types of K⁺ channels of 31, 89, 109 and 220 pS, while the basolateral has a fifth type of 26 pS. Furthermore, channels also differ in open probabilities and kinetic parameters, demonstrating a strict asymmetry. We also observe that basolateral channels are silent in the cell-attached configuration, but become active upon excising the patch.

Polarized Distribution of Chloride Channels in Epithelial Cells

Cl-currents in cultured epithelial cells (MDCK) are inhibited by 4-acetamido-4 ’ -isothiocyano-2,2’-disulfonic acid stilbene; 4,4 ’ -diisothiocyanatostilbene-2,2<i

Sodium and potassium channels in epithelial cells from thymus glands and thymomas of myasthenia gravis patients

In both normal and neoplastic epithelial cells from human thymus glands and thymomas, respectively, we found voltage-gated sodium and potassium channels that resemble the adult-type Na channel and the delayed outward rectifier K channel, respectively, of human skeletal muscle and mammalian nervous system. These voltage-gated ion channels might be part of a communication system between epithelial cells and other components of the microenvironment of the thymus.

Absence of acetylcholine‐induced current in epithelial cells from thymus glands and thymomas of myasthenia gravis patients

We investigated the activity of ion channels in epithelial cells from human thymus glands and thymomas kept in short-term cell culture by clamping the membrane potential of the cells at -85 mV and determining the membrane current flowing on application of acetylcholine, glycine, or gamma-aminobutyric acid. In concentrations of up to 10(-3) M, none of the neurotransmitters induced any detectable current. This suggests (1) that there are no acetylcholine receptors (AChRs) or other products of the AChR gene family having ion-channel properties in the membranes of these epithelial cells, and (2) that the alpha-bungarotoxin-binding protein of thymus and thymoma has no AChR-like ion-channel property. These results support the hypothesis that the cross-reacting structures that elicit the anti-AChR autoimmune response in thymoma-associated myasthenia gravis are antigens having only limited homology with the AChR. Myasthenia gravis not associated with thymoma might have a different pathogenesis.

Volume-Regulatory Ion Channels in Epithelial Cells

Patch-clamp studies showed that the regulatory volume decrease process is closely associated with functions of K+, Cl-, and Ca2+-permeable channels in epithelial cells. Na+-permeable channels in the regulatory volume increase process may also be involved, as deduced from whole cell recordings using intestinal epithelial cells.

Ion channels in epithelial cells from thymus and thymomas of myasthenia gravis patients

Ion channels in epithelial cells from thymus and thymomas of myasthenia gravis patients

Chloride current activated by cyclic AMP and parathyroid hormone in rat osteoblasts

In primary cultures of rat osteoblasts, studied with the whole-cell configuration of the patch-clamp technique, 8-bromo-cyclic AMP (8BrcAMP) forskolin (FS) and 1-34 parathyroid hormone (PTH) were shown to activate a Cl conductance. This conductance shows a pronounced outward rectification, even with symmetrical Cl concentrations. It is blocked partially and reversibly by 4,4'-diisothiocyanatostilbene 2,2'-disulfonic acid (DIDS) or diphenylcarboxylate (DPC). The blockade induced by DIDS is time- and voltage-dependent. The Cl responses to FS and PTH develop slowly, after a delay of several seconds and are very slowly reversible. These responses were observed only in a fraction of the cells tested and their detection was favoured by cell dialysis. This Cl current should be taken into account for studying possible modulations of the voltage-gated Ca currents of osteoblasts. It is suggested that its physiological role may be related to the well-known morphological changes induced by PTH in osteoblasts. The cyclic AMP-sensitivity, the outward rectification and the sensitivity to dialysis of this Cl current are reminiscent of the properties of the cystic fibrosis-sensitive Cl channels of epithelial cells.