Intermediate-conductance Ca2+-activated K+ channel KCa3.1 and its related molecules in T-lymphocytes

The intermediate-conductance Ca2+-activated K+ channel KCa3.1 (also called IKCa, IK1 and KCNN4) plays an essential role for the positive-feedback mechanism required for the enhancement of Ca2+ signaling in activated T-lymphocytes, and regulates the T cell activation, proliferation and differentiation. Recent reports have suggested that T-lymphocyte KCa3.1 K+ channel is an attractive target for the therapeutic strategies of inflammatory bowel disease (IBD). In addition, the potential KCa3.1 regulators also play critical roles in the T cell functions: phosphoinositide-3-kinase, class 2, beta polypeptide (PI3K-C2B), nucleoside diphosphate kinase B (NDPK-B), phosphohistidine phosphatase 1 (PHPT-1) and myotubularin related protein 6 (MTMR-6). We recently described that the up-regulation of KCa3.1 and NDPK-B might constitute an initiation step in CD4+ T-lymphocyte proliferation in acute IBD and might be one of important mechanisms underlying the pathogenesis of IBD (Ohya et al., Am J Physiol Gastrointest Liver Physiol. 306:G873-G885). KCa3.1 K+ channel and its regulators may be potential therapeutic targets for inflammatory diseases such as IBD

Molecular cloning of a novel gene involved in serotonin receptor-mediated signal transduction in rat stomach

AbstractIn Xenopus oocytes injected with small size mRNAs (500–700 b), obtained from rat stomach by fractionation, application of 10 μM 5-HT induced a substantial Ca2+-activated Cl− current (ICl-Ca). ICl-Ca was not elicited by 5-HT in native oocytes. Consistent results from this assay in the oocyte expression system motivated cDNA cloning experiments. A novel cDNA (named r at s tomach s erotonin receptor-related cDNA: RSS cDNA) which encodes a small protein involved in specific 5-HT receptor-mediated ICl-Ca activation was identified. The molecular weight of RSS protein in the reticulocyte lysate translation system (∼10 kDa) is identical to that calculated from the amino acid sequence. Computer-aided analysis of the predicted protein does not show any obvious sequence homologies (<18%) to any other proteins including G protein-coupled receptors. Northern analysis revealed that RSS mRNA is ubiquitously expressed at varying levels in a number of different tissues. Furthermore, the binding of [3H]spiperone, a 5-HT2 receptor antagonist, was examined in CHO cells, which highly expressed RSS transcripts (named CHO-RSS). Specific binding of [3H]spiperone was not clearly observed in native CHO but was detected in CHO-RSS. The dissociation constant was 10.3 nM in CHO-RSS. These results suggest that RSS protein may be a factor which facilitates 5-HT receptor expression or, alternatively, an enhancer of the affinity of native 5-HT receptor to 5-HT

STIM/Orai-mediated calcium entry elicits spontaneous TSLP overproduction in epidermal cells of atopic dermatitis mice

Aim: Atopic dermatitis (AD) is a pruritic, chronic inflammatory skin disease. Thymic stromal lymphopoietin (TSLP) is highly expressed in the epidermis of patients with AD and induces T helper 2 (Th2) immune responses and itching. Although the mechanism underlying the stimulus-induced TSLP production in normal keratinocytes has been intensively studied, whether the production capability of TSLP is naturally enhanced in epidermal cells in AD conditions remains unclear. Previous studies demonstrated that a deficiency of polyunsaturated fatty acid (PUFA) causes AD-like pruritic skin inflammation in special diet-fed hairless mice. The aim of the study was to examine the TSLP production capability of epidermal cells isolated from diet-induced AD mouse model and its mechanism. Methods: Epidermal cells were isolated from normal and AD mice and incubated under unstimulated culture conditions to assess spontaneous TSLP production. Messenger ribonucleic acid (mRNA) and protein levels of TSLP were determined by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Results: TSLP level was markedly increased in the skin of AD mice. When epidermal cells were isolated from AD mice and cultured without stimulation, Tslp gene expression was upregulated, and a large amount of TSLP protein was extracellularly released. Such TSLP overproduction was not observed in the epidermal cells of normal mice. TSLP overproduction in AD epidermal cells was almost completely inhibited by extracellular calcium chelation, interference with plasma membrane interaction of stromal interaction molecule 1 (STIM1), blockade of the calcium release-activated calcium (CRAC) channels Orai1 and Orai2, or treatment with a PUFA γ-linolenic acid (GLA). Conclusions: Epidermal cells isolated from AD mice can spontaneously produce TSLP through STIM/Orai-mediated calcium entry, and GLA may negatively regulate this TSLP production

Serotonin Augments Gut Pacemaker Activity via 5-HT3 Receptors

Serotonin (5-hydroxytryptamine: 5-HT) affects numerous functions in the gut, such as secretion, muscle contraction, and enteric nervous activity, and therefore to clarify details of 5-HT's actions leads to good therapeutic strategies for gut functional disorders. The role of interstitial cells of Cajal (ICC), as pacemaker cells, has been recognised relatively recently. We thus investigated 5-HT actions on ICC pacemaker activity. Muscle preparations with myenteric plexus were isolated from the murine ileum. Spatio-temporal measurements of intracellular Ca2+ and electric activities in ICC were performed by employing fluorescent Ca2+ imaging and microelectrode array (MEA) systems, respectively. Dihydropyridine (DHP) Ca2+ antagonists and tetrodotoxin (TTX) were applied to suppress smooth muscle and nerve activities, respectively. 5-HT significantly enhanced spontaneous Ca2+ oscillations that are considered to underlie electric pacemaker activity in ICC. LY-278584, a 5-HT3 receptor antagonist suppressed spontaneous Ca2+ activity in ICC, while 2-methylserotonin (2-Me-5-HT), a 5-HT3 receptor agonist, restored it. GR113808, a selective antagonist for 5-HT4, and O-methyl-5-HT (O-Me-5-HT), a non-selective 5-HT receptor agonist lacking affinity for 5-HT3 receptors, had little effect on ICC Ca2+ activity. In MEA measurements of ICC electric activity, 5-HT and 2-Me-5-HT caused excitatory effects. RT-PCR and immunostaining confirmed expression of 5-HT3 receptors in ICC. The results indicate that 5-HT augments ICC pacemaker activity via 5-HT3 receptors. ICC appear to be a promising target for treatment of functional motility disorders of the gut, for example, irritable bowel syndrome

Castration Induces Down-Regulation of A-Type K+ Channel in Rat Vas Deferens Smooth Muscle

A-type K+ channels contribute to regulating the propagation and frequency of action potentials in smooth muscle cells (SMCs). The present study (i) identified the molecular components of A-type K+ channels in rat vas deferens SMs (VDSMs) and (ii) showed the long-term, genomic effects of testosterone on their expression in VDSMs. Transcripts of the A-type K+ channel &alpha; subunit, Kv4.3L and its regulatory &beta; subunits, KChIP3, NCS1, and DPP6-S were predominantly expressed in rat VDSMs over the other related subtypes (Kv4.2, KChIP1, KChIP2, KChIP4, and DPP10). A-type K+ current (IA) density in VDSM cells (VDSMCs) was decreased by castration without changes in IA kinetics, and decreased IA density was compensated for by an oral treatment with 17&alpha;-methyltestosterone (MET). Correspondingly, in the VDSMs of castrated rats, Kv4.3L and KChIP3 were down-regulated at both the transcript and protein expression levels. Changes in Kv4.3L and KChIP3 expression levels were compensated for by the treatment with MET. These results suggest that testosterone level changes in testosterone disorders and growth processes control the functional expression of A-type K+ channels in VDSMCs

Intermediate-conductance Ca2+-activated K+ channel KCa3.1 and its related molecules in T-lymphocytes: DOI: 10.14800/ics.327

The intermediate-conductance Ca2+-activated K+ channel KCa3.1 (also called IKCa, IK1 and KCNN4) plays an essential role for the positive-feedback mechanism required for the enhancement of Ca2+ signaling in activated T-lymphocytes, and regulates the T cell activation, proliferation and differentiation. Recent reports have suggested that T-lymphocyte KCa3.1 K+ channel is an attractive target for the therapeutic strategies of inflammatory bowel disease (IBD). In addition, the potential KCa3.1 regulators also play critical roles in the T cell functions: phosphoinositide-3-kinase, class 2, beta polypeptide (PI3K-C2B), nucleoside diphosphate kinase B (NDPK-B), phosphohistidine phosphatase 1 (PHPT-1) and myotubularin related protein 6 (MTMR-6). We recently described that the up-regulation of KCa3.1 and NDPK-B might constitute an initiation step in CD4+ T-lymphocyte proliferation in acute IBD and might be one of important mechanisms underlying the pathogenesis of IBD (Ohya et al., Am J Physiol Gastrointest Liver Physiol. 306:G873-G885). KCa3.1 K+ channel and its regulators may be potential therapeutic targets for inflammatory diseases such as IBD