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Differential expression of genes related to purinergic signaling in smooth muscle cells, PDGFR?-positive cells, and interstitial cells of Cajal in the murine colon.

ABSTRACT: Purinergic signaling provides regulation of colonic motility. Smooth muscle cells (SMC), interstitial cells of Cajal (ICC), and platelet-derived growth factor receptor ?-positive (PDGFR?(+) ) cells are electrically coupled and form a functional (SIP) syncytium that constitutes the receptive field for motor neurotransmitters in the tunica muscularis. Each cell type in the SIP syncytium has specialized functions in mediating motor neurotransmission. We compared gene transcripts for purinergic receptors and membrane-bound enzymes for purine degradation expressed by each cell type of the SIP syncytium.Fluorescence-activated cell sorting (FACS) was used to purify SMC, ICC, and PDGFR?(+) cells from mixed cell populations of colonic muscles dispersed from reporter strains of mice with constitutive expression of green fluorescent proteins. Differential expression of functional groups of genes related to purinergic signaling was determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR).We detected marked phenotypic differences among SMC, ICC, and PDGFR?(+) cells. Substantial numbers of genes of importance in purinergic neurotransmission were enriched in PDGFR?(+) cells in relation to SMC and ICC. Notably, genes related to mediating effects and extracellular biotransformation of enteric purinergic inhibitory neurotransmitters were strongly expressed by PDGFR?(+) cells.Our results demonstrate differential expression of genes for proteins involved in purinergic signaling in the SIP syncytium. These results may further clarify the specific functions of each cell type, identify novel biomarkers for postjunctional cells, and provide hypotheses for further studies to understand the physiological roles of cells of the SIP syncytium.

SUBMITTER: Peri LE

PROVIDER: S-EPMC3735650 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

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Differential expression of genes related to purinergic signaling in smooth muscle cells, PDGFRα-positive cells, and interstitial cells of Cajal in the murine colon.

Peri L E LE Sanders K M KM Mutafova-Yambolieva V N VN

Neurogastroenterology and motility 20130630 9

<h4>Background</h4>Purinergic signaling provides regulation of colonic motility. Smooth muscle cells (SMC), interstitial cells of Cajal (ICC), and platelet-derived growth factor receptor α-positive (PDGFRα(+) ) cells are electrically coupled and form a functional (SIP) syncytium that constitutes the receptive field for motor neurotransmitters in the tunica muscularis. Each cell type in the SIP syncytium has specialized functions in mediating motor neurotransmission. We compared gene transcripts ...[more]

PMID: 23809506

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Project description:Key pointsColonic intramuscular interstitial cells of Cajal (ICC-IM) exhibit spontaneous Ca2+ transients manifesting as stochastic events from multiple firing sites with propagating Ca2+ waves occasionally observed. Firing of Ca2+ transients in ICC-IM is not coordinated with adjacent ICC-IM in a field of view or even with events from other firing sites within a single cell. Ca2+ transients, through activation of Ano1 channels and generation of inward current, cause net depolarization of colonic muscles. Ca2+ transients in ICC-IM rely on Ca2+ release from the endoplasmic reticulum via IP3 receptors, spatial amplification from RyRs and ongoing refilling of ER via the sarcoplasmic/endoplasmic-reticulum-Ca2+ -ATPase. ICC-IM are sustained by voltage-independent Ca2+ influx via store-operated Ca2+ entry. Some of the properties of Ca2+ in ICC-IM in the colon are similar to the behaviour of ICC located in the deep muscular plexus region of the small intestine, suggesting there are functional similarities between these classes of ICC.AbstractA component of the SIP syncytium that regulates smooth muscle excitability in the colon is the intramuscular class of interstitial cells of Cajal (ICC-IM). All classes of ICC (including ICC-IM) express Ca2+ -activated Cl- channels, encoded by Ano1, and rely upon this conductance for physiological functions. Thus, Ca2+ handling in ICC is fundamental to colonic motility. We examined Ca2+ handling mechanisms in ICC-IM of murine proximal colon expressing GCaMP6f in ICC. Several Ca2+ firing sites were detected in each cell. While individual sites displayed rhythmic Ca2+ events, the overall pattern of Ca2+ transients was stochastic. No correlation was found between discrete Ca2+ firing sites in the same cell or in adjacent cells. Ca2+ transients in some cells initiated Ca2+ waves that spread along the cell at ∼100 µm s-1 . Ca2+ transients were caused by release from intracellular stores, but depended strongly on store-operated Ca2+ entry mechanisms. ICC Ca2+ transient firing regulated the resting membrane potential of colonic tissues as a specific Ano1 antagonist hyperpolarized colonic muscles by ∼10 mV. Ca2+ transient firing was independent of membrane potential and not affected by blockade of L- or T-type Ca2+ channels. Mechanisms regulating Ca2+ transients in the proximal colon displayed both similarities to and differences from the intramuscular type of ICC in the small intestine. Similarities and differences in Ca2+ release patterns might determine how ICC respond to neurotransmission in these two regions of the gastrointestinal tract.

Project description:Key pointsRhythmic action potentials and intercellular Ca2+ waves are generated in smooth muscle cells of colonic longitudinal muscles (LSMC). Longitudinal muscle excitability is tuned by input from subserosal ICC (ICC-SS), a population of ICC with previously unknown function. ICC-SS express Ano1 channels and generate spontaneous Ca2+ transients in a stochastic manner. Release of Ca2+ and activation of Ano1 channels causes depolarization of ICC-SS and LSMC, leading to activation of L-type Ca2+ channels, action potentials, intercellular Ca2+ waves and contractions in LSMC. Nitrergic neural inputs regulate the Ca2+ events in ICC-SS. Pacemaker activity in longitudinal muscle is an emergent property as a result of integrated processes in ICC-SS and LSMC.AbstractMuch is known about myogenic mechanisms in circular muscle (CM) in the gastrointestinal tract, although less is known about longitudinal muscle (LM). Two Ca2+ signalling behaviours occur in LM: localized intracellular waves not causing contractions and intercellular waves leading to excitation-contraction coupling. An Ano1 channel antagonist inhibited intercellular Ca2+ waves and LM contractions. Ano1 channels are expressed by interstitial cells of Cajal (ICC) but not by smooth muscle cells (SMCs). We investigated Ca2+ signalling in a novel population of ICC that lies along the subserosal surface of LM (ICC-SS) in mice expressing GCaMP6f in ICC. ICC-SS fired stochastic localized Ca2+ transients. Such events have been linked to activation of Ano1 channels in ICC. Ca2+ transients in ICC-SS occurred by release from stores most probably via inositol trisphosphate receptors. This activity relied on influx via store-operated Ca2+ entry and Orai channels. No voltage-dependent mechanism that synchronized Ca2+ transients in a single cell or between cells was found. Nitrergic agonists inhibited Ca2+ transients in ICC-SS, and stimulation of intrinsic nerves activated nitrergic responses in ICC-SS. Cessation of stimulation resulted in significant enhancement of Ca2+ transients compared to the pre-stimulus activity. No evidence of innervation by excitatory, cholinergic motor neurons was found. Our data suggest that ICC-SS contribute to regulation of LM motor activity. Spontaneous Ca2+ transients activate Ano1 channels in ICC-SS. Resulting depolarization conducts to SMCs, depolarizing membrane potential, activating L-type Ca2+ channels and initiating contraction. Rhythmic electrical and mechanical behaviours of LM are an emergent property of SMCs and ICC-SS.

Dataset Information

Differential expression of genes related to purinergic signaling in smooth muscle cells, PDGFR?-positive cells, and interstitial cells of Cajal in the murine colon.

Publications

Differential expression of genes related to purinergic signaling in smooth muscle cells, PDGFRα-positive cells, and interstitial cells of Cajal in the murine colon.

Similar Datasets

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