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Characteristics of cultured subepithelial fibroblasts in the rat small intestine. II. Localization and functional analysis of endothelin receptors and cell-shape-independent gap junction permeability

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Abstract

Subepithelial fibroblasts form a cellular network with gap junctions under the epithelium of the gastrointestinal tract. Previously, we have reported their unique characteristics, such as reversible rapid cell-shape changes from a flat to a stellate configuration induced by dBcAMP and endothelins (ETs), and Ca2+ responses to, for example, ETs, ATP, and substance-P. We have now investigated the subtypes of ET receptors both in the rat small intestine and in primary cultured subepithelial fibroblasts isolated from rat duodenal villi. Their properties were compared between wild-type and endothelin-B-receptor-mutant sl/sl rats. Light- and electron-microscopic immunohistochemistry showed intense ETA immunoreactivity in the subepithelial fibroblasts from the small intestine and colon of both wild-type and sl/sl rats. In culture, immunocytochemistry, reverse transcription/polymerase chain reaction analysis, Ca2+ response measurements, and cell-shape change analysis indicated functional ETA and ETB receptors in the wild-type cells, but only ETA in the sl/sl cells. However, wild-type cells were more sensitive to ET-1 than to ET-3 by about one order of magnitude. ETA seemed to be dominant both in vivo and in vitro. The relationship between cell-shape change and gap junction permeability was examined by fluorescence recovery after photobleaching; the gap junctions were usually open but were blocked by carbenoxolone. Permeability did not change significantly with cell-shape change. This network of differentiated subepithelial fibroblasts may maintain intercellular communication via gap junctions to transduce signals evoked in the local network to the whole network. The cell-shape change of the cells through ETA activation may play an important role as a barrier and for intercellular signaling in the intestinal villi.

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Acknowledgments

We thank Drs. Yuchio Yanagawa and Tosie Ebihara for teaching us the method of the RT-PCR analysis. We also thank Dr. Seiji Hori, Ms. Katsuyo Tsuchiya, and Mr. Noboru Yamaguchi.

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Authors and Affiliations

  1. Section of Brain Structure, Center for Brain Experiment, National Institute for Physiological Sciences, Myodaiji, Okazaki, 444-8585, Japan

    Sonoko Furuya

  2. The Graduate University for Advanced Studies, Hayama, Kanagawa, Japan

    Sonoko Furuya & Tsuyoshi Ozaki

  3. Cell Mechano-sensing Project, JST, ICORP, Nagoya University, Tsurumai, Nagoya, 466-8550, Japan

    Kishio Furuya & Masahiro Sokabe

  4. Department of Physiology, School of Medicine, Nagoya University, Tsurumai, Nagoya, 466-8550, Japan

    Masahiro Sokabe

  5. Center for Experimental Animals, National Institute for Physiological Sciences, Myodaiji, Okazaki, 444-8585, Japan

    Takeshi Hiroe & Tsuyoshi Ozaki

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  1. Sonoko Furuya
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  2. Kishio Furuya
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  3. Masahiro Sokabe
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Correspondence to Sonoko Furuya.

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Furuya, S., Furuya, K., Sokabe, M. et al. Characteristics of cultured subepithelial fibroblasts in the rat small intestine. II. Localization and functional analysis of endothelin receptors and cell-shape-independent gap junction permeability. Cell Tissue Res 319, 103–119 (2005). https://doi.org/10.1007/s00441-004-0958-7

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