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Cellular mechanisms in activation of Na-K-Cl cotransport in nasal gland acinar cells of guinea pigs

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Abstract

The cellular regulation mechanism of Na-K-Cl cotransport was studied in dispersed acinar cells of the guinea pig nasal gland by a microfluorimetric imaging method using the Na+-sensitive dye sodium-binding benzofuran isophthalate. Addition of 1 μm acetylcholine (ACh) induced an immediate increase in intracellular Na+ concentration ([Na+]i) by 36.7±9.9 mm, which was almost completely abolished by the addition of atropine. The increased [Na+]i after cholinergic stimulation was due to the external Cl−-dependent cotransport system (about 80% of the total Na+ influx) and the dimethyl amiloride-sensitive Na+-H+ exchange system (of about 20%). The ACh-induced increase in [Na+]i was dependent on extracellular Ca2+ and was prevented by pretreatment with 8-(N, N-diethylamino)octyl-3,4,5-trimethoxybenzoate or O-O′-bis(2-aminophenyl)ethyleneglycol-N, N, N′, N′-tetraacetic acid tetraacetoxymethyl ester. Addition of 1μm ionomycin mimicked the ACh-induced increase in [Na+]i which was dependent on external Cl−. Moreover, both a calmodulin antagonist trifluoperazine and a myosin light chain kinase inhibitor ML-7 reduced the ACh-induced response in [Na+]i. However, the following treatment did not affect the basal [Na+]i nor the ACh-induced increase in [Na+]i: (i) addition of dibutyryl cAMP, 8-Br-cGMP, or phorbol 12-myristate 13-acetate, (ii) pretreatment of protein kinase inhibitors, H-89, H-8, H-7 or chelerythrine, (iii) prevention of cytosolic Cl− efflux by the addition of diphenylamine-2-carboxylic acid or, (iv) prevention of cytosolic K+ efflux by the addition of charybdotoxin. The present results suggest that the ACh-induced increase in [Na+]i, mainly responsible for activation of Na-K-Cl cotransport, is mediated by a Ca2+/calmodulin-dependent phosphorylation.

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Authors and Affiliations

  1. Department of Otolaryngology, Tohoku University School of Medicine, 980, Sendai, Japan

    K. Ikeda, D. Wu & T. Takasaka

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  1. K. Ikeda
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  2. D. Wu
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  3. T. Takasaka
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The authors are grateful to Ms. Michiko Yokoyama for her assistance in the preparations of this manuscript. This work was supported by Grant-In-Aid 05807156 and 06807131 to K. Ikeda.

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Ikeda, K., Wu, D. & Takasaka, T. Cellular mechanisms in activation of Na-K-Cl cotransport in nasal gland acinar cells of guinea pigs. J. Membarin Biol. 146, 307–314 (1995). https://doi.org/10.1007/BF00233950

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  • DOI: https://doi.org/10.1007/BF00233950

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