Receptor-mediated activation of a Cl- current by LPA and S1P in cultured corneal keratocytes

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Abstract

PURPOSE. This study was designed to examine the effects of lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) on Cl- currents (IClLPA) in cultured corneal keratocytes isolated from the corneas of New Zealand White rabbits. METHODS. IClLPA and resting voltages were recorded with the amphotericin perforated-patch technique. Phenotype was determined with antibodies to α-smooth muscle actin. RESULTS. Keratocytes cultured in serum have a phenotype (myofibroblast) and ionic currents similar to those of keratocytes isolated directly from corneas during wound healing. LPA and S1P both activated IClLPA in a dose-dependent manner, and the LPA receptor-specific antagonist dioctyl-glycerol pyrophosphate (DGPP) blocked the LPA response, but not the S1P response. In addition, a relatively inactive form of LPA (LPA 8:0) was relatively ineffective in activating IClLPA. Activation of IClLPA significantly depolarized the cells, and this depolarization was reversed by blocking IClLPA with 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) or 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB). CONCLUSIONS. These results demonstrate that activation of IClLPA by LPA in cultured corneal keratocytes is receptor mediated and that IClLPA can also be activated by S1P. From a functional standpoint, this work confirms that the current, which is typically thought of as purely volume-activated, can be activated through a receptor. In addition, activation of IClLPA results in depolarization of the keratocyte. Finally, this work demonstrates that cultured corneal keratocytes can act as a model for the study of ion channel function in keratocytes during corneal wound healing.

Original languageEnglish (US)
Pages (from-to)3202-3208
Number of pages7
JournalInvestigative Ophthalmology and Visual Science
Volume43
Issue number10
StatePublished - Oct 1 2002
Externally publishedYes

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Corneal Keratocytes
Wound Healing
Cornea
Lysophosphatidic Acid Receptors
Phenotype
Myofibroblasts
Amphotericin B
Ion Channels
Glycerol
Smooth Muscle
lysophosphatidic acid
sphingosine 1-phosphate
Actins
Rabbits
Acids
Antibodies
Serum

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

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title = "Receptor-mediated activation of a Cl- current by LPA and S1P in cultured corneal keratocytes",
abstract = "PURPOSE. This study was designed to examine the effects of lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) on Cl- currents (IClLPA) in cultured corneal keratocytes isolated from the corneas of New Zealand White rabbits. METHODS. IClLPA and resting voltages were recorded with the amphotericin perforated-patch technique. Phenotype was determined with antibodies to α-smooth muscle actin. RESULTS. Keratocytes cultured in serum have a phenotype (myofibroblast) and ionic currents similar to those of keratocytes isolated directly from corneas during wound healing. LPA and S1P both activated IClLPA in a dose-dependent manner, and the LPA receptor-specific antagonist dioctyl-glycerol pyrophosphate (DGPP) blocked the LPA response, but not the S1P response. In addition, a relatively inactive form of LPA (LPA 8:0) was relatively ineffective in activating IClLPA. Activation of IClLPA significantly depolarized the cells, and this depolarization was reversed by blocking IClLPA with 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) or 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB). CONCLUSIONS. These results demonstrate that activation of IClLPA by LPA in cultured corneal keratocytes is receptor mediated and that IClLPA can also be activated by S1P. From a functional standpoint, this work confirms that the current, which is typically thought of as purely volume-activated, can be activated through a receptor. In addition, activation of IClLPA results in depolarization of the keratocyte. Finally, this work demonstrates that cultured corneal keratocytes can act as a model for the study of ion channel function in keratocytes during corneal wound healing.",
author = "Jia Wang and Carbone, {Laura D} and Watsky, {Mitchell Aaron}",
year = "2002",
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pages = "3202--3208",
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T1 - Receptor-mediated activation of a Cl- current by LPA and S1P in cultured corneal keratocytes

AU - Wang, Jia

AU - Carbone, Laura D

AU - Watsky, Mitchell Aaron

PY - 2002/10/1

Y1 - 2002/10/1

N2 - PURPOSE. This study was designed to examine the effects of lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) on Cl- currents (IClLPA) in cultured corneal keratocytes isolated from the corneas of New Zealand White rabbits. METHODS. IClLPA and resting voltages were recorded with the amphotericin perforated-patch technique. Phenotype was determined with antibodies to α-smooth muscle actin. RESULTS. Keratocytes cultured in serum have a phenotype (myofibroblast) and ionic currents similar to those of keratocytes isolated directly from corneas during wound healing. LPA and S1P both activated IClLPA in a dose-dependent manner, and the LPA receptor-specific antagonist dioctyl-glycerol pyrophosphate (DGPP) blocked the LPA response, but not the S1P response. In addition, a relatively inactive form of LPA (LPA 8:0) was relatively ineffective in activating IClLPA. Activation of IClLPA significantly depolarized the cells, and this depolarization was reversed by blocking IClLPA with 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) or 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB). CONCLUSIONS. These results demonstrate that activation of IClLPA by LPA in cultured corneal keratocytes is receptor mediated and that IClLPA can also be activated by S1P. From a functional standpoint, this work confirms that the current, which is typically thought of as purely volume-activated, can be activated through a receptor. In addition, activation of IClLPA results in depolarization of the keratocyte. Finally, this work demonstrates that cultured corneal keratocytes can act as a model for the study of ion channel function in keratocytes during corneal wound healing.

AB - PURPOSE. This study was designed to examine the effects of lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) on Cl- currents (IClLPA) in cultured corneal keratocytes isolated from the corneas of New Zealand White rabbits. METHODS. IClLPA and resting voltages were recorded with the amphotericin perforated-patch technique. Phenotype was determined with antibodies to α-smooth muscle actin. RESULTS. Keratocytes cultured in serum have a phenotype (myofibroblast) and ionic currents similar to those of keratocytes isolated directly from corneas during wound healing. LPA and S1P both activated IClLPA in a dose-dependent manner, and the LPA receptor-specific antagonist dioctyl-glycerol pyrophosphate (DGPP) blocked the LPA response, but not the S1P response. In addition, a relatively inactive form of LPA (LPA 8:0) was relatively ineffective in activating IClLPA. Activation of IClLPA significantly depolarized the cells, and this depolarization was reversed by blocking IClLPA with 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) or 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB). CONCLUSIONS. These results demonstrate that activation of IClLPA by LPA in cultured corneal keratocytes is receptor mediated and that IClLPA can also be activated by S1P. From a functional standpoint, this work confirms that the current, which is typically thought of as purely volume-activated, can be activated through a receptor. In addition, activation of IClLPA results in depolarization of the keratocyte. Finally, this work demonstrates that cultured corneal keratocytes can act as a model for the study of ion channel function in keratocytes during corneal wound healing.

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