Alpha adrenergic receptor subtypes involved in prostaglandin synthesis are coupled to Ca++ channels through a pertussis toxin-sensitive guanine nucleotide-binding protein

C. Nebigil, Kafait Malik

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49 Citations (Scopus)

Abstract

Previously, we have shown that alpha-2C and alpha-1A adrenergic receptors (AR) stimulate prostacyclin (PGI2) synthesis through a pertussis toxin- sensitive guanine nucleotide-binding protein (G protein) in vascular smooth muscle cells (VSMC). The purpose of this study was to assess the role of Ca++ in PGI2 production elicited by alpha-AR activation and to investigate the modulation of the Ca++ channel by G proteins coupled to these alpha-AR in VSMC. PGI2 was measured as immunoreactive 6-keto-PGF(1α) by radioimmunoassay and cytosolic calcium ([Ca++](i)) by spectrofluorometry using fura-2. Norepinephrine, methoxamine and UK-14304 enhanced 6-keto- PGF(1α) production and [Ca++](i), which was inhibited by depletion of extracellular Ca++ and by Ca++ channel antagonists (verapamil, nifedipine and PN 200-110). Moreover, the Ca++ channel activator Bay K 8644 increased 6-keto-PGF(1α) production in a nifedipine-sensitive manner, indicating the involvement of dihydropyridine-sensitive Ca++ channels in VSMC. Pertussis toxin inhibited AR agonist-induced 6-keto-PGF(1α) production and the increase in [Ca++](i). Alpha AR agonists increase Ca++ influx in the presence of guanosine 5'-O-(2-thiodiphosphate) (GTP-γ-S), and this effect was blocked in the presence of guanine 5'-O-(2-thiodiphosphate) (GDP-β-S) and antiserum against G(iα1-2) protein in reversibly permeabilized cells with β-escin. VSMC of rabbit aortae contain a G protein(s) that was recognized by G(iα1-2) but not G(iα3) or G(o) antibodies at 1:200 dilution. The calmodulin inhibitor W-7 blocked AR agonist and Bay K 8644-stimulated 6- keto-PGF(1α) production. The phospholipase A2 inhibitors 7,7- dimethyleicosadienoic acid and oleoyloxyethyl phosphocholine but not phospholipase C inhibitor U-73122 reduced 6-keto-PGF(1α) production in VSMC. These data suggest that a pertussis toxin-sensitive G protein, probably G(iα1-2), coupled to alpha AR regulates Ca++ influx, which, in turn, by interacting with calmodulin, increases phospholipase A2 activity to release arachidonic acid for PGI2 synthesis in VSMC of rabbit aortae.

Original languageEnglish (US)
Pages (from-to)1113-1124
Number of pages12
JournalJournal of Pharmacology and Experimental Therapeutics
Volume266
Issue number2
StatePublished - Jan 1 1993

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Receptors, Adrenergic, alpha
Guanine Nucleotides
Pertussis Toxin
Prostaglandins F
Vascular Smooth Muscle
Prostaglandins
Smooth Muscle Myocytes
Epoprostenol
Carrier Proteins
GTP-Binding Proteins
3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester
Adrenergic Agonists
Calmodulin
Nifedipine
Aorta
Escin
Phospholipase A2 Inhibitors
Adrenergic alpha-1 Receptors
Isradipine
Adrenergic alpha-Agonists

All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Pharmacology

Cite this

@article{9b67f158f8884b03b65ec695c274d8a3,
title = "Alpha adrenergic receptor subtypes involved in prostaglandin synthesis are coupled to Ca++ channels through a pertussis toxin-sensitive guanine nucleotide-binding protein",
abstract = "Previously, we have shown that alpha-2C and alpha-1A adrenergic receptors (AR) stimulate prostacyclin (PGI2) synthesis through a pertussis toxin- sensitive guanine nucleotide-binding protein (G protein) in vascular smooth muscle cells (VSMC). The purpose of this study was to assess the role of Ca++ in PGI2 production elicited by alpha-AR activation and to investigate the modulation of the Ca++ channel by G proteins coupled to these alpha-AR in VSMC. PGI2 was measured as immunoreactive 6-keto-PGF(1α) by radioimmunoassay and cytosolic calcium ([Ca++](i)) by spectrofluorometry using fura-2. Norepinephrine, methoxamine and UK-14304 enhanced 6-keto- PGF(1α) production and [Ca++](i), which was inhibited by depletion of extracellular Ca++ and by Ca++ channel antagonists (verapamil, nifedipine and PN 200-110). Moreover, the Ca++ channel activator Bay K 8644 increased 6-keto-PGF(1α) production in a nifedipine-sensitive manner, indicating the involvement of dihydropyridine-sensitive Ca++ channels in VSMC. Pertussis toxin inhibited AR agonist-induced 6-keto-PGF(1α) production and the increase in [Ca++](i). Alpha AR agonists increase Ca++ influx in the presence of guanosine 5'-O-(2-thiodiphosphate) (GTP-γ-S), and this effect was blocked in the presence of guanine 5'-O-(2-thiodiphosphate) (GDP-β-S) and antiserum against G(iα1-2) protein in reversibly permeabilized cells with β-escin. VSMC of rabbit aortae contain a G protein(s) that was recognized by G(iα1-2) but not G(iα3) or G(o) antibodies at 1:200 dilution. The calmodulin inhibitor W-7 blocked AR agonist and Bay K 8644-stimulated 6- keto-PGF(1α) production. The phospholipase A2 inhibitors 7,7- dimethyleicosadienoic acid and oleoyloxyethyl phosphocholine but not phospholipase C inhibitor U-73122 reduced 6-keto-PGF(1α) production in VSMC. These data suggest that a pertussis toxin-sensitive G protein, probably G(iα1-2), coupled to alpha AR regulates Ca++ influx, which, in turn, by interacting with calmodulin, increases phospholipase A2 activity to release arachidonic acid for PGI2 synthesis in VSMC of rabbit aortae.",
author = "C. Nebigil and Kafait Malik",
year = "1993",
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language = "English (US)",
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journal = "Journal of Pharmacology and Experimental Therapeutics",
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T1 - Alpha adrenergic receptor subtypes involved in prostaglandin synthesis are coupled to Ca++ channels through a pertussis toxin-sensitive guanine nucleotide-binding protein

AU - Nebigil, C.

AU - Malik, Kafait

PY - 1993/1/1

Y1 - 1993/1/1

N2 - Previously, we have shown that alpha-2C and alpha-1A adrenergic receptors (AR) stimulate prostacyclin (PGI2) synthesis through a pertussis toxin- sensitive guanine nucleotide-binding protein (G protein) in vascular smooth muscle cells (VSMC). The purpose of this study was to assess the role of Ca++ in PGI2 production elicited by alpha-AR activation and to investigate the modulation of the Ca++ channel by G proteins coupled to these alpha-AR in VSMC. PGI2 was measured as immunoreactive 6-keto-PGF(1α) by radioimmunoassay and cytosolic calcium ([Ca++](i)) by spectrofluorometry using fura-2. Norepinephrine, methoxamine and UK-14304 enhanced 6-keto- PGF(1α) production and [Ca++](i), which was inhibited by depletion of extracellular Ca++ and by Ca++ channel antagonists (verapamil, nifedipine and PN 200-110). Moreover, the Ca++ channel activator Bay K 8644 increased 6-keto-PGF(1α) production in a nifedipine-sensitive manner, indicating the involvement of dihydropyridine-sensitive Ca++ channels in VSMC. Pertussis toxin inhibited AR agonist-induced 6-keto-PGF(1α) production and the increase in [Ca++](i). Alpha AR agonists increase Ca++ influx in the presence of guanosine 5'-O-(2-thiodiphosphate) (GTP-γ-S), and this effect was blocked in the presence of guanine 5'-O-(2-thiodiphosphate) (GDP-β-S) and antiserum against G(iα1-2) protein in reversibly permeabilized cells with β-escin. VSMC of rabbit aortae contain a G protein(s) that was recognized by G(iα1-2) but not G(iα3) or G(o) antibodies at 1:200 dilution. The calmodulin inhibitor W-7 blocked AR agonist and Bay K 8644-stimulated 6- keto-PGF(1α) production. The phospholipase A2 inhibitors 7,7- dimethyleicosadienoic acid and oleoyloxyethyl phosphocholine but not phospholipase C inhibitor U-73122 reduced 6-keto-PGF(1α) production in VSMC. These data suggest that a pertussis toxin-sensitive G protein, probably G(iα1-2), coupled to alpha AR regulates Ca++ influx, which, in turn, by interacting with calmodulin, increases phospholipase A2 activity to release arachidonic acid for PGI2 synthesis in VSMC of rabbit aortae.

AB - Previously, we have shown that alpha-2C and alpha-1A adrenergic receptors (AR) stimulate prostacyclin (PGI2) synthesis through a pertussis toxin- sensitive guanine nucleotide-binding protein (G protein) in vascular smooth muscle cells (VSMC). The purpose of this study was to assess the role of Ca++ in PGI2 production elicited by alpha-AR activation and to investigate the modulation of the Ca++ channel by G proteins coupled to these alpha-AR in VSMC. PGI2 was measured as immunoreactive 6-keto-PGF(1α) by radioimmunoassay and cytosolic calcium ([Ca++](i)) by spectrofluorometry using fura-2. Norepinephrine, methoxamine and UK-14304 enhanced 6-keto- PGF(1α) production and [Ca++](i), which was inhibited by depletion of extracellular Ca++ and by Ca++ channel antagonists (verapamil, nifedipine and PN 200-110). Moreover, the Ca++ channel activator Bay K 8644 increased 6-keto-PGF(1α) production in a nifedipine-sensitive manner, indicating the involvement of dihydropyridine-sensitive Ca++ channels in VSMC. Pertussis toxin inhibited AR agonist-induced 6-keto-PGF(1α) production and the increase in [Ca++](i). Alpha AR agonists increase Ca++ influx in the presence of guanosine 5'-O-(2-thiodiphosphate) (GTP-γ-S), and this effect was blocked in the presence of guanine 5'-O-(2-thiodiphosphate) (GDP-β-S) and antiserum against G(iα1-2) protein in reversibly permeabilized cells with β-escin. VSMC of rabbit aortae contain a G protein(s) that was recognized by G(iα1-2) but not G(iα3) or G(o) antibodies at 1:200 dilution. The calmodulin inhibitor W-7 blocked AR agonist and Bay K 8644-stimulated 6- keto-PGF(1α) production. The phospholipase A2 inhibitors 7,7- dimethyleicosadienoic acid and oleoyloxyethyl phosphocholine but not phospholipase C inhibitor U-73122 reduced 6-keto-PGF(1α) production in VSMC. These data suggest that a pertussis toxin-sensitive G protein, probably G(iα1-2), coupled to alpha AR regulates Ca++ influx, which, in turn, by interacting with calmodulin, increases phospholipase A2 activity to release arachidonic acid for PGI2 synthesis in VSMC of rabbit aortae.

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