Purinoceptor-stimulated phosphoinositide hydrolysis in Madin-Darby canine kidney (MDCK) cells

Chuen Mao Yang, Yih Jeng Tsai, Shiow Lin Pan, Chuen Tao Tsai, Wen Bin Wu, Chi Tso Chiu, Shu Fen Luo, Jonathan T. Ou

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Abstract

Extracellular nucleotides, acting through P2-purinoceptors, have been implicated in the regulation of ion transport in epithelia, including Madin-Darby canine kidney (MDCK) cells. In this study, experiments were conducted to characterize the P2-purinoceptor subtype on MDCK cells responsible for stimulating inositol phosphate (U?) accumulation using a range of nucleotide analogues. In Ca2+- and Mg2+-free Krebs-Henseleit solution (KHS), ATP, UTP, and ATPγS caused an increase in IP accumulation as a function of concentration with comparable kinetics. The order of potency for the nucleotide analogues was UTP = ATPγS > ATP = 2-chloro ATP (Cl-ATP) >> α,β-methylene ATP (α,β-MeATP) = 2-methylthio ATP (2MeSATP). Selective agonists for P1-, P(2X)- and P(2Y)-purinoceptors, such as N6-cyclopentyl adenosine, AMP, α,β-MeATP, and 2MeSATP, had little effect. Stimulation of MDCK cells with maximally effective concentrations of ATP and UTP showed no additive effect and furthermore, ATP, UTP, and ATPγS induced cross-desensitization of the IP response, suggesting that ATP and UTP act upon a common nucleotide receptor, i.e. a P(2U)-purinoceptor. In Ca2+- and Mg2+-containing KHS, the concentration-response curves of ATP, UTP, and ATPγS were Shifted to the right of those obtained in Ca2+- and Mg2+-free buffer, and asymptotic maxima were not reached, indicating that ATP4- and not MgATP2- or CaATP2- was the active agonist. Pretreatment of MDCK cells with pertussis toxin (PTX) inhibited ATP- and UTP-induced IP accumulation in a concentration-dependent fashion but did not completely abolish the UP accumulation, indicating that a PTX-sensitive G protein was partially involved in the IP response. In conclusion, ATP- and UTP-stimulated IP accumulation in MDCK cells appears to be mediated through the activation of P(2U)-purinoceptors coupled to a G protein that is partially sensitive to PTX. A form of nucleotide uncomplexed with divalent ions such as ATP4- seems to be the preferential agonist form for the purinoceptors on MDCK cells.

Original languageEnglish
Pages (from-to)1-7
Number of pages7
JournalNaunyn-Schmiedeberg's Archives of Pharmacology
Volume356
Issue number1
DOIs
Publication statusPublished - 1997
Externally publishedYes

Fingerprint

Purinergic Receptors
Madin Darby Canine Kidney Cells
Uridine Triphosphate
Phosphatidylinositols
Hydrolysis
Adenosine Triphosphate
Nucleotides
Pertussis Toxin
Purinergic P2 Receptors
GTP-Binding Proteins
Purinergic Agonists
Inositol Phosphates
Ion Transport
Adenosine Monophosphate
Adenosine
Buffers
Epithelium
Ions

Keywords

  • Inositol phosphate accumulation
  • Kidney cells
  • Madin-Darby canine
  • P(2U)-purinoceptor
  • Pertussis toxin
  • Phosphoinositide

ASJC Scopus subject areas

  • Pharmacology

Cite this

Purinoceptor-stimulated phosphoinositide hydrolysis in Madin-Darby canine kidney (MDCK) cells. / Yang, Chuen Mao; Tsai, Yih Jeng; Pan, Shiow Lin; Tsai, Chuen Tao; Wu, Wen Bin; Chiu, Chi Tso; Luo, Shu Fen; Ou, Jonathan T.

In: Naunyn-Schmiedeberg's Archives of Pharmacology, Vol. 356, No. 1, 1997, p. 1-7.

Research output: Contribution to journalArticle

Yang, Chuen Mao ; Tsai, Yih Jeng ; Pan, Shiow Lin ; Tsai, Chuen Tao ; Wu, Wen Bin ; Chiu, Chi Tso ; Luo, Shu Fen ; Ou, Jonathan T. / Purinoceptor-stimulated phosphoinositide hydrolysis in Madin-Darby canine kidney (MDCK) cells. In: Naunyn-Schmiedeberg's Archives of Pharmacology. 1997 ; Vol. 356, No. 1. pp. 1-7.
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AU - Wu, Wen Bin

AU - Chiu, Chi Tso

AU - Luo, Shu Fen

AU - Ou, Jonathan T.

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