Cloning and pharmacological characterization of the equine adenosine A 2A receptor

A potential therapeutic target for the treatment of equine endotoxemia

C. I. Brandon, M. Vandenplas, H. Dookwah, J. Linden, Thomas F. Murray

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

The aim of the current study was to clone the equine adenosine A 2A receptor gene and to establish a heterologous expression system to ascertain its pharmacologic profile via radioligand binding and functional assays. An eA2A-R expression construct was generated by ligation of the eA2A cDNA into the pcDNA3.1 expression vector, and stably transfected into human embryonic kidney cells (HEK). Binding assays identified those clones expressing the eA2A-R, and equilibrium saturation isotherm experiments were utilized to determine dissociation constants (K D), and receptor densities (Bmax) of selected clones. Equilibrium competition binding revealed a rank order of agonist potency of ATL > CV-1808 > NECA > 2-CADO > CGS21680, and a rank order of antagonist potency as ZM241385 > 8-phenyltheophylline > p-sulfophenyltheophylline > caffeine. Furthermore, adenylate cyclase assays using selective A 2A-R agonists revealed that the eA2A-R functionally coupled to Gαs as indicated by an increase in intracellular [3H]cAMP upon receptor activation. Finally, NF-κB reporter gene assays revealed a CGS21680 concentration-dependent inhibition of NF-κB activity. These results indicate that the heterologously expressed eA 2A-R has a pharmacological profile similar to that of other mammalian A2A receptors and thus can be utilized for further characterization of the eA2A-R to ascertain whether it can serve as a suitable pharmacological target for equine inflammatory disease.

Original languageEnglish
Pages (from-to)243-253
Number of pages11
JournalJournal of Veterinary Pharmacology and Therapeutics
Volume29
Issue number4
DOIs
StatePublished - Aug 2006
Externally publishedYes

Fingerprint

Adenosine A2A Receptors
endotoxemia
Endotoxemia
adenosine
Horses
Organism Cloning
molecular cloning
Clone Cells
Pharmacology
horses
therapeutics
receptors
assays
clones
Cyclic AMP Receptors
Adenosine-5'-(N-ethylcarboxamide)
Horse Diseases
agonists
Caffeine
Reporter Genes

All Science Journal Classification (ASJC) codes

  • Pharmacology
  • veterinary(all)

Cite this

Cloning and pharmacological characterization of the equine adenosine A 2A receptor : A potential therapeutic target for the treatment of equine endotoxemia. / Brandon, C. I.; Vandenplas, M.; Dookwah, H.; Linden, J.; Murray, Thomas F.

In: Journal of Veterinary Pharmacology and Therapeutics, Vol. 29, No. 4, 08.2006, p. 243-253.

Research output: Contribution to journalArticle

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