Functional assessment of recombinant human alpha(2)-adrenoceptor subtypes with cytosensor microphysiometry

Eur J Pharmacol. 1999 Dec 3;385(2-3):247-53. doi: 10.1016/s0014-2999(99)00715-3.


We applied the Cytosensor Microphysiometry system to study the three human alpha(2)-adrenoceptor subtypes, alpha(2A), alpha(2B) and alpha(2C), expressed in Chinese hamster ovary (CHO) cells, and assessed its potential in the quantitative monitoring of agonist activity. The natural full agonist, (-)-noradrenaline, was used to define agonist efficacy. The imidazole derivative dexmedetomidine was a potent full agonist of all three receptor subtypes. The imidazolines clonidine and UK 14,304 (5-bromo-N-(4, 5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine) appeared to be partial agonists at alpha(2B)-adrenoceptors (E(max) approximately 60% of (-)-noradrenaline) but full agonists at alpha(2A)- and alpha(2C)-adrenoceptors. The responses mediated by all three alpha(2)-adrenoceptor subtypes were partly inhibited by the sodium-hydrogen (Na(+)/H(+)) exchange inhibitor, MIA (5-(N-methyl-N-isobutyl)-amiloride). The agonist responses were totally abolished by pretreatment with pertussis toxin in cells with alpha(2A)- and alpha(2C)-adrenoceptors, and partly abolished in cells with alpha(2B)-adrenoceptors. The residual signal in alpha(2B)-cells was sensitive to the intracellular Ca(2+)chelator, BAPTA (1,2-bis(2-aminophenoxy)ethane-N,N,N,N-tetraacetic acid acetoxymethyl ester). Cholera toxin (which acts on G(s)-proteins) had no effect on the agonist responses. The results suggest that the extracellular acidification responses mediated by all three human alpha(2)-adrenoceptor subtypes are dependent on Na(+)/H(+)exchange and G(i/o) pathways, and that alpha(2B)-adrenoceptors are capable of coupling to another, G(i/o)-independent and Ca(2+)-dependent signaling pathway.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adrenergic alpha-Agonists / metabolism
  • Adrenergic alpha-Agonists / pharmacology
  • Amiloride / analogs & derivatives
  • Amiloride / pharmacology
  • Animals
  • Binding, Competitive / drug effects
  • Biosensing Techniques*
  • Brimonidine Tartrate
  • CHO Cells
  • Cholera Toxin / pharmacology
  • Clonidine / metabolism
  • Clonidine / pharmacology
  • Cricetinae
  • Dexmedetomidine / metabolism
  • Dexmedetomidine / pharmacology
  • Egtazic Acid / analogs & derivatives
  • Egtazic Acid / pharmacology
  • Humans
  • Hydrogen-Ion Concentration
  • Norepinephrine / metabolism
  • Norepinephrine / pharmacology
  • Pertussis Toxin
  • Quinoxalines / metabolism
  • Quinoxalines / pharmacology
  • Radioligand Assay
  • Receptors, Adrenergic, alpha-2 / genetics
  • Receptors, Adrenergic, alpha-2 / metabolism
  • Receptors, Adrenergic, alpha-2 / physiology*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Recombinant Fusion Proteins / physiology
  • Signal Transduction / drug effects
  • Virulence Factors, Bordetella / pharmacology


  • ADRA2A protein, human
  • ADRA2B protein, human
  • ADRA2C protein, human
  • Adrenergic alpha-Agonists
  • Quinoxalines
  • Receptors, Adrenergic, alpha-2
  • Recombinant Fusion Proteins
  • Virulence Factors, Bordetella
  • Brimonidine Tartrate
  • Egtazic Acid
  • Dexmedetomidine
  • Amiloride
  • Cholera Toxin
  • 5-(N-methyl-N-isobutyl)amiloride
  • Pertussis Toxin
  • 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid
  • Clonidine
  • Norepinephrine