title: Pharmacological characterization of purinergic inhibitory neuromuscular transmission in the human colon.
abstract: BACKGROUND
  In the present study, we further characterize the purinergic receptors mediating the inhibitory junction potential (IJP) and smooth muscle relaxation in the human colon using a new, potent and selective agonist (MRS2365), and antagonists (MR2279 and MRS2500) of the P2Y(1) receptor. The P2Y(12) antagonist AR-C66096 was tested as well. Using this pharmacological approach, we tested whether β-nicotinamide adenine dinucleotide (β-NAD) fulfilled the criteria to be considered an inhibitory neurotransmitter in the human colon.
  
  
  METHODS
  We carried out muscle bath and microelectrode experiments on circular strips from the human colon and calcium imaging recordings on HEK293 cells, which constitutively express the human P2Y(1) receptor.
  
  
  RESULTS
  Both the fast component of IJP and non-nitrergic relaxation was concentration-dependently inhibited by MRS2279 and MRS2500. This antagonism was confirmed in HEK293 cells. However, AR-C66096 did not modify either inhibitory response. Adenosine 5'-Ο-2-thiodiphosphate and MRS2365 caused a smooth muscle hyperpolarization and transient inhibition of spontaneous motility that was antagonized by MRS2279 and MRS2500. β-Nicotinamide adenine dinucleotide inhibited the spontaneous motility (IC(50) = 3.3 mmol L(-1) ). Nevertheless, this effect was not antagonized by high concentrations of P2Y(1) antagonists.
  
  
  CONCLUSIONS
  Inhibitory purinergic neuromuscular transmission in the human colon was pharmacologically assessed by the use of new P2Y(1) receptor antagonists MRS2179, MRS2279, and MRS2500. The rank order of potency of the P2Y(1) antagonists is MRS2500 > MRS2279 > MRS2179. We found that β-NAD partially fulfills the criteria to be considered an inhibitory neurotransmitter in the human colon, but the relative contribution of each purine (ATP/ADP vsβ-NAD) requires further studies.
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, (CIBERehd), Instituto de Salud Carlos III, Barcelona, Spain.
authors.forename: D
authors.initials: D
authors.lastname: Gallego
authors.forename: V
authors.initials: V
authors.lastname: Gil
authors.forename: J
authors.initials: J
authors.lastname: Aleu
authors.forename: M
authors.initials: M
authors.lastname: Martinez-Cutillas
authors.forename: P
authors.initials: P
authors.lastname: Clavé
authors.forename: M
authors.initials: M
authors.lastname: Jimenez
mesh_terms: D000244:Adenosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D003106:Colon / Q000033:anatomy & histology / Q000187:drug effects* / Q000502:physiology*; D003838:Deoxyadenine Nucleotides / Q000494:pharmacology; D004594:Electrophysiology; D005769:Gastrointestinal Motility / Q000187:drug effects / Q000502:physiology; D057809:HEK293 Cells; D006801:Humans; D008875:Middle Aged; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology; D009126:Muscle Relaxation / Q000187:drug effects / Q000502:physiology; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology; D009243:NAD / Q000031:analogs & derivatives; D009469:Neuromuscular Junction / Q000187:drug effects* / Q000502:physiology; D058919:Purinergic P2 Receptor Antagonists / Q000494:pharmacology*; D058912:Purinergic P2Y Receptor Agonists / Q000494:pharmacology*; D058472:Receptors, Purinergic P2Y1 / Q000378:metabolism*; D009435:Synaptic Transmission / Q000187:drug effects* / Q000502:physiology
chemical_list: C511689:(N)-methanocarba-2MeSADP; C465253:2-chloro-N6-methyl-(N)-methanocarba-2'-deoxyadenosine-3',5'-bisphosphate; C495475:2-iodo-N(6)-methyl-(N)-methanocarba-2'-deoxyadenosine-3',5'-bisphosphate; C092108:2-propylthio-D-beta,gamma-difluoromethylene ATP; D003838:Deoxyadenine Nucleotides; D058919:Purinergic P2 Receptor Antagonists; D058912:Purinergic P2Y Receptor Agonists; D058472:Receptors, Purinergic P2Y1; D009243:NAD; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate