Adenine nucleotides inhibit recombinant N-type calcium channels via G protein-coupled mechanisms in HEK 293 cells; involvement of the P2Y13 receptor-type

Br J Pharmacol. 2004 Jan;141(1):141-51. doi: 10.1038/sj.bjp.0705588. Epub 2003 Dec 8.


1. N-type Ca(2+) channel modulation by an endogenous P2Y receptor was investigated by the whole-cell patch-clamp method in HEK 293 cells transfected with the functional rabbit N-type calcium channel. 2. The current responses (I(Ca(N))) to depolarizing voltage steps were depressed by ATP in a concentration-dependent manner. Inclusion of either guanosine 5'-O-(3-thiodiphosphate) or pertussis toxin into the pipette solution as well as a strongly depolarizing prepulse abolished the inhibitory action of ATP. 3. In order to identify the P2Y receptor subtype responsible for this effect, several preferential agonists and antagonists were studied. Whereas the concentration-response curves of ADP and adenosine 5'-O-(2-thiodiphosphate) indicated a higher potency of these agonists than that of ATP, alpha,beta-methylene ATP, UTP and UDP were considerably less active. The effect of ATP was abolished by the P2Y receptor antagonists suramin and N(6)-(2-methylthioethyl)-2-(3,3,3-trifluoropropylthio)-beta,gamma-dichloromethylene-ATP, but not by pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid, 2'deoxy-N(6)-methyladenosine-3',5'-diphosphate or 2-methylthio AMP. 4. Using reverse transcription and polymerase chain reaction, mRNA for the P2Y(1), P2Y(4), P2Y(6), P2Y(11) and P2Y(13) receptor subtypes, but not the P2Y(2), and P2Y(12) subtypes, was detected in HEK 293 cells. 5. Immunocytochemistry confirmed the presence of P2Y(1), and to a minor extent that of P2Y(4), but not of P2Y(2) receptors. 6. Hence, it is tempting to speculate that P2Y(13) receptors may inhibit N-type Ca(2+) channels via the betagamma subunits of the activated G(i) protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenine Nucleotides / pharmacology*
  • Adenosine Diphosphate / analogs & derivatives*
  • Adenosine Diphosphate / pharmacology
  • Adenosine Triphosphate / pharmacology
  • Animals
  • Calcium Channels, N-Type / drug effects*
  • Calcium Signaling / drug effects
  • Calcium Signaling / physiology
  • Cell Line
  • Dose-Response Relationship, Drug
  • GTP-Binding Proteins / drug effects
  • GTP-Binding Proteins / physiology
  • Humans
  • Membrane Proteins / drug effects
  • Membrane Proteins / physiology
  • Patch-Clamp Techniques / methods
  • Purinergic P2 Receptor Agonists
  • Purinergic P2 Receptor Antagonists
  • Rabbits
  • Receptors, G-Protein-Coupled / drug effects
  • Receptors, G-Protein-Coupled / physiology*
  • Receptors, Purinergic P2 / drug effects
  • Receptors, Purinergic P2 / physiology*
  • Receptors, Purinergic P2Y12
  • Recombinant Proteins / antagonists & inhibitors*
  • Recombinant Proteins / biosynthesis
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Thionucleotides / pharmacology
  • Transfection / methods


  • Adenine Nucleotides
  • Calcium Channels, N-Type
  • Membrane Proteins
  • P2RY12 protein, human
  • P2RY13 protein, human
  • Purinergic P2 Receptor Agonists
  • Purinergic P2 Receptor Antagonists
  • Receptors, G-Protein-Coupled
  • Receptors, Purinergic P2
  • Receptors, Purinergic P2Y12
  • Recombinant Proteins
  • Thionucleotides
  • adenosine 5'-O-(1-thiodiphosphate)
  • Adenosine Diphosphate
  • Adenosine Triphosphate
  • GTP-Binding Proteins