Diadenosine tetraphosphate activates P2Y1 receptors that cause smooth muscle relaxation in the mouse colon

P2Y(1) receptors play an essential role in inhibitory neuromuscular transmission in the gastrointestinal tract. The signalling pathway involves the opening of small conductance calcium activated potassium-channels (K(ca)2 family) that results in smooth muscle hyperpolarization and relaxation. Inorganic polyphosphates and dinucleotidic polyphosphates are putative neurotransmitters that potentially act on P2Y(1) receptors. A pharmacological approach using both orthosteric (MRS2500) and allosteric (BPTU) blockers of the P2Y(1) receptor and openers (CyPPA) and blockers (apamin) of K(ca)2 channels was used to pharmacologically characterise the effect of these neurotransmitters. Organ bath and microelectrodes were used to evaluate the effect of P1,P4-Di (adenosine-5') tetraphosphate ammonium salt (Ap(4)A), inorganic polyphosphates (PolyP) and CyPPA on spontaneous contractions and membrane potential of mouse colonic smooth muscle cells. PolyP neither modified contractions nor membrane potential. In contrast, Ap(4)A caused a concentration-dependent inhibition of spontaneous contractions reaching a maximum effect at 100 mu M Ap(4)A response was antagonised by MRS2500 (1 mu M), BPTU (3 mu M) and apamin (1 mu M). CyPPA (10 mu M) inhibited spontaneous contractions and this response was antagonised by apamin but it was not affected by MRS2500 or BPTU. Both CyPPA and Ap(4)A caused smooth muscle hyperpolarization that was blocked by apamin and MRS2500 respectively. We conclude that Ap(4)A but not PolyP activates P2Y(1) receptors causing smooth muscle hyperpolarization and relaxation. Ap(4)A signalling causes activation of K(ca)2 channels through activation of P2Y(1) receptors. In contrast, CyPPA acts directly on K(ca)2 channels. Further studies are needed to evaluate if dinucleotidic polyphosphates are released from inhibitory motor neurons.