Purinergic neurogenic intestinal mucosal secretion.

摘要

Electric stimulation of interganglionic fiber tracts in the submucosal plexus of guinea-pig small intestine evoked synaptic responses that included slow excitatory postsynaptic potentials (slow EPSPs). The submucosal slow EPSPs were subdivided into three categories consisting of "pure-purinergic", "partial-purinergic" and "non-purinergic" according to their sensitivity to a specific P2Y1 receptor antagonist, MRS2179 (10 muM). Most of the purinergic slow EPSPs were found in submucosal secretomotor neurons associated with stimulus-evoked slow IPSPs, which were known electrophysiological markers for VIPergic secretomotor neurons. Application of known purinergic agonists including ATP, 2-methio-ATP etc evoked slow EPSP-like responses in the submucosal neurons with a potency order consistent with that of P2Y1 receptors in other systems. The excitatory action of ATP was suppressed by MRS2179 in a competitive manner. Pharmacological analysis suggested that the phospholipase C → IP 3 → calmodulin kinase/protein kinase C post receptor signal transduction cascade mediated the purinergic P2Y1 receptor-evoked depolarizing responses in submucosal secretomotor neurons. Neurons that supplied P2Y 1 purinergic slow excitatory synaptic input to the secretomotor neurons in the submucosal plexus were found in the myenteric and submucosal plexuses and in prevertebral sympathetic ganglia. Full-length guinea-pig P2Y1 receptor cDNA cloned from the submucosal plexus was used to transfect HEK293 cells. ATP or its analogs induced intracellular calcium release in transfected HEK293 cells, which was suppressed by MRS2179. ATP evoked neurogenic mucosal secretion by activating the P2Y1 receptor in guinea-pig small intestine. The presence of MRS2179 in the Ussing chambers also suppressed the secretory responses evoked by electrical field stimulation.; The results of the project provide for the first evidence for establishing ATP as a neurotransmitter for slow synaptic excitation in the submucosal plexus of guinea-pig small intestine. The signal transduction pathway for purinergic slow synaptic excitation includes activation of phospholipase C and intracellular IP3 receptors. Protein kinase C and Calmodulin-dependent protein kinases are also involved. Ussing chamber experiment revealed the role of the P2Y1 receptor in neurogenic mucosal secretion into the gut lumen. Identification of the functional purinergic component of enteric neurotransmission provides a basis for therapeutic drug development by targeting the P2Y 1 receptor and purine metabolic pathways.