Study Of The Pain Actions Mechanisms Of Peripheral Harmful Stimulating Mediated By The P2X₃ Receptors

Objective: P2X₃ receptor is a new type of pain receptor. The previous study shows that P2X₃ receptors exist on the sensory neuron. It possibly plays a leading role in transmitting t pain information. The object is to inject the agonists of P2X₃ receptors to observe the harmful reaction of the rats and measure the expression of the positive cells of P2X₃ receptors , of NK1 and the glutamic acid (NDMR₁) receptor on spinal dorsal horn . By intrathecally injecting antagons of both SP and glutamic acid , we progrssively observe the harmful reaction of the rats and measure the expression amount of the positive cells of NDMR₁ and NK₁ receptor of neuron cell and astrocyte on spinal dorsal horn in order to explore the role played by the P2X₃ receptors to mediate the pain reaction caused by the peripheral harmful stimulating.Method: 1 The healthy male adult rats weighting 250-300g, were randomly divided into five groups. (n=6 each).A group:blank control group(injecting 0.9%N.S), B group: the agonists of P2X₃ (ATP), C group: the potent agonists of P2X₃ (a,βmeATP), D group: the potent agonists and extraordinary antagons of P2X₃ (a,β-meATP+TNP-ATP) E group: the agonists and antagons of P2X₃ (ATP+TNP-ATP). All drugs were concocted by 0.9%N.S and injected into the left foot bottom of rats by 26 bore needle. The number of withdrawing hindpaws caused by pain has been observed for one hour. Every observing unit is five minutes. The DRG of rat is urgently dissected to extract. The expression of P2X3 receptors of DRG and the expression of both NK1 and NDMR1 on spinal dorsal horn have measured. 2 The health adult rats weighting 250-300g were randomly divided into five groups (n=6 each). A blank control group: (injecting 0.9%N.S); B group: TNP-ATP ; C group: ( L 701, 3142); D group: (S3144); E group: (L701, 3142+S3144). After a,β-meATP was injected into the left foot bottoms of rats, the above drugs were intrathecally injected including DMSO 10ul. The way of observing the harmful reaction of animals is the same as the experiment 1. The expression of both NK1 and NDMR1 on spinal dorsal horn has been measured by immunohistochemistry.Results: 1 (1)The change of behavior of rats : No harmful reaction appears in group A. But the apparently harmful reaction exists in group B and group C. The harmful reactions appears after five minutes, the peak of harmful reaction appears after 20 minutes such as bitting tail, flinching hindpaws and so on. The distinguishable reaction appears in the group C, There are significant difference between group B and group C(P<0.01). The harmful reaction and behavior are inhibited after the application of TNP–ATP . There is significant difference between group B and group E(P<0.01). the same results appear in group C and group D.(2) The P2X3 receptors are expressed on the cell body of both small and middle cells of DRG. The expression of P2X3 receptors highly increased in group B and group C, and decreased in group D and group E . There is difference between group A and group B (P < 0.05). The difference between group A and group C is significant(P<0.01) , the same as group B and group E; group C and group D.(3)The expression of NK1 and NDMR1 on spinal dorsal horn. The C group holds the highest expression percentage among the four groups, There is difference between group A and group B(P < 0.05). The difference between group A and group C is significant (P<0.01), the same as group B and group E; group C and group D. 2 (1) The change of behavior of rats: The harmful reaction and behavior appear in both A and B group such biting tail, liping foot, flinching hindpaws and so on. But there is no difference between group A and B(P>0.05). Compared with the A group, the harmful reaction of C and D group are inhibited. There is difference between A and C or D(P<0.05). The difference between group C and group E is significant (P<0.01), the same as group D and group E; group A and group E.(2) The expression of NK1 and NDMR1 on spinal dorsal horn: There is no difference between group A and B(P>0.05), There is difference between A and C or D(P<0.05). The difference between group C and group E is significant (P<0.01), the same as group D and group E; group A and group E.Conclusion: 1 P2X3 receptors were expressed on the small and middle-size cells of DRG .2 the pain reactions of rats mediated by P2X3 receptors of DRG have more to do with the receptors of both NK₁ and NDMR₁ .3 the activation of P2X3 receptors of DRG makes SP and glutamic acid of of spinal dorsal horn release to combine their corresponding receptors to play a leading role in transferring the pain signaling and transmitting the neurotransmitter of pain. 4.What's more, astrocyte of spinal dorsal horn is possibly involved in the course.