| The P2X3 receptor is a trimeric ATP-gated ion channel that is highly selectively expressed in nociceptive sensory neurons and plays an important role in neuropathic pain,inflammatory pain,and visceral pain.Pain is one of the most common clinical symptoms.The commonly used analgesics in clinical practice are mainly opioids and nonsteroidal drugs,but the toxic and side effects of these drugs have not been improved very well.Therefore,the development of new,high-efficiency and low-toxic pain treatment drugs targeting P2X3 receptor has gradually become one of the hot spots in the field of pain research.Peptides have received extensive attention because of their low toxicity,high selectivity,high activity,and low immunogenicity.Spider venom contains polypeptide toxins that perform various functions,and some polypeptides have high selectivity and activity for multiple ion channels and receptors.The peptide toxins found in spider venoms that act on ion channels can be used as tools and reagents for studying the structure and function of ion channels,as well as candidate molecules for drug development.As a tool for studying ion channels,spider peptide toxins are more used in sodium channels,potassium channels,and calcium channels,but less for P2X3 receptors.Most of the P2X3 receptor inhibitors found so far are small molecule compounds,lack Peptide inhibitor.EK36 is a peptide toxin derived from the spider Alopecosa,with 36 amino acids and 4 pairs of disulfide bonds.In this study,the polypeptide and a series of mutants were obtained by prokaryotic expression,and the activity was identified.EK36 has no inhibitory effect on P2X2,P2X4 and P2X7 receptors,but it can selectively inhibit P2X3 receptor current.1?M can completely inhibit P2X3 receptor current.Its half effective concentration(IC50)for P2X3 receptor is 108.3 n M.In order to explore the role of P2X3 receptors in pain treatment,this study constructed a mouse model of acute pain,chronic inflammatory pain and neuropathic pain to explore the therapeutic effect of EK36 on pain.The results show that EK36 can reduce acute pain,chronic inflammatory pain and neuropathic pain in a dose-dependent manner.In the formalin pain model,EK36 has a good analgesic effect in both phase I and phase II pain.In the phase I pain period,the average foot licking time of the 2 mg / kg,4 mg / kg,and 8 mg / kg EK36 groups decreased by 36.2%,54.5%,and 61.3%,respectively.During the phase II pain period,the average foot licking time of mice in the 2 mg / kg EK36 and 4 mg / kg EK36 groups decreased by 43.7% and 52.1%,respectively,and the mice in the 8 mg / kg EK36 group showed better analgesic effects.The average foot licking time of mice was reduced by 67.4%,and it had an analgesic effect comparable to 1 mg / kg morphine.In the acetic acid writhing mouse pain model,the number of writhing in the EK36 group of 1 mg / kg,2 mg / kg,4 mg / kg,and 8 mg / kg mice was reduced by 24.9%,32.0%,45.3%,and 59.7%,respectively.The 1 mg / kg twisting frequency of the drug was reduced by 58.6%,and it showed a comparable analgesic effect to 8 mg / kg EK36.In addition,in the hot plate mouse pain model,8 mg / kg EK36 showed better analgesic effect than 1 mg / kg morphine.In the SNI model of partial sciatic nerve ligation pain,compared with before treatment,the pain threshold of mice increased significantly after treatment with EK36.In the CFA-induced chronic inflammatory mouse pain model,the analgesic effect of 8 mg / kg EK36 is better than 2 mg / kg morphine.In summary,the selective inhibitor of P2X3 receptor EK36 can effectively reduce chronic inflammatory pain,acute pain and neuropathic pain,and is the most effective in treating inflammatory pain.Therefore,EK36 may provide tools and reagents for the study of the pharmacological properties of P2X3 receptors,and also provide new strategies for the pathogenesis of pain associated with P2X3 receptors and the treatment of pain. |
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