| Pain is a kind of unpleasant subjective feeling,which not only makes patients suffer physical pain,but also brings mental torture and affects their normal physiological activities.In order to reduce the pain and improve their quality of life,clinical analgesics are often used.However,most of these analgesics are opioids,which are addictive and have certain toxic and side effects.Therefore,it is urgent to develop a new analgesic with small toxic and side effects.Ericaceae is a family of plants with various medicinal values.Rhododendron amura is often used for relieving headache,attenuating bruises,and anti-rheumatoid,etc.It was found that diterpenoids from Rhododendron family are pharmacological ingredients and have anti-inflammatory and analgesic effects.Previous studies showed that the three diterpenoids(ZJF1-34-1,ZGJ2-12-2,ZGJ4-22-3)extracted from the family Azalea have inhibitory effects on the acute visceral pain induced by glacial acetic acid in mice.Therefore,there may be effective analgesic components in these three monomers,which is of great help to the research and development of new national analgesic drugs.The aim of this study was to screen out the effective analgesic constituents of these three diterpenes in Ericaceae and explore their analgesic mechanisms.In the behavioral experiment,the monomer ZJF1-34-1 with obvious analgesic effect was successfully screened out by using three different acute pain models.Intraperitoneal injection of 5mg/Kg of the monomer can effectively alleviate these three kinds of pain response including heat pain induced by hot plate,inflammatory pain induced by formalin,and acute pain induced by capsaicin.By reducing the concentration of ZJF1-34-1,it was found that ZJF1-34-1 of low concentration had no distinct effect in hot plate test,but had obvious analgesic effect in formalin and capsaicin test,and the analgesic effect decreased with the decrease of the concentration.In order to verify the behavioral experimental results of the three monomers and explore the analgesic mechanism of ZJF1-34-1 in the central nervous system.First,we took cerebral cortical neurons from neonatal mice cultured in vitro and incubated them with three monomer solutions respectively at different concentrations for 60 min.Then the MIPSCs of the cultured neurons were recorded by Whole-Cell patch-Clamp.Results showed that only two monomers,ZJF1-34-1 and ZGJ4-22-3,could enhance the spontaneous inhibitory synaptic response of cerebral cortical neurons.In the concentration and time dependence experiments of ZJF1-34-1 and ZGJ4-22-3,it was found that the two monomers had significant effect on the spontaneous inhibitory synaptic response of the cultured neurons only when the incubation concentration was20 μm and the incubation time was more than 60 min.After incubated with the two kinds of monomers above,m EPSCs of the neurons were recorded.It was found that the two monomers did not change the frequency and amplitude of the m EPSCs.In conclusion,this study demonstrated that ZJF1-34-1 and ZGJ4-22-3 could only increase the spontaneous inhibitory synaptic response of the neurons,but had no effect on the spontaneous excitatory synaptic response.In order to explore the possible analgesic mechanism of ZJF1-34-1,we extracted Dorsal root ganglia(DRG)neurons from rats and found that ZJF1-34-1 could effectively reduce the peak value of sodium channel current activated by DRG cells.The results suggests that ZJF1-34-1 may block excitatory signal transduction by inhibiting the sodium channel current of the peripheral neurons and finally achieve analgesic effect.The analgesic effect of diterpene ZJF1-34-1 in the family of Rhododendron was identified in this study,and its analgesic effect was verified in three different acute pain models.Furthermore,it was found that both the central nervous system and the peripheral nervous system were involved in this analgesic mechanism. |
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