The Effects Of Different Ratios Of Chuanxiong And Baiji On The Pain Threshold Of Migraine Model And C-fos And PCREB In The Trigeminal Vascular Pathway

Research backgroundMigraine is a common and recurrent diseases with high prevalence rate,and social economic burden,and it is the seventh reason why people lost their life due to disability in the world.The most common subtype of migraine is migraine without aura,the main symptoms are headache,nausea,vomiting,photophobia and tone of fear.At present,the trigeminalvascular reflex theory is the mainstream theory of migraine pathogenesis.The theory thinks that the pain conduction pathway is divided into three parts.In this pathway,the primary neuron is trigeminal ganglion,the second neuron is trigeminocervical complex,and the third neuron is thalamus.C-fos proto oncogene is an immediate early gene,and the expression of FOS protein or c-fos is increased in the ascending region of central nervous system when pain stimulation occurs.c-AMP response element binding protein(CREB)mediates the transcriptional regulation of multiple genes.CREB phosphorylation(p-CREB)leads to the initiation of early gene c-fos while c-fos gene can be used as a biomarker of nociceptive conduction pathway in brainstem and spinal cord,and is widely used in migraine research.In recent years,Chinese medicine has achieved good clinical effects in treating migraine,and has fewer side effects compared with western medicine.Ligusticum Chuanxiong has been widely used in treatment of migraine and has a good effect on migraine.As the saying goes,the secret of Chinese medicine is dosage,the large dose of Ligustrazine is widely used in headache,and often achieves well effects.However,few patients used Chuanxiong alone to treat headache,and large dose of Ligusticum chuanxiong could damage yin.Therefore,large dose use of Ligusticum Chuanxiong should be combined to restrict dryness.Radix Paeoniae Alba is bitter,acid and slightly cold,and compared with Ligusticum Chuanxiong in four-ingredient decoction.Radix Paeoniae Alba is also used int headache treatment,someone concluded that the average dosage of Paeonia lactiflora in treating headache is about 18 grams.ObjectiveIn this study,Radix Paeoniae Alba is used to restrict Ligusticum Chuanxiong.The base of compatibility of Ligusticum Chuanxiong and Radix Paeoniae Alba is 1:1.Then we observed effects of Ligusticum Chuanxiong and Radix Paeoniae Alba with different compatibility on hyperalgesia and c-fos and pCREB in trigeminal vascular pathway of migraine model induced by nitroglycerin(NTG),and explorer the curative effect and mechanism,then support the clinical application.MethodSeventy-two male SPF SD rats were randomly divided into nine groups,include control group,migraine group,Ligusticum Chuanxiong and Radix Paeoniae Alba with different proportion(90:30,60:30,30:30,45:15,30:15,15:15)group(A,B,C,D,E,F),and sumatriptan group.Each decoction group were added 8 times amount of water,and decocted for three times,then separately decompressed and concentrated to 1.2g/ml,0.9g/ml,0.6g/ml,0.6g/ml,0.45g/ml,0.3g/ml.The control group was injected subcutaneously saline solution(2ml/kg)for three times,once a week,and began intragastric administration of saline solution(10ml/kg/d)after the second injection.The model group was injected subcutaneously nitroglycerin solution(10mg/kg,5mg/ml)for three times,once a week,and began intragastric administration of saline solution(10mg/kg,5mg/ml)for a week after the second injection.The sumatriptan group began intragastric administration of sumatriptan solution(6mg/kg/d)for a week after the second injection of nitroglycerin solution(10mg/kg,5mg/ml),and treated in a same way with migraine group in other dispose.The Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)began intragastric administration of Ligusticum Chuanxiong and Radix Paeoniae Alba solution(12g/kg/d,9g/kg/d,6g/kg/d,6g/kg/d,4.5g/kg/d,3g/kg/d)for a week after the second injection of nitroglycerin solution(10mg/kg,5mg/ml),and treated in a same way with migraine group in other dispose.The dosage of Ligusticum Chuanxiong and Radix Paeoniae Alba of rat was six times as large as that in adult.Measure the pain threshold two hours later after last injection.Then the rats were perfusion fixed and their brain was collected for the immunohistochemistry staining.Observing the expression of c-fos,CREB and pCREB in brain slices under microscope,then count it with Image Pro Plus 6.0 and analysis statistically.Resultpain thresholdThe pain threshold of migraine group were decreased compared with control group.The mechanical withdrawal threshold of Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)and sumatriptan group was increased compared with migraine group,and B,C,D group and sumatriptan group increased significantly(p<0.05),besides,C group increased most(p<0.01).The heat pain threshold of Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)and sumatriptan group was increased compared with migraine group,and A,B,C,D,E group and sumatriptan group increased significantly(p<0.05),besides,C group increased most..cfosThe expression of c-fos in thalamus of migraine group decreased significantly(p<0.01)and increased significantly(p<0.01)in trigeminal nucleus compared with control group.The expression of c-fos in thalamus of Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)and sumatriptan group increased compared with migraine group,and D group increased most(p<0.01).The expression of c-fos in spinal trigeminal nucleus of Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)and sumatriptan group decreased compared with migraine group,and D group decreased significantly(p<0.01).CREBThe expression of CREB in thalamus and spinal trigeminal nucleus of migraine group has no significantly changes(p>0.05)compared with control group.The expression of CREB in thalamus and spinal trigeminal nucleus of Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)and sumatriptan group has no significantly changes(p>0.05)compared with migraine group.pCREBThe expression of pCREB in thalamus of migraine group has no significantly changes(p>0.05)compared with control group.The expression of pCREB in thalamus of Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)and sumatriptan group has no significantly changes(p>0.05)compared with migraine group.The expression of pCREB in spinal trigeminal nucleus of migraine group increased significantly(p<0.01)compared with control group.The expression of pCREB in spinal trigeminal nucleus of Ligusticum Chuanxiong and Radix Paeoniae Alba group(A,B,C,D,E,F)and sumatriptan group decreased compared with migraine group,and C group decreased significantly(p<0.01).Conclusion1.The pain threshold of migraine rats can be increased by the treatment of Ligusticum Chuanxiong and Radix Paeoniae Alba with different compatibility,and the sensitization behavior of pain can be improved too.The effects of C group(30g:30g)better than others.2.The expression of c-fos,and pCREB in thalamus and spinal trigeminal nucleus of migraine rats can be regulated by the treatment of Ligusticum Chuanxiong and Radix Paeoniae Alba with different compatibility.3.As a upstream factor of c-fos,CREB activate the expression of c-fos by phosphorylated CREB in spinal trigeminal nucleus and take part in migraine.4.We provided a reference of application of Ligusticum Chuanxiong and Radix Paeoniae Alba with different compatibility in treatment of migraine by the study.