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Objective To explore the analgesic effect of intra-articular botulinum neurotoxin type A (BoNTA) injection in rats with adjuvant-arthritis pain,to quantify the expression of calcitonin gene-related peptide (CGRP) in the dorsal root ganglia (DRG) associated with arthritis pain,and to investigate the retrograde axonal transport of BoNT-A into the DRG after peripheral injection.Methods Ninety Sprague-Dawley rats were randomly divided into groups A,B,C,D and E,each of 18.A murine model of adjuvant-arthritis pain was established by injecting 50 μL of complete Freund's adjuvant into the left ankle in all the mice except those in group A.The control group A was treated with intra-articular injection of 50 μL of saline solution.Three weeks later,groups A and B were treated with a 20 μL intra-articular saline injection,while groups C,D and E received an intra-articular injection of BoNT-A at 1 U/20 μL,3 U/20 μL or 10 U/20 μL respectively.Pain threshold and muscle strength were graded before and 1,5,15 and 21 days after the modelling,as well as at 1,3,5 and 14 days after the BoNT-A treatments.Protein expression and the CGRP-positive cell number were observed,as well as any BoNT-A-cleaved synaptosomal-associated 25 kDa protein (cl-SNAP-25) in the DRG using Western blotting and immunofluorescence.Results Compared with group A,there was a significant decrease in the average mechanical withdrawal threshold and muscle strength and a significant increase in the protein expression and the CGRP-positive cell number in the other 4 groups.Compared with group B,the mechanical withdrawal threshold had increased significantly more in groups D and E at 5 days after the BoNT-A injection and in group C at 14 days after the treatment.Compared with group B,the protein expression and the number of CGRP-positive cells were significantly lower in groups D and E at 3 days after the BoNT-A injection.The decrease in group C was significant after 14 days.No significant differences were found between groups D and E in any measurement at any time point.There was no significant difference among groups B,C and D in terms of muscle strength.Five days after the BoNT-A injection,significantly decreased muscle strength was observed in group E.In addition,BoNT-A cleaved-SNAP-25 was detected in the DRG.Conclusion BoNT-A can reduce arthritis pain through inhibiting the expression of CGRP in the DRG.Its analgesic effect has a dose response.A peripheral injection of BoNT-A can arrive at the DRG through retrograde axonal transport.
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The purpose of this research work is to find out the longitudinal distribution of the preganglionic neurons which project to the superior cervical sympathetic ganglion (SCSG). Experiments were performed on 12 adult rabbits and a monkey. Under sodium pentobarbital anesthesia, 20 microliters of 10% HRP were injected slowly into the rabbit's SCSG. In the monkey, 20 microliters of 15% HRP was injected.After a postoperative survival time of 3～6 days, the animals were perfused through,the ascending aorta with a cold fixative mixture composed of 2% paraformaldehyde and 1.25% glutaraldyhyde in 0.1mol phosphatebuffer at pH 7.4. The spinal cord segments C_1～L_3 were cut serially in transverse plane on a cryostat at 48 micra.The HRP reaction product was demonstrated according to Mesulam's (1976) benzidine blue reaction method, and counterstained with neutral red.HRP labeled neurons in the spinal cord were located exclusively on the side ipsilateral to the injected SCSG. The total number of labeled cells were 7908 in 12 rabbits, but the number of labeled cells varied from animal to animal. The highest amount was 1690 (423~#) and the lowest amount was 71 (425~#). The longitudinal distribution of the labeled cells in 12 rabbits was 12 segaments of the spinal cord (C_6～T_9), but the largest proportion (86.18%) of them was concentrated from T_1 to T_4, especially at the level of T_2 and T_3 (56.22%), and with a peak at T_2 (29.10%).In cross section of the spinal cord. HRP-labeled cells were concentrated in four cell groups, they are: nucleus intermediolateralis pars principalis (ILp), nucleus intermediolateralis pars funicularis (ILf), nucleus intercalatus (IC) and nucleus intercalatus pars paraependymalis (ICpe). The latter is subdivided into dorsal portion and ventral portion. HRP positive cells were mainly located in the ILp, In 12 rabbits about 92.99% cells were located in it. A small portion of labeled cells(6.25%) were seen in the ILf. A few labeled neurons could be detected within,the IC (0.68%) and ICpe (0.08%). Furthermore, occasionally, very few labeled cells were found at the dorsol portion of the anterior horn.In the monkey, generally speaking, the pattern of the distribution of labeled cells was the same as the rabbit.