This project was sponsored by National Natural Science Foundation of China (81873365 and 81603676), Zhejiang Provincial Natural Science Funds for Distinguished Young Scholars (LR17H270001) and research funds from Zhejiang Chinese Medical University (Q2019J01, 2018ZY37, 2018ZY19).

The animal protocols were approved by Zhejiang Chinese Medical University Animal Ethics Committee.
1. Animals
<ol>
	<li>Obtain male Sprague-Dawley (SD) rats (280&#8211;320 g, 8-10 weeks of age) from Shanghai Laboratory Animal Center. House the animals in Zhejiang Chinese Medical University Laboratory Animal Center. Note that the breeding conditions should include 12 h/ 2h light/dark cycles and keep temperature constant at 24 &#176;C. Provide water and food ad libitum. Note that a to...

<ol>
	<li>Goh, E. L., Chidambaram, S., Ma, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complex+regional+pain+syndrome:+a+recent+update.">Complex regional pain syndrome: a recent update.</a> Burns, Trauma. 5 (1), 2 (2017).</li><li>Birklein, F., Ajit, S. K., Goebel, A., Rsgm, P., Sommer, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complex+regional+pain+syndrome+-+phenotypic+characteristics+and+potential+biomarkers.">Complex regional pain syndrome - phenotypic characteristics and potential biomarkers.</a> Nature Reviews Neurology. 14 (5), (2018).</li><li>Shim, H., Rose, J., Halle, S., Shekane, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complex+regional+pain+syndrome:+a+narrative+review+for+the+practising+clinician.">Complex regional pain syndrome: a narrative review for the practising clinician.</a> British Journal of Anaesthesia. , (2019).</li><li>Urits, I., Shen, A. H., Jones, M. R., Viswanath, O., Kaye, A. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complex+Regional+Pain+Syndrome,+Current+Concepts+and+Treatment+Options.">Complex Regional Pain Syndrome, Current Concepts and Treatment Options.</a> Current Pain, Headache Reports. 22 (2), 10 (2018).</li><li>Helyes, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transfer+of+complex+regional+pain+syndrome+to+mice+via+human+autoantibodies+is+mediated+by+interleukin-1-induced+mechanisms.">Transfer of complex regional pain syndrome to mice via human autoantibodies is mediated by interleukin-1-induced mechanisms.</a> Proceedings of National Academy Sciences of the United States of America. 116 (26), 13067-13076 (2019).</li><li>Coderre, T. J., Xanthos, D. N., Francis, L., Bennett, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+post-ischemia+pain+(CPIP):+a+novel+animal+model+of+complex+regional+pain+syndrome-Type+I+(CRPS-I;+reflex+sympathetic+dystrophy)+produced+by+prolonged+hindpaw+ischemia+and+reperfusion+in+the+rat.">Chronic post-ischemia pain (CPIP): a novel animal model of complex regional pain syndrome-Type I (CRPS-I; reflex sympathetic dystrophy) produced by prolonged hindpaw ischemia and reperfusion in the rat.</a> Pain. 112 (1), 94-105 (2004).</li><li>Coderre, T. J., Bennett, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+hypothesis+for+the+cause+of+complex+regional+pain+syndrome-type+I+(reflex+sympathetic+dystrophy):+pain+due+to+deep-tissue+microvascular+pathology.">A hypothesis for the cause of complex regional pain syndrome-type I (reflex sympathetic dystrophy): pain due to deep-tissue microvascular pathology.</a> Pain Medicine. 11 (8), 1224-1238 (2010).</li><li>Klafke, J. Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+and+chronic+nociceptive+phases+observed+in+a+rat+hind+paw+ischemia/reperfusion+model+depend+on+different+mechanisms.">Acute and chronic nociceptive phases observed in a rat hind paw ischemia/reperfusion model depend on different mechanisms.</a> Pflugers Archiv European Journal of Physiology. 468 (2), 229-241 (2015).</li><li>Tang, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interaction+between+astrocytic+colony+stimulating+factor+and+its+receptor+on+microglia+mediates+central+sensitization+and+behavioral+hypersensitivity+in+chronic+post+ischemic+pain+model.">Interaction between astrocytic colony stimulating factor and its receptor on microglia mediates central sensitization and behavioral hypersensitivity in chronic post ischemic pain model.</a> Brain Behavioral Immunology. 68, 248-260 (2018).</li><li>Kim, J. H., Kim, Y. C., Nahm, F. S., Lee, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Therapeutic+Effect+of+Vitamin+C+in+an+Animal+Model+of+Complex+Regional+Pain+Syndrome+Produced+by+Prolonged+Hindpaw+Ischemia-Reperfusion+in+Rats.">The Therapeutic Effect of Vitamin C in an Animal Model of Complex Regional Pain Syndrome Produced by Prolonged Hindpaw Ischemia-Reperfusion in Rats.</a> International Journal of Medical Sciences. 14 (1), 97-101 (2017).</li><li>Yin, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transcriptome+profiling+of+dorsal+root+ganglia+in+a+rat+model+of+complex+regional+pain+syndrome+type-I+reveals+potential+mechanisms+involved+in+pain.">Transcriptome profiling of dorsal root ganglia in a rat model of complex regional pain syndrome type-I reveals potential mechanisms involved in pain.</a> Journal of Pain Research. 12, 1201-1216 (2019).</li><li>Hu, Q., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=TRPV1+Channel+Contributes+to+the+Behavioral+Hypersensitivity+in+a+Rat+Model+of+Complex+Regional+Pain+Syndrome+Type+1.">TRPV1 Channel Contributes to the Behavioral Hypersensitivity in a Rat Model of Complex Regional Pain Syndrome Type 1.</a> Frontiers in Pharmacology. 10, 453 (2019).</li><li>Dixon, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficient+analysis+of+experimental+observations.">Efficient analysis of experimental observations.</a> Annual Review of Pharmacology and Toxicology. 20, 441-462 (1980).</li><li>Chai, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electroacupuncture+Alleviates+Pain+Responses+and+Inflammation+in+a+Rat+Model+of+Acute+Gout+Arthritis.">Electroacupuncture Alleviates Pain Responses and Inflammation in a Rat Model of Acute Gout Arthritis.</a> Evidence-Based Complementary and Alternative Medicine. 2018, 2598975 (2018).</li><li>Chaplan, S. R., Bach, F. W., Pogrel, J. W., Chung, J. M., Yaksh, T. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+assessment+of+tactile+allodynia+in+the+rat+paw.">Quantitative assessment of tactile allodynia in the rat paw.</a> Journal of Neuroscience Methods. 53 (1), 55-63 (1994).</li><li>Fang, J. Q., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Parameter-specific+analgesic+effects+of+electroacupuncture+mediated+by+degree+of+regulation+TRPV1+and+P2X3+in+inflammatory+pain+in+rats.">Parameter-specific analgesic effects of electroacupuncture mediated by degree of regulation TRPV1 and P2X3 in inflammatory pain in rats.</a> Life Sciences. 200, 69-80 (2018).</li><li>Tai, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Involvement+of+Transient+Receptor+Potential+Cation+Channel+Member+A1+activation+in+the+irritation+and+pain+response+elicited+by+skin-lightening+reagent+hydroquinone.">Involvement of Transient Receptor Potential Cation Channel Member A1 activation in the irritation and pain response elicited by skin-lightening reagent hydroquinone.</a> Scientific Reports. 7 (1), 7532 (2017).</li><li>Liu, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=TRPM8+is+the+principal+mediator+of+menthol-induced+analgesia+of+acute+and+inflammatory+pain.">TRPM8 is the principal mediator of menthol-induced analgesia of acute and inflammatory pain.</a> Pain. 154 (10), 2169-2177 (2013).</li><li>Liu, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oxidized+Phospholipid+OxPAPC+Activates+TRPA1+and+Contributes+to+Chronic+Inflammatory+Pain+in+Mice.">Oxidized Phospholipid OxPAPC Activates TRPA1 and Contributes to Chronic Inflammatory Pain in Mice.</a> PLoS One. 11 (11), 0165200 (2016).</li><li>Terkelsen, A. J., Gierthmuhlen, J., Finnerup, N. B., Hojlund, A. P., Jensen, T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bilateral+hypersensitivity+to+capsaicin,+thermal,+and+mechanical+stimuli+in+unilateral+complex+regional+pain+syndrome.">Bilateral hypersensitivity to capsaicin, thermal, and mechanical stimuli in unilateral complex regional pain syndrome.</a> Anesthesiology. 120 (5), 1225-1236 (2014).</li><li>Drummond, P. D., Morellini, N., Finch, P. M., Birklein, F., Knudsen, L. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complex+regional+pain+syndrome:+intradermal+injection+of+phenylephrine+evokes+pain+and+hyperalgesia+in+a+subgroup+of+patients+with+upregulated+alpha1-adrenoceptors+on+dermal+nerves.">Complex regional pain syndrome: intradermal injection of phenylephrine evokes pain and hyperalgesia in a subgroup of patients with upregulated alpha1-adrenoceptors on dermal nerves.</a> Pain. 159 (11), 2296-2305 (2018).</li><li>Minert, A., Gabay, E., Dominguez, C., Wiesenfeld-Hallin, Z., Devor, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spontaneous+pain+following+spinal+nerve+injury+in+mice.">Spontaneous pain following spinal nerve injury in mice.</a> Experimental Neurology. 206 (2), 220-230 (2007).</li><li>Kingery, W. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Capsaicin+sensitive+afferents+mediate+the+development+of+heat+hyperalgesia+and+hindpaw+edema+after+sciatic+section+in+rats.">Capsaicin sensitive afferents mediate the development of heat hyperalgesia and hindpaw edema after sciatic section in rats.</a> Neuroscience Letters. 318 (1), 39-43 (2002).</li><li>Xu, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activation+of+cannabinoid+receptor+2+attenuates+mechanical+allodynia+and+neuroinflammatory+responses+in+a+chronic+post-ischemic+pain+model+of+complex+regional+pain+syndrome+type+I+in+rats.">Activation of cannabinoid receptor 2 attenuates mechanical allodynia and neuroinflammatory responses in a chronic post-ischemic pain model of complex regional pain syndrome type I in rats.</a> European Journal of Neuroscience. 44 (12), 3046-3055 (2016).</li><li>Coderre, T. J., Xanthos, D. N., Francis, L., Bennett, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+post-ischemia+pain+(CPIP):+a+novel+animal+model+of+complex+regional+pain+syndrome-type+I+(CRPS-I;+reflex+sympathetic+dystrophy)+produced+by+prolonged+hindpaw+ischemia+and+reperfusion+in+the+rat.">Chronic post-ischemia pain (CPIP): a novel animal model of complex regional pain syndrome-type I (CRPS-I; reflex sympathetic dystrophy) produced by prolonged hindpaw ischemia and reperfusion in the rat.</a> Pain. 112 (1-2), 94-105 (2004).</li><li>Weissmann, R., Uziel, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pediatric+complex+regional+pain+syndrome:+a+review.">Pediatric complex regional pain syndrome: a review.</a> Pediatric Rheumatology Online Journal. 14 (1), 29 (2016).</li><li>Kim, H., Lee, C. H., Kim, S. H., Kim, Y. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiology+of+complex+regional+pain+syndrome+in+Korea:+An+electronic+population+health+data+study.">Epidemiology of complex regional pain syndrome in Korea: An electronic population health data study.</a> PLoS One. 13 (6), 0198147 (2018).</li><li>Tang, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sex+differences+in+complex+regional+pain+syndrome+type+I+(CRPS-I)+in+mice.">Sex differences in complex regional pain syndrome type I (CRPS-I) in mice.</a> Journal of Pain Research. 10, 1811-1819 (2017).</li></ol>

The authors declare no conflicts of interest in this work.

This protocol describes the detailed methods for establishing a rat CPIP model by applying ischemia/reperfusion to hind limbs of the rats. It involves the evaluation of hind limb appearance, edema, mechanical/thermal hypersensitivities, and acute nocifensive behaviors in response to capsaicin injection.
Limb ischemia/reperfusion is a common factor contributing to CRPS-I in human patients12. This protocol describes how to establish the rat CPIP model, which is a commonly...

Complex regional pain syndrome (CRPS) reprents complex and chronic pain symptoms resulting from fractures, trauma, surgery, ischemia or nerve injury1,2,3. CRPS is classified into 2 subcategories: CRPS type-I and type-II (CRPS-I and CRPS-II)4. Epidemiological studies revealed that the prevalence of CRPS was approximately 1:20005. CRPS-I, which shows no obvious nerve damage, can result in chronic pain and dramatically affects the life quality of the patients. Current available treatments show inadequate therapeutic effects. Therefore, CRPS-I still remains an important and challenging clinical problem that needs to be addressed.
Establishing a preclinical animal model mimicking CRPS-I is crucial for exploring the mechanisms underlying CRPS-I. In order to address this issue, Coderre et al. designed a rat model by applying prolonged ischemia and reperfusion to the hind limb to recapitulate CRPS-I6. It is known that ischemia/reperfusion injury is among one of the major causes of CRPS-I7. The rat CPIP model exhibits many CRPS-I-like symptoms, which include hind limb edema and hyperemia in the early stage after model establishment, followed with persistent thermal and mechanical hypersensitivities6. With the aid from this model, it is proposed that central pain sensitization, peripheral TRPA1 channel activation and reactive oxygen species generation, etc. contribute to CRPS-I8,9,10. We recently successfully established the CPIP rat model and performed RNA-sequencing of the dorsal root ganglia (DRGs) that innervate the affected hind paw11. We discovered some potential mechanisms that are possibly involved in mediating the pain hypersensitivities of CRPS-I11. We further identified transient receptor potential vanilloid 1 (TRPV1) channel in DRG neurons as an important contributor to the mechanical and thermal hypersensitivities of CRPS-I12.
In this study, we described the detailed procedures involved in the establishment of the rat model of CPIP. We further evaluated the rat CPIP model by measuring the mechanical and thermal hypersensitivities as well as its responsiveness to acute capsaicin challenge. We propose that the rat CPIP model can be a reliable animal model for further investigation of the mechanisms involved in CRPS-I.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1.5 ml Eppendorf tube</td>
			<td>Eppendorf</td>
			<td>22431021</td>
			<td></td>
		</tr>
		<tr>
			<td>DMSO</td>
			<td>Sigma-Aldrich</td>
			<td>D1435</td>
			<td></td>
		</tr>
		<tr>
			<td>Capsaicin</td>
			<td>APEXBIO</td>
			<td>A3278</td>
			<td></td>
		</tr>
		<tr>
			<td>Digital caliper</td>
			<td>Meinaite</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>O-ring</td>
			<td>O-Rings West</td>
			<td>Nitrile 70 Durometer</td>
			<td>7/32 in. 
			internal diameter</td>
		</tr>
		<tr>
			<td>Plantar Test Apparatus</td>
			<td>UGO Basile, Italy</td>
			<td>37370</td>
			<td></td>
		</tr>
		<tr>
			<td>von Frey filaments</td>
			<td>UGO Basile, Italy</td>
			<td>NC12775</td>
			<td></td>
		</tr>
	</tbody>
</table>

chronic post-ischemia pain model for complex regional pain syndrome type-i in rats

Complex regional pain syndrome type-I (CRPS-I) is a neurological disease that causes severe pain among patients and remains an unresolved medical condition. However, the underlying mechanisms of CRPS-I have yet to be revealed. It is known that ischemia/reperfusion is one of the leading factors that causes CRPS-I. By means of prolonged ischemia and reperfusion of the hind limb, the rat chronic post-ischemia pain (CPIP) model has been established to mimic CRPS-I. The CPIP model has become a well-recognized animal model for studying the mechanisms of CRPS-I. This protocol describes the detailed procedures involved in the establishment of the rat model of CPIP, including anesthesia, followed by ischemia/reperfusion of the hind limb. Characteristics of the rat CPIP model are further evaluated by measuring the mechanical and thermal hypersensitivities of the hind limb as well as the nocifensive responses to acute capsaicin injection. The rat CPIP model exhibits several CRPS-I-like manifestations, including hind limb edema and hyperemia in the early stage after establishment, persistent thermal and mechanical hypersensitivities, and increased nocifensive responses to acute capsaicin injection. These characteristics render it a suitable animal model for further investigation of the mechanisms involved in CRPS-I.

After placing the O-ring on the ankle, the ipsilateral hind paw skin showed cyanosis, an indication of tissue hypoxia (Figure 1A). After cutting the O-ring, the ipsilateral hind paw began to fill with blood and showed robust swelling, which demonstrated an intense sign of hyperemia (Figure 1A). The paw swelling gradually diminished and returned to normal 48 h after the ischemic/reperfusion procedure (two-way ANOVA with Sidak pos...

Watch this Scientific Journal Video about Chronic Post-Ischemia Pain Model for Complex Regional Pain Syndrome Type-I in Rats at JoVE.com

Chronic Post-Ischemia Pain Model for Complex Regional Pain Syndrome Type-I in Rats

Provided here is a protocol that details steps to establish an animal model of chronic post-ischemia pain (CPIP). This is a well-recognized model mimicking human complex regional pain syndrome type-I. Mechanical and thermal hypersensitivities are further evaluated, as well as capsaicin-induced nocifensive behaviors observed in the CPIP rat model.

Complex regional pain syndrome type-I (CRPS-I) is a neurological disease that causes severe pain among patients and remains an unresolved medical ...

The mechanism underlying CRPS I still remain unclear. Establishing a preclinical animal model mimicking CRPS I will be crucial for exploring the mechanisms underlying CRPS I.The rat CPIP model exhibits many CRPS I like simitance, which include hindlimb edema and hyperemia in the early stage. After model establishment, followed by persistent thermal and mechanical hypersensitivities.Begin by anesthetizing all rats with sodium phenobarbital. Make sure that the rats are properly anesthetized by pinching their hind paw or tail with forceps. And place vet ointment on the eyes.Keep the rat on a 37 degree Celsius heated pad during the procedure. Lubricate the right hind paw and ankle with glycerol. Then slide a nitrile 70 durometer O ring into the larger side of a one point five milliliter tube with the snap cap cut off.Carefully insert the hind paw into the hollow tube until it reaches the bottom. Gradually slide the O ring from the tube to the right hindlimb near the ankle joint. And leave it there for three hours.Apply the same treatment to the sham group of rats, except with a broken O ring that should not induce ischemia. After three hours, cut off the O ring and monitor the rat until it regains enough consciousness to maintain sternal recumbency. Do not place the rat in the company of other rats until it is fully recovered from anesthesia.Place the rat in a transparent plexiglass chamber on a mesh floor, and habituate the rat to the environment for 30 minutes before any behavioral testing. Use von frey filaments to test the rat for mechanical allodynia. Begin the test from the middle filament by vertically applying it to the middle plantar surface of the hind paw.Apply suitable force to bend the filament for up to five seconds. A sudden retraction of the paw is considered a nocifensive behavior. Conduct the mechanical allodynia test on days three, two, one, and every other day until day 13.Apply the up down testing method to test the threshold and a dickson method for calculating the 50 percent paw withdrawal threshold. To test for thermal hyperalgesia, use hargreaves method by directly aiming the light beam emitted from a 50 watt bulb to the hind paw. Measure the paw withdrawal latency, setting 20 seconds as the cutoff threshold to avoid excessive injury.Repeat each test three times in five minute intervals for each hind paw, and take the average as the paw withdrawal latency. Conduct the thermal hyperalgesia test on days three, two, one, and every other day until day 13. To test for capsaicin induced acute nocifensive behavior, prepare the 200 millimolar capsaicin stock solution with DMSO, and further dilute it one to 1, 000 in PBS for injection.Capsaicin is a reagent, which can cause joint irritation to skin, eyes, and respiration. It's important to wear a mask and goggle when preparing capsaicin solution. Then inject 50 microliters of the capsaicin or a vehicle into the hind paw using a 30 gauge needle attached to a one milliliter syringe.Record nocifensive behavior, such as licking, biting, or flinching of the injected paw using a video camera for 10 minutes after the injection. Evaluate hind paw edema by measuring the increase in paw diameter at 15 minutes, 24 hours, 48 hours, and 72 hours after model establishment. After placing the O ring on the ankle, the ipsilateral hind paw skin showed cyanosis, an indication of tissue hypoxia.After cutting the O ring, the paw began to fill with blood and showed robust swelling, indicating intense hyperemia. The swelling gradually diminished and returned to normal 48 hours after the procedure. One day after model establishment, the ipsilateral hind paw of the CPIP group exhibited obvious mechanical allodynia that persisted for 13 days of the observation time frame.The contralateral hind paw of the CPIP group also displayed mechanical hyperalgesia for 13 days. When thermal hyperalgesia was measured, bilateral hind paws of CPIP rats exhibited significantly reduced withdrawal latency in response to noxious thermal stimuli, compared to the sham group rats. The thermal hyperalgesia of the ipsilateral hind paw persisted until the end of the observation time frame, whereas the thermal hyperalgesia of the contralateral hind paw lasted for seven days.The sham and CPIP groups were tested for nocifensive behavior with the capsaicin injection. When a vehicle was injected, the sham group showed a slight nocifensive response and the CPIP group showed a significantly higher response. More importantly, CPIP rats showed significantly higher responses to capsaicin injection than the sham group, which suggests that the CPIP rats exhibit a phenomenon similar to human patients with CRPS I.By this procedure, the rats are anesthetized before and during model establishment.The CPIP rat model described in this protocol established by ischemia and Human CRPS I like simitance. Families of these models, more mechanism about CRPS I can be explored.

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7.5K Görüntüleme.  Key Laboratory of Acupuncture and Neurology of Zhejiang Province. CRPS I'in altında yatan mekanizma hala belirsizliğini koruyor. CRPS'yi taklit eden bir preklinik hayvan modelinin oluşturulması CRPS I.The rat CPIP modelinin altında yatan mekanizmaları keşfetmek için çok önemli olacaktır. Model ininden sonra kalıcı termal ve mekanik hipersensitiviteler takip eder.Sodyum fenobarbital ile tüm sıçanlar anestezi ile başlayın. Fareler düzgün kendi arka pençe veya kuyruk forceps ile pinching tarafından anestezi olduğundan emin olun. V...