Anesthesia & Analgesia:
Pain and Analgesic Mechanisms: Research Reports
The Effects of Electroacupuncture on the Extracellular Signal-Regulated Kinase 1/2/P2X3 Signal Pathway in the Spinal Cord of Rats with Chronic Constriction Injury
Yu, Jianbo PhD*; Zhao, Cong MD*†; Luo, Xiaoqin MD‡
From the *Department of Anesthesiology, Tianjin Nankai Hospital, Tianjin Medical University, Tianjin; †Department of Anesthesiology, Chengdu Fifth People’s Hospital, Sichuan Province; and ‡Department of Pathology, Xiangyang First People’s Hospital, Hubei Province, China.
Accepted for publication August 02, 2012.
Published ahead of print December 7, 2012
J. Y. and C. Z. contributed equally to the manuscript.
This study was supported by the Science and Technology Support Project of Tianjin (grant no. 12ZCZDSY03300) and the Application Foundation and Frontier Technology Research Project of Tianjin (grant no. 11JCYBJC11000).
The authors declare no conflicts of interest.
Reprints will not be available from the authors.
Address correspondence to Jianbo Yu, PhD, Department of Anesthesiology, Tianjin Nankai Hospital, Tianjin Medical University, Sanwei Road 122, Nankai District, Tianjin, People’s Republic of China 300100. Address e-mail to email@example.com.
BACKGROUND: Electroacupuncture (EA), as a traditional clinical method, is widely accepted in pain clinics, but the analgesic effect of EA has not been fully demonstrated. In the present study, we investigated the effect of EA on chronic pain and expression of P2X3 receptors in the spinal cord of rats with chronic constriction injury (CCI).
METHODS: The study was conducted in 2 parts. In part 1, Sprague Dawley rats were divided into 6 groups (n = 10): sham-CCI, CCI, LEA; CCI + 2 Hz EA at acupoints), HEA; CCI + 15 Hz EA at acupoints), NA-LEA (CCI + 2 Hz EA at nonacupoints), and NA-HEA (CCI + 15 Hz EA at nonacupoints). EA treatment was performed once a day on days 4 to 9 after CCI. Nociception was assessed using von Frey filaments and a hotplate apparatus. The protein and the messenger RNA (mRNA) levels of P2X3 receptors in the spinal cord were assayed by Western blotting and real-time polymerase chain reaction, respectively. In part 2, rats were divided into 5 groups (n = 10): sham-CCI, CCI, EA (CCI + EA at acupoints), NA-EA (CCI + EA at nonacupoints), and U0126 (CCI + intrathecal injection of U0126). EA treatment was conducted similar to part 1. Rats were given 5 µg U0126 in the U0126 group and 5% dimethyl sulfoxide intrathecally. Ten microliters was used as a vehicle for the other 4 groups twice a day on days 4 to 9 after CCI. Extracellular signal-regulated kinase 1/2 (ERK1/2) and ERK1/2 phosphorylation in the spinal cord were also assayed by Western blotting.
RESULTS: EA treatment exhibited significant antinociceptive effects and reduced the CCI-induced increase of both protein and mRNA expression of P2X3 receptors in the spinal cord. Furthermore, 2 Hz EA had a better analgesic effect than 15 Hz EA, and the protein and mRNA level of P2X3 receptor in spinal cord were lower in rats treated with 2 Hz EA at acupoints than 15 Hz EA at acupoints. Either EA at acupoints or intrathecal injection of U0126 relieved allodynia and hyperalgesia and reduced the expression of P2X3 receptors and ERK1/2 phosphorylation in the spinal cord.
CONCLUSIONS: The data demonstrated that EA alleviates neuropathic pain behavior, at least in part, by reducing P2X3 receptor expression in spinal cord via the ERK1/2 signaling pathway. Low frequency EA has a better analgesic effect than high frequency HEA on neuropathic pain.
Neuropathic pain (NP), characterized by spontaneous pain, hyperalgesia, and allodynia, is often caused by nerve injury or other diseases.1 Because conventional analgesics, such as opiates and nonsteroidal antiinflammatory drugs, do not have good efficacy against NP, the choice of treatment has been largely limited. Anticonvulsants and antidepressants are prescribed for the management of NP, but their effects are limited.2,3
P2X3, a subtype of P2X receptors, which is a nonselective cationic channel, is expressed mostly in sensory neurons and is involved in transduction of pain stimuli.4 Adenosine triphosphate (ATP), a fast neurotransmitter, by activating P2 receptors especially P2X3 receptor, can modulate pain transmission under NP conditions.5–7 Rats receiving P2X3 gene knockout or P2X3 antisense receptor gene disruption have been shown to be insensitive to noxious stimulus. The acute administration of a highly selective nonnucleotide P2X3 antagonist, A317491, can fully block NP. These findings support the significant role for P2X3 receptors in the expression of pain.
Both animal experiments and clinical pain investigations have shown that electroacupuncture (EA) has an effective analgesic effect on chronic pain.8–12 In addition, studies have shown that different frequencies of EA may exhibit different degrees of analgesia.13–15 Although EA has been shown to stimulate the release of endorphins and other neurotransmitters,16,17 little is known about its effect on the P2X3 receptor.
Extracellular signal-regulated kinase 1/2(ERK1/2), a member of the mitogen-activated protein kinase (MAPK) family, is implicated in neural plasticity, which contributes to development of chronic pain.18,19 ERK1/2 activation in the spinal cord induced by nociceptive stimulation plays a critical role in central sensitization.20 The inhibition of ERK1/2 phosphorylation (p-ERK1/2) by EA in the spinal cord dorsal horn, the activation of which contributes to the induction and maintenance of chronic constriction injury (CCI)–induced NP, is possibly involved in acupuncture’s analgesic effect.21 On the other hand, a previous study showed that the enhanced fast-inactivating current induced by α,β-methylene ATP in chronic compression of dorsal root ganglion (DRG)–operated DRG neurons was significantly suppressed by a specific inhibitor of the ERK1/2, U0126.22 In addition, in peripheral inflammation and sciatic nerve transaction rat model, p-ERK1/2 labeling occurs through the P2X3 receptors in the terminals of DRG neurons after the mechanical stimuli into the inflamed tissue.23 However, whether ERK1/2 is involved in mediating EA’s effect on P2X3 receptors in the spinal cord after nerve injury has not been fully demonstrated.
In the present study, we sought to evaluate the effects and mechanisms of EA on P2X3 receptor expression in spinal cords of rats with nerve injury–induced NP and investigated the involvement of the ERK1/2 signaling pathway in regulating the effect of EA on P2X3 receptor for better understanding of EA on persistent pain.
All experiments were approved by the local animal ethics committee of the Tianjin Medical University and conducted in accordance with the Guide for the Care and Use of Laboratory Animals from the National Institutes of Health and the International Association for the Study of Pain. Male Sprague Dawley rats weighing 200 ± 20 g, supplied by Tianjin Medical University (Tianjin, China), were given free access to food and water.
The study consisted of the following 2 experiments: Experiment 1: Effects of EA on P2X3 receptor expression—CCI rats were divided into 6 groups (n = 10 each): sham-CCI group, CCI group, low-frequency EA (LEA) group (CCI + 2 Hz EA at Zusanli-Yanglingquan acupoints [ST36-GB34]), high-frequency EA (HEA; CCI + 15 Hz EA at ST36-GB34), NA-LEA (CCI + 2 Hz EA at nonacupoints), and NA-HEA (CCI + 15 Hz EA at nonacupoints); Experiment 2: Involvement of ERK1/2 signal pathway in EA regulating P2X3 receptor expression—CCI rats with an indwelling catheter placed in the subarachnoid space were divided into 5 groups (n = 10 each): sham-CCI group, CCI group, EA group (CCI + 2 Hz EA at ST36-GB34), NA-EA group (CCI + 2 Hz EA at nonacupoints), and U0126 group (CCI + U0126 intrathecal [IT] injection). U0126 dissolved in 5% dimethyl sulfoxide (DMSO) was administered to the U0126 group; the other 4 groups received the same volume of the vehicle (5% DMSO). The administration of U1026 (5 µg/10 µL IT) or DMSO (5%/10 µL IT) was started 4 days after CCI and continued twice a day until day 9. Thirty-minute EA treatment was given on days 4 to 9 after CCI. Thermal withdrawal latency (TWL) and mechanical withdrawal threshold (MWT) were determined at baseline and on days 3, 5, 7, and 9 after CCI. Lumbar 4 and lumbar 5 spinal cord was removed under deep anesthesia on day 10 after CCI. P2X3 receptor protein and messenger RNA (mRNA) levels were measured by Western blotting and real-time polymerase chain reaction, respectively. Another total ERK1/2 and ERK1/2 phosphorylation were assayed by Western blotting in experiment 2.
Chronic Constriction Injury
CCI, as described by Bennett and Xie,24 was performed on anesthetized rats. Briefly, after skin incision and blunt dissection of subcutaneous tissue and muscle in the right midthigh, the right common sciatic nerve was exposed, and 4 ligatures were loosely placed (4-0 silk thread) with 1-mm intervals to cause CCI. In sham-CCI rats, the right sciatic nerve was exposed but not ligated.
Intrathecal Drug Delivery
As described by Yaksh and Rudy,25 catheterization of the rat spinal subarachnoid space was performed on anesthetized rats 2 days before CCI. Briefly, rats were implanted with a PE-10 polyethylene catheter (8 cm) in the lumbar subarachnoid space. Rats with no postoperative neurologic deficits after surgery were kept for experiment 2. In experiment 2, rats in the U0126 group were injected intrathecally twice daily at 12-hour intervals (days 4–9) with selective ERK1/2 inhibitor U0126 (5 µg; Sigma, St. Louis, MO) dissolved in 5% DMSO (10 µL; Sigma), and rats in sham-CCI, CCI, EA, and NA-EA groups were injected intrathecally with 10 µL of 5% DMSO as a vehicle, followed by 10 µL of normal saline flush.
Rats received EA stimulation for 30 minutes once a day on days 4 to 9 after CCI. Rats without any anesthetic remained quiet and awake in tubular acrylic holders during EA treatment. Two stainless acupuncture needles of 0.3 mm diameter and 50 mm long connected to a pair of electrodes were inserted perpendicularly approximately 6 mm into the ipsilateral acupoints on the leg of the rat corresponding to the Zusanli (ST36) and Yanglingquan (GB34) acupoints in humans. In humans, ST 36 is located on the anterior aspect of the lower leg, 4 finger breadths (middle finger) below the knee joint, and 1 finger breadth lateral to the anterior crest of the tibia, while GB34 is located in the depression anterior and inferior to the head of the fibula. Two nonacupoints are located 0.5 cm horizontal and lateral to ST36 and GB34 acupoints, respectively, at nonmeridian points.26,27 The other ends of electrodes were connected to the output channel of the electrostimulator (Electronic pulse therapeutic apparatus G6805-1A; Huayi Medical Instruments, Shanghai, China). The constant frequency (2 Hz or 15 Hz) and the intensity of less than 1.5 mA to induce moderate muscle contraction of the hind limb was applied to perform EA stimulation. Rats of no-EA groups were kept in holders for 30 minutes as control.
Paw withdrawal to thermal and mechanical stimulation was tested on the day before CCI surgery and on days 3, 5, 7, and 9 after CCI. The behavioral test was performed 20 minutes after EA on days 5, 7, and 9. Animals were placed on an elevated wire grid for acclimation for 30 minutes before testing. Mechanical allodynia was determined by the 50% withdrawal threshold for the up–down method using a set of von Frey filaments (Stoelting, North Coast).28 Thermal hyperalgesia was assessed by the method described previously,27 using a hotplate with Plexiglas walls (model YLS-6B; Zhenghua Bio-instruments, Anhui, China). Rats were placed on a hotplate with a stable temperature of 52°C ± 0.2°C. The average value of 3 sessions at 10-minute intervals for the latency of ipsilateral paw withdrawal testing was used as TWL.
The right lumbar enlargement (lumbar 4 and lumbar 5) was quickly dissected under deep anesthesia on day 10 after CCI and immediately frozen in liquid nitrogen, then stored at –80°C until use.
Tissues were homogenized in lysis buffer that contained proteinase inhibitor mixture (Santa Cruz Biotechnology, Santa Cruz, CA) and centrifuged at 12,000g for 15 minutes at 4°C. Protein concentration was determined by BCA assay (Pierce, Rockford, IL), and 60-µg protein samples were separated using SDS-–PAGE gel (5%–10% gradient gel) then transferred to nitrocellulose membranes (Bio-Rad, Hercules, CA). Membranes were blocked in 5% low-fat milk with buffer containing 0.1% Tween-20 for 2 hours at room temperature, then incubated overnight at 4°C with anti-P2X3 antibody (antirabbit, 1:800; Merck Millipore, Germany) or anti-ERK1/2 antibody (1:1000; Cell Signaling Technology, Beverly, MA), p-ERK1/2 (1:1000; Cell Signaling Technology), or anti-β-actin (1:5000; Sigma), followed by incubation with horseradish peroxidase–conjugated secondary antibody (1:2000; Santa Cruz Biotechnology) for 2 hours. Protein-antibody complexes conjugated with enhanced chemiluminescence reagent (Millipore) were visualized on the Chemidoc XRS System (Bio-Rad).
Quantitative Real-Time Polymerase Chain Reaction
Trizol reagent (Invitrogen, Carlsbad, CA) was used to extract total RNA. One microgram of total RNA was reverse-transcribed to complementary DNA by the protocol of reverse transcription reaction kit (Invitrogen).The P2X3 receptor sequence-specific primers (5-TGGCGTTCTGGGTATTAAGATCGG-3/5-CAGTGGCCTGGTCACTGGCGA-3) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) sequence-specific primers (5-GACAACTTTGGCATCGTGGA-3/5-ATGCAGGGATGATGTTCTGG-3) were designed and synthesized in the Tianjin Institute of Biochemistry (China). The melting temperature for GAPDH was 56°C and 61°C for P2X3. Quantitative reverse RT-PCR amplification was performed on a DNA Engine thermal cycler (Bio-Rad) using SYBR Green I (Invitrogen). Each sample was run in triplicate for each gene. The P2X3 relative mRNA levels were calculated by the method described previously.29
Data are expressed as the mean ± SEM and statistically analyzed by the Statistical Package for the Social Sciences (SPSS) (Version 16.0; SPSS Inc., Chicago, IL). Data from all the measurements were analyzed using a one-way analysis of variance followed by post hoc analysis. P < 0.05 was considered statistically significant.
EA at ST36-GB34 Acupoints, but Not at Nonacupoints, Attenuated CCI-Induced Mechanical Allodynia and Thermal Hyperalgesia
Compared with the sham-CCI group, TWL and MWT of ipsilateral hindpaws decreased significantly on day 3 after CCI in rats treated with CCI (P < 0.001; Fig. 1, A and B), which indicated that CCI induced a prominent mechanical allodynia and thermal hyperalgesia. EA at SB36-GB34 acupoints significantly increased TWL and MWT from day 5 (P = 0.017 LEA versus CCI for MWT, P = 0.020 HEA versus CCI for MWT; P = 0.033 LEA versus CCI for TWL, P = 0.026 HEA versus CCI for TWL) to day 9 (P < 0.001, LEA and HEA versus CCI). Compared with the HEA group, TWL and MWT tested on day 9 in the LEA group increased more significantly (P = 0.023 for MWT; P = 0.041 for TWL). The value of TWL and MWT in rats from nonacupoint EA groups after CCI did not show a significant change when compared with CCI groups (P > 0.05).
EA Reduced P2X3 Receptor Synthesis at both mRNA and Protein Level in the Spinal Cord of Rats After CCI with Acupoint Specificity
The P2X3 protein level in the spinal cord is shown in Figure 2A. CCI nerve injury significantly increased P2X3 receptor protein levels in the spinal cord (P < 0.001 compared with sham-CCI), and this increase was significantly attenuated by EA stimulation of ST36-GB34 acupoints (P < 0.001, CCI versus LEA and HEA). Figure 2B shows that a CCI-induced increase in mRNA synthesis of P2X3 receptors in the spinal cord was significantly attenuated in EA (at acupoints) treatment groups (P < 0.001, CCI versus LEA and HEA). Furthermore, the suppressive effects of EA stimulation on the P2X3 mRNA and protein levels were more remarkable in the LEA group (P = 0.045 for protein level, P = 0.047 for mRNA level, LEA versus HEA). However, the P2X3 receptor protein and mRNA levels in rats of nonacupoint EA groups did not change significantly when compared with CCI groups (P > 0.05).
These data demonstrated that EA at SB36-GB34 acupoints significantly alleviates the mechanical allodynia and thermal hyperalgesia of the ipsilateral paws of rats with CCI, at least partly, by suppressing P2X3 receptors in the spinal cord. LEA has a better analgesic effect than HEA on CCI-induced pain.
Administration of U0126 Intrathecally and Treatment of EA at Acupoint: Each Attenuated CCI-Induced Mechanical Allodynia and Thermal Hyperalgesia
In experiment 2, to investigate whether ERK1/2 activation would regulate P2X3 receptor expression in the spinal cord, a group of rats (n = 10) was given U0126 (5 µg/10 µL IT) twice a day on post-CCI days 4 to 9 to mimic the effect of EA. Five percent DMSO (10 µL IT) was given as vehicle control in the other 4 groups. Our results showed that mechanical allodynia and thermal hyperalgesia occurred on day 3 after CCI (P < 0.001, when compared with sham-CCI; Fig. 3, 3A and 3B). Both administration of U0126 and EA at treatment acupoints significantly increased TWL and MWT of ipsilateral hindpaws from day 5 (P < 0.001, U0126 versus CCI; P = 0.007 for MWT and P = 0.002 for TWL, EA versus CCI) to day 9 (P < 0.001 when compared with CCI). Compared with the CCI group, the NA-EA group did not show a statistically significant analgesic effect.
Inhibition of ERK1/2 Activation May Be Involved in the Effect of EA at Acupoint Treatments on Downregulating the P2X3 Receptor Expression in the Spinal Cord of Rats with CCI
The activation of ERK1/2 and P2X3 receptor protein level was significantly higher in the spinal cord in the CCI group than in the sham-CCI group (P < 0.001; Fig. 4, 4A and 4B). Compared with the CCI group, ERK1/2 activation and P2X3 receptor protein level in the EA group and U0126 group were significantly decreased (P < 0.001 EA versus CCI for P2X3 protein expression; P = 0.004 EA versus CCI for p-ERK1/2 expression; P < 0.001 U0126 versus CCI). The mRNA expression of P2X3 receptor in the spinal cord was significantly increased in the CCI group compared with the sham-CCI (P < 0.001; Fig. 4C). Compared with the CCI group, the mRNA expression of P2X3 receptor in the EA group and in the U0126 group was significantly decreased (P < 0.001). These data demonstrated that inhibition of ERK1/2 activation may be partly involved in the effect of EA at acupoint treatment on downregulating P2X3 receptor expression in the spinal cord of rats with CCI.
Our findings showed that 2 Hz or 15 Hz EA at acupoints significantly attenuates CCI-induced hyperalgesia and allodynia. Moreover, we demonstrated that EA at SB36-GB34 acupoints significantly attenuated CCI-induced upregulation of protein and mRNA of P2X3 receptor in the spinal cord. In addition, EA administration and U0126 IT injection exhibited potency in decreasing ERK1/2 activation as well as protein and mRNA of P2X3 receptor in the spinal cord. The data indicated that the effect of EA on NP behavior may be mediated by the attenuation of ERK1/2 activation and P2X3 receptor synthesis in the spinal cord.
ATP released from injured tissue or the spinal cord is implicated in the initiation of pain by activating P2X3 receptors on sensory nerve terminals.30,31 In the present study, the protein and mRNA level of P2X3 receptor significantly increased in the spinal cord of rats with CCI injury, which is consistent with previous studies.32 Persistent nerve injury causes long-lasting excitement to nociceptive neurons, which relates to ATP-induced P2X3 receptors activation and could cause activation of the neurokinin 1 receptor in the spinal cord and glutamate release from presynaptic nerve endings.33 These mechanisms participate in changing neuron plasticity and the maintenance of NP. Our results showed that CCI-induced hyperalgesia and allodynia and an increase in protein and mRNA expression of P2X3 receptor in the spinal cord were significantly attenuated by EA stimulation. These results suggest that EA treatment enhances the analgesic effect at least partly by decreasing the expression of P2X3 receptor in the spinal cord of CCI rats. Notably, it was recently reported that EA can relieve NP by increasing P2X3 receptor expression in the lateral periaqueductal gray, the upregulation of which inhibits the progression of NP.34 All in all, these studies delineated the crucial role of P2X3 receptor in EA’s analgesic effect.
Several studies have shown various responses to different frequencies of EA stimulation. Two hertz had a better analgesic effect on NP than 100 Hz.35 Fifteen hertz EA had a significant analgesic effect on hyperalgesia induced by arthritis in rats compared with 100 Hz EA treatment.36 In a rat model of complete Freund adjuvant-induced hyperalgesia, HEA-induced potent and shorter-lasting analgesic effects were observed, whereas LEA induced moderate and longer-lasting effects.37 In addition, EA at different frequencies induces the release of different types of endogenous opioids.38 In the present study, we observed 2 Hz EA had a better analgesic effect by increasing TWL and MWT significantly on day 9 after CCI compared with 15 Hz EA treatment. The expression of P2X3 receptor in rats treated with 2 Hz EA was significantly lower than that in rats with 15 Hz EA treatment. These findings may have implications for the use of EA in various clinical practice settings.
Furthermore, in the spinal cord, intense peripheral noxious stimulation induces ERK activation, which contributes to central sensitization.39 The inhibition of ERK activation reduces the behavioral measures of activity-dependent central sensitization.40 Consistent with behavioral pain hypersensitivities, partial sciatic nerve ligation, spinal nerve ligation, and CCI of the sciatic nerve–induced persistent ERK activation in the spinal cord.41–43 Studies have shown that activation of ERK, not p38 MAPK in the spinal cord, contributes to the induction phase and early maintenance phase of long-lasting allodynia induced by IT administration of ATP.44 It was also shown that the depression of ERK activation in the spinal cord may contribute to EA’s analgesic effect.21 In our results, ERK1/2 activation and P2X3 receptor expression were increased in the spinal cord of rats after CCI, and EA administration decreased these effects. Moreover, the IT administration of U0126 increased TWL and MWT and decreased the ERK1/2 activation as well as P2X3 receptor protein and mRNA expression in the spinal cord of rats after CCI, which confirmed the effect of ERK1/2 on regulating P2X3 receptor expression.
In addition, several studies have demonstrated that EA exhibited therapeutic effects on specific acupoints.45,46 Modulation of the limbic system may differentiate between specific and nonspecific components of acupuncture.47 Acupuncture at acupoints induces specific patterns of brain activity.48 In the present study, however, there were no statistically significant changes in nociceptive behaviors and P2X3 protein and mRNA expression in EA between the nonacupoint and the CCI groups.
In conclusion, the current study demonstrated that EA produces a significant analgesic effect on nerve injury–induced NP, and this effect may be mediated through regulation of spinal ERK1/2/P2X3 signaling pathway.
Name: Jianbo Yu, PhD.
Contribution: This author helped design and conduct the study, analyze the data, and write the manuscript.
Attestation: Jianbo Yu has seen the original study data, reviewed the analysis of the data, approved the final manuscript, and is the author responsible for archiving the study files.
Name: Cong Zhao, MD.
Contribution: This author helped design and conduct the study, analyze the data, and write the manuscript.
Attestation: Cong Zhao has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
Name: Xiaoqin Luo, MD.
Contribution: This author helped analyze the data and write the manuscript.
Attestation: Xiaoqin Luo has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
This manuscript was handled by: Jianren Mao, MD, PhD.
1. Ji RR, Strichartz G. Cell signaling and the genesis of neuropathic pain. Sci STKE. 2004;2004:reE14
2. McCleane G. Pharmacological management of neuropathic pain. CNS Drugs. 2003;17:1031–43
3. Colombo B, Annovazzi PO, Comi G. Medications for neuropathic pain: current trends. Neurol Sci. 2006;27 Suppl 2:S183–9
4. Burnstock G. Purine and pyrimidine receptors. Cell Mol Life Sci. 2007;64:1471–83
5. Sawynok J. Adenosine and ATP receptors. Handb Exp Pharmacol. 2007;177:309–28
6. Chizh BA, Illes P. P2X Receptors and Nociception. Pharmacol Rev. 2001;53:4553–68
7. Wirkner K, Sperlagh B, Illes P. P2X3 receptor involvement in pain states. Mol Neurobiol. 2007;36:165–83
8. Park JH, Han JB, Kim SK, Park JH, Go DH, Sun B, Min BI. Spinal GABA receptors mediate the suppressive effect of electroacupuncture on cold allodynia in rats. Brain Res. 2010;1322:24–9
9. Aloe L, Manni L. Low-frequency electro-acupuncture reduces the nociceptive response and the pain mediator enhancement induced by nerve growth factor. Neurosci Lett. 2009;449:173–7
10. Park JH, Kim SK, Kim HN, Sun B, Koo S, Choi SM, Bae H, Min BI. Spinal cholinergic mechanism of the relieving effects of electroacupuncture on cold and warm allodynia in a rat model of neuropathic pain. J Physiol Sci. 2009;59:291–8
11. Lau WK, Lau YM, Zhang HQ, Wong SC, Bian ZX. Electroacupuncture versus celecoxib for neuropathic pain in rat SNL model. Neuroscience. 2010;170:655–61
12. Astin JA, Marie A, Pelletier KR, Hansen E, Haskell WL. A review of the incorporation of complementary and alternative medicine by mainstream physicians. Arch Intern Med. 1998;158:2303–10
13. Huang C, Wang Y, Han JS, Wan Y. Characteristics of electroacupuncture-induced analgesia in mice: variation with strain, frequency, intensity and opioid involvement. Brain Res. 2002;945:20–5
14. Lao L, Zhang RX, Zhang G, Wang X, Berman BM, Ren K. A parametric study of electroacupuncture on persistent hyperalgesia and Fos protein expression in rats. Brain Res. 2004;1020:18–29
15. Lin JG, Lo MW, Wen YR, Hsieh CL, Tsai SK, Sun WZ. The effect of high and low frequency electroacupuncture in pain after lower abdominal surgery. Pain. 2002;99:509–14
16. Han JS. Acupuncture: neuropeptide release produced by electrical stimulation of different frequencies. Trends Neurosci. 2003;26:17–22
17. Kim SK, Park JH, Bae SJ, Kim JH, Hwang BG, Min BI, Park DS, Na HS. Effects of electroacupuncture on cold allodynia in a rat model of neuropathic pain: mediation by spinal adrenergic and serotonergic receptors. Exp Neurol. 2005;195:430–6
18. Johnson GL, Lapadat R. Mitogen-activated protein kinase pathways mediated by ERK, JNK, and p38 protein kinases. Science. 2002;298:1911–2
19. Obata K, Noguchi K. MAPK activation in nociceptive neurons and pain hypersensitivity. Life Sci. 2004;74:2643–53
20. Ji R, Gereau RW IV. MAP kinase and pain. Brain Res Rev. 2009;60:135–48
21. Wang CL, Wang SX, Xu XY. Effects of electroacupuncture at jiajion phosphorylated ERK and NK-1 signal conduction pathway in dorsal hornin complete Freund’s adjuvant arthritis rats. JTCM. 2006;47:348–51
22. Xiang Z, Xiong Y, Yan N, Li X, Mao Y, Ni X, He C, LaMotte RH, Burnstock G, Sun J. Functional up-regulation of P2X 3 receptors in the chronically compressed dorsal root ganglion. Pain. 2008;140:23–34
23. Noguchi K, Obata K, Dai Y. Changes in DRG neurons and spinal excitability in neuropathy. Novartis Found Symp. 2004;261:103–10; discussion 110–5, 149–54
24. Bennett GJ, Xie YK. A peripheral mononeuropathy in rat that produces disorders of pain sensation like those seen in man. Pain. 1988;33:87–107
25. Yaksh TL, Rudy TA. Chronic catheterization of the spinal subarachnoid space. Physiol Behav. 1976;17:1031–6
26. Lai M, Wang SM, Wang Y, Tang CL, Kong LW, Xu XY. [Effects of electroacupuncture of “Zusanli” (ST 36), “Hegu” (LI 4) and/or “Sanyinjiao” (SP 9) on immunofunction in gastric carcinectomy rats]. Zhen Ci Yan Jiu. 2008;33:245–9
27. Wang K, Wu H, Wang G, Li M, Zhang Z, Gu G. The effects of electroacupuncture on TH1/TH2 cytokine mRNA expression and mitogen-activated protein kinase signaling pathways in the splenic T cells of traumatized rats. Anesth Analg. 2009;109:1666–73
28. Hayes AG, Skingle M, Tyers MB. Reversal by beta-funaltrexamine of the antinociceptive effect of opioid agonists in the rat. Br J Pharmacol. 1986;88:867–72
29. Pfaffl MW. A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 2001;29:2002–7
30. Chen CC, Akopian AN, Sivilotti L, Colquhoun D, Burnstock G, Wood JN. A P2X purinoceptor expressed by a subset of sensory neurons. Nature. 1995;377:428–31
31. Burnstock G. Purine-mediated signalling in pain and visceral perception. Trends Pharmacol Sci. 2001;22:182–8
32. Novakovic SD, Kassotakis LC, Oglesby IB, Smith JA, Eglen RM, Ford AP, Hunter JC. Immunocytochemical localization of P2X3 purinoceptors in sensory neurons in naive rats and following neuropathic injury. Pain. 1999;80:273–82
33. Gu JG, MacDermott AB. Activation of ATP P2X receptors elicits glutamate release from sensory neuron synapses. Nature. 1997;389:749–53
34. Xiao Z, Ou S, He WJ, Zhao YD, Liu XH, Ruan HZ. Role of midbrain periaqueductal gray P2X3 receptors in electroacupuncture-mediated endogenous pain modulatory systems. Brain Res. 2010;1330:31–44
35. Xing G, Liu F, Wan Y, Yao L, Han J. [Electroacupuncture of 2 Hz induces long-term depression of synaptic transmission in the spinal dorsal horn in rats with neuropathic pain]. Beijing Da Xue Xue Bao. 2003;35:453–7
36. Cao W, Deng Y, Dong X, Wang Y, Lu Z. [Effects of electroacupuncture at different frequencies on the nociceptive response and central contents of GABA and glutamic acid in arthritic rats]. Zhen Ci Yan Jiu. 1993;18:48–52
37. Xiao Z, Ou S, He WJ, Zhao YD, Liu XH, Ruan HZ. A parametric study of electroacupuncture on persistent hyperalgesia and Fos protein expression in rats. Brain Res. 2004;1020:18–29
38. Han JS, Sunven-names SL. Differential release of enkephalin and dynorphin by low and high frequency electroacupuncture in the central nervous system. Acupunct Sci Int J. 1990;1:19–27
39. Ji RR, Baba H, Brenner GJ, Woolf CJ. Nociceptive-specific activation of ERK in spinal neurons contributes to pain hypersensitivity. Nat Neurosci. 1999;2:1114–9
40. Karim F, Hu HJ, Adwanikar H, Kaplan D, Gereau RW 4th. Impaired inflammatory pain and thermal hyperalgesia in mice expressing neuron-specific dominant negative mitogen activated protein kinase kinase (MEK). Mol Pain. 2006;2:2
41. Ma W, Quirion R. Partial sciatic nerve ligation induces increase in the phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) in astrocytes in the lumbar spinal dorsal horn and the gracile nucleus. Pain. 2002;99:175–84
42. Zhuang ZY, Gerner P, Woolf CJ, Ji RR. ERK is sequentially activated in neurons, microglia, and astrocytes by spinal nerve ligation and contributes to mechanical allodynia in this neuropathic pain model. Pain. 2005;114:149–59
43. Song XS, Cao JL, Xu YB, He JH, Zhang LC, Zeng YM. Activation of ERK/CREB pathway in spinal cord contributes to chronic constrictive injury-induced neuropathic pain in rats. Acta Pharmacol Sin. 2005;26:789–98
44. Nakagawa T, Wakamatsu K, Maeda S. Differential contribution of spinal mitogen-activated protein kinases to the phase of long-lasting allodynia evoked by intrathecal administration of ATP in rats. Biol Pharm Bull. 2008;31:1164–8
45. Cha MH, Choi JS, Bai SJ, Shim I, Lee HJ, Choi SM, Lee BH. Antiallodynic effects of acupuncture in neuropathic rats. Yonsei Med J. 2006;47:359–66
46. Choi EM, Jiang F, Longhurst JC. Point specificity in acupuncture. Chin Med. 2012;7:4
47. Dhond RP, Kettner N, Napadow V. Do the neural correlates of acupuncture and placebo effects differ? Pain. 2007;128:8–12
48. Yan B, Li K, Xu J, Wang W, Li K, Liu H, Shan B, Tang X. Acupoint-specific fMRI patterns in human brain. Neurosci Lett. 2005;383:236–40
© 2013 International Anesthesia Research Society
What does "Remember me" mean?
By checking this box, you'll stay logged in until you logout. You'll get easier access to your articles, collections,
media, and all your other content, even if you close your browser or shut down your
To protect your most sensitive data and activities (like changing your password),
we'll ask you to re-enter your password when you access these services.
What if I'm on a computer that I share with others?
If you're using a public computer or you share this computer with others, we recommend
that you uncheck the "Remember me" box.
Data is temporarily unavailable. Please try again soon.
Readers Of this Article Also Read