Journal of Traditional Chinese Medicine ›› 2024, Vol. 44 ›› Issue (4): 660-669.DOI: 10.19852/j.cnki.jtcm.20240626.003
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Adenosine triphosphate mediates the pain tolerance effect of manual acupuncture at Zusanli (ST36) in mice
LI Zhongzheng1,2, ZHAO Yadan3, Ma Weigang1,2, Zhang Yonglong1,2, XU Zhifang1,2, XI Qiang5, LI Yanqi5, QIN Siru1,2, ZHANG Zichen1,2, WANG Songtao1,2, ZHAO Xue4, LIU Yangyang4, GUO Yi1,2(
), GUO Yongming4()
- 1 Research Center of Experimental Acupuncture Science, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China
2 National Clinical Research Center for Chinese Medicine Acupuncture and Moxibustion, Tianjin 301617, China
3 Department of Acupuncture and Moxibustion, Qingdao Hiser Hospital Affiliated of Qingdao University (Qingdao Traditional Chinese Medicine Hospital), Qingdao 266033, China
4 School of Acupuncture and Moxibustion and Tuina, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China
5 School of Traditional Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China
-
Received:2023-01-11Accepted:2023-07-24Online:2024-08-15Published:2024-06-26 -
Contact:GUO Yi, Research Center of Experimental Acupuncture Science, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China; National Clinical Research Center for Chinese Medicine Acupuncture and Moxibustion, Tianjin 301617, China.guoyi_168@163.com Telephone: +86-13920921016; GUO Yongming, School of Acupuncture and Moxibustion and Tuina, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China,guoymxr@163.com Telephone: +86-13920619116 -
Supported by:National Key R&D Program of China: Biological Mechanisms of Acupoint Function-Effect Associations(2019YFC1709003);National Natural Science Foundation of China (NSFC) Top-level Project: Study on the Neuroimmunological Mechanism of Macrophage Phenotypic Polarisation for Anti-inflammation Regulated by Acupuncture(81873369);National Natural Science Foundation of China Young Science Fund Project: Study on the Neuromodulation Mechanism of Electroacupuncture to Improve Neutropenia after Chemotherapy for Lung Cancer(81704146);National Natural Science Foundation of China Key Project: Research on the Initial Kinetic Regulation Mechanism of Acupuncture Effect Based on the Physicochemical Coupling Network of Acupuncture Point Microenvironment(82030125)
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LI Zhongzheng, ZHAO Yadan, Ma Weigang, Zhang Yonglong, XU Zhifang, XI Qiang, LI Yanqi, QIN Siru, ZHANG Zichen, WANG Songtao, ZHAO Xue, LIU Yangyang, GUO Yi, GUO Yongming. Adenosine triphosphate mediates the pain tolerance effect of manual acupuncture at Zusanli (ST36) in mice[J]. Journal of Traditional Chinese Medicine, 2024, 44(4): 660-669.
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Table 1 Comparison of pain tolerance threshold before and after acupuncture in mice of every group (s, $\bar{x}±s$)
| Group | n | PTT before MA | PTT after MA |
|---|---|---|---|
| A | 9 | 7.4±1.7 | 9.8±2.3a |
| B | 9 | 6.5±1.3 | 8.2±2.2a |
| C | 9 | 6.4±0.7 | 7.9±2.2a |
| D | 9 | 7.0±1.2 | 10.1±2.0a |
| E | 9 | 7.0±1.1 | 8.4±1.3a |
Table 1 Comparison of pain tolerance threshold before and after acupuncture in mice of every group (s, $\bar{x}±s$)
| Group | n | PTT before MA | PTT after MA |
|---|---|---|---|
| A | 9 | 7.4±1.7 | 9.8±2.3a |
| B | 9 | 6.5±1.3 | 8.2±2.2a |
| C | 9 | 6.4±0.7 | 7.9±2.2a |
| D | 9 | 7.0±1.2 | 10.1±2.0a |
| E | 9 | 7.0±1.1 | 8.4±1.3a |
Figure 1 Adenylate content at Zusanli (ST36) was increased after MA in a dose-dependent manner A: flow chart of experimental intervention. Intervention stages are represented by different symbols: ○: determination of basal PTT; ■: detection of adenylate at Zusanli (ST36); ?: MA intervention; ▲: needle removal. After measurements of basal PTT, different MA interventions were performed in mice. B: ATP content at Zusanli (ST36) in each group; C: Scatter plot depicting the correlation between PTT and ATP concentration at Zusanli (ST36); D: ADP content at Zusanli (ST36) in each group; E: AMP content at Zusanli (ST36) in each group. F: Adenylate EC content at Zusanli (ST36) in each group. G: Total adenylate pool content at Zusanli (ST36) in each group. Control (CTL) group: direct detection of adenylate content at Zusanli (ST36). Group A: needle twisted immediately for 2 min. Group B: needle twisted immediately for 2 min and retained for 8 min. Group C: needle twisted immediately for 2 min and retained for 18 min. Group D: needle twisted immediately for 2 min and retained for 28 min. Group E: needle twisted immediately for 2 min and retained for 28 min before removal followed by a 10-minute rest. Adenylate content at Zusanli (ST36) was measured after MA intervention. MA: manual acupuncture; PTT: pain tolerance threshold; ATP: adenosine triphosphate; ADP: adenosine diphosphate; AMP: adenosine monophosphate; EC: energy charge. Data are presented as mean ± standard deviation (n = 10 per group). The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group D, aP < 0.01 and bP < 0.05; Compared with group B, cP < 0.05.
Figure 1 Adenylate content at Zusanli (ST36) was increased after MA in a dose-dependent manner A: flow chart of experimental intervention. Intervention stages are represented by different symbols: ○: determination of basal PTT; ■: detection of adenylate at Zusanli (ST36); ?: MA intervention; ▲: needle removal. After measurements of basal PTT, different MA interventions were performed in mice. B: ATP content at Zusanli (ST36) in each group; C: Scatter plot depicting the correlation between PTT and ATP concentration at Zusanli (ST36); D: ADP content at Zusanli (ST36) in each group; E: AMP content at Zusanli (ST36) in each group. F: Adenylate EC content at Zusanli (ST36) in each group. G: Total adenylate pool content at Zusanli (ST36) in each group. Control (CTL) group: direct detection of adenylate content at Zusanli (ST36). Group A: needle twisted immediately for 2 min. Group B: needle twisted immediately for 2 min and retained for 8 min. Group C: needle twisted immediately for 2 min and retained for 18 min. Group D: needle twisted immediately for 2 min and retained for 28 min. Group E: needle twisted immediately for 2 min and retained for 28 min before removal followed by a 10-minute rest. Adenylate content at Zusanli (ST36) was measured after MA intervention. MA: manual acupuncture; PTT: pain tolerance threshold; ATP: adenosine triphosphate; ADP: adenosine diphosphate; AMP: adenosine monophosphate; EC: energy charge. Data are presented as mean ± standard deviation (n = 10 per group). The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group D, aP < 0.01 and bP < 0.05; Compared with group B, cP < 0.05.
Figure 2 Effects of acupuncture on PTT were mediated by ATP and the P2X3 receptor at Zusanli (ST36) A: flow chart of experimental intervention. Intervention stages are represented by different symbols; ?: Acupoint injection of saline; ▲: Acupoint injection of ATP; ●: Determination of PTT after intervention; ?: MA intervention; B: comparison of PTT in each group. PTT of mice in the Control (CTL) group was measured to indicate the basal level. PTT of mice in the Sal, ATP-1, ATP-2, and ATP-3 groups was measured after acupoint injection of saline, ATP-1 (26 μg/2.6 μL), ATP-2 (52 μg/2.6 μL), and ATP-3 (104 μg/2.6 μL), respectively. PTT of mice in the Acu group was measured after MA intervention (needle was twisted for 2 min and retained for 28 min). Mice in the Acu + Sal, Acu + ATP-1, Acu + ATP-2, and Acu + ATP-3 groups were administered MA (intervention method was the same as that in the Acu group) after acupoint injection of saline, ATP-1 (26 μg/2.6 μL), ATP-2 (52 μg/2.6 μL), and ATP-3 (104 μg/2.6 μL), respectively, followed by measurement of PTT. ATP: adenosine triphosphate; PTT: pain tolerance threshold; P2X3: peripheral purinergic P2X receptor 3; MA: manual acupuncture. Data are presented as mean ± standard deviation (n = 10 per group). The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group Acu, aP < 0.05.
Figure 2 Effects of acupuncture on PTT were mediated by ATP and the P2X3 receptor at Zusanli (ST36) A: flow chart of experimental intervention. Intervention stages are represented by different symbols; ?: Acupoint injection of saline; ▲: Acupoint injection of ATP; ●: Determination of PTT after intervention; ?: MA intervention; B: comparison of PTT in each group. PTT of mice in the Control (CTL) group was measured to indicate the basal level. PTT of mice in the Sal, ATP-1, ATP-2, and ATP-3 groups was measured after acupoint injection of saline, ATP-1 (26 μg/2.6 μL), ATP-2 (52 μg/2.6 μL), and ATP-3 (104 μg/2.6 μL), respectively. PTT of mice in the Acu group was measured after MA intervention (needle was twisted for 2 min and retained for 28 min). Mice in the Acu + Sal, Acu + ATP-1, Acu + ATP-2, and Acu + ATP-3 groups were administered MA (intervention method was the same as that in the Acu group) after acupoint injection of saline, ATP-1 (26 μg/2.6 μL), ATP-2 (52 μg/2.6 μL), and ATP-3 (104 μg/2.6 μL), respectively, followed by measurement of PTT. ATP: adenosine triphosphate; PTT: pain tolerance threshold; P2X3: peripheral purinergic P2X receptor 3; MA: manual acupuncture. Data are presented as mean ± standard deviation (n = 10 per group). The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group Acu, aP < 0.05.
Figure 3 Systemici pre-treatment wth ATP ameliorated PTT in mice A: dynamic adenylate EC content of mice after intraperitoneal (i.p.) injections of ATP within 70 min (n = 5 per group). B: flow chart of experimental intervention. Intervention stages are represented by different symbols: ▲: i.p. injection of ATP; ?: i.p. injection of saline; ?: MA intervention; ●: determination of PTT after intervention; C: comparison of PTT in each group (n = 15 per group). PTT of mice in the Control (CTL) group was measured after 30 min. PTT of mice in the Sal and ATP-5(4 μg/g) groups was measured 30 min after i.p. administration of saline and ATP-5, respectively. PTT of mice in the Acu group was measured 30 min after MA intervention (needle was twisted for 2 min and retained for 28 min). Mice in the Acu + Sal and Acu + ATP-5 groups received i.p. injections of saline and ATP, respectively. Mice were subjected to MA 30 min later (the intervention method was the same as that in the Acu group) followed by measurement of PTT. ATP: adenosine triphosphate; PTT: pain tolerance threshold; MA: Manual acupuncture; EC: energy charge. Data are presented as mean ± standard deviation. The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group Acu, aP < 0.05.
Figure 3 Systemici pre-treatment wth ATP ameliorated PTT in mice A: dynamic adenylate EC content of mice after intraperitoneal (i.p.) injections of ATP within 70 min (n = 5 per group). B: flow chart of experimental intervention. Intervention stages are represented by different symbols: ▲: i.p. injection of ATP; ?: i.p. injection of saline; ?: MA intervention; ●: determination of PTT after intervention; C: comparison of PTT in each group (n = 15 per group). PTT of mice in the Control (CTL) group was measured after 30 min. PTT of mice in the Sal and ATP-5(4 μg/g) groups was measured 30 min after i.p. administration of saline and ATP-5, respectively. PTT of mice in the Acu group was measured 30 min after MA intervention (needle was twisted for 2 min and retained for 28 min). Mice in the Acu + Sal and Acu + ATP-5 groups received i.p. injections of saline and ATP, respectively. Mice were subjected to MA 30 min later (the intervention method was the same as that in the Acu group) followed by measurement of PTT. ATP: adenosine triphosphate; PTT: pain tolerance threshold; MA: Manual acupuncture; EC: energy charge. Data are presented as mean ± standard deviation. The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group Acu, aP < 0.05.
Figure 4 Pretreatment with adenosine disodium ameliorated the effects of acupuncture A: flow chart of experimental intervention. Intervention stages are represented by different symbols: ?: intraperitoneal (i.p.) injection of saline; ▲: i.p. injection of adenosine disodium (represented by AD); ▲: i.m. injection of saline; ?: i.m. injection of AD; ●: determination of PTT after intervention; ?: MA intervention; B: comparison of PTT in each group. PTT of mice in the Control (CTL) group was measured to indicate the basal level. PTT of mice in the Sal (i.p.), Sal (i.m.), AD (i.p.), and AD (i.m.) groups was measured after i.p. or i.m. injections of saline or AD. PTT of mice in the Acu group was measured after MA intervention (needle was twisted for 2 min and retained for 28 min). Mice in the Acu + AD (i.p.) and Acu + AD (i.m.) groups were subjected to MA (the intervention method was the same as that in the Acu group) after i.p. or i.m. injections of AD, respectively, followed by measurement of PTT. MA: Manual acupuncture; PTT: pain tolerance threshold; AD: adenosine disodium. Data are presented as mean ± standard deviation (n = 10 per group). The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group Acu, aP < 0.05; compared with group Acu +AD (i.p.), cP < 0.05; compared with group AD (i.m.), bP < 0.05.
Figure 4 Pretreatment with adenosine disodium ameliorated the effects of acupuncture A: flow chart of experimental intervention. Intervention stages are represented by different symbols: ?: intraperitoneal (i.p.) injection of saline; ▲: i.p. injection of adenosine disodium (represented by AD); ▲: i.m. injection of saline; ?: i.m. injection of AD; ●: determination of PTT after intervention; ?: MA intervention; B: comparison of PTT in each group. PTT of mice in the Control (CTL) group was measured to indicate the basal level. PTT of mice in the Sal (i.p.), Sal (i.m.), AD (i.p.), and AD (i.m.) groups was measured after i.p. or i.m. injections of saline or AD. PTT of mice in the Acu group was measured after MA intervention (needle was twisted for 2 min and retained for 28 min). Mice in the Acu + AD (i.p.) and Acu + AD (i.m.) groups were subjected to MA (the intervention method was the same as that in the Acu group) after i.p. or i.m. injections of AD, respectively, followed by measurement of PTT. MA: Manual acupuncture; PTT: pain tolerance threshold; AD: adenosine disodium. Data are presented as mean ± standard deviation (n = 10 per group). The single factor analysis of variance of completely randomized design was used for parameter comparison between groups. Compared with group Acu, aP < 0.05; compared with group Acu +AD (i.p.), cP < 0.05; compared with group AD (i.m.), bP < 0.05.
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