Effect of manual acupuncture on bowel motility in normal kunming mouse

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1 Online Submissions: J Trdit Chin Med 215 April 15; 5(2): info@journltcm.com ISSN JTCM. All rights reserved. EXPERIMENTAL STUDY TOPIC Effect of mnul cupuncture on bowel motility in norml kunming mouse Zho Yuxue, Cui Chngxing, Qin Qinggung, Ben Hui, Go Junhong, Yu Xiochun, Zhu Bing Zho Yuxue, Cui Chngxing, Qin Qinggung, Ben Hui, Go Junhong, Yu Xiochun, Zhu Bing, Institute of Acupuncture nd Moxibustion, Chin Acdemy of Chinese Medicl Sciences, Beijing 17, Chin Supported by Ntionl Bsic Reserch Progrm of Chin: Mechnisms of Dul Effects of Acupuncture on Functionl Intestinl Disese (97 Progrm, No. 211CB552), the Ntionl Nturl Science Foundtion of Chin Reserch Grnts: Investigtions on Effect of Acupuncture In Functionl Bowel Disorders Vi CRF Signling Pthwy (No ) Correspondence to: Prof. Zhu Bing, Institute of Acupuncture nd Moxibustion, Chin Acdemy of Chinese Medicl Sciences, Beijing 17, Chin. zhubing@mil.cintcm.c.cn Telephone: Accepted: June 22, 214 Abstrct OBJECTIVE: To observe the effects of cupuncture on motility of jejunum nd distl colon t different cupoints on norml Kunming mice. METHODS: The jejunl nd colorectl motor ctivity of forty Kunming mice were recorded by the intr-intestinl cpsule. The cpsules contined with distilled wter were plced into the cvities of jejunum nd distl colon. The chnges of bowel were compred with the bckground ctivity recorded before ny stimultion. The chosen cupoints were s follows: Zusnli (ST 6) on the lower legs, Tinshu (ST 25) on the mid-lower bdomen. RESULTS: Mnul cupuncture (MA) t Zusnli (ST 6) significntly promoted the motility of jejunum nd distl colon (mnifested s incresed chnge rte of verge mplitude nd incresed chnge rte of men re under the contrctile curve) of norml Kunming mice (P <.1). MA t Tinshu (ST 25) significntly inhibited the motility of jejunum (mnifested s decresed chnge rte of verge mplitude, decresed chnge rte of men re under the contrctile curve, nd the reduced frequency) of norml Kunming mice (P <.1), while significntly incresed the distl colonic motility (mnifested s incresed mplitude nd incresed men re under the contrctile curve) of norml Kunming mice (P <.1). CONCLUSION: MA t specific cupoints [Zusnli (ST 6), Tinshu (ST 25)] hs different effects on the motility of jejunum nd distl colon, which might involve the segmentl innervtions of the relted nerve. 215 JTCM. All rights reserved. Key words: Hnd cupuncture; Point ST 6 (Zusnli); Point ST 25 (Tinshu); Gstrointestinl motility INTRODUCTION Acupuncture hs been used in Chin for more thn yers. 1,2 It hs been tried ginst vrious diseses, including gstrointestinl (GI) disorders. A lrge mount of experimentl nd clinicl evidence indicte the effectiveness of cupuncture in treting GI diseses, such s dirrhe,,4 constiption, 5,6 irritble bowel syndrome (IBS), 7-9 nd functionl bowel disorder (FBD). 1 Though there were vst reserches on the cupuncture tretment of bowel diseses, the evidence bout the role of cupuncture on bowel motility of rodents in norml stte, especilly for the mice, did not bound. Previous studies hve demonstrted the effects of cupuncture stimultion on gstric motility in nesthe- JTCM www. journltcm. com 227 April 15, 215 Volume 5 Issue 2

2 Zho YX et l. / Experimentl Study tized rts. 11,12 Nevertheless, rre experiments were crried out to record the motility of jejunum nd colon of rodents in vivo, nd even less studies relting to the effects of cupuncture on tht kind of bowel motility were reported. The ims of this study were to investigte the effects nd underlying mechnisms of cupuncture Zusnli (ST 6) or Tinshu (ST 25) on intestinl nd colorectl motility of Kunming mice in norml stte. MATERIALS AND METHODS Animls Adult mle mice of specific pthogen-free grde [Kunming mice, (2 ± 2) g] were obtined from Chin Acdemy of Militry Science [Certificte of qulity No. SCXK-(Army) 212-4]. Animls were housed -4/cges nd mintined under controlled conditions of temperture (22-25) nd light (6 AM on, 6 PM off) nd fed d libitum with stndrd rodent chow nd tp wter. All protocols were pproved by the Institutionl Animl Cre nd Use Committee of Chin Acdemy of Chinese Medicl Sciences. Drugs nd Instruments Urethne ws purchsed from Sinophrm Chemicl Regent Co., Ltd., (Shnghi, Chin). Miniturized solid-stte pressure trnsducer ctheter TSD14A (Biopc Inc., Golet, CA, USA) nd trnsducer mplifier MP15CE were from Biopc Inc (Golet, CA, USA). Trnsducer control unit 141, Micro 141 interfce, nd Spike 2 version 7 dt cquisition softwre were from Cmbridge Electronic Design Limited (Cmbridge, Englnd). Acupuncture needles (. mm in dimeter) were from Suzhou Medicl Applince Fctory (Suzhou, Chin). Animl preprtion Mice were nesthetized with urethne [1%, 2 g/kg, intrperitonelly (ip)]. A miniturized solid-stte pressure trnsducer ctheter with wter cpsule in the end ws inserted into the lumen of the jejunum through the incision on the intestinl surfce. Another ctheter of the sme type ws inserted into the distl colon through the nus. The cpsules were plced 1.5 nd 1 cm bove the incision of jejunum nd the nus, respectively. Dt cquisition nd nlysis Ech miniture pressure trnsducer ctheter ws connected to trnsducer control unit, whose output signl ws subsequently mplified further using trnsducer mplifier in differentil mode. This output ws cquired vi Micro 141 interfce nd recorded using Spike 2 version 7 dt cquisition softwre. Experimentl protocols All experimentl studies were conducted in dult overnight fsted mice which were nesthetized with urethne (1%, 2 g/kg, ip) nd there fter kept on thermostticlly controlled electric heting pd for experimentl niml to mintin body temperture within Thirty mice were rndomly designted into two groups: Jejunum recording group (n = 15), nd Distl colon recording group (n = 15). Mice were left undisturbed for t lest min fter intr-intestinl plcement of the pressure trnsducer ctheter nd the wter cpsule. At the end of the experiment, ll of the mice were euthnized by n overdose of ip urethne (4%). Acupuncture procedure in nesthetized mice After recording of bsl intestinl contrctions for t lest min, cupuncture ws performed. Needles were inserted to depth of 5 mm into the skin nd underlying muscles t either Zusnli (ST 6) or Tinshu (ST 25) points. Zusnli (ST 6) is locted t 5 mm lterl nd lower from the nterior tubercle of the tibi in mice. 1 Tinshu (ST 25) is locted 2 mm bove the symphysis pubis nd 5 mm lterl from the midline in mice. 14 For ech mouse in both two groups (Jejunum recording group nd Distl colon recording group), cupuncture needles were inserted unilterlly into both Zusnli (ST 6) nd Tinshu (ST 25) cupoints, nd were twisted, lifted nd thrust mnully t frequency of 2 Hz 15 for s (Jejunum recording group) or 6 s (Distl colon recording group). It's importnt to note tht cupuncture stimultion in Zusnli (ST 6) nd Tinshu (ST 25) ws not operted in the sme time. We stimulte one point [Zusnli (ST 6) or Tinshu (ST 25)] for s, then stop stimultion for while, nd stimulte the other point fter the intestinl contrctions returned to the bsl level. Repet such opertion three times. Sttisticl nlysis All the dt in the present study were expressed s mens ± stndrd devition ( xˉ ± s ), nd nlyzed by two-tiled Student's t-test or one-wy nlysis of vrince with SPSS softwre 1. (IBM, Armonk, NY, USA). The lest significnt difference test ws used to compre men vlues mong groups. P <.5 ws considered significnt. RESULTS Effect of mnul cupuncture (MA) on the contrctility of jejunum Spontneous jejunl pressure ws monitored for 2 h fter insertion of the mnometric ctheter into the jejunum cvity in totl of 15 urethne nesthetized Kunming mice (Figure 1). MA t Zusnli (ST 6) significntly promoted the jejunum motility of norml Kunming mice. Both the chnge rte of verge mplitude nd the chnge rte of men re under the contrctile curve (AUC) during JTCM www. journltcm. com 228 April 15, 215 Volume 5 Issue 2

3 Zho YX et l. / Experimentl Study MA ST-6 Stop stimultion A Before MA During MA 6 After MA 9 MA ST-25 Stop stimultion B Chnge rte of mplitude (%) Chnge rte of men AUC (%) Before MA During MA 6 After MA 9 C 25 Bsl vlue ST-6 ST-25 Bsl vlue ST-6 ST-25 Bsl vlue ST-25 Figure 1 Effect of MA on the jejunl contrctility A: the peristltic wves of jejunum were promoted by MA t Zusnli (ST 6) from to 6 s, which is mnifested s incresed chnge rte of verge mplitude nd incresed chnge rte of men re under the contrctile curve; B: the peristltic wves of jejunum were inhibited by MA t Tinshu (ST 25) from to 6 s, which is mnifested s decresed chnge rte of verge mplitude, decresed chnge rte of men re under the contrctile curve, nd the reduced frequency; C: MA Zusnli (ST 6) significntly incresed the chnge rte of verge mplitude. MA Tinshu (ST 25) significntly inhibited the chnge rte of verge mplitude; D: the men frequency ws not significntly chnged during MA t Zusnli (ST 6), compred with tht before MA; E: MA Zusnli (ST 6) significntly incresed the chnge rte of men AUC. MA Tinshu (ST 25) significntly inhibited the chnge rte of men AUC; F: the men frequency ws significntly reduced by MA t Tinshu (ST 25) in contrct to the frequency before MA. P <.1 vs bsl vlue; b P <.5 vs bsl vlue. MA: mnul cupuncture; AUC: re under the contrctile curve. E Frenquence (peks/ s) Frenquence (peks/ s) Bsl vlue ST-6 b D F MA were significntly incresed thn the bsl vlues before MA (2.46% ±.46% vs 1.% ±.%, P =.9; 2.17% ±.19% vs 1.% ±.%, P <.1] The men frequency ws not significntly chnged during MA compred with tht before MA [(21.5 ± 1.44) vs (19.91 ±.98) peks/ s, P =.67]. MA t Tinshu (ST 25) significntly inhibited the jejunum motility of norml Kunming mice. Both the chnge rte of verge mplitude nd the men AUC during MA were significntly decresed thn bsl vlues before MA (.47% ±.% vs 1.% ±.%, P <.1;.49% ±.5% vs 1.% ±.%, P <.1), the men frequency ws significntly reduced JTCM www. journltcm. com 229 April 15, 215 Volume 5 Issue 2

4 Zho YX et l. / Experimentl Study by MA in contrct to the frequency before MA [(14.81 ± 1.6) vs (19.1 ±.72) peks/ s, P =.8]. Effect of MA on the contrctility of distl colon Spontneous colonic pressure ws monitored for 2 h fter insersion of the mnometric ctheter into the distl colon through nus in totl of 15 urethne nesthetized Kunming mice (Figure 2). MA t Zusnli (ST 6) significntly incresed the colonic motility of norml Kunming mice. Both the chnge rte of verge mplitude nd the men AUC during MA were significntly incresed thn bsl vlues before MA (2.52% ±.18% vs 1.% ±.%, P <.1; 1.91% ±.22% vs 1.% ±.%, P =.1), the men frequency ws not significntly chnged during MA compred with tht before MA [(9.7 ± 1.47) vs (8.5 ± 1.9) peks/ s, P =.7]. MA t Tinshu (ST 25) significntly promoted the colonic motility of norml Kunming mice. Both the chnge rte of verge mplitude nd the men AUC MA ST-6 Stop stimultion A Before MA 6 During MA 12 After MA 18 MA ST-25 Stop stimultion B Chnge rte of mplitude (%) Chnge rte of men AUC (%) Before MA 6 During MA 12 After MA 18 C 12 Bsl vlue ST-6 ST-25 E Frenquence (peks/ s) Frenquence (peks/ s) Bsl vlue ST-6 ST-25 Bsl vlue ST-25 Figure 2 Effect of MA on the colonrectl contrctility A: the peristltic wves of distl colon were promoted by MA t Zusnli (ST 6) from 6 to 12 s, which is mnifested s incresed mplitude nd incresed men re under the contrctile curve; B: the peristltic wves of distl colon were promoted by MA t Tinshu (ST 25) from 6 to 12 s, which is mnifested s incresed mplitude nd incresed men re under the contrctile curve; C: both MA t Zusnli (ST 6) nd MA t Tinshu (ST 25) significntly incresed the chnge rte of verge mplitude; D: the men frequency ws not significntly chnged during MA t Zusnli (ST 6); E: both MA t Zusnli (ST 6) nd MA t Tinshu (ST 25) significntly incresed the chnge rte of men AUC; F: the men frequency ws not significntly chnged during MA t Tinshu (ST 25). MA: mnul cupuncture; AUC: re under the contrctile curve. P <.1 vs bsl vlue. JTCM www. journltcm. com 2 April 15, 215 Volume 5 Issue Bsl vlue ST-6 D F

5 Zho YX et l. / Experimentl Study during MA were significntly incresed thn bsl vlues before MA (.1% ±.45% vs 1.% ±.%, P =.1; 2.5% ±.57% vs 1.% ±.%, P =.24), the men frequency ws not significntly chnged during MA compred with tht before MA [(7. ±.4) vs (7. ±.57) peks/ s, P =.552]. DISCUSSION Acupuncture is n empiricl medicine tht hs been prcticed in Estern countries for few thousnd yers 16,17 nd it hs been rpidly ccepted throughout Western countries in recent few decdes. Acupuncture is simple, noticebly effective, nd side-effect-free "green tretment" involving stimultion of the cupoints in the skin by thin, metllic needles tht re mnipulted mnully or combined with electricl stimultion. 18 It hs been pplied to tret vrious diseses, especilly for the tretment of GI disorders. 19 A lot of clinicl trils hve proven the effectiveness of cupuncture in treting GI diseses, such s dirrhe, constiption, IBS, nd FGD. -1 The most commonly used cupuncture points (cupoints) for focused tretment of gstrointestinl symptoms re the Tinshu (ST 25) nd the Zusnli (ST 6) points. Acupuncture t the Tinshu (ST 25) cupoint hs been documented to hve moderting effects for ptients with functionl constiption, 2,21 dirrhe, 22 chronic colitis, 2 nd IBS. 24 Acupuncture t the Zusnli (ST 6) cupoint hs been reported to tret GI diseses s constiption, gstroesophgel reflux, nd IBS Experimentl studies demonstrted the effects of cupuncture stimultion of the vrious segmentl res on gstric motility in nesthetized rts. 11 Acupuncture t the hind limbs stimultes gstric motility, 11,29 while cupuncture t the bdomen inhibits gstric motility. 12 Our previous studies on rts indicted tht cupuncture the points on bdomen produced moderte-strong suppressive responses on gstric motility. Acupuncture the points in hind limb evoked moderte ugmenttion of gstric motility. The cupuncture stimultion with the electric current of -4 ma t the cupoints of Quchi (LI 11) nd Zusnli (ST 6) elicited mildmoderte fcilittion on the gstric motility, wheres the electrocupuncture stimultion with the sme intensity to the cupoints on bdomen brought moderte depression of gstric motility. 1 We proposed tht cupuncture provoked reflex tht hd cutneous nd muscle nerves s its fferent pthwy, nd the sympthetic gstric brnches s its efferent pthwy. Acupuncture the points in bdomen inhibited gstric motility by incresing the ctivity of the efferent nerves of sympthetic gstric brnches. Acupuncture the point in hind limb stimulted gstric motility by incresed vgl efferent. In the present study, prticulr ttention is pid to the effect of MA on intestinl nd colorectl motility of norml Kunming mice, which ws rrely reported before. We chose Zusnli (ST 6) nd Tinshu (ST 25) s the points to stimulte. The results showed tht MA t Zusnli (ST 6) significntly promoted the jejunum motility of norml Kunming mice, with the ugmented chnge rte of verge mplitude nd incresed men AUC. MA t Tinshu (ST 25) significntly inhibited the jejunum motility of norml Kunming mice, with the decresed chnge rte of verge mplitude nd reduced men AUC. MA t Zusnli (ST 6) or Tinshu (ST 25) significntly promoted the colonic motility of norml Kunming mice, with the ugmented chnge rte of verge mplitude nd men AUC. Results of present study indicted tht MA t Zusnli (ST 6) nd MA t Tinshu (ST 25) plyed opposite roles in jejunum motility of norml Kunming mice, wheres the effects of MA t Zusnli (ST 6) nd MA t Tinshu (ST 25) in modulting the colorectl motility were similr. The underlying mechnism involves the innervtions of the cupoint re nd relted role of centrl nervous system. We nmed it s "heterotopic points" nd "homotopic points". The term of "homotopic points" refers to the cupoints linked to the viscerl orgns with the sme segmentl innervtion. And the term of "heterotopic points" refers to the cupoints linked to the viscerl orgns with heterotopic segmentl innervtion. There is closely connection between the exterior of the humn body nd internl orgns, which reltes to the somites. Segmenttion of vertebrtes is initited very erly in the developing embryo through the formtion of segments clled somites, which lter give rise to vertebre nd skeletl muscle, s well s to some dermis 1 The stimultion of locl skin could induce viscerl responses of internl orgns supplied by corresponding segmentl nerves. Our previous studies demonstrted tht the viscerl nociception nd the gstric motility could be modulted by cupuncture.,2, The underlying mechnism for these effects involves the reflex pthwys of "somto-sympthetic nerve" nd "somto-vgus nerve". The gstrointestinl trct is innervted both by sympthetic nd prsympthetic nerves. Gstrointestinl motility could be suppressed by incresed sympthetic tone, nd promoted by excittion of prsympthetic nerves. Prsympthetic innervtion of jejunum rises from dorsl motor nucleus of vgus, nmely vgus nerve. Sympthetic innervtion of jejunum rises from T9-T12 levels of thorcic spinl cord. 4 The somtic fferent inputs from Tinshu (ST 25) project to T9-T11. The sympthetic innervtions of jejunum nd the sensory fferents from ST 25 project to reltively similr segments in spinl cord (Figure ). Thus, Tinshu (ST 25) is the "homotopic point" for jejunum, nd MA t Tinshu (ST 25) could inhibit the motility of jejunum vi somto-sympthetic reflex pthwy. The somtic fferent inputs from Zusnli (ST 6) project to L4-L5. The sympthetic innervtions of jejunum nd the sen- JTCM www. journltcm. com 21 April 15, 215 Volume 5 Issue 2

6 Zho YX et l. / Experimentl Study DMV C1 C2 C C4 C5 C6 C7 C8 T1 T2 T T4 T5 T6 Vgus nerve T7 T8 T9 T1 Jejunum ST 25 Somtic fferents T11 T12 L1 Sympthetic nerve Colon L2 Rectum L ST 6 Somtic fferents L5 S1-S5 L4 Prsympthetic nerve Co4 Figure Regultory mechnism of MA t Zusnli (ST 6) or MA t Tinshu (ST 25) on the motility of jejunum nd distl colon in the present study might be s follows: sympthetic innervtions of jejunum rises from T9-T12 levels of thorcic spinl cord; prsympthetic innervtions of jejunum rises from DMV, nmely vgus nerve Sympthetic innervtions of distl colon project to L1 nd L2; prsympthetic innervtions of distl colon rise from scrl level of spinl cord (S1). Somtic fferent inputs from Tinshu (ST 25) project to T9-T11. Somtic fferent inputs from Zusnli (ST 6) project to L4-L5. Tinshu (ST 25) is the "homotopic point" for jejunum, thus, MA t Tinshu (ST 25) could inhibit the motility of jejunum vi somto-sympthetic reflex pthwy. Zusnli (ST 6) is the "heterotopic point" for jejunum, thus, MA t ST6 could promote the motility of jejunum vi incresed prsympthetic tone. both Tinshu (ST 25) nd Zusnli (ST 6) re the "heterotopic points" for distl colon, nd the incresed colorectl motility is medited by somto-prsympthetic reflex pthwy. MA: mnul cupuncture; DMV: dorsl motor nucleus of vgus. sory fferents from ST6 project to different segments in the spinl cord (Figure ). Thus, Zusnli (ST 6) is the "heterotopic point" for jejunum, nd MA t Zusnli (ST 6) could promote the motility of jejunum vi incresed prsympthetic tone. The results of present study indicted tht both MA t Tinshu (ST 25) nd MA t Zusnli (ST 6) could promote the colorectl motility. The nervous innervtions of distl colon project to L1, L2, nd S1, which re different from the projecting segments of Tinshu (ST 25) nd Zusnli (ST 6) (Figure ). Thus, both of the two points re the "heterotopic points" for distl colon, nd the incresed colorectl motility is medited by somto-prsympthetic reflex pthwy. REFERENCES 1 Hsu DT. Acupuncture. A review. Reg Anesth 1996; 21(4): JTCM www. journltcm. com 22 April 15, 215 Volume 5 Issue 2

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