Initiation mechanisms of acupuncture effect: a literature review of basic studies

Zhang Kuo (张阔), Ding Sha-sha (丁沙沙), Gong Yi-nan (公一囡), Zhao Xue (赵雪),4, Li Kai (李凯)

1 Academy of Medical Engineering and Translational Medicine, Tianjin University, Tianjin 300072, China

2 Department of Acupuncture and Physical Therapy, Tianjin Nankai Hospital, Tianjin 300100, China

3 Research Center of Experimental Acupuncture Science, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China

4 Acu-moxibustion and Tuina Department, Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China

5 Department of Gynaecology, First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Tianjin 300381, China

Abstract

Keywords: Acupuncture Therapy; Acupuncture Points; Research on Acupoints; Acupuncture Effect; Review; Literature

According to traditional Chinese medicine (TCM)theory, acupoints are the particular points where meridian qi transfuses from inner body to the surface.Acupoints are considered as the critical spots to sense stimulation, initiate the effect and amplify acupuncture signals[1].Therefore, it is crucial to clarify the initiation mechanism of acupoints.

Acupoint is a stereochemical structure that consists of nerves, veins, connective tissues, muscles and tendons, etc.[2]The alternation of local microenvironment around the acupoints is closely related to nerve, endocrine and immune systems, which could be the vital biological foundation of acupuncture effect initiation[3].Therefore, in this article, we reviewed the related researches about local microenvironment of acupoints and acupuncture effect, in order to provide a basis for clarifying the initiation mechanism of acupuncture effect.

1 Materials and Methods

The studies were identified from PubMed, Web of Science Core Collection, China National Knowledge Infrastructure (CNKI), Wanfang Academic Journal Full-text Database (Wanfang), Chongqing VIP Database(CQVIP).All searches were performed up to December 31, 2018.The search terms used were: acupuncture AND acupoint; electroacupuncture AND acupoint.The screening process is summarized in the flow diagram(Figure 1).

Figure 1.Flow diagram of study selection process

2 Results

2.1 Local neuroendocrine system in the initiation of acupuncture effect

As a minimally invasive stimulation to human body,acupuncture can instantly and directly affect nervous system.

2.1.1 Afferent nerve

Local area of the acupoint has more concentrated distribution of receptors, free nerve endings, nerve bundles and nerve plexuses[4].The primary information of acupoints concludes generator potential evoked by the activation of peripheral nerve endings or needling sensation sensory receptors and afferent impulses of various types of codes[5].Researchers have confirmed that acupuncture can change afferent electrical signals that control acupoints[6].In addition, analgesic effect of needling Zusanli (ST 36) on the nociceptive responses of the rat nucleus raphe magnus neurons could be significantly inhibited if the nerves or nerves proximal to the heart in the same nerve trunk have been blocked or cut off[7].It suggests that the afferent nerve excitation of the acupoints was important in initiating the acupuncture effect.What’s more, researches have shown that analgesia induced by acupuncture at Zusanli(ST 36) has close relationship with the excitement of local thick nerve fibers, especially neural fibers Ⅱ,among which, the distribution of myelinated fibers is more dominant than that of unmyelinated fibers (with the ratio of 2.7: 1), which indicates that acupuncture effect may be related to the category and amount of local afferent nerves[8].

Besides, different acupuncture stimulation intensity may excite the different category of afferent nerves,generating different effect mechanism.Some researches indicated low-intensity (2 V) electroacupuncture mainly excited types Ⅰ, Ⅱ and part of Ⅲ(Aδ) thick afferent nerves, and achieved localized and transient analgesic effects through spinal cord integration[9].While high-intensity (18 V) electroacupuncture could fully excite the Ⅲ (Aδ) and Ⅳ(C)-type thin afferent fibers, activate the negative feedback regulation mechanism of brainstem above the spinal cord, and exert a systemic, wide-range and long-acting analgesic effect[10].

2.1.2 Neurotransmitter and endocrine hormone

As amine neurotransmitters, 5-hydroxytryptamine(5-HT) and histamine play a certain role in the initiation of acupuncture effect.Studies have demonstrated that acupuncture can significantly up-regulate the expression of 5-HT and Histamine in Zusanli (ST 36), and the analgesic effect of acupuncture on rats with adjuvant-induced arthritis is histamine-dependent[11].

Some researchers found that endocrine hormones were involved in the acupuncture effects.Ogay V,et al[12]found that normal rabbits’ skin in Zhongwan (CV 12)contains cells that store catecholamines, and acupuncture can release high levels of norepinephrine.Our previous studies have shown that acupuncture can not only relieve pain, but also increase the concentration of follicle stimulating hormone (FSH),thyroid stimulating hormone (TSH) and corticosterone in Zusanli (ST 36) of complete Freund’s adjuvant (CFA)rats.

Local purine signals are indispensable in acupuncture effect.Adenosine triphosphate (ATP) is an excitatory neurotransmitter.Geoffrey Burnstock firstly proposed that purine signal may reveal the effect principle of acupuncture[13]: mechanical stimulation of acupuncture can cause skin deformation, and promote subcutaneous cells such as skin keratinocytes to release ATP, which will activate ATP-specific receptors (such as P2X3, P2X2/3,P2X7 receptors) located in the sensory nerve endings of the skin, and transmit acupuncture signals to dorsal root ganglia of the spinal cord, then finally integrate in the cerebral cortex, transmitting information and relieving pain.After that, Goldman N,et al[14]reported the role of adenosine A1 receptor in the analgesic effect induced by acupuncture at Zusanli (ST 36) in CFA rats,suggesting that local analgesic effect of acupuncture is due to the local release of adenosine after noxious stimulation, which then binds with adenosine A1 receptors at near-by afferent nerve endings, and finally blocks the transmission of pain signals.Afterwards,other scholars repeated part of the research of Goldman N,et al, and found that PAP, which promotes the conversion of adenosine monophosphate (AMP) to adenosine, can significantly prolong the analgesia effect of acupuncture[15].P2X3 receptor mediates acupuncture analgesia[16], and local ATP in Zusanli (ST 36) may be involved in the peripheral time response of acupuncture analgesia in CFA rats[17], and acupuncture may activate histamine receptor 1 (H1) or A1receptor by local mast cells[18].Takano T,et al[19]have also reported changes of local adenosine levels in acupuncture at Zusanli (ST 36)in healthy people, which further proved animal experiment results and provided supplementary evidence for local adenosine mediating acupuncture analgesia at acupoints.

2.2 Local immune system in the initiation of acupuncture effect

Increasing evidences have shown that the status of local immune cell function, immune-related cytokines,inflammatory substances and immune-related signaling pathways can change after acupuncture, which plays an important role in the initiation of acupuncture effect.

2.2.1 Immune cell function

Mast cells (MCs): MCs are one of the key factors of acupuncture signal amplification[20].And MCs degranulation is a crucial step in its biological function.In 1977, some researchers found that the number of MCs in the Quchi (LI 11), Hegu (LI 4), Waiguan (TE 5)and other 15 acupoints is significantly higher than that in the surrounding non-acupoints in human amputation specimens[21].After that, Zhu ZX,et al[22]observed that the number of MCs in the layers of the low-resistance meridians of rats was significantly higher than that near the meridians.Wu JL,et al[23]found that MCs often distributed along small blood vessels and nerve bundles tracing the meridian line.There are successive studies on the number, degranulation rate and main distribution of MCs after acupuncture under normal or pathological conditions, and evidences have showed that after acupuncture, MCs near Dazhui (GV 14) in rats highly increased and transformed with obvious degranulation, especially those around acupuncture needles[24].And for hepatic fibrosis rats, acupuncture can increase the number of local MCs and promote its degranulation rate, while causing local directional distribution of MCs in subcutaneous tissue of Ganshu(BL 18)[25], suggesting that the number and degranulation rate of MCs in acupoints may have a certain correlation with acupuncture effect.

This correlation mechanism is related to both the neuroendocrine and immune systems.Zhang BZ[26]believes that there are synaptic-like joints with MCs as the mediator in the end of the adjacent ganglion nerves in the skin, and MCs transmit information between two nerve terminals, which constitutes the ‘axle reflex relay linkage’, and may be the material basis of propagated sensation along meridians (PSM).The numerous neuropeptides released by MCs degranulation and the cytokine cascade triggered by it can regulate acupuncture effects through neurohumoral mechanism[27].The bioactive substances released by MCs degranulation after acupuncture interact with each other to further promote the degranulation of MCs and the transmission of acupuncture information[28], and the regulation of local bioactive substance content and MCs functional status will cause changes in the acupuncture effect[29].Moreover, the mechanical force signal generated by acupuncture can directly act on MCs by means of the transient receptor potential vanilloid 2 (TRPV2) channel on the cell membrane,exciting MCs’ function and generating a subsequent effect[30].

Neutrophils and cell lysosomes: Studies have shown that acupuncture twirling at Zusanli (ST 36) can cause local tissue injury like muscle fiber entanglement,breakage and broken capillaries in normal rat.Six hours later, acupuncture can also cause degeneration and necrosis of local tissues, as well as inflammatory infiltration, neutrophils mainly.It is suggested that acupuncture can trigger ‘self-generated’ inflammation locally, which may be one of the initiation mechanisms of its immune regulation[31].Moreover, some people used electron microscope to observe the local ultrastructure of Dazhui (GV 14), Baihui (GV 20),Jiaochao [it is the same as Changqiang (GV 1) of human body] in normal pigs, and they found that after acupuncture, lysosomes with single cell membrane increased in myocytes cytoplasm[32].However, whether locally increased neutrophils and cytolysosomes participate in the initiation of acupuncture effect requires further researches.

2.2.2 Expression of immune-related cytokines and inflammatory substances

There are growing evidence indicating that the expressions of local immune-related cytokines and inflammatory substances change significantly after acupuncture.For example, some researchers found that acupuncture can activate nerve-related cell function and promote the release of proinflammatory cytokines such as interleukin (IL)-1β and IL-6 in Zusanli (ST 36) of normal rats[33-34].The local concentration of IL-8, tumor necrosis factor alpha (TNF-α), E-selectin and L-selectin in Zusanli (ST 36) of normal rats also increased after acupuncture[31].Some other researchers found that acupuncture at Zusanli (ST 36) produced biological effect by up-regulating local neuropeptides such as neuropeptide Y (NPY), calcitonin gene-related peptide(CGRP) as well as inflammatory factors such as IL-18,and down-regulated those under chronic atrophic gastritis conditions[35].Our previous studies have shown that except for analgesia function, acupuncture can increase the local concentration of chemokines at Zusanli (ST 36) like CXCL1, monocyte chemoattractant protein-1 (MCP-1) as well as proinflammatory factors like IL-1β and IL-6 in CFA rats simultaneously.Thus, local cytokines and inflammatoryrelated substances near acupoints may be involved in the initiation of acupuncture effect.

2.2.3 Immune-related signaling pathway status

The status of local immune-related signaling pathways in acupoints is associated with acupuncture effect, especially inflammatory response-related pathways.Some researchers have indicated that for CFA rats, the biological processes of differentially expressed genes significantly enrich around acupoints after needling Zusanli (ST 36) include inflammatory responses,defense responses, etc.[36]As for normal rats, the concentration of local Toll-like receptor 4 (TLR4), high mobility group box 1 (HMGB1) and TNF-α increased after needling Zusanli (ST 36), suggesting that the local TLR4-mediated signaling pathway may be one of the inflammation-related pathways after acupuncture[37].And for rats with myocardial ischemia-reperfusion injury,acupuncture plays a therapeutic role by inhibiting the local TLR4/MyD88/NF-κB pathway in Neiguan (PC 6).

The role of MAPK signaling pathway in the initiation of acupuncture effect has received certain attention.Studies have shown that acupuncture can significantly increase the protein expression of ERK1/2 in the local superficial fascia under nomal rat’s Zusanli (ST 36)acupoints[38].Park JY,et al[39]discovered that there are at least 7 immune and inflammation-related signaling pathways in acupoints, including MAPK and Toll-like receptors (TLR), which are involved in the analgesic and anti-inflammatory processes of acupuncture at Zusanli(ST 36) in arthritis rats, and blocking the local MAPK/ERK signaling pathway could inhibit acupuncture analgesia.Other researchers have indicated that inflammatory pain can increase the expression of p38MAPK and phospho-p38MAPK in the skin of Zusanli(ST 36), and the expression can be down-regulated by acupuncture[40].Moreover, acupuncture can inhibit the activation of p38MAPK pathway and reduce allergy and inflammation caused by allergic contact dermatitis in rats[41].

2.3 Local neuro-endocrine-immune (NEI) network in the initiation of acupuncture effect

NEI network represents the complex interrelation among nervous system, endocrine system and immune system, which together maintain the normal balance and steady state of organism[42].Presently, studies have shown that skin, as an important anatomical structure of acupoints, is a NEI organ[43].There is a closed loop between brain and skin, through which may acupuncture trigger a series of central and local stress reactions.

Further studies have shown that the three systems above may share some common chemical signal molecules (common signal molecule), including cytokines, neuropeptide endocrine hormone and their receptors, and they work as common languages to achieve information exchange in NEI network[44].The mentioned various neurotransmitters, endocrine hormones, and immune cytokines that locally increased in acupoints are common or related signal molecules of NEI network, and they are regulated by acupuncture and participate in the initiation of acupuncture effect.Acupuncture can promote the interactions between the signal molecules, amplify the acupuncture information cascade, and then mobilize the human body to regulate the NEI network.Through NEI regulation, the acupuncture effect information can be output to the target organ, adjust the disease network, and produce acupuncture effect.This way of acupuncture regulation is more complex and extensive than that of the individual nervous, endocrine and immune systems.Under the analgesic and anti-inflammatory effect of acupuncture, we used liquid-phase chip and other methods to detect the expression amount of 55 kinds of common NEI signaling molecules, serum and inflammatory footpad tissue in Zusanli (ST 36) of CFA rats, including 13 neurotransmitters or neuropeptides,18 endocrine hormones, and 24 cytokines, for example,substance P (SP), NPY, dopamine, β-endorphin,corticosterone, α-MSH, orexin A, somatostatin, IL-1α,IL-6, IL-10, IL-17, etc.and we applied complex network method to establish a network among the common signal molecules in each part, based on which we found the key signal molecules such as MIP-3α and TNF-α in acupoints, and the result further affirmed the regulation effect of acupuncture on local NEI network in acupoint.

2.4 Local connective tissues in the initiation of acupuncture effect

It is believed that connective tissues movement can greatly affect local effect of acupuncture, that the displacement of tissue can be five-times amplified by needle manipulation[45].Acupuncture needle movements pull and deform the connective tissues[46],causing muscle sleeve traction and forceful reflex muscle contraction which affect small blood vessels,nerve branches, nerve stems and other known needlesensing structures adjacent to or away from pinholes.

Collagen fiber is one of the important fiber components of connective tissues, and it remains winding on needle after manipulation[47].And tubular structure formed by the association of collagen fibers are considered to be the material basis of acuesthesia[48].Acupuncture can significantly increase the related gene expression of local collagen fibers,elastic fibers and extracellular matrix[49].Moreover,fibroblasts and collagen fibers in subcutaneous connective tissues proliferate over time in the deformation, distortion and breakage spot of muscle fibers[50].

Collagen fibers play an extremely important role in information and energy exchange activities of meridians,and regulate the effects of acupuncture and signal transmission[51].Studies have shown that needles of different thicknesses can cause shiftiness of subcutaneous collagen fibers quantity in acupoints and torsion degree of connective tissues, which will affect the intensity of acupuncture effects[52].Langevin HM[53]believed that acupuncture can be effective because of the distortion and rotation of connective tissues(collagen fibers), through which the mechanical force signals pass through and even spread to distant distances.Distortion of connective tissues caused by acupuncture can stimulate cellular signal transmission and form a specific pathway to achieve therapeutic effects[54].

Moreover, collagen fibers and MCs are closely related that acupuncture mechanical force in subdermal loose connective tissues can activate the degranulation of MCs[55].Furthermore, after destroying the local collagen fibers in Zusanli (ST 36), the analgesic effect of acupuncture was inhibited in CFA rats[56].

2.5 Local ions in the initiation of acupuncture

Acupuncture may increase the ion permeability of local cell membranes, accelerate the activation of pathways, and change the concentration of local ions and neurotransmitters in Zusanli (ST 36) in experimental stomachache rats[57].And variation in electrical communication induced by changes in ion concentration is considered to be the key factor of communication changes between nerve endings and cells, the physiological basis for acupuncture effects as well.

Studies have shown that after acupuncture, the content variation of K+and Na+was negatively-related at local Zusanli (ST 36) in normal rabbits, suggesting that acupuncture may promote or enhance the operation of‘sodium pump’, so that longer-lasting excitement spreads in acupoints, which activates the regulation of nerves and body fluid pathways, thus creating complex acupuncture effects; and the remarkable drop of H+may be related to tension contractions of muscle fibers in CFA rat’s Zusanli (ST 36)[58].Other studies have indicated that the local extracellular concentration of Ca2+, Cl-and tissue fluid concentration of K+in Zusanli(ST 36) are up-regulated by acupuncture, pain threshold in CFA rats as well[59].

The function of Ca2+is vital in nerve electrical signal transmission.We have investigated the correlation between local Ca2+and acupuncture effects, and results showed that the Ca2+concentration in acupoints was significantly higher than that in non-acupoints[60], and the concentration of Ca2+in acupoints exceeded that in side points.Moreover, after complexation of Ca2+in Zusanli (ST 36), the effect of acupuncture in inhibiting gastric hypermotility in rabbits would be weakened or lost, and this common regularity could be observed in different animals or different disease models[61].We excavated the internal mechanism between the local Ca2+in Zusanli (ST 36) and acupuncture effect and relative experiments indicated that after the complexation of Ca2+, the discharge frequency and amplitude were significantly reduced in spinal dorsal root nerve filaments, which made a further explanation that Ca2+should be the key factor in acupuncture effect and an important material basis for meridian activity.

3 Conclusion

Acupuncture is an important traditional Chinese medicine treatment.It has been widely used in 183 countries and regions.The World Health Organization(WHO) and the National Institutes of Health (NIH)recommend acupuncture for low back pain, arthritis,female infertility, etc.However, the mechanism of acupuncture effect has not been fully elucidated[62].The microenvironment changes of acupoints play an important role in the initiation and amplification of acupuncture information.The study on initiation mechanism of acupuncture effect will contribute to revealing the key basic factors of the complex mechanism and the principle of acupuncture.

In conclusion, acupuncture can excite local afferent nerves, activate cell function, promote the release of neuropeptides, hormones, cytokines and other chemical substances, cause mechanical deformation of connective tissues, and changes in chemical ions and ion channels in the acupoints.However, at present,researches on the initiation mechanism of acupuncture effect mostly only focus on a single level about genes,proteins and cells, which lacks integral and networked analysis of the local acupoints microenvironment.

It is of great practical significance to clarify the initiation mechanism of acupuncture effect, and to explore effective ways and modes of acupuncture regulation, so as to improve acupuncture application and achieve better promotion and development of acupuncture.In the future, we should hold a holistic,dynamic and disciplinary perspective to conduct a comprehensive and thorough research on the initiation mechanism of acupuncture effects, in order to provide more evidence for the mechanism of acupuncture.