Acupuncture at Zu San Li increases the serum levels of superoxide dismutase, one of the body’s strongest antioxidants, thus reducing damage by free radicals, which is one of the main theories on ageing (2). 

Acupuncture at Zu San Li not only reduces the shortening of the telomeres but increases them by up to 200% (3). When the telomeres get too short, replication of that cell line is either destroyed or mutated. This is known as senescence and is one of the other main theories on ageing. The shorter the telomeres the faster you age and more likely you are to suffer from chronic disease. Further research suggests that the increase in telomere length was mediated by an increase in brain derived neurotrophic factor, stimulated via acupuncture, and its downstream pathways (4). It should be said this research was on a small sample and with one researcher and has not been replicated since.

Research has shown that stimulating the Zu San Li acupoint may have anti-nociceptive effects. One study found that acupuncture at Zu San Li significantly reduced Mechanical and cold allodynia in rats with nerve damage (7). Another study found that electroacupuncture at Zu San Li inhibited neuropathic pain in rats by reducing the expression of pro-inflammatory cytokines and increasing the expression of anti-inflammatory cytokines at microglial cells. This was mediated by suppressing P2X7R expression and thus reducing abnormal dendritic spine/synaptic reconstruction and inflammation (8).

A study was comparing the effect of manual, sham and electroacupuncture, just after and 24hrs after stimulation at ST36 and Feng long (ST40), they found that Compared to sham acupuncture, EA significantly increased both single pain threshold (SPT) and temporal summation/repeated pain thresholds (TST) immediately after the treatment on the treatment leg as well as 24h after treatment on both the treatment and non-treatment legs, this effect was not as strong in manual acupuncture, not reaching statistical significance versus sham. The researchers surmised that; EA induces bilateral, segmentally distributed and prolonged analgesia on both SPT and TST, indicating a non-centrally specific effect.’ (9)

Acupuncture at Zusanli (ST36) causes significant improvements in pain tolerance thresholds (PTT) and significant increases in ATP levels. The effects of acupuncture were further augmented by administration of different doses of ATP at Zusanli (ST36), and pretreatment with a P2X3 receptor antagonist decreased PTT. Adenylate energy charge peaked at 30 min after intraperitoneal injection of ATP, and pretreatment with various doses of i.p. ATP 30 min prior to acupuncture increased PTT in a dose-dependent manner. Additionally, pretreatment with an i.p. or intramuscular injection of adenosine disodium enhanced the effects of acupuncture. Thus ATP is involved in the regulation of PTT through acupuncture at Zu san li (10).

In another study on EA for analgesia during surgery, researchers found that Electroacupuncture at ST36 and SP6 can mitigate the adrenal cortical inhibition induced by etomidate and can reduce the secretion of catecholamines during surgery (11).

EA at ST36 could attenuate cancer-induced pain, at least in part, through suppressing transient receptor potential vanilloid subfamily 1 mRNA and protein upregulation in the dorsal root ganglia (12).

In an interesting study, researchers looked at collagen fibres and their involvement in acupuncture pain relief, especially with manipulation techniques. ‘When the structure of the collagen fibers at Zusanli (ST36) was destroyed by injection of type I collagenase, the needle force caused by the acupuncture declined and the analgesic effects of rotation or lift-thrusting manipulations was attenuated accompanying the restraint of the degranulation ratios of mast cells. We propose that collagen fibers play an important role in acupuncture-induced analgesia, and they participate in signal transmission and transform processes.’ (13)

Several studies have suggested that stimulating the Zu San Li acupoint may have an effect on the autonomic nervous system. For example, one study found that acupuncture at Zu San Li increased parasympathetic nervous system activity and cerebral blood flow in healthy subjects (14). Another study found that acupuncture at Zu San Li activated the vagus nerve, which led to an increase in the release of endogenous opioids and a reduction in pain perception in mice (15).

Several studies have suggested that stimulating the Zu San Li acupoint may regulate cardiovascular function. For instance, one study found that electroacupuncture at Zu San Li reduced blood pressure in a rat model of hypertension (16). 

EA at Zu san li to Feng long can benefit hypertension, in rats, regulating sympathetic discharge and blood pressure. This may be via its effects on upregulating the level of neuropeptide Y mRNA and protein expression in the paraventricular nucleus (17).

EA at ST36 significantly improved cardiac function (pressures, volumes, output and heart rate and heart rate variability) and exercise tolerance in chronic heart failure mice. The underlying mechanism may be related to the activation of vagus efferent nerve and the regulation of autonomic nerve balance at which the number of choline acetyltransferase (ChAT)+/c-fos+co-labeled neurons in the dorsal motor nucleus of the vagus were significantly raised (18). 

Unilateral electroacupuncture was shown to induce  neuronal nitric oxide synthase expression in the gracile nucleus and medial nucleus tractus solitarius, and enhanced nNOS-NO in the nuclei may modify central cardiovascular regulation, which contribute to hypotensive effects of acupuncture (19).

EA at Zu san li regulates the activities of creatine kinase-MB, TNF-α, nitric oxide, and myeloperoxidase and the rate of water content were significantly lower than those of the other groups 6h after cecal ligation and puncture (CLP). EA after vagotomy showed less anti-inflammatory and protective effects. The results indicated that EA obviously reduced the increased levels of the proinflammatory factors at 6h after CLP, and vagotomy could weaken or eliminate the effects of EA. Cholinergic anti-inflammatory pathway is one of the main mechanisms of cardioprotective effect of EA in sepsis model of rats (20).

A meta-analysis involving 16 studies and 401 right-handed healthy patients showed that ‘acupuncture on ST36 positively activates the opercular part of the right inferior frontal gyrus (IFG.R), left superior temporal gyrus (STG.L), and right median cingulate/paracingulate gyri (MCG.R) regions. Needle retention time in an acupuncture session positively correlates with the activation of the left olfactory cortex, as shown in meta-regression analysis. Subgroup analysis revealed that EA stimulation may be a source of heterogeneity in the pooled results. Functional network mappings showed that the activated areas were mapped to the auditory network and salience network. Further functional decoding analysis showed that acupuncture on ST36 was associated with pain, secondary somatosensory, sound and language processing, and mood regulation.’ (21)

Electroacupuncture at Shang ju xu and Zu san li (ST36) can reduce mossy fiber sprouting, spike population, and high-frequency hippocampal oscillations in kainic acid (KA)-induced epileptic seizure rats. It can also reduce COX-2 levels in the hippocampus and the number of COX-2 immunoreactive cells in the hippocampal CA1 region, which plays a critical role in postseizure brain inflammation and neuronal hyperexcitability (22). Additionally, long term EA at these points can increase this anti-inflammatory effect long term and as such can be a beneficial option for seizures 

EA at ST36 and ST37 and PC6 (Nei guan) and PC7 (Da ling) exert an analgesic effect, EA at PC6 and PC7 can enhance heart function via upregulation of adrenaline and noradrenaline, and EA at ST36 and ST37 modulates the cerebral cortex via upregulating glutamate (23).

‘Acupuncture at ST36 increased motor cortical excitation and had an effect on the remaining needle phase. Deqi sensation was correlated with MEP amplitude. Acupuncture at ST36 also decreased motor cortical inhibition.’ (24)

Acupuncture at Bai hui (DU20) and ST36 in rats after cerebral ischemia reperfusion injury can adjust the expression of heat shock protein 70 and TNF-α in the peripheral serum, which might be one of the mechanisms of acupuncture’s attenuation of cerebral ischemia reperfusion injury (25). 

‘EA at ST36 with low-intensity may improve neuroinflammatory response and systemic inflammatory cognitive dysfunction by increasing the choline level in the hippocampus and reducing the release of inflammatory factors through up-regulating the expression of α7 nicotinic acetylcholine receptor in hippocampal microglia. As such anti-inflammatory factor interleukin (IL)-10 and M2 macrophage marker arginase 1 (Arg1) was increased, hippocampal ionized calcium-binding adapter molecule 1 (Iba-1), the contents of M1 macrophage marker CD206, inducible nitric oxide synthase (iNOS), the content of pro-inflammatory factor IL-1β and the mRNA expression of TNF-α were decreased (26)

EA at ST36 can help intracerebral haemorrage, speeding up recovery by increasing angiogenesis (new blood vessel growth). The mechanism, shown in a study was post haemorrage, ‘HIF-1α protein levels increased, but HIF-1α mRNA levels did not change. EA increased BrdU-labeled nuclei in cerebral endothelial cells and up-regulated the expression of HIF-1α protein (27)

Similarly EA of Bai hui and Zu san li was shown to improve neurological deficit and reduce cerebral infarct volume in CI rats, which may be associated with its functions in promoting angiogenesis in ischemic penumbra area and in up-regulating the activities of HIF/VEGF/Notch signaling pathway and related factors. (28)

Acupuncture at Zu san li can increase reaction time and alertness and executive function by increasing the amplitude of the event related potentials of N2 and P300, thus improving cognitive ability (29)

Researchers looked into the mechanism behind acupuncture benefits for cognitive decline in vascular dementia, particularly at dopamine’s involvement in acupuncture benefits to neuroplasticity. They found that, ‘acupuncture remarkably reversed cognitive deficits in 2-vessel occlusion model (2VO) rats, and the acupuncture points Zusanli (ST36) and Baihui (DU20) were confirmed to be the most effective combination. Electrophysiological recording data showed that 2VO-induced impairments of long-term potentiation were prevented by acupuncture. In addition, acupuncture promoted the release of dopamine and its major metabolites in the hippocampus of 2VO rats. The immunofluorescence experiment showed that the decrease of D1R and D5R in hippocampal dentate gyrus region of 2VO rats was reversed by acupuncture. Furthermore, we found that the effects of acupuncture against 2VO-induced impairments in cognition and synaptic plasticity were abolished by SCH23390 [a D1 dopamine receptor antagonist]’ (30). 

In a pretty intense study of post stroke reperfusion injury, researchers studies EA at Qu chi (LI11) and Zu san li, they found, ‘at Day 3 attenuated neurological deficits and cerebral infarction volume in ischemia and reperfusion (I/R) injured rats. Animal behavior assessments found that the speed of Catwalk gait, equilibrium and coordination of Rotarod test were improved. Furthermore, EA treatment exerted neuroprotective effects via activation of glial fibrillary acidic protein (GFAP), vimentin and nestin positive cells. Simultaneously, an obvious increase in GFAP/vimentin, GFAP/nestin and GFAP/BrdU co-labeling appeared in the peri-infract cortex and striatum, suggesting EA can promote the proliferation of GFAP/vimentin/nestin-positive reactive astrocytes. The expression of cell cycle-associated proteins Cyclin Dl, CDK4 and phospho-Rb were increased in the peri-infract cortex and striatum, indicating proliferated reactive astrocytes-mediated CyclinDl/CDK4 regulation of the transition of the G1-to-S cell cycle phases. In addition, EA enhanced the localized expression of brain-derived neurotrophic factor (BDNF) in the peri-infract cortex and striatum. These results demonstrated that EA treatment at the LI11 and ST36 acupoints on Day 3 exerted neuroprotection via proliferation of GFAP/vimentin/nestin-positive reactive astrocytes and, potentially, secretion of reactive astrocytes-derived BDNF in I/R injured rats. (31)

Researchers found that EA at Quchi (LI11) and Zusanli (ST36) acupoints on the contralateral paralyzed limb significantly improved neurological deficits and cerebral infarction. In addition, electroacupuncture profoundly activated PI3K/Akt signaling in ischemic cerebral tissues. As such, electroacupuncture resulted in the inhibition of cerebral apoptosis. EA also increased the serum secretion levels of the PI3K activators brain derived neurotrophic factor  and glial cell line-derived neurotrophic factor, as well as upregulating the anti-apoptotic Bcl-2/Bax ratio in ischemic cerebrum, or post stroke recovery (32)

Zu san li is one of the most commonly used points for gastrointestinal conditions, indeed researchers found it to be:

• First for post-operative gastrointestinal dysfunction (33). 
• Similarly so for post intestinal cancer surgery (34, 35)
• Again, for gastric dysfunction in patients with gastric cancer (36)
• And, among the most commonly used for functional constipation (37).

Zu san li can be used for gastric discomfort and pain (visceral hyperplasia):

• Acupuncture at Zu san li, Shang ju xu (ST37) and Tian shu (ST25) is more effective than Qu chi (LI11), Da chang shu (BL25) and Ci liao (BL32) for visceral hypersensitivity. It also reduced elevated c-fos levels in the anterior cingulate cortex, hypothalamus, spinal dorsal horn and colorectum (38).
• EA at Shang ju xu and Zu san li can reduce pain and distention and may be mediated by the ion channel and TrpV1 expression in colorectum as well as ERK1/2 MAPK pathway activation in peripheral and central nerve system (39). 
• EA at Shang ju xu and Zu san li reduces serum protein and mRNA Epac1 and Piezo2, the upstream of 5-HT, as well as dorsal root ganglia L5-S2 levels (40). 
• The signal of gastric distention and acupuncture could converge in lateral hypothalamus area (LHA) and fastigial nuclear (FN) circuit; acupuncture (at Zu san li) presented different regulation effects on identical type of GD-sensitive neurons in different nuclear groups; LHA-FN circuit might participate in central integration mechanism of acupuncture on gastric function (41).

For Ulcerative colitis:

• Zu san li regulates HMGB1,7 nAChR and NF-κB mRNA, and effectively inhibits the immune response, reduces the inflammatory response of colon and improves pathological changes of colonic mucosa. The researchers showed these effects were greater with Shang ju xu than (in order of effect) Zu San Li (ST36), Xia ju xu (ST39), Yang Ling Quan (GB34) (42). 
• Modulation of the microbiome was shown in a group of mice with colitis, this time through the mitogen activated protein kinase signaling pathway (43)

For Chron’s disease:

• Acupuncture at ST36 (Zu san li), ST37 (Shang ju xu), LI11 (Qu chi), and LI4 (He gu) and herb partitioned moxa at  ST25 (Tian shu), REN6 (Qi hai), and REN9 (Shui fen) can repair intestinal epithelial barrier lesions and relieve inflammation by upregulating the expression of tight junction proteins and their mRNAs (44).

For constipation:

• One study found that electroacupuncture at Zu San Li increased the motility of the small intestine and improved gastrointestinal transit in rats (45). 
• Acupuncture stimulation at ST 36 for 5 days increased the intestinal transit and down-regulated the concentration of VIP and up-regulated the concentrations of motilin, ghrelin and gastrin, whereas acupuncture stimulation at LI 11 did not change them significantly (46). 

For IBS-diarrhoea

• Daily acupuncture for 20 mins at Bai hui (DU20) Tian shu (ST25), Shang ju xu (ST37), Zu san li (ST36), and Tai chong (LR3) for two weeks can improve the general and emotional state, inflammatory response, and neuropeptide expression in rats with IBS-D, and alleviate the symptoms of IBS-D, which may be related to the regulation of neuropeptides and inflammatory factors levels, notably; interleukin-10, tumor necrosis factor-α, calcitonin gene-related peptide in the hypothalamus and colon tissue and corticotropin-releasing factor(CRF) in the hypothalamus and colon tissue (47). 
• Acupuncture and moxibustion at Zu san li, Guan yuan (REN4) and Nei guan (PC6) can effectively improve the symptoms (spontaneous activities, visceral hypersensitivity) of IBS-D rats. This may be due to its functions in regulating autonomic nervous activities, and down-regulating blood norepinepherine and calcitonin gene-related peptide contents and colonic 5-HT protein expression and immunoactivity, and the effects of the above acupoints are superior Zu san li, Tian shu (ST25) and Shang ju xu (ST37). (48)
• EA at He gu vs Zu san li (2/100Hz for 20 mins, tolerable level, every other day for 5 times) were compared for their impacts on pain and motility. It was found that, ‘EA of ST36 was significantly superior… in lowering Bristol fecal scale score and colonic 5-hydroxytryptamine 3A receptor (5-HT3AR) expression in the muscular layer, but obviously inferior… in increasing the latency of muscular initial contraction wave and down-regulating muscular contraction waves and 5-HT3AR expression in the mucosal layer.’ Thus LI4 is better for intestinal sensitivity and ST36 for motility (49). 
• EA (10Hz, 1mA) at ST36 for mice with IBS-D significantly improves the macroscopic/microscopic characteristics and the weight and length of the colon. Reversing inflammation and leukocyte infiltration and normalising the elevated levels of IL-1β, IL-18, and NLRP3. Furthermore, EA improved the expression levels of ZO-1, occludin, and claudin 1, exhibiting normalization of the colon’s tight junctions. The researchers theorised that this slew of actions was enacted by regulating the pathological phenotype of the NLRP3 inflammasome pathway (50)

For functional dyspepsia:

• A study found that acupuncture at Zu San Li improved the symptoms of functional dyspepsia in rats (51). 
• Another study showed improved intestinal muscosal healing by increasing the trefoil factor in rats with peptic ulcer disease as well as improving the microbiota (52).’
• EA Acupuncture at 6 differnet acupoints were compared for FD rats; Zu san li, Shang ju xu (ST37), Xia ju xu (ST39), Tian shu (ST25), Zhong wan (REN12) and Guan yuan (REN4). The researchers noted that, as others have, that abdominal points didn’t seem to increase gastric motility, but nor did they improve markers of inflammatory and mucosal injury. ST37 and ST39 could reduce the immunoreactivity of  eosinophil major basic protein and also ameliorated FD-tight junction protein in the expression of Claudin3 and ZO1, reducing inflammation and leaky gut. But only ST36 could regulate gut motility, the above markers and CD45 and occludin1, thus reducing immune cell infiltration and preventing gut membrane permeability, respectively (53).
• EA at ST36 can promote gastrointestinal motility in FD rats, which is possibly mediated by inhibiting excessive autophagy of interstitial cells of Cajal via down-regulating AMPK/ULK1 signaling. As shown by the therapeutic effect of down-regulating the expression of LC3Ⅱ/LC3Ⅰ， Beclin 1, p-AMPK and p-ULK1 proteins and in up-regulating the expression of c-kit protein (54)
• EA of ST36 can suppress visceral hypersensitivity and increase the gastric compliance in FD rats, which may be related with its effects in inhibiting the activation of gastric mast cells, and down-regulating the expression of gastric proteinase activated receptor 2 (PAR2) and transient receptor potential vanilloid type1 (TRPV1) proteins and substance P and CGRP contents (55).

Zu san li is commonly combined with Feng long for hyperlipidemia and related conditions, and has a wide range of mechanisms;

• EA at Feng long and Zu san li (ST36), in rats, increases serum HDL-C, decreases contents of LDL-C, total cholesterol, triglycerides, and alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity, improves hepatocyte morphology and reduced steatosis. Interestingly it also increased expression of SIRT1, FOXO1, and ABCA1 proteins and mRNA and decreased AC-FOXO1 protein expression. The SIRT1/FOXO1 signaling pathway is a central regulator of oxidative stress resistance, inflammation control, and metabolic homeostasis. Its activation supports cell survival and anti-aging processes (by preventing DNA and cellular degradation), is cardio and neuro protective and has a role in glucose metabolism. (56)
• Additionally, like [a] SIRT1 agonist, EA [at Zu san li, Zhong wan, Guan yuan and Feng long] suppressed body weight gain and insulin resistance levels in obese rats, but this was only partially blocked by the SIRT1 antagonist. EA could upregulate protein expression of hypothalamic SIRT1 and downregulate the acetylation level of FOXO1 in the hypothalamic arcuate nucleus (ARC), which decreased gene expression of neuropeptide Y and increased that of POMC. The agonist targeted the hypothalamic SIRT1gene, unlike EA, which targeted posttranscriptional regulation. (57)
• Another study concluded; ‘EA [at the above points] enhanced high-fat diet-induced insulin resistance by activating SIRT1 and GLUT4 in the quadriceps femoris. These results provide powerful evidence supporting the beneficial effects of EA on HFD-induced insulin resistance.’ (58)
• EA, at Guan yuan (REN4), Zhong wan (REN12),  Zu san li (ST36), and Feng long (ST40), for 10 minutes 3 times a week, can effectively regulate intestinal Toll-like receptor 4 (TLR4), inhibit the interaction between TLR4 and NF-κB p65, and reduce the activity of NF-κB p65, which may be a potential mechanism of EA in reducing body weight and blood glucose in obese rats. Regulating this pathway can be beneficial for chronic inflammatory gut conditions, maintaining gut integrity and regulating the microbiome (59) 
• Zu san li to Feng long and Zhong wan to Guan yuan as well as Tian shu EA is superior to abdominal EA in improving lipid metabolism in obesity rats, which may be related to its stronger effectiveness in down-regulating hepatic TLR4/NF-κB p65 signaling. (60)
• Electroacupuncture at, Zu san li (ST36),  Zhong wan (REN12), Guan yuan (REN4) and “Fenglong”(ST40), was able to reverse the modeling induced increase of body mass, Lee’s index, contents of serum TNF-α, IL-6 and free fatty acid  (NEFA), expression levels of hypothalamic TNF-α, IL-6 and iNOS proteins, the immunoactivity level of Iba-1 and the content of Evans blue, and decrease of serum content and expression level of IL-10 in the hypothalamus. Thus ‘EA can treat obesity, improve body inflammation caused by obesity, and relieve central inflammation by repairing the blood-brain barrier.’ (61)
• ‘EA at ST40 and ST36 can significantly improve liver function in [non-alcoholic fatty liver disease] rats, and its mechanism of action may involve inhibiting PERK expression thereby targeting the downstream ATF4/CHOP signaling pathway to suppress endoplasmic reticulum stress, exerting a liver protective effect；the optimal effect was observed with EA intensity of 4 mA [20 minutes, 5 times a week] for 4 weeks.’ (62)
• EA of ST36-ST40 and REN4-REN12, can alleviate oxidative stress and mitochondrial dysfunction in diabetic neuropathic rats, which may be associated with its functions in up-regulating SIRT1/PGC-1α signaling, and decreasing renal fibrosis. Additionally the levels of fasting blood glucose, serum glycosylated hemoglobin, urea nitrogen, creatinine, urine in 24 hours, triglycerides, total cholesterol, LDL-C and malondialdehyde, and the expressions of α-smooth muscle actin, renal collagen Ⅰ, collage Ⅳ and fibronec-tin proteins were markedly down-regulated, and those of body weight, HDL-C, superoxide dismutase, glutathione, catalase expressions of SIRT1 and PGC-1α mRNAs and proteins significantly up-regulated. The researchers also showed that the mesangial dilatation, glomerular hypertrophy, mesangial matrix accumulation, glomerular extracellular matrix deposition and enhancement of glomerular fibrosis and interstitial space expansion, foot process fusion, basement membrane thickening and organelle injury of the model group was mitigated by electroacupuncture (63)
• ‘EA at ST36 effectively reduced body weight, reversed Olanzapine-induced adipocyte hypertrophy and hepatocyte damage, and restored disturbances in free fatty acids (FFA) and liver lipid metabolism. These effects also involved the restoration of olanzapine-induced disruptions in respiratory exchange rate (RER) during the dark cycle. We further explored the potential regulatory mechanisms, and we successfully demonstrated that EA stimulation significantly deactivated Agouti-related protein (AGRP)-expressing neurons in the lateral septal nucleus (LS), nucleus accumbens (NAc), and pallidum, suggesting that AGRP+ neurons in these brain regions are involved in the liver-brain interaction during EA treatment. These findings highlight the critical role of metabolism-regulating neurons in the acupuncture-mediated liver-brain axis and provide insights into the mechanisms of information integration from the brain to the liver.’ (64)

Post-operative ileus (temporary bowel impairment post surgery)

• EA (10 or 30Hz, but not 2 or 100) at ST36, ST37, ST39 (Xia ju xu), or REN4 (Guan yuan) significantly inhibited local MPO activity, immune cells infiltration, and increased α-SMA, thus regulating inflammation in the intestinal musculature and was also able to improve post operative dysmotility (65). 
• Lower He-sea points, Zu san li and Shang ju xu, are key points for post operative ileus. A review found the mechanisms involved include; 
• Activating the Janus kinase 2/signal transducer and activation of transcription 3 signaling pathway mediated by α7 nicotinic acetylcholine receptor in macrophages, leading to a reduction in the production of inflammatory cytokines
• inhibit the expression of gamma absorptiometry aminobutyric acid receptors in the dorsal motor nucleus of the vagus nerve, thereby activating the vagus nerve and improving inflammation
• activating the α7 nicotinic acetylcholine receptor signaling pathway mediated by the vagus nerve, independent of the spleen
• stimulation of the vagus nerve by Th1 cells and the inhibition of inflammatory responses. 
• stimulating neurons of the solitary tract nucleus, rather than activating the cholinergic anti-inflammatory pathway.
• regulating the macrophage interleukin-6/microRNA-19a/C-kit signaling pathway (66).

Acute gastrointestinal injury (in critically ill patients)

• Acupuncture, daily, primarily at Hegu (LI4), Zhongwan (CV 12), Tianshu (ST 25), Zusanli (ST 36), Shangjuxu (ST 37), and Xiajuxu (ST 39), is able to normalize the gut on ultrasound imaging, controlling gastrointestinal secretion, improving gastrointestinal motility, and minimizing side effects in critically ill patients. Acute gastrointestinal injury in these patients is a predictor of mortality (67). 

Microbiome cont.

• Acupuncture, where Zu san li is a key point, can help to regulate the gut microbiome and benefit a variety of conditions, including; ulcerative colitis, Chron’s, IBS functional constipation, chronic acute gastritis as well as mental ill health, metabolic disorders, cardiac and cognitive decline, cancer related fatigue, sleep, osteoarthritis and even premature ovarian failure (68, 69). 
• A meta-analysis showed that acupuncture increases the Firmicutes and decreases the Bacteroidetes in treatment groups, and increases the Lactobacillus and Ruminococcus genera (70). 

Adhesive intestinal obstruction

• A review found acupuncture to be an effective therapy for the life threatening condition with Zu san li, Tian shu and Shang ju xu among the most commonly used points (71).

Gastric ulcers

• ‘EA using different acupoint combinations [He-Mu (ST36-CV12), Shu-Mu (BL21-CV12), and Yuan-Luo (ST42- ST40)] differentially alleviated gastric mucosal injury in GU rats, with the He-Mu group exhibiting superior tissue damage alleviation, as well as inflammation and oxidative stress reductions. A Venn diagram transcriptome analysis revealed a shared central pathway among the three groups, corresponding to focal adhesion. Quantitative validation confirmed that the mRNA, protein, and phosphorylated protein expression of FAK, VCL, and EGFR—the core signal transduction factors of the focal adhesion pathway activated in gastric tissue after EA treatment—were upregulated, consistent with their therapeutic efficacy.’
• FAK (focal adhesion kinase) is central to signal transduction and cellular repair
• VCL (Vinculin) supports cell adhesion and structural integrity
• EGFR (epidermal growth factor receptor) triggers cell growth and tissue repair (72)
• EA at ST36 and Zhong wan (REN12) in comparison with the model group showed that, the ulcer index and lesion score, the levels of malonaldehyde and myeloperoxidase, contents of serum TNF-α and IL-6, expression levels of TLR4 and MyD88 proteins in positive drug and He-Sea-Front-Mu point combination groups were significantly decreased, while glutathione peroxidase and IκB-α were significantly increased. Thus EA can prevent stress induced gastric ulcers, which may be related to its effects in anti-oxidant, anti-inflammatory and regulating TLR4/MyD88/IκB signaling pathway. (73)

Gastric reperfusion injury

• EA (2/100Hz, 2mA) at Zu san li reduces inflammation, showing lower tumor necrosis factor alpha and interleukin 8 in the blood, lung, gut and liver. It also decreased intestinal permeability. This effect was mediated through the vagus nerve and the cholinergic anti-inflammatory dependent pathway, as severing the verve or blocking acetylcholine receptor blocked these effects (74).

Type 2 Diabetes Mellitus

• EA at Zu san li increases the survival rate in diabetic dementia rats. Following 4 weeks’ interventions, the enhanced levels of tau phosphorylation and GSK-3β activity in pancreas and hippocampus were partly reversed in the EA group compared to the model group, thus improving metabolic and neuronal degradation (75). 
• EA of ST36 and SP6 can improve glucose and lipid metabolism and insulin resistant, obese diabetic rats, which may be related to its function in suppressing PKCβ/P66shc signaling and oxidative stress. Researchers showed significant reductions in the levels of fasting blood glucose, fasting serum insulin, homeostasis model assessment of IR, total cholesterol, triglycerides, low density lipoprotein cholesterol, malondialdehyde, and reactive oxygen species. Additionally expressions of P66shc mRNA and PKCβ mRNA were remarkably down-regulated and insulin sensitive index, high density lipoprotein cholesterol, superoxide dismutase, and serum glutathione peroxidase were up-regulated (76)

It is also one of the most commonly used points in fatty liver, along with Feng long (ST40) and Tai chong (LV3) (77). For which it is shown to inhibit inflammatory responses, regulate lipid metabolism disorder, lowering glucose levels and treating insulin resistance, ease oxidative stress injury, and endoplasmic reticulum stress and thus improving hepatic function and mitigating liver fibrosis (78, 79). 

EA at Feng long (ST40) – Zu san li (ST36) and Zhongwan (REN12) – Guan yuan (REN4) can reduce the 24-hour food intake, body weight and serum insulin in the EA group were, while the glucose infusion rate was significantly increased. The Bacteroides abundance was increased, the pachytene abundance was decreased and the functional clustering of the above functional cog protein in the EA group was increased. It was also noted that intestinal flora, at the genus level, was riveted back to normal. Thus acupuncture can reduce appetite, regulate weight loss and may be related to its benefits on the intestinal microflora (80). 

EA, at Zu san li and Feng long (2Hz, 1mA, 20 minutes, 3 times a week for 11 weeks), can reduce the number of tumors and inflammation reaction in colorectal tissue and improve the body condition in mice with colorectal cancer, which may be related to its functions in activating the expression of intestinal SIRT1, and then facilitating cellular autophagy. (81)

An interesting review of acupuncture mechanisms for obesity summed up the above effects as such; ‘In addition, as a kind of mechanical stimulation for skin and subcutaneous tissue, the effect of acupuncture was considered to be related to the somatosensory autonomic reflex. Experimental evidence has shown that the neural circuit between peripheral and central nervous system is involved in the mechanism by which EA improves inflammation via driving the vagus nerve-adrenal axis. Similarly, as mentioned above, the peripheral appetite signal mediated by the vagus nerve is also an important mechanism by which EA participates in regulating energy metabolism. Further, our study also shows that EA-mediated activity of GLP-1 neurons in the solitary tract nucleus can affect peripheral adipose metabolism through central regulation, but the neural circuit mechanism that links the central to peripheral nervous system has not yet been elucidated. Recent studies suggest that the metabolism of adipocytes may be linked to sympathetic excitability regulated by the hypothalamus. Exploration of the role played by sympathetic nerves in mediating adipocyte metabolism is a promising avenue for advancing the understanding of acupuncture’s effects on obesity.’ (82)

An interesting study found that looked into the effects of Zu san li on intragastric pressure and the core modulating tissue as well as its pathway and found that, ‘Zusanli acupuncture therapy effectively elevated intragastric pressure, but inhibited expression of c-fos in oxytocinergic neurons of the paraventricular nucleus in upper cervical cord injured rats. These Zusanli acupuncture effects remained even after complete dorsal cord transection. However, after complete transection of the spinal cord or dorsolateral funiculus, the effects were significantly attenuated and even disappeared. These findings suggest that the paraventricular nucleus is responsible for pooling and integrating signals from the Zusanli acupuncture and sensory information from the intragastric pressure variation, thereby contributing to the regulation of intragastric pressure. The upper cervical cord serves as the key link between ascending and descending pathways, which conveys afferent inputs to the paraventricular nucleus through the dorsolateral funiculus.’ (83)

EA stimulation at ST36 can have a bidirectional effect on gastric motility shown in the frequency and average peak amplitude. Additionally, EA stimulation at ST36 regulated the expression of some genes in the PKC and MAPK signaling pathways, and it regulated the expression of the CaD and CaP proteins. EA serum induced SMC contractility. Promotion of gastric motility may correlate with up-regulation of MAPK6 (ERK3), MAPK13, and Prostaglandin-endoperoxide synthase 2 (PTGS2) gene expression, and the down-regulation of the collagen, type I, alpha 1 (COL1A1) gene and CaD and CaP protein expression. Inhibition of gastric motility may correlate with down-regulation of the Interleukin-1 receptor type 2 (IL1R2) and Matrix metalloproteinase-9 (MMP9) genes, and up-regulation of CaD and CaP protein expression. (84)

EA at the Zusanli point significantly promoted the intestinal impelling ratio and increased the amount of mucosal blood flow after scald injury. The plasma diamine oxidase (DAO) and intestinal permeability decreased significantly after scald injury in the EA group compared with others. However, EA after atropine injection or cervical vagotomy failed to improve intestinal motility and mucosa blood flow suggesting that the mechanism of EA may be related to the activation of the cholinergic nerve pathway. (85)

Another study looked at the enteroendocrine effects of EA at ST36, as noted in some of the other studies, if found that, ‘EA stimulus at St36 has the potential to influence gastric mucous substances and enteroendocrine cells (gastrin, serotonin, CGRP, insulin, and PP) that subsequently modulate digestive functions.’ (86)

In its first of a kind, researchers looked at a comprehensive genomic response to EA at Zu san li (2Hz, to the level of muscle twitching) for 15 minutes. It is worth reading the study in full. The broad strokes are that, ‘EA [at Zu san li] affected the expression of genes not only in the acupunctured site but also in the internal organs. EA commonly affected biological networks involved in cytoskeleton and cell adhesion, and also regulated unique process networks in specific organs, such as γ-aminobutyric acid-ergic neurotransmission in brain and inflammation process in lung. In addition, EA affected the expression of genes related to various diseases, such as neurodegenerative diseases in the brain and obstructive pulmonary diseases in the lung.’ (87)

Another gene study tried to find out which genes are responsible for ageing in rats and whether acupuncture can slow down the aging process. The researchers found 4052 genes in rats, 3079 were expressed differently between the faster ageing rats and the controls. EA at Guan yuan (REN4), Zu san li and Bai hui (DU20) regulated 983 of those genes and was more similar to the control than the ageing group. (88)

Zu san li is a potent point for sepsis patients:

• Electroacupuncture at Zu san li and Shang ju xu can reduce inflammatory markers TNF-α and IL-1β and intraabdominal pressure (89)
• EA has clinical benefits in relieving septic inflammation, improving immune function, and attenuating related multi-organ injury through several mechanisms, such as activation of the cholinergic anti-inflammatory pathway (CAP), vagal-adrenal axis, inhibition of the nuclear factor Kappa-B (NF-κB) signaling pathway, signal transducers and activators of transcription (STAT) signaling pathway, and improvement of immune cell function. Zu san li (ST36), Tian shu (ST25), Shang ju xu (ST37) and Xiajuxu (ST39) was the second most used prescription in this review, after Zu san li and Guan yuan (90). 
• EA (twice a day for 5 days) at ST36-ST37 can reduce inflammatory reaction and has protective effects on intestinal function in patients with sepsis-induced intestinal dysfunction. Acupuncture with medicines reduced PCT, TNF-α, I-FABP, D-lactate more strongly than the medication alone and citrulline was significantly increased (91). 
• A review pointed to all of these pathways, ‘mechanistically, a change in the acupoint microenvironment is the initial response link for acupuncture to take effect, whereas PROKR2 neurons, high-threshold thin nerve fibres, cannabinoid CB2 receptor (CB2R) activation, and Ca2+ influx are the key material bases. The cholinergic anti-inflammatory pathway of the vagus nervous system, the adrenal dopamine anti-inflammatory pathway, and the sympathetic nervous system are key to the transmission of acupuncture information and the inhibition of systemic inflammation. In MODS, acupuncture protects against septic organ damage by inhibiting excessive inflammatory reactions, resisting oxidative stress, protecting mitochondrial function, and reducing apoptosis and tissue or organ damage.’ (92)
• An interesting study had many groups of rats to compare what benefit acupuncture at Zu san li, pre or post sepsis modelling, could have. The researchers concluded that; ‘ST36 acupuncture pretreatment significantly attenuated the [sepsis model] kidney injury and the increases in renal nitric oxide concentration and inducible nitric oxide synthase expression. However, ST36 acupuncture pretreatment did not affect the LPS-induced liver injury and increases in hepatic NO concentration or iNOS expression. Furthermore, ST36 acupuncture performed after LPS did not affect the LPS-induced organ injuries or increases in NO concentration and iNOS expression.’ (93)
• Another study found ST36 would mitigate and mediate damage through the cholinergic anti-inflammatory pathway, lowering tumor necrosis factor alpha, and that severing the cervical vagal pathway would prevent this beneficial effect (94). 
• One study compared Qu chi, Zu san li and Tian shu to see their mechanisms in sepsis rats with or without spleenectomy. They found that; ‘EA at ST36 and LI11, respectively, could alleviate inflammation reaction, protect the intestinal barrier, and maintain intestinal T-cell function in septic rats. Spleen participated in the protective effect of EA at ST36 in sepsis.’ The rats with spleenectomy did not reduce inflammatory markets, but did increase CD3+CD4+/CD3+CD8+cells and Treg/Th17 cells. (95) 
• ‘EA at Zusanli (ST36) could inhibit the expression of HMGB1 in the jejunum of septic rats, and promote the expression of Ghrelin. The expression of HMGB1 was inhibited by Ghrelin receptor blocking agent, which suggested that the anti-inflammation of EA at Zusanli (ST36) might be associated with Ghrelin.’ (96)
• ‘EA preconditioning at ST36 obviously ameliorated CLP-induced intestinal injury and high permeability and reduced the mortality of CLP-induced sepsis rats. Moreover, electroacupuncture (EA) pretreatment exerted protective effects on intestinal mucosal immune barrier by increasing the concentration of sIgA and the percentage of CD3+, γ/δ, and CD4+ T cells and the ratio of CD4+/CD8+ T cells’ (97)
• EA at ST36 can reduce death rate and intestinal bacteria translocation incidence in sepsis rats, which may be related to its functions in regulating the expression of intestinal Bcl-2 and Bax proteins and inhibiting the apoptosis of intestinal mucosal T lymphocytes, thereby protecting the immune barrier function of intestinal mucosa to reduce the intestinal permeability (98)
• EA at ST36 reduced the expression of IL-1β, IL-6, and TNF-α and increased the expression of IL-10 to inhibit the inflammatory response. EA at ST36 also inhibited apoptosis and decreased the Bax/Bcl2 ratio and levels of caspase-3 and cleaved caspase-3, as well as lactate dehydrogenase release. Thus, alleviation of sepsis may correlate with the downregulation of levels of TLR4, NF-κB, and MyD88. Additionally, EA at ST36 improved the diversity of the intestinal flora and increased the abundance of Firmicutes and Actinobacteria. (99)

An interesting study on cancer causing toxin injected mice found that daily electroacupuncture at Zu san li for 12 days not only significantly reduced the inflammatory markers (IL-1β, IL-6, TNF-α and IL-17A in serum and IL-17A, CXCL1 and IL-23 in colon tissue – all of which were positively correlated with colon cancer ‘burden’). The mice in the acupuncture group had fewer and smaller tumors. Researchers also noted lower Proliferating Cell Nuclear Antigen, which is higher in tumor growth, and higher apoptosis (of the cancer cells) (100). 

EA at Zusanli was also found to have a protective effect on hepatic ischemia refusion injury in mice by alleviating oxidative stress, hepatocyte death, and inflammation response. Nuclear factor E2-related factor 2 (Nrf2) as a crucial target was regulated by EA and was then successfully validated. The Nrf2 inhibitor ML385 and cervical vagotomy eliminated the protective effect in the EA treatment group (101).

EA of ST36 can enhance the effect of capeOX in inhibiting colorectal cancer growth in nude mice with CRC, which may be related with its functions in promoting tumor cell apoptosis, inhibiting ferroptosis, and modulating immune tolerance. In addition, EA can lower the side effects of capeOX (combined administration of fluorouracil, oxaliplatin and capecitabine) in hematopoietic and immune, liver, and kidney functions. This was shown in ‘the tumor volume difference, tumor weight, and contents of serum malondialdehyde (MDA), alanine aninotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), and creatinine (Cr) in the EA+capeOX group were markedly lower than in the capeOX group, while the spleen coefficient, proportion of apoptotic cells and GPX4 expression level in the EA+capeOX group were markedly higher than those in the capeOX group’ (102)

In chronic psychological stress (CPS) rats, EA at ST36 could improve food intake, weight, visceral hypersensitivity, and immunity; in CPS rats, in small intestine, the morphology of interstitial cells of Cajal (ICC) was abnormal and the number was decreased, which may be part causes of gastrointestinal motility dysfunction. EA at ST36 showed useful therapeutic effects. The mechanisms may be partially related to its repairing effects on ICCs damages and its immunomodulation (103). 

This was similarly shown in another study with spinal cord injury (SCI) rats, where, ‘abnormal ICCs in colon tissues and the downregulated expression of c-Kit could be observed after SCI. EA at Zusanli (ST36) could improve the colon function by repairing the morphology and increasing the number of ICCs and upregulating c-Kit expression (104)

EA at Zu san li improves the blood pressure and raises the early survival rate of hemorrhagic shock rats, maintains the intestinal barrier function, and improves the degree of intestinal ischemia (105)

A study looking into the effects of acupuncture for those suffering with chronic fatigue syndrome. They compared single acupuncture, to dual, control, sham, and with moxibustion and found that; ‘Acupuncture excelled in regulating the immediate effects of heart rate variability, while moxibustion excelled in regulating long-term effects. Acupuncture targeting both the Zusanli and Guanyuan points proved more effective than acupuncture targeting a single point, with this difference reflected in the distinct regulatory effects on the sympathetic and parasympathetic nervous systems. The intervention mechanism of Zusanli may involve the parasympathetic nervous system, whereas Guanyuan may operate through the sympathetic nervous system.’ (106)

Acupuncture at Ming men (GV4) and Zu san li (ST36) can help lupus sufferers. It triggers inhibition of transforming growth factor-β1 expression in lupus nephritis model mice, which may help with healing, including neuroprotection and gut lining health. (107).

EA at ST-36 and PC6 acupoints can attenuate endotoxin shock-induced myocardial injury in rabbits and upregulation of HO-1 expression and inhibition of inflammatory responses may be involved in the mechanism (108).

Research has shown that acupuncture at ST36 can influence microcirculation. A study found that acupuncture at ST36 increased skin and muscle blood flow in healthy individuals (109).

EA at Zu san li acupoint (3Hz) can improve blood coagulation during intestinal I/R in rats,and the mechanism is related to activating the cholinergic anti-inflammatory pathway. Significantly decreasing TNFα, tissue factor (TF), tissue plasminogen activator (tPA),fiber plasminogen activator inhibitor-1 (PAl-l) and D-dimer, whilst the plasma antithrombin (AT) concentration and platelet count (PLT) were increased (110).  

The antidepressant effect of EA at ST36 and ST25 is related to regulating intestinal flora and the neurotransmitter system. EA significantly increased the intestine propulsion rate, the relative abundance of Bacteroidetes, Proteobacteria, and Actinobacteria at the phylum level, Bifidobacterium and Streptococcus at the genus level, and vasoactive intestinal peptide (VIP),  and CGRP in plasma, and significantly decreased Firmicutes, the ratio of Firmicutes to Bacteroidetes at the phylum level, somatostatin (SST), and adrenocorticotropic hormone (ACTH) in plasma, and SST mRNA in the colon (111)

‘EA at ST36 ameliorated the locomotor activity, social behavior, spatial learning and memory and repetitive behavior compared with autistic spectrum disorder rats. It decreased oxidative stress markers in the tissues of prefrontal cortex, enhanced translocation of nuclear factor erythroid2-related factor2 (Nrf2) from cytoplasm to nucleus, and up-regulated the levels of NADP(H) quinone oxidoreductase (NQO1) and heme oxygenase (HO-1). However, these effects induced by electroacupuncture at ST36 were abolished after injection of Ad-siRNA-Nrf2’, which silences the Nrf2 gene (112).

Additionally, researchers found that, ‘after ST36 acupuncture, pain sensitivity, autonomous activity, sociability index, sociability preference index, and learning and memory were improved in the autism model rats. Levels of TXNIP, NLRP3, IL-1β, and caspase 1 were decreased after acupuncture. Interference with TXNIP alleviated the behavior disorders and inhibited NLRP3, caspase 1, and IL-1β levels. In summary, ST36 acupuncture reduced TXNIP expression, inhibited the activation of the NLRP3 inflammasome, and alleviated the behavior disorder related to the prefrontal cortex of the autistic rats.’ (113)

A study compared the effects of acupuncture at ST36, GB34, and ST36+GB34 on obese rats who were developing osteoarthritis. It has been shown the acupuncture can mitigate the pathogenesis of osteoarthritis, but not whether it can mitigate obesogenic osteoarthritis, which is triggered by lower HDL and higher oxidised LDL that can trigger release of MMPs  (a family of compounds that are catabolic/destructive to cartilage tissue). ‘The results showed that the three acupuncture protocols both prevented obesity-induced cartilage matrix degradation and MMP expression and mitigated obesity-induced systemic and local inflammation but had different regulatory effects on lipid metabolism and gut microbiota disorder of obese-induced OA rats. Furthermore, the three acupuncture protocols increased the microbial diversity and altered the structure of the community of feces in obese rats. We found that ST36 and GB34 could inhibit proinflammatory shift in the gut microbiome with an increase in the ratio of Bacteroidetes/Firmicutes and promote the recovery of relative abundance of Clostridium, Akkermansia, Butyricimonas, and Lactococcus. Although both ST36 and GB34 had an anti-inflammatory effect on serum inflammatory mediators, only the acupuncture protocol with both ST36 and GB34 could effectively inhibit LPS-mediated joint inflammation in obesity rats. Therefore, relieving obesity-related chronic inflammation, lipid metabolism disorder, and gut microbiota disorder may be an important mechanism for acupuncture with ST36 and GB34 to promote OA recovery’ (114)

Electroacupuncture at ST35 and ST36 can significantly alleviate the hyperalgesia and cartilage degeneration as well as reducing nerve sprouting in OA knee joint. Moreover,

acupuncture treatment may inhibit the MCP1/CCR2 axis as well as down-regulate inflaming factor and NGF in cartilage and synovial tissue (115).

‘EA at [Zu san li and San yin jiao] treatment enhanced the A2AR activity and inhibited osteoclast formation by inhibition of RANKL, RANK, TRAF6, p38α, NF-κB, and NFATc1. SCH58261 reversed the effect of EA. These results suggest that EA regulated p38α-MAPK signaling by increasing A2AR activity, which inhibited osteoclastogenesis.’ (116)

EA at ST36 can correct the imbalance of mitochondrial fission (too much is linked to neurodegeneration, heart disease, and inflammation) and fusion (too much may impair removal of damaged mitochondria) in skeletal muscle of rats with spleen qi deficiency, thereby improving the damage of mitochondrial structure and function, and leading to an increase of energy metabolism. (117)

Needling-ST36 benefits rats with lower-limb ankle arthritis. The mechanism, at least in part, relies on NTPDase1 activities in the sciatic nerve. In addition to facilitating adenosine signaling in conjunction with ecto-5′-nucleotidase (NT5E), most importantly, NTPDase1 may provide an appropriate low-level ATP milieu for the activation of P2Y2R in the sciatic nerve, particularly in Schwann cells. (118). 

Researchers found that electroacupuncture at Zusanli (ST36) could improve muscle healing following jumping-induced muscle injury, by upregulating Heat shock protein beta-7 (Hspb7) and myozenin2 (Myoz2) proteins (119).

Additional research found that, ‘Both EA-ST36 and EA-Ashi promoted myofiber regeneration and restoration of neuromuscular junctions. EA-Ashi was more effective at earlier stages, whereas EA-ST36 played a more important role at later stages.’ They increased the levels of  expressions of acetylcholinesterase (AChE), Neuregulin 1 (NGR1), and muscle-specific kinase (MuSK) and decreased growth differentiation factor-8 (GDF-8) (120)

TRPA1 (Transient receptor potential ankyrin 1) plays a significant role in the analgesic effects produced via acupuncture at the ST36 (121).

‘EA of ST36 and REN4 can promote the apoptosis of synoviocytes and the expressions of Fas and FasL proteins in adjuvant-induced arthritis rats, which may contribute to its role in relieving synovitis and the related inflammatory markers through activating Fas/FasL signaling’ (122). 

Researchers also found that ‘We found that EA [at ST36 and GB39] significantly decreased arthritis index scores, paw volume, and HE staining scores. EA could significantly inhibit the expression of CD34, promoting apoptosis of synovial cells in the joint synovial tissue of adjuvant arthritis rats. The expression of Notch1 signaling pathway proteins and mRNAs (Notch1, Hes1, TGF-β, and bFGF) were [also] markedly downregulated by EA treatment.’ (123)

A systematic review looked at whether acupuncture can exert an excitatory effect on the spinal cord and thus improve motor function conditions. They found that both EA and MA could effectively and safely enhance corticospinal excitement, bringing the corticospinal pathway closer to the threshold for firing, which may ultimately improve motor function. LI4, ST36, LI11, SJ5 and GB34 are the most commonly used acupoints (124). 

Zu san li for rheumatoid arthritis rats effectively corrected the related metabolic disorders. Plasma metabolomics identified a total of 10 differential metabolites primarily associated with arachidonic acid and pentose phosphate metabolic pathways (125)

Additionally, acupuncture markedly decreased heat hyperalgesia and paw swelling in acquired arthritis rats, decreased levels of pro-inflammatory cytokines (tumor necrosis factor (TNF)-α, interleukin (IL)-1β) and increased anti-inflammatory cytokines (IL-10, transforming growth factor (TGF)-β1) in the ankle joints at protein and mRNA levels. CCC network analysis confirmed that macrophages are of critical importance and are potential therapeutic targets in RA. Repeated treatment with MA triggered a macrophage phenotypic switch in the paws, with fewer M1 macrophages. Prominent increases in the Treg cell population and TGF-β1 in the popliteal lymph nodes demonstrated the immunomodulatory effects of MA. Furthermore, a selective TGF-β1-receptor inhibitor, SB431542, attenuated the anti-inflammatory effects of MA and MA-induced suppression of the levels of M1-released cytokines (126)

‘The therapeutic effect of MA on AIA is achieved primarily through the regulation of steroid hormone biosynthesis, cell metabolism, and tissue repair processes. MA at ST36 can increase the gene contents of tissue repair growth factors, including PEG3, GADD45A, GDF5, FGF5, SOX2, and ATP6V1C2 in the inflammatory side joints of rheumatoid arthritis mice, as well as the gene expression of the anti-inflammatory cytokine IL-10.’ (127)

Further researchers look at immune cells and their mediators with manual acupuncture at ST36 showing anti-inflammatory and analgesic effects as well as inhibition of immune cell communication networks in inflamed joints of AIA rats. The surmised that inhibiting the polarization of macrophages to the M1-phenotype in inflamed joints may be one of the key mechanisms of MA anti-inflammatory action. (128)

Acupuncture at LI4 (Hegu), SJ6 (Zhigou), ST36 (Zusanli) and GB34 (Yanglingquan) for patients with traumatic rib fracture can significantly lower pain levels at rest and deep breathing (129). 

Electro-acupuncture can regulate the expression of inhibition of DNA binding protein 2 (Id2) and myelin basic protein (MBP) after compressive spinal cord injury.The down-regulation of Id2 which controls MBP negatively and the up-regulation of MBP may contribute to remyelination in the injured spinal cord (130).

Several studies have suggested that the effect of stimulating the Zu San Li acupoint may be mediated by the neuroendocrine system. For example, one study found that electroacupuncture at Zu San Li decreased the expression of neuropeptide Y (NPY) in the hypothalamus and plasma of diabetic rats (131). NPY is involved in the regulation of food intake and energy metabolism. Another study found that acupuncture at Zu San Li increased the levels of β-endorphin, dynorphin A, and enkephalin in the hypothalamus and plasma, which are endogenous opioids that play a role in pain regulation and stress responses (132). 

Acupuncture at ST36 can inhibit ethanol withdrawal-induced hyperactivation of the hypothalamic–pituitary–adrenal axis, and it may be mediated via the modulation of hypothalamic corticotropin-releasing factor, which in turn regulates corticosterone and adrenocorticotropic hormone, both of which were also reduced by ST36 (133).

EA at ST36 and ST37 and PC6 and PC7 exerted an analgesic effect, EA at PC6 and PC7 can enhance heart function by modulating dopamine, adrenaline and noradrenaline in the extra cellular fluid, while EA at ST36 and ST37 modulates the cerebral cortex, by modulating glutamate and GABA in cerebrospinal fluid (134)

EA at PC6 causes downregulation of adenosine, adrenaline, γ-aminobutyric acid, glycine, and glutamate majorly in the hippocampus, and then in the cerebral cortex. EA at ST36 resulted in upregulation of adrenaline and arginine in hippocampus, and all stimulations showed barely change of identified neurotransmitters in hypothalamus. These changes show PC6 stimulation reducing neural excitability, especially in memory- and emotion-related areas like the hippocampus. It fits with sedative, anti-anxiety, or cardioprotective effects. Whilst ST36 may boost neural and metabolic activity, consistent with its known tonifying effects. (135)

EA (2/100Hz, 0.5-1.5 mA, 30 min/day for 7 days) at the acupoints Zu san li and San yin jiao reduced morphine tolerance in rats. Morphine ED50 curves were remarkably leftward shifted on day 8. Additionally, upregulated cAMP was reduced by acupuncture, which is part of the mechanism for drug tolerance. Phosphorylation levels of PKA, ERK, and CREB was also reversed (136).

A very interesting study was conducted to try to understand how a mechanical stimulation (acupuncture), could lead to neurological signalling. The researchers tested three pathways; transient receptor potential vanilloid receptors 1 and 4 (TRPV1 and TRPV4) and the acid-sensing ion channel 3 (ASIC3); as well as using agonists of those pathways to mimic acupuncture, all at Zu san li. The researchers found that there is an abundance of TRPV1, TRPV4 and ASIC3 at ST36, in different layers and neural and non-neural cells, but only the TRPV1 agonist mimicked the effects of Zu san li. The researchers concluded,’ these findings suggest TRPV1 might act as an acupuncture-responding channel by sensing physical stimulation from acupuncture and conducting the signaling via calcium wave propagation to nerve terminals.’ (137)

Another study noted that acupuncture at Zu san li can enhance peripheral perfusion and parasympathetic activity (138). 

EA ST36 induces nitric oxide release in the gracile nucleus, which contributes to improvement of sensory neuropathies in diabetic rats (139)

EA stimulation at Zu san li up-regulates the expression of ICAM-1 mRNA and enhances mast cell release from the stimulated regional subcutaneous tissue in healthy rats, suggesting ICAM-1 played an important role in affecting the movement and recruitment of mas cells towards acupoint during EA. (140)

Electro acupuncture at ST36 prevents nicotine damage in the lung tissue of neonatal rats. Researchers found that lung function; peak inspiratory flow (PIF), peak expiratory flow (PEF), lung resistance (RL), exhalation resistance (RE) and lung dynamic compliance (Cdyn), were all improved compared to nicotine exposed neonates that received no acupuncture. Additionally, congestion and edema of the alveolar wall, alveolar deformation, rupture and fusion, and reduction of the number of the pulmonary alveoli were evidently milder in those receiving acupuncture. Yang ling quan did not have this effect (141, 142)

Electroacupuncture at Lieque (LU7), Chize (LU5), and Zusanli (ST36) can lessen the lung injury induced by severe acute pancreatitis, increasing blood oxygenation, and the mechanism may be related to the decreased TNF-α and increased IL-10 value (143).

EA at Zu san li and Fei shu (BL13) improved lung function of rats with COPD and had an anti-inflammatory effect (reducing tumor necrosis factor a and interleukin 1beta), which may be related to down-regulation of Orexin alpha (OXA) and its receptors. Orexins (OXs), found in peripheral plasma, are neuropeptides that regulate respiration and their levels are related to COPD (144). 

A systematic review concluded that Fei shu and Zu san li should be the basis of any acupuncture prescription for COPD and that its mechanisms are enacted via the regulation of relevant anti-inflammatory pathways; ‘nuclear factor-κB (NF-κB) (e.g., myeloid differentiation, primary response 88/NF-κB, toll-like receptor-4/NF-κB, silent information regulatortranscript-1/NF-κB), mitogen-activated protein kinase signalling pathways (extracellular signal-regulated kinase 1/2, p38 and c-Jun NH2-terminal kinase), cholinergic anti-inflammatory pathway, and dopamine D2 receptor pathway.’ (145)

EA at ST36 can improve COPD symptoms in smoking triggered COPD in rats. Showing decreased lung resistance and increased lung compliance, reduced bronchi and bronchiole airway obstruction and lower inflammation (TNF-α, IL-1β, and malonaldehyde in bronchoalveolar lavage fluid were lowered, albeit not to control levels (146). 

In mice with chronic asthma acupuncture at BL13 does not decrease IL-17 significantly but a combination of acupuncture at BL 13 and ST 36 does (147)

A very curious study used metabolomics to study the hypotensive effects of Tai chong (LR3) and Zu san li. Interestingly they found that ‘abnormal bile acid metabolism may be an independent risk factor the development of hypertension.’ LR3 and ST36 effectively modulated bile acids metabolism in spontaneously hypertensive rats renal cortex tissues to exert a hypotensive effect. Cholic acid, was one of the acids acupuncture regulated and may be a new target for the treatment of hypertension (148)

There is also research showing that acupuncture at Zu San Li can benefit contact dermatitis by modulating local pro and anti inflammatory processes (149).

EA at ST36 may ameliorate inflammation associated with DNFB-induced allergic contact dermatitis via triggering local (to the ear in this study) anti-inflammatory IL-10 production, reducing IgE, attenuating CD4+IFN-γ+ and CD4+IL-4+ T cells and inhibiting p38 MAPK activation (150) 

EA at ST36 relieved the pathological progression of delayed-type hypersensitivity responses by reducing footpad swelling, infiltration of inflammatory cells, IgG and IgE, IFN-γ and TNF-α in homogenized footpad tissue. Moreover, EA attenuated the percentage of CD4+IFN-γ+ T cells and prevented Th cells differentiation into Th1 cells, and this results from inhibiting secretion of IFN-γ and suppressing expression of T-bet, an IFN-γ transcription factor. Thus EA improved Th1-mediated allergic skin inflammation via restoring Th1/Th2 balance by curbing Th1 differentiation. Suggesting EA at ST36 might be a useful and promising therapeutic for allergic inflammatory as well as Th1-mediated inflammation response (151).

Acupuncture at Zu san li significantly increased both non-rapid eye movement and rapid eye movement sleep improves pain-induced sleep disturbances by increasing total sleep time and reducing wakefulness. Naloxone microinjection into the NTS blocked acupuncture’s sleep-promoting and analgesic effects, suggesting involvement of the endogenous opioid system. The researchers also noted pain thresholds, measured through paw withdrawal tests, were significantly elevated in acupuncture-treated rats, confirming the analgesic effects of acupuncture (152).

A study looked at the effects of acupuncture on anaesthesia, hypothesizing that the benzodiazapine-GABA pathway may be involved. They found that The drop of Narcotrend Index (which shows lowered brain activity) induced by EA didn’t reverse after administration of flumazenil (a blocker of the GABA-A receptor). Thus the effect of EA [at Zu san li and Nei guan] in deepening anesthesia may not be mediated by the benzodiazepines-GABA signaling pathway (153).

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