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To study the effect of electroacupuncture (EA) on the cholinergic anti-inflammatory pathway (CAP) by measurement of vagal activity in rats with high-fat diet (HFD)-induced obesity.Diet-induced obesity (DIO) was induced in 30 rats by feeding them a HFD for 12 weeks. A further 10 rats fed normal food comprised the lean diet (LD) control group. DIO rats were further subdivided into three groups that received a HFD only (HFD group, n=10), a HFD plus electroacupuncture (HFD+EA group, n=10) or a HFD plus minimal acupuncture (HFD+MA group, n=10). EA and MA treatments were continued for 8 weeks. Heart rate variability (HRV) was used to measure the function of the autonomic nervous system before and after treatment. ELISA was used to determine acetylcholine (ACh) and tumour necrosis factor (TNF)-α levels in the serum. Real-time PCR was used to assess the mRNA expression of α7-subtype nicotinic acetylcholine cholinergic receptors (α7nAChRs) and TNF-α in the mesenteric white adipose tissues (MWAT).EA but not MA significantly reduced rats' bodyweight. No difference was found in the low frequency (LF), high frequency (HF) and the balance between LF and HF (LF/HF) components of HRV before treatment. After the EA intervention, HF was elevated and LF/HF was reduced in the HFD+EA group comparedwith the HFD group. TNF-α in the serum and MWAT were increased in the HFD group, but were reduced in the HFD+EA group. Furthermore, EA promoted expression of α7nAChRs and ACh in the MWAT. There was no difference between the HFD and HFD+MA groups for any indices.EA enhanced vagal activity, promoted ACh release and activated α7nAChRs in the MWAT, leading to inhibition of proinflammatory cytokine production.