Electroacupuncture (EA) can effectively alleviate anxiety disorders and memory impairments caused by various neurodegenerative diseases; however, the molecular mechanisms underlying its neuroprotective effects are unclear. Previous studies have shown that the renin-angiotensin system (RAS) comprises of two axes with mutual antagonism: the classical angiotensin converting enzyme/angiotensin II/angiotensin II type 1 receptor (ACE/Ang II/AT1R) axis and the protective angiotensin converting enzyme 2/angiotensin-(1-7)/Mas receptor (ACE2/Ang-(1-7)/MasR) axis. In this study, we observed that chronic cerebral hypoperfusion (CCH) mediated anxiety-like behavior and memory impairments in spontaneously hypertensive rats (SHR) via upregulation of the hippocampal classical axis (ACE/Ang II/AT1R) and the partial hippocampal protective axis (ACE2/Ang-(1-7)). However, Ang II levels were much higher than those of Ang-(1–7), indicating that the ACE/Ang II/AT1R axis plays a dominant role in the comorbidity of CCH and hypertension. Moreover, candesartan cilexetil (Canc) and perindopril (Peril) were used as positive control drugs. We found that EA, Canc, and Peril attenuated CCH-induced anxiety-like behavior and memory impairments in SHR, potentially via downregulation of the hippocampal classical axis (ACE/Ang II/AT1R) and upregulation of the whole hippocampal protective axis (ACE2/Ang-(1-7)/MasR). These results suggest that EA therapy for CCH with hypertension may be mediated by two hippocampal RAS axes.

Background: Clinical patients suffering from pain usually exhibit aversion to pain-associated environments (pain aversion). Electro-acupuncture (EA) has been proven to be effective for the treatment of pain aversion in our previous studies. The amygdala could have substantial consequences on emotion and pain consolidation as well as general pain aversion behavior, however, the underlying mechanism remains unclear. Purpose: The current study was performed to investigate Isobaric tags for relative and absolute quantitation (iTRAQ) based quantitative proteomic analysis of the amygdala in rats with complete Freund’s adjuvant (CFA)-induced pain aversion, and comprehensive analysis of protein expression were performed to explore the underlying mechanism by which EA affects pain aversion. Materials and methods: Inflammatory pain was induced with an intraplantar injection of 100 μL of CFA in the plantar surface of the left hind paw of the male Spragure-Dawley (SD) rats. Then the CFA-induced conditioned place aversion (C-CPA) test was performed. EA stimulation on the bilateral Zusanli and Sanyinjiao acu-points was used for 14 days and the EA stimulation frequency is 2 Hz. Based on iTRAQ-based proteomics analysis, we investigated the protein expression in the amygdala. Results: EA can increase the paw withdrawal threshold in inflammatory pain induced by noxious stimulation. A total of 6319 proteins were quantified in amygdala. Of these identified proteins, 123 were identified in the pain aversion group relative to those in the saline group, and 125 significantly altered proteins were identified in the pain aversion + EA group relative to the pain aversion group. A total of 11 proteins were found to be differentially expressed in the amygdala of pain aversion and EA-treated rats. The expression of three proteins, glyceraldehyde-3-phosphate dehydrogenase, glutamate transporter-1, and p21-activated kinase 6, were confirmed to be consistent with the results of the proteome. Conclusion: Our investigation demonstrated the possible mechanism of central nerve system by which EA intervetion on pain aversion.

Comorbid chronic neuropathic pain and anxiety is a common disease that represents a major clinical challenge. The underlying mechanisms of chronic neuropathic pain and anxiety are not entirely understood, which limits the exploration of effective treatment methods. Glutamatergic neurons in the ventrolateral periaqueductal gray (vlPAG) have been implicated in regulating pain, but the potential roles of the vlPAG in neuropathic pain-induced anxiety have not been investigated. Herein, whole-cell recording and immunofluorescence showed that the excitability of CamkIIα neurons in the vlPAG (vlPAG^CamkIIα+neurons) was decreased in mice with spared nerve injury (SNI), while electroacupuncture (EA) activated these neurons. We also showed that chemogenetic inhibition of vlPAG^CamkIIα+neurons resulted in allodynia and anxiety-like behaviors in naive mice. Furthermore, chemogenetic activation of vlPAG^CamkIIα+neurons reduced anxiety-like behaviors and allodynia in mice with SNI, and EA had a similar effect in alleviating these symptoms. Nevertheless, EA combined with chemogenetic activation failed to further relieve allodynia and anxiety-like behaviors. Artificial inhibition of vlPAG^CamkIIα+neurons abolished the analgesic and anxiolytic effects of EA. Overall, our study reveals a novel mechanism of neuropathic pain-induced anxiety and shows that EA may relieve comorbid chronic neuropathic pain and anxiety by activating vlPAG^CamkIIα+neurons.

Our previous studies have confirmed that electroacupuncture (EA) can effectively intervene in pain memory, but the neural mechanism involved remains unclear. In this study, we observed the effects of EA in regulating pain memory-related behaviors and synchronous neural oscillations in the rostral anterior cingulate cortex (rACC). During nociceptive behavioral testing, pain memory induced a nonpain stimulus that spurred a neural oscillatory reaction similar to that caused by pain stimuli in the rACC. After EA, nonpain stimuli did not induce decreased neural oscillatory activity in the rACC until the presentation of pain stimuli. During aversive behavioral testing, EA, through the downregulation of theta power, inhibited the retrieval of aversive memory and relieved pain memory-induced aversive behaviors. These changes of oscillatory activity may be the hallmarks of EA therapy for pain memory.

Pain-related aversive memory is common in chronic pain patients. Electroacupuncture has been demonstrated to block pain-related aversive memory. The insular cortex is a key region closely related to aversive behaviors. In our study, a potential mechanism underlying the effect of electroacupuncture treatment on pain-related aversive memory behaviors relative to the insular cortex was investigated. Our study used the chemogenetic method, pharmacological method, electroacupuncture intervention, and behavioral detection. Our study showed that both inhibition of gamma-aminobutyric acidergic neurons and activation of the kappa opioid receptor in the insular cortex blocked the pain-related aversive memory behaviors induced by 2 crossover injections of carrageenan in mice; conversely, both the activation of gamma-aminobutyric acidergic neurons and inhibition of kappa opioid receptor in the insular cortex play similar roles in inducing pain-related aversive memory behaviors following 2 crossover injections of carrageenan. In addition, activation of gamma-aminobutyric acidergic neurons in the insular cortex reversed the effect of kappa opioid receptor activation in the insular cortex. Moreover, electroacupuncture effectively blocked pain-related aversive memory behaviors in model mice, which was reversed by both activation of gamma-aminobutyric acidergic neurons and inhibition of kappa opioid receptor in the insular cortex. The effect of electroacupuncture on blocking pain-related aversive memory behaviors may be related to the activation of the kappa opioid receptor and inhibition of gamma-aminobutyric acidergic neurons in the insular cortex.

Research shows that across life, the incidence of mental illness is highest in the young. In the context of the COVID-19 pandemic, mental health issues of the young in particular have received global attention. The rostral anterior cingulate cortex (rACC) plays an important role in psychiatric disorders and chronic pain-psychiatric comorbidities. However, it remains unknown whether or how the afferent and efferent circuits of the rACC change with aging. In this study, we microinjected a retrograde tracer virus and an anterograde trans-monosynaptic virus into the rACC of young and middle-aged mice (both male and female), and systematically and quantitatively analyzed the whole-brain afferent and efferent connections of rACC at different ages and sexes. Notably, in young and middle-aged mice, afferents of the rACC belong to four groups of brain structures arising mainly from the amygdala [mainly basolateral amygdaloid nucleus (BLA)] and cerebral cortex (mainly orbital cortex), with a small part originating from the basal forebrain and thalamus. In contrast, efferents of the rACC belong to four groups of brain structures mainly projecting to the thalamus (mainly ventral anterior-lateral/ventromedial thalamic nucleus (VAL/VM)], with a very small part projecting to the amygdala, basal forebrain, and cerebral cortex. Compared with young mice, the BLA-rACC circuit in middle-aged mice (male and female) did not change significantly, while the rACC-VAL/VM circuit in middle-aged mice (male and female) decreased significantly. In conclusion, this study comprehensively analyzed the input-output neural projections of rACC in mice of different ages and sexes and provided preliminary evidence for further targeted research.