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Abstract Chronic pain is often accompanied by anxiety, and gradually increasing anxiety makes the pain itself more protracted. Berberine has been found to be able to cross the blood–brain barrier to treat psychiatric disorders, but its neurocirculatory mechanisms remain unclear. Here, we found that neurons in cingulate area 2 (Cg2) of the caudal anterior cingulate cortex (cACC), but not in Cg1 of the cACC, projected to the ventral lateral thalamus (VLT). Next, we induced chronic inflammatory pain by plantar injection of complete Freund’s adjuvant (CFA) and observed stable anxiety-like behaviors until two weeks postinjection. We specifically activated VLT-projecting cACC (Cg2) neurons in one-week-old CFA-induced mice without anxiety-like behaviors and in normal control mice to induce anxiety-like behaviors. We inhibited the activation of VLT-projecting cACC (Cg2) neurons in two-week-old CFA-treated mice with anxiety-like behaviors and observed that their anxiety-like behaviors were alleviated. On this basis, we further screened the effective dose of berberine for anxiolysis in two-week-old CFA-treated mice. We observed that the effective dose of berberine obtained above decreased the activity of VLT-projecting cACC (Cg2) neurons. The activation of VLT-projecting cACC (Cg2) neurons abrogated the anxiolytic effect of berberine in two-week-old CFA-treated mice.

Abstract Chronic pain easily leads to concomitant mood disorders, and the excitability of anterior cingulate cortex (ACC) pyramidal neurons (PNs) is involved in chronic pain-related anxiety. However, the mechanism by which PNs regulate pain-related anxiety is still unknown. The GABAergic system plays an important role in modulating neuronal activity. In this paper, we aimed to study how the GABAergic system participates in regulating the excitability of ACC PNs, consequently affecting chronic inflammatory pain-related anxiety. A rat model of CFA-induced chronic inflammatory pain displayed anxiety-like behaviors, increased the excitability of ACC PNs, and reduced inhibitory presynaptic transmission; however, the number of GAD65/67 was not altered. Interestingly, intra-ACC injection of the GABAAR agonist muscimol relieved anxiety-like behaviors but had no effect on chronic inflammatory pain. Intra-ACC injection of the GABAAR antagonist picrotoxin induced anxiety-like behaviors but had no effect on pain in normal rats. Notably, chemogenetic activation of GABAergic neurons in the ACC alleviated chronic inflammatory pain and pain-induced anxiety-like behaviors, enhanced inhibitory presynaptic transmission, and reduced the excitability of ACC PNs. Chemogenetic inhibition of GABAergic neurons in the ACC led to pain-induced anxiety-like behaviors, reduced inhibitory presynaptic transmission, and enhanced the excitability of ACC PNs but had no effect on pain in normal rats. We demonstrate that the GABAergic system mediates a reduction in inhibitory presynaptic transmission in the ACC, which leads to enhanced excitability of pyramidal neurons in the ACC and is associated with chronic inflammatory pain-related anxiety.

Summary: Opiates produce a strong rewarding effect, but abstinence from opiate use emerges with severe negative emotions. Depression is one of the most frequent emotion disorders associated with opiate abstinence, which is thought to be a main cause for relapse. However, neurobiological bases of such an aversive emotion processing are poorly understood. Here, we find that morphine abstinence activates κ-opioid receptors (KORs) by increasing endogenous KOR ligand dynorphin expression in the amygdala, which in turn facilitates glutamate transporter 1 (GLT1) expression by activation of p38 mitogen-activated protein kinase (MAPK). Upregulation of GLT1 expression contributes to opiate-abstinence-elicited depressive-like behaviors through modulating amygdalar glutamatergic inputs to the nucleus accumbens (NAc). Intra-amygdala injection of GLT1 inhibitor DHK or knockdown of GLT1 expression in the amygdala significantly suppresses morphine-abstinence-induced depressive-like behaviors. Pharmacological and pharmacogenetic activation of amygdala-NAc projections prevents morphine-abstinence-induced behaviors. Overall, our study provides key molecular and circuit insights into the mechanisms of depression associated with opiate abstinence.

Abstract Background The effect of electroacupuncture (EA) is affected by both the acupuncture point selection and the frequency of stimulation. However, little is known regarding acupuncture point and simulation frequency selection. Neuronal activation of the nucleus of the solitary tract (NTS) is one of the important targets of EA for modulating gastrointestinal function. This study investigated the effects of various combinations of EA frequencies and acupuncture points on NTS neurons. Methods Rats were randomly divided into normal, 2 Hz EA, 100 Hz EA and the alternate 2/100 Hz EA groups. Then rats in each group were randomly divided into the following two subgroups according to the acupuncture point: ST 36 group and ST 25 group. All the rats underwent electrode implantation surgery. Rats in all EA groups received one treatment with EA (a constant square wave at, 2 Hz,100 Hz or 2/100 Hz frequencies with intensities ranging from 1 to 2 mA), and NTS neuronal activation was recorded before and after EA treatment. Finally, to confirm the effect of EA on the NTS, minimal acupuncture was administered and its effect on NTS was detected. Results ST 36 stimulated with 2 Hz EA significantly increased the population of excited NTS neurons and spike frequency. However, ST 36 stimulated with 100 Hz or 2/100 Hz EA produced only a transient effect on the activity of NTS neurons and did not induce any effect on the spike frequency. Furthermore, the excitatory effect of 100 Hz or 2/100 Hz EA on NTS neurons in the ST 36 group was lower than 2 Hz EA at the same point. When applied to ST 25, 2 Hz EA had no significant excitatory effect on NTS neurons or spike frequency. However, 100 Hz EA or 2/100 Hz EA at ST 25 decreased both NTS neuronal excitability and spike frequency. By comparing the effects of different EA combinations, it was shown 2 Hz EA applied to ST 36 had the strongest excitatory effect on NTS neurons, while 100 Hz EA applied to ST 25 had the greatest inhibitory effect. Minimal acupuncture stimulation produced no effect on NTS neurons. Conclusion EA’s effects on NTS were mainly affected by the acupuncture point selection, but the frequency of EA also played a role. Different combinations of acupuncture points and frequency selection may lead to different EA effects on NTS neuronal excitability.

Hypothesis: Electroacupuncture (EA) has been used as an alternative analgesic therapy for hundreds of years, yet its analgesic potency and therapeutic advantage against bone cancer pain (BCP) in comparison with morphine remains unclear. This study aimed to investigate the effects of EA on mechanical allodynia and cellular immunity of BCP rats, and to further explore the potential mechanism. Methods: The BCP model was established by implanting Walker 256 mammary gland carcinoma cells into the left tibia of adult female Sprague-Dawley rats. EA (dilatational wave, 2/100 Hz, 0.5 mA–1mA–1.5 mA for 10 minutes each intensity) was applied bilaterally to Zusanli (ST 36) and Kunlun (BL 60) for 30 minutes. Both EA stimulation and morphine (10 mg/kg, intraperitoneally) was given once every other day. Naloxone (0.3 mg/kg, intraperitoneally) was injected at 30 minutes prior to EA. Mechanical allodynia were demonstrated by paw withdrawal thresholds (PWTs) which measured by dynamic plantar aesthesiometer. T cell proliferation, percentage of CD3 + , CD4 + and CD8 + T lymphocytes in spleen as well as expression of interleukin-2 (IL-2) in plasma were detected by WST-8, flow cytometry, and enzyme-linked immunosorbent assay technique, respectively. Results: An intratibial inoculation of Walker 256 mammary gland carcinoma cells significantly decreased PWTs to mechanical stimuli. EA stimulation alleviated mechanical allodynia in BCP rats, and the analgesic potency of EA was weaker than that of morphine. In contrast to morphine, EA stimulation of BCP rats increased splenic concanavalin A (Con A)-induced T cell proliferation and plasma IL-2 content, as well as increased the percentages of splenic CD 3 + CD 4 + and CD 3 + CD 8 + T cell subsets. Moreover, both the analgesic effect and the partial immunomodulation of EA were suppressed by an intraperitoneal injection of naloxone. Conclusion: EA could significantly alleviate BCP-induced mechanical allodynia. Although the analgesic effect of EA was weaker than that of morphine, EA had an immunomodulation effect on cellular immunity. Both analgesic and immunomodulatory effect of EA might share the same mechanism via the opioid-mediated pathway, which needs further investigation.

Pain memory is considered as endopathic factor underlying stubborn chronic pain. Our previous study demonstrated that electroacupuncture (EA) can alleviate retrieval of pain memory. This study was designed to observe the different effects between EA and indomethacin (a kind of nonsteroid anti-inflammatory drugs, NSAIDs) in a rat pain memory model. To explore the critical role of protein kinase A (PKA) in pain memory, a PKA inhibitor was microinjected into anterior cingulate cortex (ACC) in model rats. We further investigated the roles of the cyclic adenosine monophosphate (cAMP), PKA, cAMP response element-binding protein (CREB), and cAMP/PKA/CREB pathway in pain memory to explore the potential molecular mechanism. The results showed that EA alleviates the retrieval of pain memory while indomethacin failed. Intra-ACC microinjection of a PKA inhibitor blocked the occurrence of pain memory. EA reduced the activation of cAMP, PKA, and CREB and the coexpression levels of cAMP/PKA and PKA/CREB in the ACC of pain memory model rats, but indomethacin failed. The present findings identified a critical role of PKA in ACC in retrieval of pain memory. We propose that the proper mechanism of EA on pain memory is possibly due to the partial inhibition of cAMP/PKA/CREB signaling pathway by EA.

The influence of electroacupuncture (EA), a traditional Chinese medical treatment, on type Ⅱ collagen-induced arthritis (CIA) was examined in DBA/1J mice in vivo. Mice were immunized intradermally twice at the 3-week interval with bovine type Ⅱ collagen(C Ⅱ). EA stimulation, begun on the 21 simultaneously with the second immunization, was applied at the acupoint equivalent to GV4 three times a week for 3 weeks. The results showed that EA delayed the onset, attenuated the severity of arthritis, and reduced the anti-collagen antibody level. Furthermore, we investigated the impact of EA on the productions of endogenous (IL-1 beta) and prostaglandin E2 (PGE2), and the levels of IL-1 beta mRNA in splenocytes and synovial tissues from C Ⅱ immunized mice on the 45 and cyclooxygenase-2 (COX-2) mRNA in lipopolysaccharide (LPS)-stimulated macrophages of normal mice by using reverse transcriptase-polymerase chain reaction (RT-PCR). EA stimulation significant inhibited the concentrations of splenic endogenous IL-1 beta and serum PGE2. The expression of IL-1 beta mRNA in spleen cells was obviously down-regulated and that in synovial tissues was modestly affected by EA. COX-2 mRNA was highly expressed in cultured peritoneal macrophages when stimulated with LPS. Previous treatment with EA also reduced LPS-stimulated induction of COX-2 mRNA. These data suggest that EA has an inhibitory effect on murine CIA, and the partial mechanism of its therapeutic result may be attributed to inhibiting the productions of IL-1 beta and PGE2 by suppression the IL-1 beta and COX-2 gene activations.