Cortical tagged synaptic long-term depression in the anterior cingulate cortex of adult mice.

The anterior cingulate cortex (ACC) is a key cortical region for pain perception and emotion. Different forms of synaptic plasticity, including long-term potentiation (LTP) and long-term depression (LTD), have been reported in the ACC. Synaptic tagging of LTP plays an important role in hippocampus-related associative memory. In this study, we demonstrate that synaptic tagging of LTD is detected in the ACC of adult male and female mice. This form of tagged LTD requires the activation of metabotropic glutamate receptor subtype 1 (mGluR1). The induction of tagged LTD is time-related with the strongest tagged LTD appearing when the interval between two independent stimuli is 30 min. Inhibitors of mGluR1 blocked the induction of tagged LTD, however, blocking N-methyl-d-aspartate (NMDA) receptors did not affect the induction of tagged LTD. Nimodipine, an inhibitor of L-type voltage-gated calcium channels (VGCCs), also blocked tagged LTD. In an animal model of amputation, we found that tagged LTD was either reduced or completely blocked. Together with our previous report of tagged LTP in the ACC, this study strongly suggests that excitatory synapses in the adult ACC are highly plastic. The biphasic tagging of synaptic transmission provides a new form of heterosynaptic plasticity in the ACC which has functional and pathophysiological significance in phantom pain.Significance Statement The anterior cingulate cortex (ACC) is a key cortical region for pain perception and chronic pain. Previous studies have reported a novel form of long-term heterosynaptic potentiation in the ACC. In this study, we discovered a long-term depression (LTD) form of synaptic tagging in the ACC of adult male and female mice. This form of tagged LTD requires the activation of metabotropic glutamate receptors (mGluRs). In an animal model of amputation, tagged LTD is reduced or completely blocked. Our results strongly suggest that brain cortices of adult mice are highly plastic and show biphasic tagging of plasticity. These findings of tagged LTD may provide a new direction for future treatment of phantom pain and amputation-related emotional disorders.

Radiogenomic analysis for predicting lymph node metastasis and molecular annotation of radiomic features in pancreatic cancer.

BACKGROUND: To provide a preoperative prediction model for lymph node metastasis in pancreatic cancer patients and provide molecular information of key radiomic features. METHODS: Two cohorts comprising 151 and 54 pancreatic cancer patients were included in the analysis. Radiomic features from the tumor region of interests were extracted by using PyRadiomics software. We used a framework that incorporated 10 machine learning algorithms and generated 77 combinations to construct radiomics-based models for lymph node metastasis prediction. Weighted gene coexpression network analysis (WGCNA) was subsequently performed to determine the relationships between gene expression levels and radiomic features. Molecular pathways enrichment analysis was performed to uncover the underlying molecular features. RESULTS: Patients in the in-house cohort (mean age, 61.3 years ± 9.6 [SD]; 91 men [60%]) were separated into training (n = 105, 70%) and validation (n = 46, 30%) cohorts. A total of 1,239 features were extracted and subjected to machine learning algorithms. The 77 radiomic models showed moderate performance for predicting lymph node metastasis, and the combination of the StepGBM and Enet algorithms had the best performance in the training (AUC = 0.84, 95% CI = 0.77-0.91) and validation (AUC = 0.85, 95% CI = 0.73-0.98) cohorts. We determined that 15 features were core variables for lymph node metastasis. Proliferation-related processes may respond to the main molecular alterations underlying these features. CONCLUSIONS: Machine learning-based radiomics could predict the status of lymph node metastasis in pancreatic cancer, which is associated with proliferation-related alterations.

Inhibition of calcium-stimulated adenylyl cyclase subtype 1 (AC1) for the treatment of pain and anxiety symptoms in Parkinson's disease mice model.

Pain and anxiety are two common and undertreated non-motor symptoms in Parkinson's disease (PD), which affect the life quality of PD patients, and the underlying mechanisms remain unclear. As an important subtype of adenylyl cyclases (ACs), adenylyl cyclase subtype 1 (AC1) is critical for the induction of cortical long-term potentiation (LTP) and injury induced synaptic potentiation in the cortical areas including anterior cingulate cortex (ACC) and insular cortex (IC). Genetic deletion of AC1 or pharmacological inhibition of AC1 improved chronic pain and anxiety in different animal models. In this study, we proved the motor deficit, pain and anxiety symptoms of PD in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice model. As a lead candidate AC1 inhibitor, oral administration (1 dose and seven doses) of NB001 (20 and 40 mg/kg) showed significant analgesic effect in MPTP-treated mice, and the anxiety behavior was also reduced (40 mg/kg). By using genetic knockout mice, we found that AC1 knockout mice showed reduced pain and anxiety symptoms after MPTP administration, but not AC8 knockout mice. In summary, genetic deletion of AC1 or pharmacological inhibition of AC1 improved pain and anxiety symptoms in PD model mice, but didn't affect motor function. These results suggest that NB001 is a potential drug for the treatment of pain and anxiety symptoms in PD patients by inhibiting AC1 target.

Inhibition of cortical synaptic transmission, behavioral nociceptive, and anxiodepressive-like responses by arecoline in adult mice.

Areca nut, the seed of Areca catechu L., is one of the most widely consumed addictive substances in the world after nicotine, ethanol, and caffeine. The major effective constituent of A. catechu, arecoline, has been reported to affect the central nervous system. Less is known if it may affect pain and its related emotional responses. In this study, we found that oral application of arecoline alleviated the inflammatory pain and its induced anxiolytic and anti-depressive-like behavior. Arecoline also increased the mechanical nociceptive threshold and alleviated depression-like behavior in naïve mice. In the anterior cingulate cortex (ACC), which acts as a hinge of nociception and its related anxiety and depression, by using the multi-electrode field potential recording and whole-cell patch-clamp recording, we found that the evoked postsynaptic transmission in the ACC of adult mice has been inhibited by the application of arecoline. The muscarinic receptor is the major receptor of the arecoline in the ACC. Our results suggest that arecoline alleviates pain, anxiety, and depression-like behavior in both physiological and pathological conditions, and this new mechanism may help to treat patients with chronic pain and its related anxiety and disorder in the future.

Enhanced long-term potentiation in the anterior cingulate cortex of tree shrew.

Synaptic plasticity is a key cellular model for learning, memory and chronic pain. Most previous studies were carried out in rats and mice, and less is known about synaptic plasticity in non-human primates. In the present study, we used integrative experimental approaches to study long-term potentiation (LTP) in the anterior cingulate cortex (ACC) of adult tree shrews. We found that glutamate is the major excitatory transmitter and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionicacid (AMPA) receptors mediate postsynaptic responses. LTP in tree shrews was greater than that in adult mice and lasted for at least 5 h. N-methyl-d-aspartic acid (NMDA) receptors, Ca2+ influx and adenylyl cyclase 1 (AC1) contributed to tree shrew LTP. Our results suggest that LTP is a major form of synaptic plasticity in the ACC of primate-like animals. This article is part of a discussion meeting issue 'Long-term potentiation: 50 years on'.

Cortical nitric oxide required for presynaptic long-term potentiation in the insular cortex.

Nitric oxide (NO) is a key diffusible messenger in the mammalian brain. It has been proposed that NO may diffuse retrogradely into presynaptic terminals, contributing to the induction of hippocampal long-term potentiation (LTP). Here, we present novel evidence that NO is required for kainate receptor (KAR)-dependent presynaptic form of LTP (pre-LTP) in the adult insular cortex (IC). In the IC, we found that inhibition of NO synthase erased the maintenance of pre-LTP, while the induction of pre-LTP required the activation of KAR. Furthermore, NO is essential for pre-LTP induced between two pyramidal cells in the IC using the double patch-clamp recording. These results suggest that NO is required for homosynaptic pre-LTP in the IC. Our results present strong evidence for the critical roles of NO in pre-LTP in the IC. This article is part of a discussion meeting issue 'Long-term potentiation: 50 years on'.

Recruitment of cortical silent responses by forskolin in the anterior cingulate cortex of adult mice.

Recent studies using different experimental approaches demonstrate that silent synapses may exist in the adult cortex including the sensory cortex and anterior cingulate cortex (ACC). The postsynaptic form of long-term potentiation (LTP) in the ACC recruits some of these silent synapses and the activity of calcium-stimulated adenylyl cyclases (ACs) is required for such recruitment. It is unknown if the chemical activation of ACs may recruit silent synapses. In this study, we found that activation of ACs contributed to synaptic potentiation in the ACC of adult mice. Forskolin, a selective activator of ACs, recruited silent responses in the ACC of adult mice. The recruitment was long-lasting. Interestingly, the effect of forskolin was not universal, some silent synapses did not undergo potentiation or recruitment. These findings suggest that these adult cortical synapses are not homogenous. The application of a selective calcium-permeable AMPA receptor inhibitor 1-naphthyl acetyl spermine (NASPM) reversed the potentiation and the recruitment of silent responses, indicating that the AMPA receptor is required. Our results strongly suggest that the AC-dependent postsynaptic AMPA receptor contributes to the recruitment of silent responses at cortical LTP.

The Pathway-Selective Dependence of Nitric Oxide for Long-Term Potentiation in the Anterior Cingulate Cortex of Adult Mice.

Nitric oxide (NO) is a key diffusible messenger in the mammalian brain. It has been proposed that NO may diffuse in retrograde into presynaptic terminals, contributing to the induction of hippocampal long-term potentiation (LTP). Here, we present novel evidence that NO is selectively required for the synaptic potentiation of the interhemispheric projection in the anterior cingulate cortex (ACC). Unilateral low-frequency stimulation (LFS) induced a short-term synaptic potentiation on the contralateral ACC through the corpus callosum (CC). The use of the antagonists of the NMDA receptor (NMDAR), or the inhibitor of the L-type voltage-dependent Ca2+ channels (L-VDCCs), blocked the induction of this ACC-ACC potentiation. In addition, the inhibitor of NO synthase, or inhibitors for its downstream signaling pathway, also blocked this ACC-ACC potentiation. However, the application of the NOS inhibitor blocked neither the local electric stimulation-induced LTP nor the stimulation-induced recruitment of silent responses. Our results present strong evidence for the pathway-selective roles of NO in the LTP of the ACC.

Alleviation of migraine related pain and anxiety by inhibiting calcium-stimulating AC1-dependent CGRP in the insula of adult rats.

BACKGROUND: Recent animal and clinical findings consistently highlight the critical role of calcitonin gene-related peptide (CGRP) in chronic migraine (CM) and related emotional responses. CGRP antibodies and receptor antagonists have been approved for CM treatment. However, the underlying CGRP-related signaling pathways in the pain-related cortex remain poorly understood. METHODS: The SD rats were used to establish the CM model by dural infusions of inflammatory soup. Periorbital mechanical thresholds were assessed using von-Frey filaments, and anxiety-like behaviors were observed via open field and elevated plus maze tests. Expression of c-Fos, CGRP and NMDA GluN2B receptors was detected using immunofluorescence and western blotting analyses. The excitatory synaptic transmission was detected by whole-cell patch-clamp recording. A human-used adenylate cyclase 1 (AC1) inhibitor, hNB001, was applied via insula stereotaxic and intraperitoneal injections in CM rats. RESULTS: The insular cortex (IC) was activated in the migraine model rats. Glutamate-mediated excitatory transmission and NMDA GluN2B receptors in the IC were potentiated. CGRP levels in the IC significantly increased during nociceptive and anxiety-like activities. Locally applied hNB001 in the IC or intraperitoneally alleviated periorbital mechanical thresholds and anxiety behaviors in migraine rats. Furthermore, CGRP expression in the IC decreased after the hNB001 application. CONCLUSIONS: Our study indicated that AC1-dependent IC plasticity contributes to migraine and AC1 may be a promising target for treating migraine in the future.

Anterior cingulate cortex and insomnia: A cingulate-striatum connection.

Insomnia is an important comorbidity of chronic pain. In this issue of Neuron, Li et al. report that chronic-pain-induced insomnia is mediated by the pyramidal neurons in the anterior cingulate cortex and their dopaminergic projections to the dorsal medial striatum.

Fear memory regulation by the cAMP signaling pathway as an index of reexperiencing symptoms in posttraumatic stress disorder.

Posttraumatic stress disorder (PTSD) is a psychiatric disorder associated with traumatic memory, yet its etiology remains unclear. Reexperiencing symptoms are specific to PTSD compared to other anxiety-related disorders. Importantly, reexperiencing can be mimicked by retrieval-related events of fear memory in animal models of traumatic memory. Recent studies revealed candidate PTSD-associated genes that were related to the cyclic adenosine monophosphate (cAMP) signaling pathway. Here, we demonstrate the tight linkage between facilitated cAMP signaling and PTSD by analyzing loss- and gain-of-cAMP signaling effects on fear memory in mice and the transcriptomes of fear memory-activated mice and female PTSD patients with reexperiencing symptoms. Pharmacological and optogenetic upregulation or downregulation of cAMP signaling transduction enhanced or impaired, respectively, the retrieval and subsequent maintenance of fear memory in mice. In line with these observations, integrative mouse and human transcriptome analysis revealed the reduced mRNA expression of phosphodiesterase 4B (PDE4B), an enzyme that degrades cAMP, in the peripheral blood of PTSD patients showing more severe reexperiencing symptoms and the mouse hippocampus after fear memory retrieval. Importantly, more severe reexperiencing symptoms and lower PDE4B mRNA levels were correlated with decreased DNA methylation of a locus within PDE4B, suggesting the involvement of methylation in the mechanism of PTSD. These findings raise the possibility that the facilitation of cAMP signaling mediating the downregulation of PDE4B expression enhances traumatic memory, thereby playing a key role in the reexperiencing symptoms of PTSD patients as a functional index of these symptoms.

Selective enhancement of fear extinction by inhibiting neuronal adenylyl cyclase 1 (AC1) in aged mice.

Adenylyl cyclase 1 (AC1) is a selective subtype of ACs, which is selectively expressed in neurons. The activation of AC1 is activity-dependent, and AC1 plays an important role in cortical excitation that contributes to chronic pain and related emotional disorders. Previous studies have reported that human-used NB001 (hNB001, a selective AC1 inhibitor) produced analgesic effects in different animal models of chronic pain. However, the potential effects of hNB001 on learning and memory have been less investigated. In the present study, we found that hNB001 affected neither the induction nor the expression of trace fear, but selectively enhanced the relearning ability during the extinction in aged mice. By contrast, the same application of hNB001 did not affect recent, remote auditory fear memory, or remote fear extinction in either adult or aged mice. Furthermore, a single or consecutive 30-day oral administration of hNB001 did not affect acute nociceptive response, motor function, or anxiety-like behavior in either adult or aged mice. Our results are consistent with previous findings that inhibition of AC1 did not affect general sensory, emotional, and motor functions in adult mice, and provide strong evidence that inhibiting the activity of AC1 may be beneficial for certain forms of learning and memory in aged mice.

Long-term plasticity of NMDA GluN2B (NR2B) receptor in anterior cingulate cortical synapses.

The anterior cingulate cortex (ACC) is a key cortical area for pain perception, emotional fear and anxiety. Cortical excitation is thought to be the major mechanism for chronic pain and its related emotional disorders such as anxiety and depression. GluN2B (or called NR2B) containing NMDA receptors play critical roles for such excitation. Not only does the activation of GluN2B contributes to the induction of the postsynaptic form of LTP (post-LTP), long-term upregulation of GluN2B subunits through tyrosine phosphorylation were also detected after peripheral injury. In addition, it has been reported that presynaptic NMDA receptors may contribute to the modulation of the release of glutamate from presynaptic terminals in the ACC. It is believed that inhibiting subtypes of NMDA receptors and/or downstream signaling proteins may serve as a novel therapeutic mechanism for future treatment of chronic pain, anxiety, and depression.

Cortical synaptic basis of consciousness.

Consciousness is one of final questions for humans to tackle in neuroscience. Due to a lack of understanding of basic brain networks and mechanisms of functions, our knowledge of consciousness mainly stays at a theoretical level. Recent studies using brain imaging in humans and modern neuroscience techniques in animal studies reveal the basic brain network for consciousness. The projection from the thalamus to different cortical regions forms a network of activities to maintain consciousness in humans and animals. These feedback and feedforward circuits maintain consciousness even in certain brain injury conditions. Pterions and ion channels that contribute to these circuit neural activities are targets for drugs and manipulations that affect consciousness such as anesthetic agents. Synaptic plasticity that trains synapses during learning and information recall modified the circuits and contributes to a high level of consciousness in a certain population.

Single-cell RNA sequencing uncovers the cell type-dependent transcriptomic changes in the retrosplenial cortex after peripheral nerve injury.

The retrosplenial cortex (RSC) is a vital area for storing remote memory and has recently been found to undergo broad changes after peripheral nerve injury. However, little is known about the role of RSC in pain regulation. Here, we examine the involvement of RSC in the pain of mice with nerve injury. Notably, reducing the activities of calcium-/calmodulin-dependent protein kinase type II-positive splenial neurons chemogenetically increases paw withdrawal threshold and extends thermal withdrawal latency in mice with nerve injury. The single-cell or single-nucleus RNA-sequencing results predict enhanced excitatory synaptic transmissions in RSC induced by nerve injury. Local infusion of 1-naphthyl acetyl spermine into RSC to decrease the excitatory synaptic transmissions relieves pain and induces conditioned place preference. Our data indicate that RSC is critical for regulating physiological and neuropathic pain. The cell type-dependent transcriptomic information would help understand the molecular basis of neuropathic pain.

Activation of the glutamatergic cingulate cortical-cortical connection facilitates pain in adult mice.

The brain consists of the left and right cerebral hemispheres and both are connected by callosal projections. Less is known about the basic mechanism of this cortical-cortical connection and its functional importance. Here we investigate the cortical-cortical connection between the bilateral anterior cingulate cortex (ACC) by using the classic electrophysiological and optogenetic approach. We find that there is a direct synaptic projection from one side ACC to the contralateral ACC. Glutamate is the major excitatory transmitter for bilateral ACC connection, including projections to pyramidal cells in superficial (II/III) and deep (V/VI) layers of the ACC. Both AMPA and kainate receptors contribute to synaptic transmission. Repetitive stimulation of the projection also evoked postsynaptic Ca2+ influx in contralateral ACC pyramidal neurons. Behaviorally, light activation of the ACC-ACC connection facilitated behavioral withdrawal responses to mechanical stimuli and noxious heat. In an animal model of neuropathic pain, light inhibitory of ACC-ACC connection reduces both primary and secondary hyperalgesia. Our findings provide strong direct evidence for the excitatory or facilitatory contribution of ACC-ACC connection to pain perception, and this mechanism may provide therapeutic targets for future treatment of chronic pain and related emotional disorders.

A PRoliferation-Inducing Ligand (APRIL) in the Pathogenesis of Immunoglobulin A Nephropathy: A Review of the Evidence.

A PRoliferation-Inducing Ligand (APRIL), the thirteenth member of the tumor necrosis factor superfamily, plays a key role in the regulation of activated B cells, the survival of long-lived plasma cells, and immunoglobulin (Ig) isotype class switching. Several lines of evidence have implicated APRIL in the pathogenesis of IgA nephropathy (IgAN). Globally, IgAN is the most common primary glomerulonephritis, and it can progress to end-stage kidney disease; yet, disease-modifying treatments for this condition have historically been lacking. The preliminary data in ongoing clinical trials indicate that APRIL inhibition can reduce proteinuria and slow the rate of kidney disease progression by acting at an upstream level in IgAN pathogenesis. In this review, we examine what is known about the physiologic roles of APRIL and evaluate the experimental and epidemiological evidence describing how these normal biologic processes are thought to be subverted in IgAN. The weight of the preclinical, clinical, and genetic data supporting a key role for APRIL in IgAN has galvanized pharmacologic research, and several anti-APRIL drug candidates have now entered clinical development for IgAN. Herein, we present an overview of the clinical results to date. Finally, we explore where more research and evidence are needed to transform potential therapies into clinical benefits for patients with IgAN.

Sound-induced analgesia cannot always be observed in adult mice.

Music seems promising as an adjuvant pain treatment in humans, while its mechanism remains to be illustrated. In rodent models of chronic pain, few studies reported the analgesic effect of music. Recently, Zhou et al. stated that the analgesic effects of sound depended on a low (5 dB) signal-to-noise ratio (SNR) relative to ambient noise in mice. However, despite employing multiple behavioral analysis approaches, we were unable to extend these findings to a mice model of chronic pain listening to the 5 dB SNR.

Silent synapses in pain-related anterior cingulate cortex.

Synaptic plasticity such as Long-term potentiation (LTP) is a key mechanism for learning in central synapses including the cortex. There are two least two major forms of LTPs: presynaptic LTP and postsynaptic LTP. For postsynaptic LTP, the potentiation of AMPA receptor-mediated responses through protein phosphorylation is thought to be a key mechanism. Silent synapses have been reported in the hippocampus, but it is thought to be mainly present in the cortex during early development, and may contribute to maturation of the cortical circuit. However, recent several lines of evidence demonstrate that silent synapses may exist in mature synapses of adult cortex, and they can be recruited by LTP-inducing protocols, as well as chemical-induced LTP. In pain-related cortical regions, silent synapses may not only contribute to cortical excitation after peripheral injury, but also the recruitment of new cortical circuits as well. Thus, it is proposed that silent synapses and modification of functional AMPA receptors and NMDA receptors may play important roles in chronic pain, including phantom pain.

Inhibiting neuronal AC1 for treating anxiety and headache in the animal model of migraine.

Migraines are a common medical condition. From a basic science point of view, the central mechanism for migraine and headache is largely unknown. In the present study, we demonstrate that cortical excitatory transmission is significantly enhanced in the anterior cingulate cortex (ACC)-a brain region which is critical for pain perception. Biochemical studies found that the phosphorylation levels of both the NMDA receptor GluN2B and AMPA receptor GluA1 were enhanced in ACC of migraine rats. Both the presynaptic release of glutamate and postsynaptic responses of AMPA receptors and NMDA receptors were enhanced. Synaptic long-term potentiation (LTP) was occluded. Furthermore, behavioral anxiety and nociceptive responses were increased, which were reversed by application of AC1 inhibitor NB001 within ACC. Our results provide strong evidence that cortical LTPs contribute to migraine-related pain and anxiety. Drugs that inhibit cortical excitation such as NB001 may serve as potential medicines for treating migraine in the future.