Activation of glycine site and GluN2B subunit of NMDA receptors is necessary for ERK/CREB signaling cascade in rostral anterior cingulate cortex in rats: Implications for affective pain

Objective The rostral anterior cingulate cortex(rACC)is implicated in processing the emotional component of pain.N-methyl-D-aspartate receptors(NMDARs)are highly expressed in the rACC and mediate painrelated affect by activating a signaling pathway that involves cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA)and/or extracellular regulated kinase(ERK)/cAMP-response element-binding protein(CREB).The present study investigated the contributions of the NMDAR glycine site and GluN2B subunit to the activation of ERK and CREB both in vitro and in vivo in rat rACC.Methods Immunohistochemistry and Western blot analysis were used to separately assess the expression of phospho-ERK(pERK)and phospho-CREB(pCREB)in vitro and in vivo.Double immunostaining was also used to determine the colocalization of pERK and pCREB.Results Both bath application of NMDA in brain slices in vitro and intraplantar injection of formalin into the rat hindpaw in vivo induced significant up-regulation of pERK and pCREB in the rACC,which was inhibited by the NMDAR antagonist DL-2-amino-5-phospho-novaleric acid.Selective blockade of the NMDAR GluN2B subunit and the glycinebinding site,or degradation of endogenous D-serine,a co-agonist for the glycine site,significantly decreased the up-regulation of pERK and pCREB expression in the rACC.Further,the activated ERK predominantly colocalized with CREB.Conclusion Either the glycine site or the GluN2B subunit of NMDARs participates in the phosphorylation of ERK and CREB induced by bath application of NMDA in brain slices or hindpaw injection of 5% formalin in rats,and these might be fundamental molecular mechanisms underlying pain affect.     

The role of the subgenual anterior cingulate cortex in dorsomedial prefrontal–amygdala neural circuitry during positive‐social emotion regulation

Positive‐social emotions mediate one's cognitive performance, mood, well‐being, and social bonds, and represent a critical variable within therapeutic settings. It has been shown that the upregulation of positive emotions in social situations is associated with increased top‐down signals that stem from the prefrontal cortices (PFC) which modulate bottom‐up emotional responses in the amygdala. However, it remains unclear if positive‐social emotion upregulation of the amygdala occurs directly through the dorsomedial PFC (dmPFC) or indirectly linking the bilateral amygdala with the dmPFC via the subgenual anterior cingulate cortex (sgACC), an area which typically serves as a gatekeeper between cognitive and emotion networks. We performed functional MRI (fMRI) experiments with and without effortful positive‐social emotion upregulation to demonstrate the functional architecture of a network involving the amygdala, the dmPFC, and the sgACC. We found that effortful positive‐social emotion upregulation was associated with an increase in top‐down connectivity from the dmPFC on the amygdala via both direct and indirect connections with the sgACC. Conversely, we found that emotion processes without effortful regulation increased network modulation by the sgACC and amygdala. We also found that more anxious individuals with a greater tendency to suppress emotions and intrusive thoughts, were likely to display decreased amygdala, dmPFC, and sgACC activity and stronger connectivity strength from the sgACC onto the left amygdala during effortful emotion upregulation. Analyzed brain network suggests a more general role of the sgACC in cognitive control and sheds light on neurobiological informed treatment interventions.     

The primary somatosensory cortex and the insula contribute differently to the processing of transient and sustained nociceptive and non‐nociceptive somatosensory inputs

Transient nociceptive stimuli elicit consistent brain responses in the primary and secondary somatosensory cortices (S1, S2), the insula and the anterior and mid‐cingulate cortex (ACC/MCC). However, the functional significance of these responses, especially their relationship with sustained pain perception, remains largely unknown. Here, using functional magnetic resonance imaging, we characterize the differential involvement of these brain regions in the processing of sustained nociceptive and non‐nociceptive somatosensory input. By comparing the spatial patterns of activity elicited by transient (0.5 ms) and long‐lasting (15 and 30 s) stimuli selectively activating nociceptive or non‐nociceptive afferents, we found that the contralateral S1 responded more strongly to the onset of non‐nociceptive stimulation as compared to the onset of nociceptive stimulation and the sustained phases of nociceptive and non‐nociceptive stimulation. Similarly, the anterior insula responded more strongly to the onset of nociceptive stimulation as compared to the onset of non‐nociceptive stimulation and the sustained phases of nociceptive and non‐nociceptive stimulation. This suggests that S1 is specifically sensitive to changes in incoming non‐nociceptive input, whereas the anterior insula is specifically sensitive to changes in incoming nociceptive input. Second, we found that the MCC responded more strongly to the onsets as compared to the sustained phases of both nociceptive and non‐nociceptive stimulation, suggesting that it could be involved in the detection of change regardless of sensory modality. Finally, the posterior insula and S2 responded maximally during the sustained phase of non‐nociceptive stimulation but not nociceptive stimulation, suggesting that these regions are preferentially involved in processing non‐nociceptive somatosensory input. Hum Brain Mapp 36:4346–4360, 2015. © 2015 Wiley Periodicals, Inc.     

The neural basis of the abnormal self‐referential processing and its impact on cognitive control in depressed patients

Persistent pondering over negative self‐related thoughts is a central feature of depressive psychopathology. In this study, we sought to investigate the neural correlates of abnormal negative self‐referential processing (SRP) in patients with Major Depressive Disorder and its impact on subsequent cognitive control‐related neuronal activation. We hypothesized aberrant activation dynamics during the period of negative and neutral SRP in the rostral anterior cingulate cortex (rACC) and in the amygdala in patients with major depressive disorder. Additionally, we assumed abnormal activation in the fronto‐cingulate network during Stroop task execution. 19 depressed patients and 20 healthy controls participated in the study. Using an event‐related functional magnetic resonance imaging (fMRI) design, negative, positive and neutral self‐referential statements were displayed for 6.5 s and followed by incongruent or congruent Stroop conditions. The data were analyzed with SPM8. In contrast to controls, patients exhibited no significant valence‐dependent rACC activation differences during SRP. A novel finding was the significant activation of the amygdala and the reward‐processing network during presentation of neutral self‐referential stimuli relative to baseline and to affective stimuli in patients. The fMRI analysis of the Stroop task revealed a reduced BOLD activation in the right fronto‐parietal network of patients in the incongruent condition after negative SRP only. Thus, the inflexible activation in the rACC may correspond to the inability of depressed patients to shift their attention away from negative self‐related stimuli. The accompanying negative affect and task‐irrelevant emotional processing may compete for neuronal resources with cognitive control processes and lead thereby to deficient cognitive performance associated with decreased fronto‐parietal activation. Hum Brain Mapp 36:2781–2794, 2015. © 2015 Wiley Periodicals, Inc.     

Structural and functional connectivity of the human brain in autism spectrum disorders and attention‐deficit/hyperactivity disorder: A rich club‐organization study

Attention‐deficit/hyperactive disorder (ADHD) and autism spectrum disorders (ASD) are two of the most common and vexing neurodevelopmental disorders among children. Although the two disorders share many behavioral and neuropsychological characteristics, most MRI studies examine only one of the disorders at a time. Using graph theory combined with structural and functional connectivity, we examined the large‐scale network organization among three groups of children: a group with ADHD (8–12 years, n = 20), a group with ASD (7–13 years, n = 16), and typically developing controls (TD) (8–12 years, n = 20). We apply the concept of the rich‐club organization, whereby central, highly connected hub regions are also highly connected to themselves. We examine the brain into two different network domains: (1) inside a rich‐club network phenomena and (2) outside a rich‐club network phenomena. The ASD and ADHD groups had markedly different patterns of rich club and non rich‐club connections in both functional and structural data. The ASD group exhibited higher connectivity in structural and functional networks but only inside the rich‐club networks. These findings were replicated using the autism brain imaging data exchange dataset with ASD (n = 85) and TD (n = 101). The ADHD group exhibited a lower generalized fractional anisotropy and functional connectivity inside the rich‐club networks, but a higher number of axonal fibers and correlation coefficient values outside the rich club. Despite some shared biological features and frequent comorbity, these data suggest ADHD and ASD exhibit distinct large‐scale connectivity patterns in middle childhood. Hum Brain Mapp 35:6032–6048, 2014. © 2014 Wiley Periodicals, Inc.     

TrkB‐mediated activation of the phosphatidylinositol‐3‐kinase/Akt cascade reduces the damage inflicted by oxygen–glucose deprivation in area CA3 of the rat hippocampus

The selective vulnerability of hippocampal area CA1 to ischemia‐induced injury is a well‐known phenomenon. However, the cellular mechanisms that confer resistance to area CA3 against ischemic damage remain elusive. Here, we show that oxygen–glucose deprivation–reperfusion (OGD‐RP), an in vitro model that mimic the pathological conditions of the ischemic stroke, increases the phosphorylation level of tropomyosin receptor kinase B (TrkB) in area CA3. Slices preincubated with brain‐derived neurotrophic factor (BDNF) or 7,8‐dihydroxyflavone (7,8‐DHF) exhibited reduced depression of the electrical activity triggered by OGD‐RP. Consistently, blockade of TrkB suppressed the resistance of area CA3 to OGD‐RP. The protective effect of TrkB activation was limited to area CA3, as OGD‐RP caused permanent suppression of CA1 responses. At the cellular level, TrkB activation leads to phosphorylation of the accessory proteins SHC and Gab as well as the serine/threonine kinase Akt, members of the phosphoinositide 3‐kinase/Akt (PI‐3‐K/Akt) pathway, a cascade involved in cell survival. Hence, acute slices pretreated with the Akt antagonist MK2206 in combination with BDNF lost the capability to resist the damage inflicted with OGD‐RP. Consistently, with these results, CA3 pyramidal cells exhibited reduced propidium iodide uptake and caspase‐3 activity in slices pretreated with BDNF and exposed to OGD‐RP. We propose that PI‐3‐K/Akt downstream activation mediated by TrkB represents an endogenous mechanism responsible for the resistance of area CA3 to ischemic damage.     

A case of recurrent epilepsy‐associated rosette‐forming glioneuronal tumor with anaplastic transformation in the absence of therapy

Rosette‐forming glioneuronal tumor (RGNT) most commonly occurs adjacent to the fourth ventricle and therefore rarely presents with epilepsy. Recent reports describe RGNT occurrence in other anatomical locations with considerable morphologic and genetic overlap with the epilepsy‐associated dysembryoplastic neuroepithelial tumor (DNET). Examples of RGNT or DNET with anaplastic change are rare, and typically occur in the setting of radiation treatment. We present the case of a 5‐year‐old girl with seizures, who underwent near total resection of a cystic temporal lobe lesion. Pathology showed morphologic and immunohistochemical features of RGNT, albeit with focally overlapping DNET‐like patterns. Resections of residual or recurrent tumor were performed 1 year and 5 years after the initial resection, but no adjuvant radiation or chemotherapy was given. Ten years after the initial resection, surveillance imaging identified new and enhancing nodules, leading to another gross total resection. This specimen showed areas similar to the original tumor, but also high‐grade foci with oligodendroglial morphology, increased cellularity, palisading necrosis, microvascular proliferation, and up to 13 mitotic figures per 10 high power fields. Ancillary studies the status by sequencing showed wild‐type of the isocitrate dehydrogenase 1 (IDH1), IDH2, and human histone 3.3 (H3F3A) genes, and BRAF studies were negative for mutation or rearrangement. Fluorescence in situ hybridization (FISH) showed codeletion of 1p and 19q limited to the high‐grade regions. By immunohistochemistry there was loss of nuclear alpha‐thalassemia mental retardation syndrome, X‐linked (ATRX) expression only in the high‐grade region. Next‐generation sequencing showed an fibroblast growth factor receptor receptor 1 (FGFR1) kinase domain internal tandem duplication in three resection specimens. ATRX mutation in the high‐grade tumor was confirmed by sequencing which showed a frameshift mutation (p.R1427fs), while the apparent 1p/19q‐codeletion by FISH was due to loss of chromosome arm 1p and only partial loss of 19q. Exceptional features of this case include the temporal lobe location, 1p/19q loss by FISH without true whole‐arm codeletion, and anaplastic transformation associated with ATRX mutation without radiation or chemotherapy.     

Widespread changes in dendritic and axonal morphology in Mecp2‐mutant mouse models of rett syndrome: Evidence for disruption of neuronal networks

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X‐linked gene MECP2. Girls with RTT show dramatic changes in brain function, but relatively few studies have explored the structure of neural circuits. Examining two mouse models of RTT (Mecp2B and Mecp2J), we previously documented changes in brain anatomy. Herein, we use confocal microscopy to study the effects of MeCP2 deficiency on the morphology of dendrites and axons in the fascia dentata (FD), CA1 area of hippocampus, and motor cortex following Lucifer yellow microinjection or carbocyanine dye tracing. At 3 weeks of age, most (33 of 41) morphological parameters were significantly altered in Mecp2B mice; fewer (23 of 39) were abnormal in Mecp2J mice. There were striking changes in the density and size of the dendritic spines and density and orientation of axons. In Mecp2B mice, dendritic spine density was decreased in the FD (～11%), CA1 (14–22%), and motor cortex (～16%). A decreased spine head size (～9%) and an increased spine neck length (～12%) were found in Mecp2B FD. In addition, axons in the motor cortex were disorganized. In Mecp2J mice, spine density was significantly decreased in CA1 (14–26%). In both models, dendritic swelling and elongated spine necks were seen in all areas studied. Marked variation in the type and extent of changes was noted in dendrites of adjacent neurons. Electron microscopy confirmed abnormalities in dendrites and axons and showed abnormal mitochondria. Our findings document widespread abnormalities of dendrites and axons that recapitulate those seen in RTT. J. Comp. Neurol. 514:240–258, 2009. © 2009 Wiley‐Liss, Inc.     

Neurosteroid dehydroepiandrosterone enhances activity and trafficking of astrocytic GLT‐1 via σ1 receptor‐mediated PKC activation in the hippocampal dentate gyrus of rats

Neurosteroid dehydroepiandrosterone (DHEA) has been reported to exert a potent neuroprotective effect against glutamate‐induced excitotoxicity. However, the underlying mechanism remains to be elucidated. One of the possible mechanisms may be an involvement of astrocytic glutamate transporter subtype‐1 (GLT‐1) that can quickly clear spilled glutamate at the synapse to prevent excitotoxicity. To examine the effect of DHEA on GLT‐1 activity, we measured synaptically induced glial depolarization (SIGD) in the dentate gyrus (DG) of adult rats by applying an optical recording technique to the hippocampal slices stained with voltage‐sensitive dye RH155. Bath‐application of DHEA for 10 min dose‐dependently increased SIGD without changing presynaptic glutamate releases, which was sensitive to the GLT‐1 blocker DHK. Patch‐clamp recordings in astrocytes showed that an application of 50 μM DHEA increased glutamate‐evoked inward currents (Iglu) by approximately 1.5‐fold, which was dependent on the GLT‐1 activity. In addition, the level of biotinylated GLT‐1 protein in the surface of astrocytes was significantly elevated by DHEA. The DHEA‐increased SIGD, Iglu, and GLT‐1 translocation to the cell surface were blocked by the σ₁R antagonist NE100 and mimicked by the σ₁R agonist PRE084. DHEA elevated the phosphorylation level of PKC in a σ₁R‐dependent manner. Furthermore, the PKC inhibitor chelerythrine could prevent the DHEA‐increased SIGD, Iglu, and GLT‐1 translocation. Collectively, present results suggest that DHEA enhances the activity and translocation to cell surface of astrocytic GLT‐1 mainly via σ₁R‐mediated PKC cascade.     

Resting state functional connectivity of the anterior cingulate cortex in veterans with and without post‐traumatic stress disorder

Post‐traumatic stress disorder (PTSD) is an anxiety disorder that is associated with structural and functional alterations in several brain areas, including the anterior cingulate cortex (ACC). Here, we examine resting state functional connectivity of ACC subdivisions in PTSD, using a seed‐based approach. Resting state magnetic resonance images were obtained from male veterans with (n = 31) and without (n = 25) PTSD, and healthy male civilian controls (n = 25). Veterans with and without PTSD (combat controls) had reduced functional connectivity compared to healthy controls between the caudal ACC and the precentral gyrus, and between the perigenual ACC and the superior medial gyrus and middle temporal gyrus. Combat controls had increased connectivity between the rostral ACC and precentral/middle frontal gyrus compared to PTSD patients and healthy civilian controls. The resting state functional connectivity differences in the perigenual ACC network reported here indicate that veterans differ from healthy controls, potentially due to military training, deployment, and/or trauma exposure. In addition, specific alterations in the combat controls may potentially be related to resilience. These results underline the importance of distinguishing trauma‐exposed (combat) controls from healthy civilian controls when studying PTSD. Hum Brain Mapp, 36:99–109, 2015. © 2014 Wiley Periodicals, Inc.     

Protease‐activated receptor 4 activation increases the expression of calcitonin gene‐related peptide mRNA and protein in dorsal root ganglion neurons

Accumulating evidence demonstrates that nociceptor activation evokes a rapid change in mRNA and protein levels of calcitonin gene‐related peptide (CGRP) in dorsal root ganglion (DRG) neurons. Although the colocalization of CGRP and protease‐activated receptor‐4 (PAR4), a potent modulator of pain processing and inflammation, was detected in DRG neurons, the role of PAR4 activation in the expression of CGRP has not been investigated. In the present study, the expression of CGRP and activation (phosphorylation) of extracellular signal‐regulated kinases 1 and 2 (ERK1/2) in rat DRG neurons were measured by immunofluorescence, real‐time PCR, and Western blotting after AYPGKF‐NH2 (selective PAR4‐activating peptide; PAR4‐AP) intraplantar injection or treatment of cultured DRG neurons. The expression of CGRP in cultured DRG neurons was also assessed after treatment with AYPGKF‐NH2 with preaddition of PD98059 (an inhibitor for ERK1/2 pathway). Results showed that PAR4‐AP intraplantar injection or treatment of cultured DRG neurons evoked significant increases in DRG cells displaying CGRP immunoreactivity and cytoplasmic and nuclear staining for phospho‐ERK1/2 (p‐ERK1/2). Percentages of total DRG neurons expressing both CGRP and PAR4 or p‐ERK1/2 also increased significantly at 2 hr after PAR4‐AP treatment. Real‐time PCR and Western blotting showed that PAR4‐AP treatment significantly increased expression of CGRP mRNA and protein levels in DRG neurons. The PAR4 activation‐evoked CGRP expression both at mRNA and at protein levels was significantly inhibited after p‐ERK1/2 was inhibited by PD98059. These results provide evidence that activation of PAR4 upregulates the expression of CGRP mRNA and protein levels in DRG neurons via the p‐ERK1/2 signal pathway. © 2013 Wiley Periodicals, Inc.     

Sex‐specific mediation effect of the right fusiform face area volume on the association between variants in repeat length of AVPR1A RS3 and altruistic behavior in healthy adults

Microsatellite variants in the arginine vasopressin receptor 1A gene (AVPR1A) RS3 have been associated with normal social behaviors variation and autism spectrum disorders (ASDs) in a sex‐specific manner. However, neural mechanisms underlying these associations remain largely unknown. We hypothesized that AVPR1A RS3 variants affect altruistic behavior by modulating the gray matter volume (GMV) of specific brain regions in a sex‐specific manner. We investigated 278 young healthy adults using the Dictator Game to assess altruistic behavior. All subjects were genotyped and main effect of AVPR1A RS3 repeat polymorphisms and interaction of genotype‐by‐sex on the GMV were assessed in a voxel‐wise manner. We observed that male subjects with relatively short repeats allocated less money to others and exhibited a significantly smaller GMV in the right fusiform face area (FFA) compared with male long homozygotes. In male subjects, the GMV of the right FFA exhibited a significant positive correlation with altruistic behavior. A mixed mediation and moderation analysis further revealed both a significant mediation effect of the GMV of the right FFA on the association between AVPR1A RS3 repeat polymorphisms and allocation sums and a significant moderation effect of sex (only in males) on the mediation effect. Post hoc analysis showed that the GMV of the right FFA was significantly smaller in male subjects carrying allele 426 than in non‐426 carriers. These results suggest that the GMV of the right FFA may be a potential mediator whereby the genetic variants in AVPR1A RS3 affect altruistic behavior in healthy male subjects. Hum Brain Mapp 37:2700–2709, 2016. © 2016 Wiley Periodicals, Inc.     

Remarkable selectivity of the in vivo binding of [3H]Ro15‐4513 to α5 subtype of benzodiazepine receptor in the living mouse brain

To evaluate the binding characteristics of [³H]Ro15‐4513 with the central benzodiazepine (BZ) receptor, inhibition experiments of [³H]Ro15‐1788 and [³H]Ro15‐4513 were performed both in vitro and in vivo, using two BZ ligands, flunitrazepam (FNP), and ethyl‐β‐carboline‐3‐carboxylate (β‐CCE). FNP inhibited the binding of [³H]Ro15‐1788 and [³H]Ro15‐4513 in a dose‐dependent manner in the mouse cerebral cortex, hippocampus, and cerebellum, both in vitro and in vivo. β‐CCE also inhibited the binding of [³H]Ro15‐1788 and [³H]Ro15‐4513 in all the aforementioned brain regions in vitro. However, in vivo, β‐CCE inhibited the binding of [³H]Ro15‐4513 in the cerebral cortex and cerebellum, but not in the hippocampus, even at an injected dose of up to 1mg/kg. In contrast, more than 50% of the in vivo binding of [³H]Ro15‐1788 was inhibited by 1 mg/kg of β‐CCE in all regions. The time‐activity curve of [³H]Ro15‐4513 in the hippocampus also showed no alteration of the peak uptake between the control group and 0.3 mg/kg of β‐CCE coinjected group. These results indicated that the binding characteristics of [³H]Ro15‐4513 with the BZ receptor differed markedly between the in vitro and in vivo condition, and the selectivity of [³H]Ro15‐4513 binding to α5 subtype of BZ receptor in the mouse brain seemed to be remarkable under the in vivo condition. Synapse 64:928–936, 2010. © 2010 Wiley‐Liss, Inc.