Regulation of depressive-like behaviours by palmitoylation: Role of AKAP150 in the basolateral amygdala.

BACKGROUND AND PURPOSE: Protein palmitoylation is involved in learning and memory, and in emotional disorders. Yet, the underlying mechanisms in these processes remain unclear. Herein, we describe that A-kinase anchoring protein 150 (AKAP150) is essential and sufficient for depressive-like behaviours in mice via a palmitoylation-dependent mechanism. EXPERIMENTAL APPROACH: Depressive-like behaviours in mice were induced by chronic restraint stress (CRS) and chronic unpredictable mild stress (CUMS). Palmitoylated proteins in the basolateral amygdala (BLA) were assessed by an acyl-biotin exchange assay. Genetic and pharmacological approaches were used to investigate the role of the DHHC2-mediated AKAP150 palmitoylation signalling pathway in depressive-like behaviours. Electrophysiological recording, western blotting and co-immunoprecipitation were performed to define the mechanistic pathway. KEY RESULTS: Chronic stress successfully induced depressive-like behaviours in mice and enhanced AKAP150 palmitoylation in the BLA, and a palmitoylation inhibitor was enough to reverse these changes. Blocking the AKAP150-PKA interaction with the peptide Ht-31 abolished the CRS-induced AKAP150 palmitoylation signalling pathway. DHHC2 expression and palmitoylation levels were both increased after chronic stress. DHHC2 knockdown prevented CRS-induced depressive-like behaviours, as well as attenuating AKAP150 signalling and synaptic transmission in the BLA in CRS-treated mice. CONCLUSION AND IMPLICATIONS: These results delineate that DHHC2 modulates chronic stress-induced depressive-like behaviours and synaptic transmission in the BLA via the AKAP150 palmitoylation signalling pathway, and this pathway may be considered as a promising novel therapeutic target for major depressive disorder.

Activation of transient receptor potential vanilloid 1 ameliorates tau accumulation-induced synaptic damage and cognitive dysfunction via autophagy enhancement.

AIMS: The autophagy-lysosomal pathway is important for maintaining cellular proteostasis, while dysfunction of this pathway has been suggested to drive the aberrant intraneuronal accumulation of tau protein, leading to synaptic damage and cognitive impairment. Previous studies have demonstrated that the activation of transient receptor potential vanilloid 1 (TRPV1) by capsaicin has a positive impact on cognition and AD-related biomarkers. However, the effect and mechanism of TPRV1 activation on neuronal tau homeostasis remain elusive. METHODS: A mouse model of tauopathy was established by overexpressing full-length human tau in the CA3 area. Mice were fed capsaicin diet (0.0125%) or normal diet for 9 weeks. The cognitive ability, synaptic function, tau phosphorylation levels, and autophagy markers were detected. In vitro, capsaicin-induced alterations in cellular autophagy and tau degradation were characterized using two cell models. Besides, various inhibitors were applied to validate the role of TRPV1-mediated autophagy enhancement in tau clearance. RESULTS: We observed that TRPV1 activation by capsaicin effectively mitigates hippocampal tau accumulation-induced synaptic damages, gliosis, and cognitive impairment in vivo. Capsaicin promotes the degradation of abnormally accumulated tau through enhancing autophagic function in neurons, which is dependent on TRPV1-mediated activation of AMP-activated protein kinase (AMPK) and subsequent inhibition of the mammalian target of rapamycin (mTOR). Blocking AMPK activation abolishes capsaicin-induced autophagy enhancement and tau degradation in neurons. CONCLUSION: Our findings reveal that capsaicin-induced TRPV1 activation confers neuroprotection by restoring neuronal tau homeostasis via modulating cellular autophagy and provides additional evidence to support the potential of TRPV1 as a therapeutic target for tauopathies.

Regulation of anxiety-like behaviors by S-palmitoylation and S-nitrosylation in basolateral amygdala.

Protein posttranslational modification regulates synaptic protein stability, sorting and trafficking, and is involved in emotional disorders. Yet the molecular mechanisms regulating emotional disorders remain unelucidated. Here we report unknown roles of protein palmitoylation/nitrosylation crosstalk in regulating anxiety-like behaviors in rats. According to the percentages of open arm duration in the elevated plus maze test, the rats were divided into high-, intermediate- and low-anxiety groups. The palmitoylation and nitrosylation levels were detected by acyl-biotin exchange assay, and we found low palmitoylation and high nitrosylation levels in the basolateral amygdala (BLA) of high-anxiety rats. Furthermore, we observed that 2-bromopalmitate (2-BP), a palmitoylation inhibitor, induced anxiety-like behaviors, accompanied with decreased amplitude and frequency of mEPSCs and mIPSCs in the BLA. Additionally, we also found that inhibiting nNOS activity with 7-nitroindazole (7-NI) in the BLA caused anxiolytic effects and reduced the synaptic transmission. Interestingly, diazepam (DZP) rapidly elevated the protein palmitoylation level and attenuated the protein nitrosylation level in the BLA. Specifically, similar to DZP, the voluntary wheel running exerted DZP-like anxiolytic action, and induced high palmitoylation and low nitrosylation levels in the BLA. Lastly, blocking the protein palmitoylation with 2-BP induced an increase in protein nitrosylation level, and attenuating the nNOS activity by 7-NI elevated the protein palmitoylation level. Collectively, these results show a critical role of protein palmitoylation/nitrosylation crosstalk in orchestrating anxiety behavior in rats, and it may serve as a potential target for anxiolytic intervention.

WDR62-deficiency Causes Autism-like Behaviors Independent of Microcephaly in Mice.

Brain size abnormality is correlated with an increased frequency of autism spectrum disorder (ASD) in offspring. Genetic analysis indicates that heterozygous mutations of the WD repeat domain 62 (WDR62) are associated with ASD. However, biological evidence is still lacking. Our study showed that Wdr62 knockout (KO) led to reduced brain size with impaired learning and memory, as well as ASD-like behaviors in mice. Interestingly, Wdr62 Nex-cKO mice (depletion of WDR62 in differentiated neurons) had a largely normal brain size but with aberrant social interactions and repetitive behaviors. WDR62 regulated dendritic spinogenesis and excitatory synaptic transmission in cortical pyramidal neurons. Finally, we revealed that retinoic acid gavages significantly alleviated ASD-like behaviors in mice with WDR62 haploinsufficiency, probably by complementing the expression of ASD and synapse-related genes. Our findings provide a new perspective on the relationship between the microcephaly gene WDR62 and ASD etiology that will benefit clinical diagnosis and intervention of ASD.

GABA-B receptors enhance GABA-A receptor currents by modulation of membrane trafficking in dentate gyrus granule cells.

Activation of postsynaptic GABA-B receptors enhances tonic inhibition mediated by high-affinity extrasynaptic GABAA receptors in dentate gyrus granule cells (DGGCs), thalamocortical neurons, and cerebellar granule cells. We investigated the mechanism(s) of GABA current modulation by GABAB receptors in DGGCs using a combination of electrophysiological and biochemical approaches. In acute hippocampal brain slices the GABAB receptor agonist baclofen increased GABA-evoked currents in ∼2/3rds of DGGCs, significantly increasing GABAA currents by 41% on average. Nonstationary noise analysis was performed to estimate the effects of baclofen on single channel conductance, mean open time, and channel number; these estimates suggest that GABAB receptor activation increases receptor number but does not modify single channel properties of GABAA receptors. To directly assess baclofen-induced changes in plasma membrane expression of GABAA receptors, biotinylated western blots were performed. Treatment of hippocampal slices with baclofen significantly increased the surface expression of GABAA receptor subunits (both δ and γ2 subunits) and this effect was inhibited by the GABAB receptor antagonist CGP55845. These data indicate that changes in membrane trafficking and increased number of GABAA receptors in plasma membrane contribute to the enhancement of GABA currents produced by GABAB receptor activation in DGGCs.

Research hotspots and trends in visceral pain research: A global comprehensive bibliometric analysis.

Background: Visceral pain is a complex and heterogeneous disorder that is considered more prominent compared to somatic pain, due to its multiple and complex causes and accompanying emotional and mood disorders. Research has become increasingly extensive over the years, but a bibliometric analysis of this field is lacking. The aim of this study was to analyze global research trends in visceral pain over the past 40 years through visual analysis. Methods: We conducted a comprehensive search of the literature from January 1981 to December 2021 using the Web of Science core database. The medical subject term 'visceral pain' was searched. We used CiteSpace and VOSviewer for bibliometric analysis and network visualization, including top-ranked authors, keywords, research collaborations, and literature co-occurrence network analysis. Results: A total of 5,047 articles were included in the analysis. The number of articles on visceral pain has continued to grow steadily over the past 40 years. The United States (1,716 articles), University of California (159 articles), and Neurogastroenterology and Motility (276 articles) were the country, institution, and journal with the most publications, respectively. Keyword analysis showed that inflammation, visceral hypersensitivity, inflammatory bowel disease (IBD), irritable bowel syndrome (IBS), anxiety, and quality of life were the research trends and priorities in this research field. Conclusion: Visceral pain-related research has received increasing attention in recent decades. However, there are still many unresolved issues in the field of visceral pain, such as the specific molecular mechanisms and clinical treatments between visceral pain and inflammation, IBD, IBS, anxiety, and quality of life, which may require further exploration based on modern scientific and technological means and more basic research, especially for the therapeutic targets of visceral pain, which may become a hot spot for future research and provide guidance for the treatment of clinical diseases related to visceral pain.

Synaptic memory requires CaMKII.

Long-term potentiation (LTP) is arguably the most compelling cellular model for learning and memory. While the mechanisms underlying the induction of LTP ('learning') are well understood, the maintenance of LTP ('memory') has remained contentious over the last 20 years. Here, we find that Ca2+-calmodulin-dependent kinase II (CaMKII) contributes to synaptic transmission and is required LTP maintenance. Acute inhibition of CaMKII erases LTP and transient inhibition of CaMKII enhances subsequent LTP. These findings strongly support the role of CaMKII as a molecular storage device.

How the brain stores information is a question that has fascinated neuroscientists for well over a century. Two general ideas have emerged. The first is that groups of neurons hold information by staying active. The second is that they hold information by strengthening their connections to one another, making it easier for them to work together in the future. Scientists call this second idea 'long-term potentiation'. One of the molecules involved in long-term potentiation is a protein called calcium-calmodulin-dependent kinase II, or CaMKII for short. Blocking CaMKII, or deleting its gene, stops the connections between neurons from becoming stronger. This suggests neurons need CaMKII to learn, but it remains unclear whether neurons also use CaMKII to maintain neuronal memories after they have been created. If CaMKII does play a role in maintaining memories, blocking it after learning should reverse the learning process, but so far, experiments have not been able to show this. Tao et al. revisited these experiments to find out more. They examined slices of brain tissue from mice that had been treated with fast-acting CaMKII inhibitors. It took tens of minutes, but the inhibitors were able to reverse long-term potentiation, both for newly acquired neuronal memories and for older memories that had formed when the mice were alive. The choice of CaMKII inhibitor and the time lag could explain why scientists have not observed the effect before. Understanding long-term potentiation is a fundamental part of understanding learning and memory. It could also reveal more about the opposite phenomenon: long-term depression. This is a type of learning where the connections between neurons become weaker. Long-term depression also takes tens of minutes to occur, suggesting that future research into CaMKII might shed light on how it works.

Mechanisms underlying the synaptic trafficking of the glutamate delta receptor GluD1.

Ionotropic glutamate delta receptors do not bind glutamate and do not generate ionic current, resulting in difficulty in studying the function and trafficking of these receptors. Here, we utilize chimeric constructs, in which the ligand-binding domain of GluD1 is replaced by that of GluK1, to examine its synaptic trafficking and plasticity. GluD1 trafficked to the synapse, but was incapable of expressing long-term potentiation (LTP). The C-terminal domain (CT) of GluD1 has a classic PDZ-binding motif, which is critical for the synaptic trafficking of other glutamate receptors, but we found that its binding to PSD-95 was very weak, and deleting the PDZ-binding motif failed to alter synaptic trafficking. However, deletion of the entire CT abolished synaptic trafficking, but not surface expression. We found that mutation of threonine (T) T923 to an alanine disrupted synaptic trafficking. Therefore, GluD1 receptors have strikingly different trafficking mechanisms compared with AMPARs. These results highlight the diversity of ionotropic glutamate receptor trafficking rules at a single type of synapse. Since this receptor is genetically associated with schizophrenia, our findings may provide an important clue to understand schizophrenia.

Signal peptide represses GluK1 surface and synaptic trafficking through binding to amino-terminal domain.

Kainate-type glutamate receptors play critical roles in excitatory synaptic transmission and synaptic plasticity in the brain. GluK1 and GluK2 possess fundamentally different capabilities in surface trafficking as well as synaptic targeting in hippocampal CA1 neurons. Here we find that the excitatory postsynaptic currents (EPSCs) are significantly increased by the chimeric GluK1(SPGluK2) receptor, in which the signal peptide of GluK1 is replaced with that of GluK2. Coexpression of GluK1 signal peptide completely suppresses the gain in trafficking ability of GluK1(SPGluK2), indicating that the signal peptide represses receptor trafficking in a trans manner. Furthermore, we demonstrate that the signal peptide directly interacts with the amino-terminal domain (ATD) to inhibit the synaptic and surface expression of GluK1. Thus, we have uncovered a trafficking mechanism for kainate receptors and propose that the cleaved signal peptide behaves as a ligand of GluK1, through binding with the ATD, to repress forward trafficking of the receptor.

Postsynaptic δ1 glutamate receptor assembles and maintains hippocampal synapses via Cbln2 and neurexin.

The δ1 glutamate receptor (GluD1) was cloned decades ago and is widely expressed in many regions of the brain. However, its functional roles in these brain circuits remain unclear. Here, we find that GluD1 is required for both excitatory synapse formation and maintenance in the hippocampus. The action of GluD1 is absent in the Cbln2 knockout mouse. Furthermore, the GluD1 actions require the presence of presynaptic neurexin 1ß carrying the splice site 4 insert (+S4). Together, our findings demonstrate that hippocampal synapse assembly and maintenance require a tripartite molecular complex in which the ligand Cbln2 binds with presynaptic neurexin 1ß (+S4) and postsynaptic GluD1. We provide evidence that this mechanism may apply to other forebrain synapses, where GluD1 is widely expressed.

LTP requires postsynaptic PDZ-domain interactions with glutamate receptor/auxiliary protein complexes.

Long-term potentiation (LTP) is a persistent strengthening of synaptic transmission in the brain and is arguably the most compelling cellular and molecular model for learning and memory. Previous work found that both AMPA receptors and exogenously expressed kainate receptors are equally capable of expressing LTP, despite their limited homology and their association with distinct auxiliary subunits, indicating that LTP is far more promiscuous than previously thought. What might these two subtypes of glutamate receptor have in common? Using a single-cell molecular replacement strategy, we demonstrate that the AMPA receptor auxiliary subunit TARP γ-8, via its PDZ-binding motif, is indispensable for both basal synaptic transmission and LTP. Remarkably, kainate receptors and their auxiliary subunits Neto proteins share the same requirement of PDZ-binding domains for synaptic trafficking and LTP. Together, these results suggest that a minimal postsynaptic requirement for LTP is the PDZ binding of glutamate receptors/auxiliary subunits to PSD scaffolding proteins.

Postsynaptic GABAB receptors enhance extrasynaptic GABAA receptor function in dentate gyrus granule cells.

Ambient GABA in the brain tonically activates extrasynaptic GABA(A) receptors, and activity-dependent changes in ambient GABA concentration can also activate GABA(B) receptors. To investigate an interaction between postsynaptic GABA(B) and GABA(A) receptors, we recorded GABA(A) currents elicited by exogenous GABA (10 µm) from dentate gyrus granule cells (DGGCs) in adult rat hippocampal slices. The GABA(B) receptor agonist baclofen (20 µm) enhanced GABA(A) currents. This enhancement was blocked by the GABA(B) receptor antagonist CGP 55845 and intracellular solutions containing the GTP analog GDP-ß-s, indicating that baclofen was acting on postsynaptic GABA(B) receptors. Modulation of GABA(A) currents by postsynaptic GABA(B) receptors was not observed in CA1 pyramidal cells or layer 2/3 cortical pyramidal neurons. Baclofen reduced the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) but did not alter sIPSC amplitude or kinetics. Thus, GABA(A) receptors activated at synapses were not modulated by postsynaptic GABA(B) receptors. In contrast, tonic GABA currents and currents activated by the GABA(A) receptor δ subunit-selective agonist THIP (10 µm) were potentiated by baclofen. Our data indicate that postsynaptic GABA(B) receptors enhance the function of extrasynaptic GABA(A) receptors, including δ subunit-containing receptors that mediate tonic inhibition in DGGCs. The modulation of GABA(A) receptor function by postsynaptic GABA(B) receptors is a newly identified mechanism that will influence the inhibitory tone of DGGCs when GABA(B) and GABA(A) receptors are both activated.

Rapid regulation of tonic GABA currents in cultured rat hippocampal neurons.

Subacute and chronic changes in tonic GABAergic inhibition occur in human and experimental epilepsy. Less is known about how tonic inhibition is modulated over shorter time frames (seconds). We measured endogenous tonic GABA currents from cultured rat hippocampal neurons to evaluate how they are affected by 1) transient increases in extracellular GABA concentration ([GABA]), 2) transient postsynaptic depolarization, and 3) depolarization of presynaptic cells. Transient increases in [GABA] (1 µM) reduced tonic currents; this reduction resulted from GABA-induced shifts in the reversal potential for GABA currents (E(GABA)). Transient depolarization of postsynaptic neurons reversed the effects of exogenous GABA and potentiated tonic currents. The voltage-dependent potentiation of tonic GABA currents was independent of E(GABA) shifts and represented postdepolarization potentiation (PDP), an intrinsic GABA(A) receptor property (Ransom CB, Wu Y, Richerson GB. J Neurosci 30: 7672-7684, 2010). Inhibition of vesicular GABA release with concanamycin A (ConA) did not affect tonic currents. In ConA-treated cells, transient application of 12 mM K(+) to depolarize presynaptic neurons and glia produced a persistent increase in tonic current amplitude. The K(+)-induced increase in tonic current was reversibly inhibited by SKF89976a (40 µM), indicating that this was caused by nonvesicular GABA release from GABA transporter type 1 (GAT1). Nonvesicular GABA release due to GAT1 reversal also occurred in acute hippocampal brain slices. Our results indicate that tonic GABA currents are rapidly regulated by GABA-induced changes in intracellular Cl(-) concentration, PDP of extrasynaptic GABA(A) receptors, and nonvesicular GABA release. These mechanisms may influence tonic inhibition during seizures when neurons are robustly depolarized and extracellular GABA and K(+) concentrations are elevated.

Neuroprotective effects of active ingredients isolated from Pegasus laternarius on cultured cerebral neurons.

Seamoth (Pegasus laternarius Cuvier) is extensively used to treat various diseases on the coastland of Guangdong Province in China, such as scrofula, cough, and diarrhea. The total extract of Pegasus laternarius (EP) was subjected to column chromatography to acquire three different constituents (EPC1, EPC2, and EPC3). Cerebral neuron injury was induced by glutamate, H2O2, and serum deprivation. After treating with or without different extracts, cell viability was assessed with the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, and cell apoptosis was analyzed with Hoechst 33258 staining and agarose gel electrophoresis. We also determined the levels of lactate dehydrogenase (LDH), maleic dialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px). The results showed that both EP and EPC2 promoted the outgrowth of cultural neurons, increased antioxidant enzyme activity, and protected neurons from neuronal injury or apoptosis induced by glutamate, H2O2, and serum deprivation. EPC1 and EPC3 had little or no effect on neurons. These results suggest that the active ingredients obtained from Pegasus laternarius have potential neuroprotective effects on injured neurons by promoting the outgrowth of cultured neurons, increasing the activity of intracellular antioxidants, and exerting antiapoptotic effects. This neuroprotection may be attributable to specific active ingredients, such as taurine, novel ceramide, and cholesterol.

New medicinal properties of mangostins: analgesic activity and pharmacological characterization of active ingredients from the fruit hull of Garcinia mangostana L.

The fruit hull of Garcinia mangostana L. contains oxygenated and prenylated phenol derivatives, such as xanthones or xanthen-9H-ones, and is used by people in Southeast Asia as a traditional medicine for the treatment of abdominal pain, dysentery, wound infections, suppuration, and chronic ulcer. We isolated the active ingredients from the crude ethanol extract of G.mangostana L. (CEM) and investigated their analgesic effects and underlying mechanisms. CEM at intragastric (i.g.) doses of 0.5, 1, and 3 g/kg clearly exhibited antinociceptive effects in the hot-plate and acetic acid-induced writhing tests in mice. Two isolated compounds, alpha-mangostin and gamma-mangostin, exhibited analgesic effects at doses of 25 and 50 mg/kg (i.g.) in the hot-plate and formalin tests, respectively. CEM at doses of 0.5, 1, and 3 g/kg significantly inhibited xylene-induced release of inflammatory mediators. CEM, alpha-mangostin, and gamma-mangostin each dose-dependently demonstrated the ability to scavenge reactive oxygen species. In conclusion, our results demonstrate that CEM and mangostins possess potent peripheral and central antinociceptive effects in mice and suggest that xanthones may be developed as novel analgesics and anti-inflammatory drugs.

Critical roles of voltage-dependent sodium channels in the process of synaptogenesis during the postnatal cortical development of rats.

The developmental changes of the sodium channel and construction of synapse connection were studied in cerebral cortical pyramidal neurons of rats at different age groups. We used whole-cell patch-clamp recordings to characterize electrophysiological properties of cortical neurons at different age stages, including the sodium currents, APs evoked by depolarizing current and short-term plasticity of the eEPSCs. The result shows that the sodium currents undergo a hyperpolarizing shift in activation process and acceleration of activation and inactivation with age. The maximal sodium current also increased with maturation, and the evident difference appeared from P7-P11 (with the day of birth as P0) to P12-P15 group. The tendency of the sodium current density changes which exhibited the same properties as that of sodium current, showed the significant increases from P19-P21 to P >or= 22 group. The APs' parameters exhibited the age-dependent changes except the threshold, including the increase of the peak amplitude from P or= 22 group, the 2nd response showed the tendency of facilitation compared with the younger age groups. Our results indicated that the cerebral cortical pyramidal neurons of rats are undergoing marked changes in the characteristics of their sodium channels with maturation, which play a critical role in synaptogenesis and construction of the neuronal network.

The effect of recombinant neurotoxins from the sea anemone Anthopleura sp. on sodium currents of rat cerebral cortical neurons.

We have investigated the action of the recombinant neurotoxins, named Hk7a and Hk2a, whose amino acid sequences differ only in two positions, isolated from the sea anemone Anthopleura sp., on neuronal sodium currents using the whole-cell voltage-clamp techniques. The rat cerebral cortical neurons in primary culture were used for this study. In our experiments, these cells all express tetrodotoxin-sensitive (TTX-S) sodium currents. Under the voltage-clamp condition, application of Hk7a and Hk2a reduced the sodium channel current amplitude and shifted the voltage dependence of activation to more positive potential; while Hk7a produced no significant effect on the voltage at which 50% of the channels were inactivated, Hk2a caused profound hyperpolarizing shift of the voltage-dependent inactivation. Also, both Hk7a and Hk2a increased the time course of recovery from inactivation. In kinetic studies, whereas application of Hk2a slows the time to peak of voltage-gated sodium channel, the time course of fast and slow inactivating component, no significant effect was observed in Hk7a. These results suggested that the difference of key amino acid between Hk7a and Hk2a might contribute to their different action; therefore, they could be used as pharmacological tool to study the structure and function of voltage-gated sodium channel.