MicroRNA-mediated regulation of neurotransmitter receptors in epilepsy: A systematic review.

BACKGROUND: Pathogenesis of epilepsy involves dysregulation of the neurotransmitter system contributing to hyper-excitability of neuronal cells. MicroRNA (miRNAs) are small non-coding RNAs known to play a crucial role in post-transcriptional regulation of gene expression. METHODS: The present review was prepared following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines, employing a comprehensive search strategy to identify and extract data from published research articles. Keywords suchas epilepsy, micro RNA (micro RNAs, miRNA, miRNAs, miR), neurotransmitters (specific names), and neurotransmitter receptors (specific names) were used to construct the query. RESULTS: A total of 724 articles were identified using the keywords epilepsy, microRNA along with select neurotransmitter and neurotransmitter receptor names. After exclusions, the final selection consisted of 17 studies, most of which centered on glutamate and gamma-aminobutyric acid (GABA) receptors. Singular studies also investigated miRNAs affecting cholinergic, purinergic, and glycine receptors. CONCLUSION: This review offers a concise overview of the current knowledge on miRNA-mediated regulation of neurotransmitter receptors in epilepsy and highlights their potential for future clinical application.

Serotonin Receptors and Acetylcholinesterase Gene Expression Alternations: The Potential Value on Tumor Microenvironment of Gastric Cancer.

INTRODUCTION: Gastric cancer is one of the common causes of cancer-related death in the world. Neurotransmitters have recently been related to the proliferation of cancer cells, but the role of neurotransmitters in the progression of gastric cancer is still unexplored. The cross-talk between the nervous system and immune cells through serotonin and its receptors in the tumor microenvironment can impact tumor progress. Our purpose is to expose probable changes in serotonin receptors, acetylcholinesterase, and monoamine oxidase A gene expression in gastric cancer. METHODS: Transcript of serotonin receptors (5-HTR2A, 5-HTR2B, 5-HTR3A, 5-HTR7) and monoamine oxidase A genes in the peripheral blood mononuclear cells (40 patients and 40 control) and tissue (21 tumors and 21 normal adjacent tissues) were assessed. The gene expression was analyzed by quantitative real-time PCR using suitable primers. Statistical analysis was performed using appropriate software (REST, Prism). RESULTS: Significantly higher amounts of 5-HTR2A, 5-HTR2B, 5-HTR3A, 5-HTR7, and acetylcholinesterase gene transcripts were found in the peripheral blood of gastric cancer patients compared with healthy individuals. The expression of 5-HTR2B and 5-HTR3A genes was significantly higher (p = 0.0250, p = 0.0005, respectively) and the acetylcholinesterase gene was lower in the tissue of patients (p = 0.0119) compared with adjacent normal tissue. CONCLUSION: This study highlights the role of serotonin receptors in gastric cancer that might have suggestions for the development of novel therapeutics and defensive approaches that target factors associated with the link between the nervous system, cancer cells, and the tumor microenvironment.

Dopamine receptors gene overexpression in the microenvironment of invasive gastric cancer and its potential implications.

BACKGROUND: Gastric cancer (GC) is the fifth most common cancer worldwide and the most commonly diagnosed cancer in Iran. The nervous system provides proximity to tumor cells by releasing neurotransmitters such as dopamine and presenting them to the corresponding receptor-bearing tumors. While nerve fibers infiltrate the tumor microenvironment, little is known about the expression levels of dopamine (DA), dopamine receptors (DRs), and catechol-O-methyltransferase (COMT) in GC patients. METHODS: DRs and COMT expression were analyzed in 45 peripheral blood mononuclear cells (PBMCs) and 20 paired tumor and adjacent tissue of GC patients by quantitative polymerase chain reaction. DA was measured in plasma specimens using enzyme-linked immunosorbent assay. Protein-protein interaction analysis was carried out to identify GC-related hub genes. RESULTS: Increased expression of DRD1-DRD3 was found in tumor specimens compared with adjacent non-cancerous specimens (P < 0.05). A positive correlation was found between DRD1 and DRD3 expression (P = 0.009); DRD2 and DRD3 expression (P = 0.04). Plasma levels of dopamine were significantly lower in patients (1298 pg/ml) than in controls (4651 pg/ml). DRD1-DRD4 and COMT were up-regulated in PBMCs of patients compared with controls (P < 0.0001). Bioinformatic analyses showed 30 hub genes associated with Protein kinase A and extracellular signal-regulated kinase signaling pathways. CONCLUSIONS: The findings indicated dysregulation of DRs and COMT mRNA expression in GC and suggest that the brain- gastrointestinal axis may mediate gastric cancer development. Network analysis revealed that combination treatments could be considered for optimizing and improving the precision treatment of GC.

Personalized brain models identify neurotransmitter receptor changes in Alzheimer's disease.

Alzheimer's disease involves many neurobiological alterations from molecular to macroscopic spatial scales, but we currently lack integrative, mechanistic brain models characterizing how factors across different biological scales interact to cause clinical deterioration in a way that is subject-specific or personalized. As important signalling molecules and mediators of many neurobiological interactions, neurotransmitter receptors are promising candidates for identifying molecular mechanisms and drug targets in Alzheimer's disease. We present a neurotransmitter receptor-enriched multifactorial brain model, which integrates spatial distribution patterns of 15 neurotransmitter receptors from post-mortem autoradiography with multiple in vivo neuroimaging modalities (tau, amyloid-ß and glucose PET, and structural, functional and arterial spin labelling MRI) in a personalized, generative, whole-brain formulation. In a heterogeneous aged population (n = 423, ADNI data), models with personalized receptor-neuroimaging interactions showed a significant improvement over neuroimaging-only models, explaining about 70% (±20%) of the variance in longitudinal changes to the six neuroimaging modalities. In Alzheimer's disease patients (n = 25, ADNI data), receptor-imaging interactions explained up to 39.7% (P < 0.003, family-wise error-rate-corrected) of inter-individual variability in cognitive deterioration, via an axis primarily affecting executive function. Notably, based on their contribution to the clinical severity in Alzheimer's disease, we found significant functional alterations to glutamatergic interactions affecting tau accumulation and neural activity dysfunction and GABAergic interactions concurrently affecting neural activity dysfunction, amyloid and tau distributions, as well as significant cholinergic receptor effects on tau accumulation. Overall, GABAergic alterations had the largest effect on cognitive impairment (particularly executive function) in our Alzheimer's disease cohort (n = 25). Furthermore, we demonstrate the clinical applicability of this approach by characterizing subjects based on individualized 'fingerprints' of receptor alterations. This study introduces the first robust, data-driven framework for integrating several neurotransmitter receptors, multimodal neuroimaging and clinical data in a flexible and interpretable brain model. It enables further understanding of the mechanistic neuropathological basis of neurodegenerative progression and heterogeneity, and constitutes a promising step towards implementing personalized, neurotransmitter-based treatments.

Neurotransmitter receptor densities are associated with changes in regional Cerebral blood flow during clinical ongoing pain.

Arterial spin labelling (ASL) plays an increasingly important role in neuroimaging pain research but does not provide molecular insights regarding how regional cerebral blood flow (rCBF) relates to underlying neurotransmission. Here, we integrate ASL with positron emission tomography (PET) and brain transcriptome data to investigate the molecular substrates of rCBF underlying clinically relevant pain states. Two data sets, representing acute and chronic ongoing pain respectively, were utilised to quantify changes in rCBF; one examining pre-surgical versus post-surgical pain, and the second comparing patients with painful hand Osteoarthritis to a group of matched controls. We implemented a whole-brain spatial correlation analysis to explore associations between change in rCBF (ΔCBF) and neurotransmitter receptor distributions derived from normative PET templates. Additionally, we utilised transcriptomic data from the Allen Brain Atlas to inform distributions of receptor expression. Both datasets presented significant correlations of ΔCBF with the µ-opioid and dopamine-D2 receptor expressions, which play fundamental roles in brain activity associated with pain experiences. ΔCBF also correlated with the gene expression distributions of several receptors involved in pain processing. Overall, this is the first study illustrating the molecular basis of ongoing pain ASL indices and emphasises the potential of rCBF as a biomarker in pain research.

The Role of DNA Methylation and Activity of Neurotransmitter Receptors in the Mechanisms of Specific Anterograde Amnesia and Memory Recovery.

Impairment of reconsolidation of conditioned food aversion memory led to the development of a specific anterograde amnesia: repeated training of amnestic snails did not induce long-term memory formation. DNA demethylation caused by injections of DNA methyltransferase inhibitor (DNAMT) during repeated training led to long-term memory formation. Injections of an NMDA glutamate receptor antagonist or a serotonin receptor antagonist prevented memory formation induced by administration of DNAMT inhibitor and repeated training. We hypothesize that methylation-dependent repression of neuronal genes underlies anterograde amnesia. Demethylation eliminated the blockade of these genes and created conditions for long-term memory formation, the induction mechanisms of which involve neurotransmitter receptors.

The post-synaptic scaffolding protein tamalin regulates ligand-mediated trafficking of metabotropic glutamate receptors.

Group I metabotropic glutamate receptors (mGluRs) play important roles in various neuronal functions and have also been implicated in multiple neuropsychiatric disorders like fragile X syndrome, autism, and others. mGluR trafficking not only plays important roles in controlling the spatiotemporal localization of these receptors in the cell but also regulates the activity of these receptors. Despite this obvious significance, the cellular machineries that control the trafficking of group I metabotropic glutamate receptors in the central nervous system have not been studied in detail. The post-synaptic scaffolding protein tamalin has been shown to interact with group I mGluRs and also with many other proteins involved in protein trafficking in neurons. Using a molecular replacement approach in mouse hippocampal neurons, we show here that tamalin plays a critical role in the ligand-dependent internalization of mGluR1 and mGluR5, members of the group I mGluR family. Specifically, knockdown of endogenous tamalin inhibited the ligand-dependent internalization of these two receptors. Both N-terminal and C-terminal regions of tamalin played critical roles in mGluR1 endocytosis. Furthermore, we found that tamalin regulates mGluR1 internalization by interacting with S-SCAM, a protein that has been implicated in vesicular trafficking. Finally, we demonstrate that tamalin plays a critical role in mGluR-mediated internalization of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, a process believed to be the cellular correlate for mGluR-dependent synaptic plasticity. Taken together, these findings reveal a mechanistic role of tamalin in the trafficking of group I mGluRs and suggest its physiological implications in the brain.

New boundaries and dissociation of the mouse hippocampus along the dorsal-ventral axis based on glutamatergic, GABAergic and catecholaminergic receptor densities.

In rodents, gene-expression, neuronal tuning, connectivity and neurogenesis studies have postulated that the dorsal, the intermediate and the ventral hippocampal formation (HF) are distinct entities. These findings are underpinned by behavioral studies showing a dissociable role of dorsal and ventral HF in learning, memory, stress and emotional processing. However, up to now, the molecular basis of such differences in relation to discrete boundaries is largely unknown. Therefore, we analyzed binding site densities for glutamatergic AMPA, NMDA, kainate and mGluR2/3 , GABAergic GABAA (including benzodiazepine binding sites), GABAB , dopaminergic D1/5 and noradrenergic α1 and α2 receptors as key modulators for signal transmission in hippocampal functions, using quantitative in vitro receptor autoradiography along the dorsal-ventral axis of the mouse HF. Beside general different receptor profiles of the dentate gyrus (DG) and Cornu Ammonis fields (CA1, CA2, CA3, CA4/hilus), we detected substantial differences between dorsal, intermediate and ventral subdivisions and individual layers for all investigated receptor types, except GABAB . For example, striking higher densities of α2 receptors were detected in the ventral DG, while the dorsal DG possesses higher numbers of kainate, NMDA, GABAA and D1/5 receptors. CA1 dorsal and intermediate subdivisions showed higher AMPA, NMDA, mGluR2/3 , GABAA , D1/5 receptors, while kainate receptors are higher expressed in ventral CA1, and noradrenergic α1 and α2 receptors in the intermediate region of CA1. CA2 dorsal was distinguished by higher kainate, α1 and α2 receptors in the intermediate region, while CA3 showed a more complex dissociation. Our findings resulted not only in a clear segmentation of the mouse hippocampus along the dorsal-ventral axis, but also provides insights into the neurochemical basis and likely associated physiological processes in hippocampal functions. Therein, the presented data has a high impact for future studies modeling and investigating dorsal, intermediate and ventral hippocampal dysfunction in relation to neurodegenerative diseases or psychiatric disorders.

Personality traits and polymorphisms of genes coding neurotransmitter receptors or transporters: review of single gene and genome-wide association studies.

BACKGROUND: The most popular tool used for measuring personality traits is the Five-Factor Model (FFM). It includes neuroticism, extraversion, openness, agreeableness and conscientiousness. Many studies indicated the association of genes encoding neurotransmitter receptors/transporters with personality traits. The relationship connecting polymorphic DNA sequences and FFM features has been described in the case of genes encoding receptors of cannabinoid and dopaminergic systems. Moreover, dopaminergic system receives inputs from other neurotransmitters, like GABAergic or serotoninergic systems. METHODS: We searched PubMed Central (PMC), Science Direct, Scopus, Cochrane Library, Web of Science and EBSCO databases from their inception to November 19, 2020, to identify original studies, as well as peer-reviewed studies examining the FFM and its association with gene polymorphisms affecting the neurotransmitter functions in central nervous system. RESULTS: Serotonin neurons modulate dopamine function. In gene encoding serotonin transporter protein, SLC6A4, was found polymorphism, which was correlated with openness to experience (in Sweden population), and high scores of neuroticism and low levels of agreeableness (in Caucasian population). The genome-wide association studies (GWASs) found an association of 5q34-q35, 3p24, 3q13 regions with higher scores of neuroticism, extraversion and agreeableness. However, the results for chromosome 3 regions are inconsistent, which was shown in our review paper. CONCLUSIONS: GWASs on polymorphisms are being continued in order to determine and further understand the relationship between the changes in DNA and personality traits.

Ion Channels as New Attractive Targets to Improve Re-Myelination Processes in the Brain.

Multiple sclerosis (MS) is the most demyelinating disease of the central nervous system (CNS) characterized by neuroinflammation. Oligodendrocyte progenitor cells (OPCs) are cycling cells in the developing and adult CNS that, under demyelinating conditions, migrate to the site of lesions and differentiate into mature oligodendrocytes to remyelinate damaged axons. However, this process fails during disease chronicization due to impaired OPC differentiation. Moreover, OPCs are crucial players in neuro-glial communication as they receive synaptic inputs from neurons and express ion channels and neurotransmitter/neuromodulator receptors that control their maturation. Ion channels are recognized as attractive therapeutic targets, and indeed ligand-gated and voltage-gated channels can both be found among the top five pharmaceutical target groups of FDA-approved agents. Their modulation ameliorates some of the symptoms of MS and improves the outcome of related animal models. However, the exact mechanism of action of ion-channel targeting compounds is often still unclear due to the wide expression of these channels on neurons, glia, and infiltrating immune cells. The present review summarizes recent findings in the field to get further insights into physio-pathophysiological processes and possible therapeutic mechanisms of drug actions.

A Critical Role for Sorting Nexin 1 in the Trafficking of Metabotropic Glutamate Receptors.

Group I metabotropic glutamate receptors (mGluRs) function as modulators of neuronal physiology and they have also been implicated in various neuropsychiatric disorders. Trafficking of mGluRs plays important roles in controlling the precise localization of these receptors at specific region of the cell, as well as it regulates the activity of these receptors. Despite this obvious significance, we know very little about the cellular machineries that control the trafficking of these receptors in the CNS. Sorting nexin 1 (SNX1) has been shown to regulate the endosomal sorting of few cell surface receptors either to lysosomes where they are downregulated or back to the cell surface. Using "molecular replacement" approach in hippocampal neurons derived from mice of both sexes, we show here that SNX1 plays critical role in the trafficking of mGluR1, a member of the group I mGluR family. Overexpression of dominant-negative SNX1 or knockdown of endogenous SNX1 resulted in the rapid recycling of the receptor. Importantly, recycling via the rapid recycling route, did not allow the resensitization of the receptors. Our data suggest that both, N-terminal and C-terminal region of SNX1 play critical role in the normal trafficking of the receptor. In addition, we also show here that SNX1 regulates the trafficking of mGluR1 through the interaction with Hrs (hepatocyte growth factor-regulated tyrosine kinase substrate), a protein that has been implicated in both signaling and vesicular trafficking. Thus, these studies reveal a mechanistic role of SNX1 in the trafficking of group I mGluRs and its physiological implications.SIGNIFICANCE STATEMENT Group I mGluRs are activated by the neurotransmitter glutamate in the CNS, and play various important roles in the brain. Similar to many other receptors, trafficking plays crucial roles in controlling the precise localization as well as activity of these receptors. Despite this obvious significance very little is known about the cellular machineries that control the trafficking of these receptors. We demonstrate here, that SNX1 plays a critical role in the trafficking of mGluR1, a member of the group I mGluR family. SNX1-mediated trafficking is critical for the resensitization of the receptor. SNX1 controls the trafficking of the receptor through the interaction with another protein, Hrs. The results suggest a role for SNX1 in the regulation of group I mGluRs.

Physiology of Astroglia.

Astrocytes are principal cells responsible for maintaining the brain homeostasis. Additionally, these glial cells are also involved in homocellular (astrocyte-astrocyte) and heterocellular (astrocyte-other cell types) signalling and metabolism. These astroglial functions require an expression of the assortment of molecules, be that transporters or pumps, to maintain ion concentration gradients across the plasmalemma and the membrane of the endoplasmic reticulum. Astrocytes sense and balance their neurochemical environment via variety of transmitter receptors and transporters. As they are electrically non-excitable, astrocytes display intracellular calcium and sodium fluctuations, which are not only used for operative signalling but can also affect metabolism. In this chapter we discuss the molecules that achieve ionic gradients and underlie astrocyte signalling.

Group I Metabotropic Glutamate Receptors (mGluRs): Ins and Outs.

Glutamate is a nonessential amino acid, known to act as a major excitatory neurotransmitter in the central nervous system. Glutamate transduces its signal by activating two types of receptors, viz., ionotropic glutamate receptors and metabotropic glutamate receptors (mGluRs). mGluR1 and mGluR5 are members of the group I mGluR family, and they belong to the G-protein-coupled receptor (GPCR) family. These receptors are involved in various forms of synaptic plasticity including learning and memory. Similar to many other GPCRs, trafficking plays a critical role in controlling the spatiotemporal localization of these receptors on the cell surface, which is critical for the normal ligand/receptor interaction. Improper targeting of GPCRs results in aberrant signaling, which often leads to various diseases. Trafficking also regulates the activity of these receptors. Thus, inappropriate trafficking of these receptors might have pathological consequences. Group I mGluRs have been implicated in various neuropsychiatric disorders like Fragile X syndrome, autism, etc. In this review, we discuss the current understanding of group I mGluR trafficking in the central nervous system and its physiological importance.

The lipid habitats of neurotransmitter receptors in brain.

Neurotransmitter receptors, the macromolecules specialized in decoding the chemical signals encrypted in the chemical signaling mechanism in the nervous system, occur either at the somatic cell surface of chemically excitable cells or at specialized subcellular structures, the synapses. Synapses have lipid compositions distinct from the rest of the cell membrane, suggesting that neurotransmitter receptors and their scaffolding and adaptor protein partners require specific lipid habitats for optimal operation. In this review we discuss some paradigmatic cases of neurotransmitter receptor-lipid interactions, highlighting the chemical nature of the intervening lipid species and providing examples of the receptor mechanisms affected by interaction with lipids. The focus is on the effects of cholesterol, glycerophospholipids and covalent fatty acid acylation on neurotransmitter receptors. We also briefly discuss the role of lipid phase states involving lateral heterogeneities of the host membrane known to modulate membrane transport, protein sorting and signaling. Modulation of neurotransmitter receptors by lipids occurs at multiple levels, affecting a wide span of activities including their trafficking, sorting, stability, residence lifetime at the cell surface, endocytosis, and recycling, among other important functional properties at the synapse.

A receptor-based analysis of local ecosystems in the human brain.

BACKGROUND: As a complex system, the brain is a self-organizing entity that depends on local interactions among cells. Its regions (anatomically defined nuclei and areas) can be conceptualized as cellular ecosystems, but the similarity of their functional profiles is poorly understood. The study used the Allen Human Brain Atlas to classify 169 brain regions into hierarchically-organized environments based on their expression of 100 G protein-coupled neurotransmitter receptors, with no a priori reference to the regions' positions in the brain's anatomy or function. The analysis was based on hierarchical clustering, and multiscale bootstrap resampling was used to estimate the reliability of detected clusters. RESULTS: The study presents the first unbiased, hierarchical tree of functional environments in the human brain. The similarity of brain regions was strongly influenced by their anatomical proximity, even when they belonged to different functional systems. Generally, spatial vicinity trumped long-range projections or network connectivity. The main cluster of brain regions excluded the dentate gyrus of the hippocampus. The nuclei of the amygdala formed a cluster irrespective of their striatal or pallial origin. In its receptor profile, the hypothalamus was more closely associated with the midbrain than with the thalamus. The cerebellar cortical areas formed a tight and exclusive cluster. Most of the neocortical areas (with the exception of some occipital areas) clustered in a large, statistically well supported group that included no other brain regions. CONCLUSIONS: This study adds a new dimension to the established classifications of brain divisions. In a single framework, they are reconsidered at multiple scales-from individual nuclei and areas to their groups to the entire brain. The analysis provides support for predictive models of brain self-organization and adaptation.

Cortical chemoarchitecture shapes macroscale effective functional connectivity patterns in macaque cerebral cortex.

The mammalian cortex is a complex system of-at the microscale level-interconnected neurons and-at the macroscale level-interconnected areas, forming the infrastructure for local and global neural processing and information integration. While the effects of regional chemoarchitecture on local cortical activity are well known, the effect of local neurotransmitter receptor organization on the emergence of large scale region-to-region functional interactions remains poorly understood. Here, we examined reports of effective functional connectivity-as measured by the action of strychnine administration acting on the chemical balance of cortical areas-in relation to underlying regional variation in microscale neurotransmitter receptor density levels in the macaque cortex. Linking cortical variation in microscale receptor density levels to collated information on macroscale functional connectivity of the macaque cortex, we show macroscale patterns of effective corticocortical functional interactions-and in particular, the strength of connectivity of efferent macroscale pathways-to be related to the ratio of excitatory and inhibitory neurotransmitter receptor densities of cortical areas. Our findings provide evidence for the microscale chemoarchitecture of cortical areas to have a direct stimulating influence on the emergence of macroscale functional connectivity patterns in the mammalian brain. Hum Brain Mapp 37:1856-1865, 2016. © 2016 Wiley Periodicals, Inc.

A gating mechanism of pentameric ligand-gated ion channels.

Pentameric ligand-gated ion channels (pLGICs) play a central role in intercellular communication in the nervous system and are involved in fundamental processes such as attention, learning, and memory. They are oligomeric protein assemblies that convert a chemical signal into an ion flux through the postsynaptic membrane, but the molecular mechanism of gating ions has remained elusive. Here, we present atomistic molecular dynamics simulations of the prokaryotic channels from Gloeobacter violaceus (GLIC) and Erwinia chrysanthemi (ELIC), whose crystal structures are thought to represent the active and the resting states of pLGICs, respectively, and of the eukaryotic glutamate-gated chloride channel from Caenorhabditis elegans (GluCl), whose open-channel structure was determined complexed with the positive allosteric modulator ivermectin. Structural observables extracted from the trajectories of GLIC and ELIC are used as progress variables to analyze the time evolution of GluCl, which was simulated in the absence of ivermectin starting from the structure with bound ivermectin. The trajectory of GluCl with ivermectin removed shows a sequence of structural events that couple agonist unbinding from the extracellular domain to ion-pore closing in the transmembrane domain. Based on these results, we propose a structural mechanism for the allosteric communication leading to deactivation/activation of the GluCl channel. This model of gating emphasizes the coupling between the quaternary twisting and the opening/closing of the ion pore and is likely to apply to other members of the pLGIC family.

ELIC-α7 Nicotinic acetylcholine receptor (α7nAChR) chimeras reveal a prominent role of the extracellular-transmembrane domain interface in allosteric modulation.

The native α7 nicotinic acetylcholine receptor (α7nAChR) is a homopentameric ligand-gated ion channel mediating fast synaptic transmission and is of pharmaceutical interest for treatment of numerous disorders. The transmembrane domain (TMD) of α7nAChR has been identified as a target for positive allosteric modulators (PAMs), but it is unclear whether modulation occurs through changes entirely within the TMD or changes involving both the TMD and the extracellular domain (ECD)-TMD interface. In this study, we constructed multiple chimeras using the TMD of human α7nAChR and the ECD of a prokaryotic homolog, ELIC, which is not sensitive to these modulators, and for which a high resolution structure has been solved. Functional ELIC-α7nAChR (EA) chimeras were obtained when their ECD-TMD interfaces were modified to resemble either the ELIC interface (EAELIC) or α7nAChR interface (EAα7). Both EAα7 and EAELIC show similar activation response and desensitization characteristics, but only EAα7 retained the unique pharmacology of α7nAChR evoked by PAMs, including potentiation by ivermectin, PNU-120596, and TQS, as well as activation by 4BP-TQS. This study suggests that PAM modulation through the TMD has a more stringent requirement at the ECD-TMD interface than agonist activation.

An internally modulated, thermostable, pH-sensitive Cys loop receptor from the hydrothermal vent worm Alvinella pompejana.

Cys loop receptors (CLRs) are commonly known as ligand-gated channels that transiently open upon binding of neurotransmitters to modify the membrane potential. However, a class of cation-selective bacterial homologues of CLRs have been found to open upon a sudden pH drop, suggesting further ligands and more functions of the homologues in prokaryotes. Here we report an anion-selective CLR from the hydrothermal vent annelid worm Alvinella pompejana that opens at low pH. A. pompejana expressed sequence tag databases were explored by us, and two full-length CLR sequences were identified, synthesized, cloned, expressed in Xenopus oocytes, and studied by two-electrode voltage clamp. One channel, named Alv-a1-pHCl, yielded functional receptors and opened upon a sudden pH drop but not by other known agonists. Sequence comparison showed that both CLR proteins share conserved characteristics with eukaryotic CLRs, such as an N-terminal helix, a cysteine loop motif, and an intracellular loop intermediate in length between the long loops of other eukaryotic CLRs and those of prokaryotic CLRs. Both full-length Alv-a1-pHCl and a truncated form, termed tAlv-a1-pHCl, lacking 37 amino-terminal residues that precede the N-terminal helix, formed functional channels in oocytes. After pH activation, tAlv-a1-pHCl showed desensitization and was not modulated by ivermectin. In contrast, pH-activated, full-length Alv-a1-pHCl showed a marked rebound current and was modulated significantly by ivermectin. A thermostability assay indicated that purified tAlv-a1-pHCl expressed in Sf9 cells denatured at a higher temperature than the nicotinic acetylcholine receptor from Torpedo californica.

Differential regulation of GABAB receptor trafficking by different modes of N-methyl-D-aspartate (NMDA) receptor signaling.

Inhibitory GABAB receptors (GABABRs) can down-regulate most excitatory synapses in the CNS by reducing postsynaptic excitability. Functional GABABRs are heterodimers of GABAB1 and GABAB2 subunits and here we show that the trafficking and surface expression of GABABRs is differentially regulated by synaptic or pathophysiological activation of NMDA receptors (NMDARs). Activation of synaptic NMDARs using a chemLTP protocol increases GABABR recycling and surface expression. In contrast, excitotoxic global activation of synaptic and extrasynaptic NMDARs by bath application of NMDA causes the loss of surface GABABRs. Intriguingly, exposing neurons to extreme metabolic stress using oxygen/glucose deprivation (OGD) increases GABAB1 but decreases GABAB2 surface expression. The increase in surface GABAB1 involves enhanced recycling and is blocked by the NMDAR antagonist AP5. The decrease in surface GABAB2 is also blocked by AP5 and by inhibiting degradation pathways. These results indicate that NMDAR activity is critical in GABABR trafficking and function and that the individual subunits can be separately controlled to regulate neuronal responsiveness and survival.