Receptor Trafficking Research Articles

Regulated N-glycosylation controls chaperone function and receptor trafficking

One-fifth of human proteins are N-glycosylated in the endoplasmic reticulum (ER) by two oligosaccharyltransferases, OST-A and OST-B. Contrary to the prevailing view of N-glycosylation as a housekeeping function, we identified an ER pathway that modulates the activity of OST-A. Genetic analyses linked OST-A to HSP90B1, an ER chaperone for membrane receptors, and CCDC134, an ER luminal protein. During its translocation into the ER, an N-terminal peptide in HSP90B1 templates the assembly of a translocon complex containing CCDC134 and OST-A that protects HSP90B1 during folding, preventing its hyperglycosylation and degradation. Disruption of this pathway impairs WNT and IGF1R signaling and causes the bone developmental disorder osteogenesis imperfecta. Thus, N-glycosylation can be regulated by specificity factors in the ER to control cell surface receptor signaling and tissue development.

Glutamate signaling and neuroligin/neurexin adhesion play opposing roles that are mediated by major histocompatibility complex I molecules in cortical synapse formation.

Although neurons release neurotransmitter before contact, the role for this release in synapse formation remains unclear. Cortical synapses do not require synaptic vesicle release for formation (Verhage et al., 2000; Sando et al., 2017; Sigler et al., 2017; Held et al., 2020), yet glutamate clearly regulates glutamate receptor trafficking (Roche et al., 2001; Nong et al., 2004) and induces spine formation (Engert and Bonhoeffer, 1999; Maletic-Savatic et al., 1999; Toni et al., 1999; Kwon and Sabatini, 2011; Oh et al., 2016). Using rat and murine culture systems to dissect molecular mechanisms, we found that glutamate rapidly decreases synapse density specifically in young cortical neurons in a local and calcium-dependent manner through decreasing N-methyl-D-aspartate receptor (NMDAR) transport and surface expression as well as co-transport with neuroligin (NL1). Adhesion between NL1 and neurexin 1 protects against this glutamate-induced synapse loss. Major histocompatibility I (MHCI) molecules are required for the effects of glutamate in causing synapse loss through negatively regulating NL1 levels in both sexes. Thus, like acetylcholine at the neuromuscular junction (NMJ), glutamate acts as a dispersal signal for NMDARs and causes rapid synapse loss unless opposed by NL1-mediated trans-synaptic adhesion. Together, glutamate, MHCI and NL1 mediate a novel form of homeostatic plasticity in young neurons that induces rapid changes in NMDARs to regulate when and where nascent glutamatergic synapses are formed.Significance Statement The role for neurotransmitter release in synaptogenesis in the central nervous system (CNS) remains unclear. Here, we reconcile conflicting results in the field by showing that glutamate plays an important role in synapse formation by acting as a dispersal signal for NMDARs that is counteracted by trans-synaptic adhesion in intact tissue, similar to the role for neurotransmitter at the NMJ. We also describe a novel form of homeostatic plasticity in young neurons that allows them to respond to changes in activity through surprisingly rapid changes in synapse density. Finally, we show that this plasticity is modulated by immune proteins-MHCI molecules-through negative regulation of NL1 levels, connecting two important synaptic signaling pathways for the first time.

First person – Vikas Tillu

ABSTRACT First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping researchers promote themselves alongside their papers. Vikas Tillu is co-first author on ‘ Precision in situ cryogenic correlative light and electron microscopy of optogenetically positioned organelles’, published in JCS. Vikas is a Postdoctoral Researcher in the lab of Nicholas Ariotti at Institute for Molecular Bioscience, The University of Queensland, Australia, investigating in situ receptor trafficking analysis using cryo-electron tomography (cryo-ET).

Ex vivo nanoscale abluminal mapping of putative cargo receptors at the blood-brain barrier of expanded brain capillaries

Receptor mediated transport of therapeutic antibodies through the blood-brain barrier (BBB) give promise for drug delivery to alleviate brain diseases. We developed a low-cost method to obtain nanoscale localization data of putative cargo receptors. We combine existing ex vivo isolation methods with expansion microscopy (ExM) to analyze receptor localizations in brain microcapillaries. Using this approach, we show how to analyze receptor localizations in endothelial cells of brain microcapillaries in relation to the abluminal marker collagen IV. By choosing the thinnest capillaries, microcapillaries for analysis, we ensure the validity of collagen IV as an abluminal marker. With this tool, we confirm transferrin receptors as well as sortilin to be both luminally and abluminally localized. Furthermore, we identify basigin to be an abluminal receptor. Our methodology can be adapted to analyze different types of isolated brain capillaries and we anticipate that this approach will be very useful for the research community to gain new insight into cargo receptor trafficking in the slim brain endothelial cells to elucidate novel paths for future drug design.

Discovery of Antinociceptive α9α10 Nicotinic Acetylcholine Receptor Antagonists by Stable Receptor Expression.

Chronic neuropathic pain is an increasingly prevalent societal issue that responds poorly to existing therapeutic strategies. The α9α10 nicotinic acetylcholine receptor (nAChR) has emerged as a potential target to treat neuropathic pain. However, challenges in expressing functional α9α10 nAChRs in mammalian cell lines have slowed the discovery of α9α10 ligands and studies into the relationship between α9α10 nAChRs and neuropathic pain. Here, we develop a cell line in the HEK293 background that stably expresses functional α9α10 nAChRs. By also developing cell lines expressing only α9 and α10 subunits, we identify distinct receptor pharmacology between homomeric α9 or α10 and heteromeric α9α10 nAChRs. Moreover, we demonstrate that incubation with nAChR ligands differentially regulates the expression of α9- or α10-containing nAChRs, suggesting a possible mechanism by which ligands may modify receptor composition and trafficking in α9- and α10-expressing cells. We then apply our α9α10 cell line in a screen of FDA-approved and investigational drugs to identify α9α10 ligands that provide new tools to probe α9α10 nAChR function. We demonstrate that one compound from this screen, diphenidol, possesses antinociceptive activity in a murine model of neuropathic pain. These results expand our understanding of α9α10 receptor pharmacology and provide new starting points for developing efficacious neuropathic pain treatments.

Measuring Calcium-Sensing Receptor Agonist-Driven Insertional Signaling (ADIS) and Trafficking by TIRF Microscopy.

The calcium-sensing receptor (CaSR), which regulates parathyroid hormone secretion by sensing serum calcium concentrations, has developed unique trafficking mechanisms to respond to constant exposure to its orthosteric ligand calcium. CaSR rapidly responds to fluctuations in serum calcium by driving forward trafficking of the receptor to cell surfaces in a mechanism known as agonist-driven insertional signaling (ADIS). This increase in CaSR at cell surfaces is counterbalanced by both constitutive and agonist-driven internalization of the receptor. Deciphering these mechanisms is important to understand how mutations in the CaSR and components of its signaling and trafficking pathways cause human disorders of calcium homeostasis.This chapter describes a protocol to measure CaSR ADIS and endocytosis in parallel using total internal reflection fluorescence (TIRF) microscopy. This utilizes a mammalian expression construct comprising the full-length human CaSR with an N-terminal bungarotoxin minimal-binding site that can be labeled with commercially available fluorescent ligands to measure endocytosis, and a super-ecliptic pHluorin (SEP) to measure total cell surface expression and exocytic events. This protocol could easily be adapted to simultaneously assess forward trafficking and endocytosis of other membrane proteins by TIRF microscopy.

Exploring the role of AMPA receptor auxiliary proteins in synaptic functions and diseases.

α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) ionotropic glutamate receptors (AMPARs) mediate rapid excitatory synaptic transmission in the mammalian brain, primarily driven by the neurotransmitter glutamate. The modulation of AMPAR activity, particularly calcium-permeable AMPARs (CP-AMPARs), is crucially influenced by various auxiliary subunits. These subunits are integral membrane proteins that bind to the receptor's core and modify its functional properties, including ion channel kinetics and receptor trafficking. This review comprehensively catalogs all known AMPAR auxiliary proteins, providing vital insights into the biochemical mechanisms governing synaptic modulation and the specific impact of CP-AMPARs compared to their calcium-impermeable AMPA receptor (CI-AMPARs). Understanding the complex interplay between AMPARs and their auxiliary subunits in different brain regions is essential for elucidating their roles in cognitive functions such as learning and memory. Importantly, alterations in these auxiliary proteins' expression, function or interactions have been implicated in various neurological disorders. Aberrant signaling through CP-AMPARs, in particular, is associated with severe synaptic dysfunctions across neurodevelopmental, neurodegenerative and psychiatric conditions. Targeting the distinct properties of AMPAR-auxiliary subunit complexes, especially those involving CP-AMPARs, could disclose new therapeutic strategies, potentially allowing for more precise interventions in treating complex neuronal disorders.

Epidermal Growth Factor Intralesional Delivery in Chronic Wounds: The Pioneer and Standalone Technique for Reversing Wound Chronicity and Promoting Sustainable Healing.

The early expectations about growth factors' (GFs') discovery as an undisputed therapeutic solution for chronic wounds progressively eclipsed when they failed to accelerate acute wound closure and restore the healing trajectory of stagnant ulcers. Critical knowledge about chronic wound biology and GF pharmacology was a conundrum at that time. Diabetes undermines keratinocytes' and fibroblasts' physiology, impairing skin healing abilities. Diabetic ulcers, as other chronic wounds, are characterized by hyperinflammation, unbalanced proteolytic activity, catabolism, and free radical cytotoxicity. This hostile scenario for the chemical stability, integrity, and functionality of GFs led to the conclusion that topical administration may jeopardize GFs' clinical effectiveness. Epidermal growth factor (EGF) has a proximal position in tissues homeostasis by activating survival and mitogenic pathways from embryonic life to adulthood. Seminal experiments disclosed unprecedented pharmacological bounties of parenterally administered EGF. Accordingly, the experience accumulated for more than 20 years of EGF intralesional infiltration of diabetic wound bottoms and edges has translated into sustained healing responses, such as low recurrences and amputation rates. This delivery route, in addition to being safe and tolerated, has shown to restore a variety of circulating biochemical markers ordinarily disturbed in diabetic conditions. EGF infiltration triggers a cascade of local fibroblast reactions, supporting its molecular integrity, prolonged mean residence time, and ultimately eliciting its receptor trafficking and nuclear translocation. The intralesional delivery route seems to warrant that EGF reaches wound fibroblasts' epigenetic core, mitigating the consequences of metabolic memory imprinting.

Redox Signaling in Endosomes Using the Example of EGF Receptors: A Graphical Review.

Early endosomes represent first-line sorting compartments or even organelles for internalized molecules. They enable the transport of molecules or ligands to other compartments of the cell, such as lysosomes, for degradation or recycle them back to the membrane by various mechanisms. Moreover, early endosomes function as signaling and scaffolding platforms to initiate or prolong distinct signaling pathways. Accordingly, early endosomes have to be recognized as either part of a degradation or recycling pathway. The physical proximity of many ligand-binding receptors with other membrane-bound proteins or complexes such as NADPH oxidases may result in an interaction of second messengers, like reactive oxygen species (ROS) and early endosomes, that promote the correct recognition of individual early endosomes. In fact, redoxosomes comprise an endosomal subsection of signaling endosomes. One example of such potential interaction is epidermal growth factor receptor (EGFR) signaling. Here we summarize recent findings on EGFR signaling as a well-studied example for receptor trafficking and trans-activation and illustrate the interplay between cellular and endosomal ROS.

International Union of Basic and Clinical Pharmacology. CXVIII. Update on the Nomenclature for Atypical Chemokine Receptors including ACKR5.

Chemokines signal through classical G protein-coupled receptors (GPCRs) to induce cell migration during development, immune homeostasis and multiple diseases. Over the last decade a subfamily of atypical chemokine receptors (ACKRs) was delineated from GPCRs based on their inability to trigger conventional G protein signaling or mediate cell migration in response to chemokines. These receptors nevertheless play an important role within the chemokine system by sequestering, transporting or internalizing chemokines thereby regulating their availability and shaping their gradients. GPR182, the recently deorphanized chemokine receptor, shares about 30% of sequence similarity with its closest relative ACKR3. GPR182 is mainly expressed on endothelial cells and was proposed to act as a scavenger regulating the availability of a large set of chemokines from the CXC, CC and XC families and to act cooperatively with ACKR3 and ACKR4. Unlike other ACKRs, GPR182 was shown to have a strong constitutive interaction with β-arrestins that is required for intracellular receptor trafficking and chemokine scavenging. Chemokine ligation of GPR182 has no additional detectable impact on β-arrestin recruitment. Genetic ablation of GPR182 affects spleen size, myelopoiesis, and serum chemokine levels, indicating its role in chemokine homeostasis and immune regulation. GPR182 was also reported to regulate immune responses to bloodborne antigens and tumorigenesis. Taken together, compelling cumulative evidence indicates that GPR182 does not trigger G protein-mediated signaling but acts as a scavenger for chemokines in vitro and in vivo strongly supporting its inclusion as ACKR5 in the systematic nomenclature of chemokine receptors. Significance Statement The summarized presented findings strongly support the designation of GPR182 as ACKR5 and its formal inclusion in the family of atypical chemokine receptors.

Postsynaptic Density Proteins and Their Role in the Trafficking of Group I Metabotropic Glutamate Receptors.

Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system that regulates multiple different forms of synaptic plasticity, including learning and memory. Glutamate transduces its signal by activating ionotropic glutamate receptors and metabotropic glutamate receptors (mGluRs). Group I mGluRs belong to the G protein-coupled receptor (GPCR) family. Regulation of cell surface expression and trafficking of the glutamate receptors represents an important mechanism that assures proper transmission of information at the synapses. There is growing evidence implicating dysregulated glutamate receptor trafficking in the pathophysiology of several neuropsychiatric disorders. The postsynaptic density (PSD) region consists of many specialized proteins which are assembled beneath the postsynaptic membrane of dendritic spines. Many of these proteins interact with group I mGluRs and have essential roles in group I mGluR-mediated synaptic function and plasticity. This review provides up-to-date information on the molecular determinants regulating cell surface expression and trafficking of group I mGluRs and discusses the role of few of these PSD proteins in these processes. As substantial evidences link mGluR dysfunction and maladaptive functioning of many PSD proteins to the pathophysiology of various neuropsychiatric disorders, understanding the role of the PSD proteins in group I mGluR trafficking may provide opportunities for the development of novel therapeutics in multiple neuropsychiatric disorders.

7096 Variants in WASHC3, a component of the WASH complex, cause distinctive facial dysmorphism, short stature, and variable neurodevelopmental abnormalities

Abstract Disclosure: Y. Jee: None. J. Lui: None. D. Marafi: None. Z. Xia: None. R. Bhatia: None. E. Zhou: None. I. Herman: None. A.G. Temnycky: None. P. Whalen: None. G. Elliot: None. E. Leschek: None. R. Wijngaard: None. R. Beek: None. A. Vreugd: None. M. De Vries: None. C. Karnebeek: None. M. Oud: None. T. Markello: None. K. Barnes: None. H. Alrohaif: None. H.H. Freeze: None. W. Gahl: None. M. Malicdan: None. J. Posey: None. J. Lupski: None. J. Baron: None. Genetic defects that impair growth plate chondrogenesis can result in a spectrum of phenotypes, ranging from skeletal dysplasia to mild short stature, sometimes accompanied by syndromic features. However, the genetic underpinnings of impaired skeletal growth often remain elusive. In our study, we investigated three unrelated families with short stature (height standard deviation score z < -2), distinctive facies (down-slanting eyes, low set ears, high arched palate, and short neck), and various neurodevelopmental abnormalities (from leaning difficulties to low intelligence quotient). In these families, exome sequencing (ES) identified rare, predicted-pathogenic variant alleles in WASHC3 that segregated with the phenotype. WASHC3 is a component of the WASH complex, which is involved in endosome receptor recycling. In the first family, a de novo-dominant missense variant (p.L69F) impaired WASHC3 participation in the WASH complex. This variant led to altered PTH1R endosomal trafficking, resulting in diminished PTH1R signaling and impairing growth plate chondrocyte hypertrophic differentiation, providing a likely explanation for the short stature. Knockdown of other WASH complex components (WASH1, SWIP, Strumpellin) also diminished PTH1R signaling, confirming the importance of an intact WASH complex. In the second and third families, a homozygous variant in the start codon (p.M1?) markedly reduced WASHC3 protein expression by approximately 75%. In conclusion, our findings strongly suggest that the WASH complex is required for normal skeletal growth and that, consequently, genetic abnormalities impairing the function of the WASH complex (WASHopathy) cause short stature as well as distinctive facies and variable neurodevelopmental abnormalities. These effects are likely mediated through disruptions in endosomal receptor trafficking. This new pathological information sheds light on the molecular basis of impaired longitudinal bone growth. Presentation: 6/3/2024

Haploinsufficiency of Syngap1 in striatal indirect pathway neurons alters motor and goal-directed behaviors in mice.

SYNGAP1 is a high-confidence autism spectrum disorder (ASD) risk gene and mutations in SYNGAP1 lead to a neurodevelopmental disorder (NDD) that presents with epilepsy, ASD, motor developmental delay, and intellectual disability. SYNGAP1 codes for Ras/Rap GTP-ase activating protein SynGAP (SynGAP). In mice, SynGAP is located in the postsynaptic density of glutamatergic synapses and regulates glutamate receptor trafficking in an activity-dependent manner. In addition to forebrain glutamatergic neurons, Syngap1 is highly expressed in the striatum, although the functions of SynGAP in the striatum have not been extensively studied. Here we show that Syngap1 is expressed in both direct and indirect pathway striatal projection neurons (dSPNs and iSPNs) in mice of both sexes. In a mouse model of Syngap1 haploinsufficiency, dendritic spine density, morphology, and intrinsic excitability are altered primarily in iSPNs, but not dSPNs. At the behavioral level, SynGAP reduction alters striatal-dependent motor learning and goal-directed behavior. Several behavioral phenotypes are reproduced by iSPN-specific Syngap1 reduction and, in turn, prevented by iSPN-specific Syngap1 rescue. These results establish the importance of SynGAP to striatal neuron function and pinpoint the indirect pathway as a key circuit in the neurobiology of SYNGAP1-related NDD.Significance statement SYNGAP1 mutations cause a neurodevelopmental disorder presenting with intellectual disability, motor problems, epilepsy, autism spectrum disorder, and a constellation of other behavioral and psychiatric conditions. SynGAP protein is highly expressed in the striatum but its functions in this brain region have not yet been explored. This study shows that loss of one copy of the Syngap1 gene from striatal indirect, but not direct, pathway neurons alters synaptic properties, cellular excitability, motor behaviors, and goal-directed responding in mice. This work provides a new perspective on the functions of SynGAP and suggests that altered activity in striatal circuits may be an important driver of the motor and learning alterations in people with SYNGAP1 disorder.

AMPA Receptors and Neuronal Plasticity

Interneuronal information transfer occurs at synapses, where AMPA receptors play a key role. With regard to physiological function, synaptic trafficking of AMPA receptors underlies memory, learning and experience. Analysis of animal models of disease and postmortem brains of patients has revealed that abnormal expression and functions of AMPA receptors may trigger various neuropsychiatric disorders. Such findings are currently being used for the development of therapeutic drugs through quantification of AMPA receptors in patients' brains in real-world practice.

Designed endocytosis-inducing proteins degrade targets and amplify signals.

Endocytosis and lysosomal trafficking of cell surface receptors can be triggered by endogenous ligands. Therapeutic approaches such as lysosome-targeting chimaeras1,2 (LYTACs) and cytokine receptor-targeting chimeras3 (KineTACs) have used this to target specific proteins for degradation by fusing modified native ligands to target binding proteins. Although powerful, these approaches can be limited by competition with native ligands and requirements for chemical modification that limit genetic encodability and can complicate manufacturing, and, more generally, there may be no native ligands that stimulate endocytosis through a given receptor. Here we describe computational design approaches for endocytosis-triggering binding proteins (EndoTags) that overcome these challenges. We present EndoTags for insulin-like growth factor 2 receptor (IGF2R) and asialoglycoprotein receptor (ASGPR), sortilin and transferrin receptors, and show that fusing these tags to soluble or transmembrane target protein binders leads to lysosomal trafficking and target degradation. As these receptors have different tissue distributions, the different EndoTags could enable targeting of degradation to different tissues. EndoTag fusion to a PD-L1 antibody considerably increases efficacy in a mouse tumour model compared to antibody alone. The modularity and genetic encodability of EndoTags enables AND gate control for higher-specificitytargeted degradation, and the localized secretion of degraders from engineered cells. By promoting endocytosis, EndoTag fusion increases signalling through an engineered ligand-receptor system by nearly 100-fold. EndoTags have considerable therapeutic potential as targeted degradation inducers, signalling activators for endocytosis-dependent pathways, and cellular uptake inducers for targeted antibody-drug and antibody-RNA conjugates.

Involvement of sphingosine 1-phosphate signaling in insulin-like growth factor-II/mannose 6-phosphate receptor trafficking from endosome to the trans-Golgi network

The insulin-like growth factor II/mannose 6-phosphate (IGF-II/M6P) receptor is a multifunctional glycoprotein not only play roles in IGF-II degradation and pro-TGFβ activation but binding to and transport M6P-bearing lysosomal enzymes from the trans-Golgi network (TGN) or the cell surface to lysosomes. At present, information regarding a retrograde transport of IGF-II/M6P receptor from endosomes to the TGN is still limited. We show here that a continuous ligand-dependent activation of sphingosine 1-phosphate receptor type 3 (S1P3R) on the endosomal membranes is required for subsequent recycling back of cargo-unloaded IGF-II/M6P receptors to the TGN. We have further clarified that Gq coupled with S1P3R plays a critical role in the activation of casein kinase 2, which phosphorylates and keeps PACS1 connector protein active for the association with IGF-II/M6P receptors, which enables transport carrier formation with the aid of other adaptor proteins toward the TGN. These findings shed light on the molecular mechanism underlying how continuous activation of the S1P receptor and subsequent downstream Gq signaling regulates the retrograde transport of the empty IGF-II/M6P receptors back to the TGN.

The SORL1 p.Y1816C variant causes impaired endosomal dimerization and autosomal dominant Alzheimer’s disease

Truncating genetic variants of SORL1, encoding the endosome recycling receptor SORLA, have been accepted as causal of Alzheimer's disease (AD). However, most genetic variants observed in SORL1 are missense variants, for which it is complicated to determine the pathogenicity level because carriers come from pedigrees too small to be informative for penetrance estimations. Here, we describe three unrelated families in which the SORL1 coding missense variant rs772677709, that leads to a p.Y1816C substitution, segregates with Alzheimer's disease. Further, we investigate the effect of SORLA p.Y1816C on receptor maturation, cellular localization, and trafficking in cell-based assays. Under physiological circumstances, SORLA dimerizes within the endosome, allowing retromer-dependent trafficking from the endosome to the cell surface, where the luminal part is shed into the extracellular space (sSORLA). Our results showed that the p.Y1816C mutant impairs SORLA homodimerization in the endosome, leading to decreased trafficking to the cell surface and less sSORLA shedding. These trafficking defects of the mutant receptor can be rescued by the expression of the SORLA 3Fn-minireceptor. Finally, we find that iPSC-derived neurons with the engineered p.Y1816C mutation have enlarged endosomes, a defining cytopathology of AD. Our studies provide genetic as well as functional evidence that the SORL1 p.Y1816C variant is causal for AD. The partial penetrance of the mutation suggests this mutation should be considered in clinical genetic screening of multiplex early-onset AD families.

An engineered trafficking biosensor reveals a role for DNAJC13 in DOR downregulation.

Trafficking of G protein-coupled receptors (GPCRs) through the endosomal-lysosomal pathway is critical to homeostatic regulation of GPCRs following activation with agonist. Identifying the genes involved in GPCR trafficking is challenging due to the complexity of sorting operations and the large number of cellular proteins involved in the process. Here, we developed a high-sensitivity biosensor for GPCR expression and agonist-induced trafficking to the lysosome by leveraging the ability of the engineered peroxidase APEX2 to activate the fluorogenic substrate Amplex UltraRed (AUR). We used the GPCR-APEX2/AUR assay to perform a genome-wide CRISPR interference screen focused on identifying genes regulating expression and trafficking of the δ-opioid receptor (DOR). We identified 492 genes consisting of both known and new regulators of DOR function. We demonstrate that one new regulator, DNAJC13, controls trafficking of multiple GPCRs, including DOR, through the endosomal-lysosomal pathway by regulating the composition of the endosomal proteome and endosomal homeostasis.

Visual mismatch negativity in Parkinson's psychosis and potential for testing treatment mechanisms

Abstract Psychosis and visual hallucinations are a prevalent non-motor symptom of Parkinson's disease, negatively affecting patients’ quality of life and constituting a greater risk for dementia. Understanding neural mechanisms associated to these symptoms is instrumental for treatment development. The mismatch negativity is an event-related potential evoked by a violation in a sequence of sensory events. It is widely considered an index of sensory change-detection. Reduced mismatch negativity response is one of the most replicated results in schizophrenia and has been suggested to be a superior psychosis marker. To understand whether this event-related potential component could be a similarly robust marker for Parkinson's psychosis, we used electroencephalography with a change-detection task to study the mismatch negativity in the visual modality in 20 participants with Parkinson's and visual hallucinations and 18 matched Parkinson's participants without hallucinations. We find that visual mismatch negativity is clearly present in participants with Parkinson’s disease without hallucinations at both parieto-occipital and frontal sites, whereas participants with Parkinson's and visual hallucinations show reduced or no differences in the two waveforms, confirming the sensitivity of mismatch negativity to psychosis, even within the same diagnostic group. We also explored the relationship between hallucination severity and visual mismatch negativity amplitude, finding a negative correlation between visual hallucinations severity scores and visual mismatch negativity amplitude at a central frontal and a parieto-occipital electrodes, whereby the more severe or complex (illusions, formed visual hallucinations) the symptoms the smaller the amplitude. We have also tested the potential role of the serotonergic 5-HT2A cascade in visual hallucinations in Parkinson's with these symptoms, following the receptor trafficking hypothesis. We did so with a pilot study in healthy controls (N = 18) providing support for the role of the Gi/o-dependent pathway in the psychedelic effect and a case series in participants with Parkinson's and visual hallucinations (N = 5) using a double-blind crossover design. Positive results on psychosis scores and mismatch amplitude add further to the potential role of serotonergic modulation of visual hallucinations in Parkinson's disease.

SARM1 is essential for NMDA receptor-dependent endocytosis of AMPA receptors in hippocampal neurons

Long-term depression (LTD) is a form of synaptic plasticity thought to be the cellular basis of experience-dependent learning and memory. LTD is caused by an activity-dependent decrease in cell surface α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptors (AMPA receptors) at the postsynaptic sites. However, the mechanism through which AMPA receptors are removed from the cell surface via neuronal activity is not fully understood. In this study, we showed that small interfering RNA (siRNA)-mediated knockdown of sterile alpha and toll/interleukin receptor motif containing 1 (SARM1) in cultured hippocampal neurons prevented the N-methyl-d-aspartate (NMDA)-induced reduction in cell surface AMPA receptors. However, the control RNA did not affect NMDA-mediated AMPA receptor trafficking. Overexpression of the siRNA-resistant form of SARM1 in SARM1-knocked-down neurons restored AMPA receptor trafficking. However, overexpression of SARM1, which lacks the mitochondrial transport signal, in the SARM1-knocked-down neurons did not restore NMDA-dependent AMPA receptor endocytosis. Moreover, the inhibition of the NADase activity of SARM1 blocked the NMDA-induced reduction of cell surface AMPA receptors. These results suggest that both the mitochondrial localization and NADase activity of SARM1 are essential for NMDA receptor-dependent AMPA receptor internalization in the hippocampal neurons.