Clinical features, functional consequences, and rescue pharmacology of missense GRID1 and GRID2 human variants.

GRID1 and GRID2 encode the enigmatic GluD1 and GluD2 proteins, which form tetrameric receptors that play important roles in synapse organization and development of the central nervous system. Variation in these genes has been implicated in neurodevelopmental phenotypes. We evaluated GRID1 and GRID2 human variants from the literature, ClinVar, and clinical laboratories and found that many of these variants reside in intolerant domains, including the amino terminal domain of both GRID1 and GRID2. Other conserved regions, such as the M3 transmembrane domain, show different intolerance between GRID1 and GRID2. We introduced these variants into GluD1 and GluD2 cDNA and performed electrophysiological and biochemical assays to investigate the mechanisms of dysfunction of GRID1/2 variants. One variant in the GRID1 distal amino terminal domain resides at a position predicted to interact with Cbln2/Cbln4, and the variant disrupts complex formation between GluD1 and Cbln2, which could perturb its role in synapse organization. We also discovered that, like the lurcher mutation (GluD2-A654T), other rare variants in the GRID2 M3 domain create constitutively active receptors that share similar pathogenic phenotypes. We also found that the SCHEMA schizophrenia M3 variant GluD1-A650T produced constitutively active receptors. We tested a variety of compounds for their ability to inhibit constitutive currents of GluD receptor variants and found that pentamidine potently inhibited GluD2-T649A constitutive channels (IC50 50 nM). These results identify regions of intolerance to variation in the GRID genes, illustrate the functional consequences of GRID1 and GRID2 variants, and suggest how these receptors function normally and in disease.

Classification of missense variants in the N-methyl-d-aspartate receptor GRIN gene family as gain- or loss-of-function.

Advances in sequencing technology have generated a large amount of genetic data from patients with neurological conditions. These data have provided diagnosis of many rare diseases, including a number of pathogenic de novo missense variants in GRIN genes encoding N-methyl-d-aspartate receptors (NMDARs). To understand the ramifications for neurons and brain circuits affected by rare patient variants, functional analysis of the variant receptor is necessary in model systems. For NMDARs, this functional analysis needs to assess multiple properties in order to understand how variants could impact receptor function in neurons. One can then use these data to determine whether the overall actions will increase or decrease NMDAR-mediated charge transfer. Here, we describe an analytical and comprehensive framework by which to categorize GRIN variants as either gain-of-function (GoF) or loss-of-function (LoF) and apply this approach to GRIN2B variants identified in patients and the general population. This framework draws on results from six different assays that assess the impact of the variant on NMDAR sensitivity to agonists and endogenous modulators, trafficking to the plasma membrane, response time course and channel open probability. We propose to integrate data from multiple in vitro assays to arrive at a variant classification, and suggest threshold levels that guide confidence. The data supporting GoF and LoF determination are essential to assessing pathogenicity and patient stratification for clinical trials as personalized pharmacological and genetic agents that can enhance or reduce receptor function are advanced. This approach to functional variant classification can generalize to other disorders associated with missense variants.

De novo GRIN variants in M3 helix associated with neurological disorders control channel gating of NMDA receptor.

N-methyl-D-aspartate receptors (NMDARs) are members of the glutamate receptor family and participate in excitatory postsynaptic transmission throughout the central nervous system. Genetic variants in GRIN genes encoding NMDAR subunits are associated with a spectrum of neurological disorders. The M3 transmembrane helices of the NMDAR couple directly to the agonist-binding domains and form a helical bundle crossing in the closed receptors that occludes the pore. The M3 functions as a transduction element whose conformational change couples ligand binding to opening of an ion conducting pore. In this study, we report the functional consequences of 48 de novo missense variants in GRIN1, GRIN2A, and GRIN2B that alter residues in the M3 transmembrane helix. These de novo variants were identified in children with neurological and neuropsychiatric disorders including epilepsy, developmental delay, intellectual disability, hypotonia and attention deficit hyperactivity disorder. All 48 variants in M3 for which comprehensive testing was completed produce a gain-of-function (28/48) compared to loss-of-function (9/48); 11 variants had an indeterminant phenotype. This supports the idea that a key structural feature of the M3 gate exists to stabilize the closed state so that agonist binding can drive channel opening. Given that most M3 variants enhance channel gating, we assessed the potency of FDA-approved NMDAR channel blockers on these variant receptors. These data provide new insight into the structure-function relationship of the NMDAR gate, and suggest that variants within the M3 transmembrane helix produce a gain-of-function.

Preclinical Evaluation of Azabenzimidazole-Based PET Radioligands for γ-8 Dependent Transmembrane AMPA Receptor Regulatory Protein Imaging.

AMPA glutamate receptors (AMPARs) play a pivotal role in excitatory neurotransmission, particularly in the hippocampus where the TARP γ-8 subunit is enriched and serves as a target for emerging anti-epileptic drugs. To enable in vivo visualization of TARP γ-8 distribution and expression by positron emission tomography (PET), this study focuses on the development of novel 18 F-labeled TARP γ-8 inhibitors and their corresponding precursors, stemming from the azabenzimidazole scaffold. The resulting radioligands [18 F]TARP-2204 and [18 F]TARP-2205 were successfully synthesized with acceptable radiochemical yield, high molar activity, and excellent radiochemical purity. In vitro autoradiography demonstrates high level of specific binding of [18 F]TARP-2205 to TARP γ-8 in both rat and nonhuman primate brain tissues. However, unexpected radiodefluorination in PET imaging studies of rodents emphasizes the need for further structural refinement. This work serves as an excellent starting point for the development of future 18 F-labeled TARP γ-8 PET tracers, offering valuable insights into medicinal chemistry design, radiosynthesis and subsequent PET evaluation.

Expression profiling and function analysis identified new genes regulating cumulus expansion and apoptosis.

Studies on the mechanisms behind cumulus expansion and cumulus cell (CC) apoptosis are essential for understanding the mechanisms for oocyte maturation. Genes expressed in CCs might be used as markers for competent oocytes and/or embryos. In this study, both in vitro (IVT) and in vivo (IVO) mouse oocyte models with significant difference in cumulus expansion and CC apoptosis were used to identify and validate new genes regulating cumulus expansion and CC apoptosis of mouse oocytes. We first performed mRNA sequencing and bioinformatic analysis using the IVT oocyte model to identify candidate genes. We then analyzed functions of the candidate genes by RNAi or gene overexpression to select the candidate cumulus expansion and CC apoptosis-regulating genes. Finally, we validated the cumulus expansion and CC apoptosis-regulating genes using the IVO oocyte model. The results showed that while Spp1, Sdc1, Ldlr, Ezr and Mmp2 promoted, Bmp2, Angpt2, Edn1, Itgb8, Cxcl10 and Agt inhibited cumulus expansion. Furthermore, Spp1, Sdc1 and Ldlr inhibited CC apoptosis. In conclusion, by using both IVT and IVO oocyte models, we have identified and validated a new group of cumulus expansion and/or apoptosis-regulating genes, which may be used for selection of quality oocytes/embryos and for elucidating the molecular mechanisms behind oocyte maturation.

Novel neuroactive steroids as positive allosteric modulators of NMDA receptors: mechanism, site of action, and rescue pharmacology on GRIN variants associated with neurological conditions.

N-methyl-D-aspartate receptors (NMDARs) play vital roles in normal brain functions (i.e., learning, memory, and neuronal development) and various neuropathological conditions, such as epilepsy, autism, Parkinson's disease, Alzheimer's disease, and traumatic brain injury. Endogenous neuroactive steroids such as 24(S)-hydroxycholesterol (24(S)-HC) have been shown to influence NMDAR activity, and positive allosteric modulators (PAMs) derived from 24(S)-hydroxycholesterol scaffold can also enhance NMDAR function. This study describes the structural determinants and mechanism of action for 24(S)-hydroxycholesterol and two novel synthetic analogs (SGE-550 and SGE-301) on NMDAR function. We also show that these agents can mitigate the altered function caused by a set of loss-of-function missense variants in NMDAR GluN subunit-encoding GRIN genes associated with neurological and neuropsychiatric disorders. We anticipate that the evaluation of novel neuroactive steroid NMDAR PAMs may catalyze the development of new treatment strategies for GRIN-related neuropsychiatric conditions.

Functional effects of disease-associated variants reveal that the S1-M1 linker of the NMDA receptor critically controls channel opening.

The short pre-M1 helix within the S1-M1 linker (also referred to as the pre-M1 linker) between the agonist-binding domain (ABD, S1) and the M1 transmembrane helix of the NMDA receptor (NMDAR) is devoid of missense variants within the healthy population but is a locus for de novo pathogenic variants associated with neurological disorders. Several de novo variants within this helix have been identified in patients presenting early in life with intellectual disability, developmental delay, and/or epilepsy. In this study, we evaluated functional properties for twenty variants within the pre-M1 linker in GRIN1, GRIN2A, and GRIN2B genes, including six novel missense variants. The effects of pre-M1 variants on agonist potency, sensitivity to endogenous allosteric modulators, response time course, channel open probability, and surface expression were assessed. Our data indicated that virtually all of the variants evaluated altered channel function, and multiple variants had profound functional consequences, which may contribute to the neurological conditions in the patients harboring the variants in this region. These data strongly suggest that the residues within the pre-M1 helix play a key role in channel gating and are highly intolerant to genetic variation.

Clinical and functional consequences of GRIA variants in patients with neurological diseases.

AMPA receptors are members of the glutamate receptor family and mediate a fast component of excitatory synaptic transmission at virtually all central synapses. Thus, their functional characteristics are a critical determinant of brain function. We evaluate intolerance of each GRIA gene to genetic variation using 3DMTR and report here the functional consequences of 52 missense variants in GRIA1-4 identified in patients with various neurological disorders. These variants produce changes in agonist EC50, response time course, desensitization, and/or receptor surface expression. We predict that these functional and localization changes will have important consequences for circuit function, and therefore likely contribute to the patients' clinical phenotype. We evaluated the sensitivity of variant receptors to AMPAR-selective modulators including FDA-approved drugs to explore potential targeted therapeutic options.

Role and action mechanisms of tPA in CRH-induced apoptosis of mouse oviductal epithelial and mural granulosa cells.

Understanding how stress hormones induce apoptosis in oviductal epithelial cells (OECs) and mural granulosa cells (MGCs) can reveal the mechanisms by which female stress impairs embryonic development and oocyte competence. A recent study showed that tissue plasminogen activator (tPA) ameliorates corticosterone-induced apoptosis in MGCs and OECs by acting on its receptors low-density lipoprotein receptor-related protein 1 (LRP1) and Annexin A2 (ANXA2), respectively. However, whether tPA is involved in corticotropin-releasing hormone (CRH)-induced apoptosis and whether it uses the same or different receptors to inhibit apoptosis induced by different hormones in the same cell type remains unknown. This study showed that CRH triggered apoptosis in both OECs and MGCs and significantly downregulated tPA expression. Moreover, tPA inhibits CRH-induced apoptosis by acting on ANXA2 in both OECs and MGCs. While ANXA2 inhibits apoptosis via phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling, LRP1 reduces apoptosis via mitogen-activated protein kinase (MAPK) signaling. Thus, tPA used the same receptor to inhibit CRH-induced apoptosis in both OECs and MGCs, however used different receptors to inhibit corticosterone-induced apoptosis in MGCs and OECs. These data helps understand the mechanism by which female stress impairs embryo/oocyte competence and proapoptotic factors trigger apoptosis in different cell types.

Structure, Function, and Pharmacology of Glutamate Receptor Ion Channels.

Many physiologic effects of l-glutamate, the major excitatory neurotransmitter in the mammalian central nervous system, are mediated via signaling by ionotropic glutamate receptors (iGluRs). These ligand-gated ion channels are critical to brain function and are centrally implicated in numerous psychiatric and neurologic disorders. There are different classes of iGluRs with a variety of receptor subtypes in each class that play distinct roles in neuronal functions. The diversity in iGluR subtypes, with their unique functional properties and physiologic roles, has motivated a large number of studies. Our understanding of receptor subtypes has advanced considerably since the first iGluR subunit gene was cloned in 1989, and the research focus has expanded to encompass facets of biology that have been recently discovered and to exploit experimental paradigms made possible by technological advances. Here, we review insights from more than 3 decades of iGluR studies with an emphasis on the progress that has occurred in the past decade. We cover structure, function, pharmacology, roles in neurophysiology, and therapeutic implications for all classes of receptors assembled from the subunits encoded by the 18 ionotropic glutamate receptor genes. SIGNIFICANCE STATEMENT: Glutamate receptors play important roles in virtually all aspects of brain function and are either involved in mediating some clinical features of neurological disease or represent a therapeutic target for treatment. Therefore, understanding the structure, function, and pharmacology of this class of receptors will advance our understanding of many aspects of brain function at molecular, cellular, and system levels and provide new opportunities to treat patients.

Channel-Blind Joint Source-Channel Coding for Wireless Image Transmission.

Joint source-channel coding (JSCC) based on deep learning has shown significant advancements in image transmission tasks. However, previous channel-adaptive JSCC methods often rely on the signal-to-noise ratio (SNR) of the current channel for encoding, which overlooks the neural network's self-adaptive capability across varying SNRs. This paper investigates the self-adaptive capability of deep learning-based JSCC models to dynamically changing channels and introduces a novel method named Channel-Blind JSCC (CBJSCC). CBJSCC leverages the intrinsic learning capability of neural networks to self-adapt to dynamic channels and diverse SNRs without relying on external SNR information. This approach is advantageous, as it is not affected by channel estimation errors and can be applied to one-to-many wireless communication scenarios. To enhance the performance of JSCC tasks, the CBJSCC model employs a specially designed encoder-decoder. Experimental results show that CBJSCC outperforms existing channel-adaptive JSCC methods that depend on SNR estimation and feedback, both in additive white Gaussian noise environments and under slow Rayleigh fading channel conditions. Through a comprehensive analysis of the model's performance, we further validate the robustness and adaptability of this strategy across different application scenarios, with the experimental results providing strong evidence to support this claim.

Differential responses of disease-related GRIN variants located in pore-forming M2 domain of N-methyl-D-aspartate receptor to FDA-approved inhibitors.

N-methyl-D-aspartate receptors (NMDAR), ionotropic glutamate receptors, mediate a slow component of excitatory synaptic transmission in the central nervous system and play a key role in normal brain function and development. Genetic variations in GRIN genes encoding NMDAR subunits that alter the receptor's functional characteristics are associated with a wide range of neurological and neuropsychiatric conditions. Pathological GRIN variants located in the M2 re-entrant loop lining the channel pore cause significant functional changes, the most consequential alteration being a reduction in voltage-dependent Mg2+ inhibition. Voltage-dependent Mg2+ block is a unique feature of NMDAR biology whereby channel activation requires both ligand binding and postsynaptic membrane depolarization. Thus, loss of NMDAR Mg2+ block will have a profound impact on synaptic function and plasticity. Here, we choose 11 missense variants within the GRIN1, GRIN2A, and GRIN2B genes that alter residues located in the M2 loop and significantly reduce Mg2+ inhibition. Each variant was evaluated for tolerance to genetic variation using the 3-dimensional structure and assessed for functional rescue pharmacology via electrophysiological recordings. Three FDA-approved NMDAR drugs-memantine, dextromethorphan, and ketamine-were chosen based on their ability to bind near the M2 re-entrant loop, potentially rectifying dysregulated NMDAR function by supplementing the reduced voltage-dependent Mg2+ block. These results provide insight of structural determinants of FDA-approved NMDAR drugs at their binding sites in the channel pore and may further define conditions necessary for the use of such agents as potential rescue pharmacology.

De novo CLPTM1 variants with reduced GABAA R current response in patients with epilepsy.

OBJECTIVE: To investigate the clinical features and potential pathogenesis mechanism of de novo CLPTM1 variants associated with epilepsy. METHODS: Identify de novo genetic variants associated with epilepsy by reanalyzing trio-based whole-exome sequencing data. We analyzed the clinical characteristics of patients with these variants and performed functional in vitro studies in cells expressing mutant complementary DNA for these variants using whole-cell voltage-clamp current recordings and outside-out patch-clamp recordings from transiently transfected human embryonic kidney (HEK) cells. RESULTS: Two de novo missense variants related to epilepsy were identified in the CLPTM1 gene. Functional studies indicated that CLPTM1-p.R454H and CLPTM1-p.R568Q variants reduced the γ-aminobutyric acid A receptor (GABAA R) current response amplitude recorded under voltage clamp compared to the wild-type receptors. These variants also reduced the charge transfer and altered the time course of desensitization and deactivation following rapid removal of GABA. The surface expression of the GABAA R γ2 subunit from the CLPTM1-p.R568Q group was significantly reduced compared to CLPTM1-WT. SIGNIFICANCE: This is the first report of functionally relevant variants within the CLPTM1 gene. Patch-clamp recordings showed that these de novo CLPTM1 variants reduce GABAA R currents and charge transfer, which should promote excitation and hypersynchronous activity. This study may provide insights into the molecular mechanisms of the CLPTM1 variants underlying the patients' phenotypes, as well as for exploring potential therapeutic targets for epilepsy.

Alive but not well: The neglected cost of air pollution.

Besides medical expenses, hospitalizations associated with air pollution will incur the welfare loss due to activity restrictions and the wage loss due to inability to work. We fill in the gap in the literature by examining the impact of air pollution on volume and intensity of hospitalizations, which allows us to incorporate the welfare loss and the wage loss. Using a data set that covers most of the inpatients in a major Chinese city during 2015-16, we find that worse air quality causes more hospital admissions, more total inpatient days, and higher total inpatient expenditure for various diseases, particularly diseases of the respiratory and circulatory systems. We also find that there would be an underestimate of the loss from air pollution if we had ignored the loss associated with activity restrictions and the wage loss during hospitalization.

Role of calcium-sensing receptor in regulating activation susceptibility of postovulatory aging mouse oocytes.

The mechanisms underlying postovulatory oocyte aging (POA) remain largely unknown. The expression of the calcium-sensing receptor (CaSR) in mouse oocytes and its role in POA need to be explored. Our objective was to observe CaSR expression and its role in the susceptibility to activating stimuli (STAS) in POA mouse oocytes. The results showed that, although none of the newly ovulated oocytes were activated, 40% and 94% of the oocytes recovered 19 and 25 h after human chorionic gonadotropin (hCG) injection were activated, respectively, after ethanol treatment. The level of the CaSR functional dimer protein in oocytes increased significantly from 13 to 25 h post hCG. Thus, the CaSR functional dimer level was positively correlated with the STAS of POA oocytes. Aging in vitro with a CaSR antagonist suppressed the elevation of STAS, and cytoplasmic calcium in oocytes recovered 19 h post hCG, whereas aging with a CaSR agonist increased STAS, and cytoplasmic calcium of oocytes recovered 13 h post hCG. Furthermore, the CaSR was more important than the Na-Ca2+ exchanger in regulating oocyte STAS, and T- and L-type calcium channels were inactive in aging oocytes. We conclude that the CaSR is involved in regulating STAS in POA mouse oocytes, and that it is more important than the other calcium channels tested in this connection.

Opportunities for Precision Treatment of GRIN2A and GRIN2B Gain-of-Function Variants in Triheteromeric N-Methyl-D-Aspartate Receptors.

N-methyl-D-aspartate receptors (NMDARs) are tetrameric assemblies of two glutamate N-methyl-D-aspartate receptor subunits, GluN1 and two GluN2, that mediate excitatory synaptic transmission in the central nervous system. Four genes (GRIN2A-D) encode four distinct GluN2 subunits (GluN2A-D). Thus, NMDARs can be diheteromeric assemblies of two GluN1 plus two identical GluN2 subunits, or triheteromeric assemblies of two GluN1 subunits plus two different GluN2 subunits. An increasing number of de novo GRIN variants have been identified in patients with neurologic conditions and with GRIN2A and GRIN2B harboring the vast majority (> 80%) of variants in these cases. These variants produce a wide range of effects on NMDAR function depending upon its subunit subdomain location and additionally on the subunit composition of diheteromeric versus triheteromeric NMDARs. Increasing evidence implicates triheteromeric GluN1/GluN2A/GluN2B receptors as a major component of the NMDAR pool in the adult cortex and hippocampus. Here, we explore the ability of GluN2A- and GluN2B-selective inhibitors to reduce excess current flow through triheteromeric GluN1/GluN2A/GluN2B receptors that contain one copy of GRIN2A or GRIN2B gain-of-function variants. Our data reveal a broad range of sensitivities for variant-containing triheteromeric receptors to subunit-selective inhibitors, with some variants still showing strong sensitivity to inhibitors, whereas others are relatively insensitive. Most variants, however, retain sensitivity to non-selective channel blockers and the competitive antagonist D-(-)-2-amino-5-phosphonopentanoic acid. These results suggest that with comprehensive analysis, certain disease-related GRIN2A and GRIN2B variants can be identified as potential targets for subunit-selective modulation and potential therapeutic gain. SIGNIFICANCE STATEMENT: Triheteromeric NMDA receptors that contain one copy each of the GluN2A and GluN2B subunits show intermediate sensitivity to GluN2A- and GluN2B-selective inhibitors, making these compounds candidates for attenuating overactive, GRIN variant-containing NMDA receptors associated with neurological conditions. We show that functional evaluation of variant properties with inhibitor pharmacology can support selection of a subset of variants for which GluN2 subunit-selective agents remain effective inhibitors of variant-containing triheteromeric NMDA receptors.

Topoisomerase II dysfunction causes metaphase I arrest by activating Aurora B, SAC and MPF and prevents PB1 abscission in mouse oocytes†.

Oocyte aneuploidy is caused mainly by chromosome nondisjunction and/or unbalanced sister chromatid pre-division. Although studies in somatic cells have shown that topoisomerase II (TOP2) plays important roles in chromosome condensation and timely separation of centromeres, little is known about its role during oocyte meiosis. Furthermore, because VP-16, which is a TOP2 inhibitor and induces DNA double strand breaks, is often used for ovarian cancer chemotherapy, its effects on oocytes must be studied for ovarian cancer patients to recover ovarian function following chemotherapy. This study showed that inhibiting TOP2 with either ICRF-193 or VP-16 during meiosis I impaired chromatin condensation, chromosome alignment, TOP2α localization, and caused metaphase I (MI) arrest and first polar body (PB1) abscission failure. Inhibiting or neutralizing either spindle assembly checkpoint (SAC), Aurora B or maturation-promoting factor (MPF) significantly abolished the effect of ICRF-193 or VP-16 on MI arrest. Treatment with ICRF-193 or VP-16 significantly activated MPF and SAC but the effect disappeared when Aurora B was inhibited. Most of the oocytes matured in the presence of ICRF-193 or VP-16 were arrested at MI, and only 11-27% showed PB1 protrusion. Furthermore, most of the PB1 protrusions formed in the presence of ICRF-193 or VP-16 were retracted after further culture for 7 h. In conclusion, TOP2 dysfunction causes MI arrest by activating Aurora B, SAC, and MPF, and it prevents PB1 abscission by promoting chromatin bridges.

Expression profiling and function analysis identified new microRNAs regulating cumulus expansion and apoptosis in cumulus cells.

Although microRNAs (miRNAs) expressed in cumulus cells (CCs) may be used to select competent oocytes/embryos, only a limited number of such miRNAs has been reported. To identify more miRNAs that regulate cumulus expansion (CE) and CC apoptosis, we first established that mouse cumulus-oocyte complexes (COCs) cultured in expansion-supporting medium supported full CE while undergoing mild apoptosis, whereas mouse oocytectomized COCs (OOXs) cultured in apoptosis-triggering medium underwent severe apoptosis while supporting no CE. RNA- and miRNA-sequencing and bioinformatics using CCs from these cultured COCs/OOXs identified candidate apoptosis- and/or CE-regulating miRNAs. Transfection of COCs/OOXs with miRNA mimic or inhibitor validated that miR-212-5p and 149-5p promoted CE by facilitating Has2 expression; miR-31-5p and 27a-3p promoted CE by increasing both Has2 and Ptx3 expression; and miR-351-5p and 503-5p inhibited CE by suppressing Ptx3 expression. Furthermore, miR-212-5p, 149-5p and Nov798 inhibited CC apoptosis, involving both Bcl2/Bax and Fas signaling. Analysis using in vivo matured COCs further verified the above apoptosis- and/or CE-regulating miRNAs, except for miR-149-5p. In conclusion, this study identified and validated new CE- and apoptosis-regulating miRNAs in CCs, which could be used as biomarkers to select competent oocytes/embryos and for elucidating how the oocyte-derived factors regulate CE and CC apoptosis.

Biased modulators of NMDA receptors control channel opening and ion selectivity.

Allosteric modulators of ion channels typically alter the transitions rates between conformational states without changing the properties of the open pore. Here we describe a new class of positive allosteric modulators of N-methyl D-aspartate receptors (NMDARs) that mediate a calcium-permeable component of glutamatergic synaptic transmission and play essential roles in learning, memory and cognition, as well as neurological disease. EU1622-14 increases agonist potency and channel-open probability, slows receptor deactivation and decreases both single-channel conductance and calcium permeability. The unique functional selectivity of this chemical probe reveals a mechanism for enhancing NMDAR function while limiting excess calcium influx, and shows that allosteric modulators can act as biased modulators of ion-channel permeation.

Antinociceptive C19-diterpenoid alkaloids from the root of Aconitum episcopale.

A chemical investigation on the roots of Aconitum episcopale afforded three undescribed aconitine-type C19-diterpenoid alkaloids, episcopalines A-C (1-3). The structures of the new compounds were elucidated by spectroscopic analysis (NMR, IR, UV, and MS). The isolated alkaloids were tested in vivo for their antinociceptive properties. As a result, episcopaline B (2) showed potent antinociceptive effect and its ID50 value (55.0 µmol/kg) was 2-fold less than those of the positive control drugs aspirin and acetaminophen.