Supralinear potentiation of NR1/NR3A excitatory glycine receptors by Zn2+ and NR1 antagonist.
Ontology highlight
ABSTRACT:
SUBMITTER: Madry C
PROVIDER: S-EPMC2527951 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:BACKGROUND:N-methyl-D-aspartate receptors (NMDARs) are the most complex of ionotropic glutamate receptors (iGluRs). Subunits of this subfamily assemble into heteromers, which - depending on the subunit combination - may display very different pharmacological and electrophysiological properties. The least studied members of the NMDAR family, the NR3 subunits, have been reported to assemble with NR1 to form excitatory glycine receptors in heterologous expression systems. The heterogeneity of NMDARs in vivo is in part conferred to the receptors by splicing of the NR1 subunit, especially with regard to proton sensitivity. RESULTS:Here, we have investigated whether the NR3B subunit is capable of assembly with each of the eight functional NR1 splice variants, and whether the resulting receptors share the unique functional properties described for NR1-1a/NR3. We provide evidence that functional excitatory glycine receptors formed regardless of the NR1 isoform, and their pharmacological profile matched the one reported for NR1-1a/NR3: glycine alone fully activated the receptors, which were insensitive to glutamate and block by Mg2+. Surprisingly, amplitudes of agonist-induced currents showed little dependency on the C-terminally spliced NR1 variants in NR1/NR3B diheteromers. Even more strikingly, NR3B conferred proton sensitivity also to receptors containing NR1b variants - possibly via disturbing the "proton shield" of NR1b splice variants. CONCLUSION:While functional assembly could be demonstrated for all combinations, not all of the specific interactions seen for NR1 isoforms with coexpressed NR2 subunits could be corroborated for NR1 assembly with NR3. Rather, NR3 abates trafficking effects mediated by the NR1 C terminus as well as the N-terminally mediated proton insensitivity. Thus, this study establishes that NR3B overrides important NR1 splice variant-specific receptor properties in NR1/NR3B excitatory glycine receptors.
| S-EPMC2669480 | biostudies-literature
Project description:NMDA receptors are typically excited by a combination of glutamate and glycine. Here we describe excitatory responses in CNS myelin that are gated by a glycine agonist alone and mediated by NR1/NR3 "NMDA" receptor subunits. Response properties include activation by d-serine, inhibition by the glycine-site antagonist CNQX, and insensitivity to the glutamate-site antagonist d-APV. d-Serine responses were abrogated in NR3A-deficient mice. Our results suggest the presence of functional NR1/NR3 receptors in CNS myelin.
| S-EPMC2941801 | biostudies-literature
Project description:GluN3A and GluN3B are glycine-binding subunits belonging to the NMDA receptor (NMDAR) family that can assemble with the GluN1 subunit to form unconventional receptors activated by glycine alone. Functional characterization of GluN1/GluN3 NMDARs has been difficult. Here, we uncover two modalities that have transformative properties on GluN1/GluN3A receptors. First, we identify a compound, CGP-78608, which greatly enhances GluN1/GluN3A responses, converting small and rapidly desensitizing currents into large and stable responses. Second, we show that an endogenous GluN3A disulfide bond endows GluN1/GluN3A receptors with distinct redox modulation, profoundly affecting agonist sensitivity and gating kinetics. Under reducing conditions, ambient glycine is sufficient to generate tonic receptor activation. Finally, using CGP-78608 on P8-P12 mouse hippocampal slices, we demonstrate that excitatory glycine GluN1/GluN3A NMDARs are functionally expressed in native neurons, at least in the juvenile brain. Our work opens new perspectives on the exploration of excitatory glycine receptors in brain function and development.
| S-EPMC6233196 | biostudies-literature
Project description:Cannabinoids enhance the function of glycine receptors (GlyRs). However, little is known about the mechanisms and behavioral implication of cannabinoid-GlyR interaction. Using mutagenesis and NMR analysis, we have identified a serine at 296 in the GlyR protein critical for the potentiation of I(Gly) by ?(9)-tetrahydrocannabinol (THC), a major psychoactive component of marijuana. The polarity of the amino acid residue at 296 and the hydroxyl groups of THC are critical for THC potentiation. Removal of the hydroxyl groups of THC results in a compound that does not affect I(Gly) when applied alone but selectively antagonizes cannabinoid-induced potentiating effect on I(Gly) and analgesic effect in a tail-flick test in mice. The cannabinoid-induced analgesia is absent in mice lacking ?3GlyRs but not in those lacking CB1 and CB2 receptors. These findings reveal a new mechanism underlying cannabinoid potentiation of GlyRs, which could contribute to some of the cannabis-induced analgesic and therapeutic effects.
| S-EPMC3388539 | biostudies-literature
Project description:Glycine receptors (GlyRs) belong to the superfamily of pentameric cys-loop receptor-operated channels and are involved in numerous physiological functions, including movement, vision, and pain. In search for compounds performing subunit-specific modulation of GlyRs we studied action of ginkgolic acid, an abundant Ginkgo biloba product. Using patch-clamp recordings, we analyzed the effects of ginkgolic acid in concentrations from 30 nM to 25 ?M on ?1-?3 and ?1/?, ?2/? configurations of GlyR and on GABAARs expressed in cultured CHO-K1 cells and mouse neuroblastoma (N2a) cells. Ginkgolic acid caused an increase in the amplitude of currents mediated by homomeric ?1 and heteromeric ?1/? GlyRs and provoked a left-shift of the concentration-dependent curves for glycine. Even at high concentrations (10-25 ?M) ginkgolic acid was not able to augment ionic currents mediated by ?2, ?2/?, and ?3 GlyRs, or by GABAAR consisting of ?1/?2/?2 subunits. Mutation of three residues (T59A/A261G/A303S) in the ?2 GlyR subunit to the corresponding ones from the ?1 converted the action of ginkgolic acid to potentiation with a distinct decrease in EC50 for glycine, suggesting an important role for these residues in modulation by ginkgolic acid. Our results suggest that ginkgolic acid is a novel selective enhancer of ?1 GlyRs.
| S-EPMC4624854 | biostudies-literature
Project description:Subunit composition of N-methyl-D-aspartate-type glutamate receptors (NMDARs) dictates their function, yet the ontogenic profiles of human NMDAR subunits from gestation to adulthood have not been determined. We examined NMDAR mRNA and protein development in human dorsolateral prefrontal cortex (DLPFC), an area in which NMDARs are critical for higher cognitive processing and NMDAR hypofunction is hypothesized in schizophrenia. Using quantitative reverse transcriptase-polymerase chain reaction and western blotting, we found NR1 expression begins low prenatally, peaks in adolescence, yet remains high throughout life, suggesting lifelong importance of NMDAR function. In contrast, NR3A levels are low during gestation, surge soon after birth, and decline progressively through adolescence and into adulthood. Because NR3A subunits uniquely attenuate NMDAR-mediated currents, limit calcium influx, and suppress dendritic spine formation, high levels during early childhood may be important for regulating neuroprotection and activity-dependent sculpting of synapses. We also examined whether subunit changes underlie reduced NMDAR activity in schizophrenia. Our results reveal normal NR1 and NR3A protein levels in DLPFC from schizophrenic patients, indicating that NMDAR hypofunction is unlikely to be maintained by gross changes in NR3A-containing NMDARs or overall NMDAR numbers. These data provide insights into NMDAR functions in the developing CNS and will contribute to designing pharmacotherapies for neurological disorders.
| S-EPMC2733318 | biostudies-literature
Project description:Both exogenous and endogenous cannabinoids can allosterically modulate glycine receptors (GlyRs). However, little is known about the molecular basis of cannabinoid-GlyR interactions. Here we report that sustained incubation with the endocannabinoid anandamide (AEA) substantially increased the amplitude of glycine-activated current in both rat cultured spinal neurons and in HEK-293 cells expressing human α1, rat α2 and α3 GlyRs. While the α1 and α3 subunits were highly sensitive to AEA-induced potentiation, the α2 subunit was relatively insensitive to AEA. Switching a serine at 296 and 307 in the TM3 (transmembrane domain 3) of the α1 and α3 subunits with an alanine (A) at the equivalent position in the α2 subunit converted the α1/α3 AEA-sensitive receptors to sensitivity resembling that of α2. The S296 residue is also critical for exogenous cannabinoid-induced potentiation of I(Gly). The magnitude of AEA potentiation decreased with removal of either the hydroxyl or oxygen groups on AEA. While desoxy-AEA was significantly less efficacious in potentiating I(Gly), desoxy-AEA inhibited potentiation produced by both Δ(9)-tetrahydrocannabinol (THC), a major psychoactive component of marijuana, and AEA. Similarly, didesoxy-THC, a modified THC with removal of both hydroxyl/oxygen groups, did not affect I(Gly) when applied alone but inhibited the potentiation of I(Gly) induced by AEA and THC. These findings suggest that exogenous and endogenous cannabinoids potentiate GlyRs via a hydrogen bonding-like interaction. Such a specific interaction likely stems from a common molecular basis involving the S296 residue in the TM3 of the α1 and α3 subunits.
| S-EPMC3334839 | biostudies-literature
Project description:Nociception and motor coordination are critically governed by glycine receptor (GlyR) function at inhibitory synapses. Consequentially, GlyRs are attractive targets in the management of chronic pain and in the treatment of several neurological disorders. High-resolution mechanistic details of GlyR function and its modulation are just emerging. While it has been known that cannabinoids such as Δ9-tetrahydrocannabinol (THC), the principal psychoactive constituent in marijuana, potentiate GlyR in the therapeutically relevant concentration range, the molecular mechanism underlying this effect is still not understood. Here, we present Cryo-EM structures of full-length GlyR reconstituted into lipid nanodisc in complex with THC under varying concentrations of glycine. The GlyR-THC complexes are captured in multiple conformational states that reveal the basis for THC-mediated potentiation, manifested as different extents of opening at the level of the channel pore. Taken together, these structural findings, combined with molecular dynamics simulations and functional analysis, provide insights into the potential THC binding site and the allosteric coupling to the channel pore.
| S-EPMC9388682 | biostudies-literature
Project description:The large intracellular loop (IL) of the glycine receptor (GlyR) interacts with various signaling proteins and plays a fundamental role in trafficking and regulation of several receptor properties, including a direct interaction with Gbetagamma. In the present study, we found that mutation of basic residues in the N-terminal region of the IL reduced the binding of Gbetagamma to 21 +/- 10% of control. Two basic residues in the C-terminal region, on the other hand, contributed to a smaller extent to Gbetagamma binding. Using docking analysis, we found that both basic regions of the IL bind in nearby regions to the Gbetagamma dimer, within an area of high density of amino acids having an electronegative character. Thereafter, we generated a 17-amino acid peptide with the N-terminal sequence of the wild-type IL (RQH) that was able to inhibit the in vitro binding of Gbetagamma to GlyRs to 57 +/- 5% of control in glutathione S-transferase pull-down assays using purified proteins. More interestingly, when the peptide was intracellularly applied to human embryonic kidney 293 cells, it inhibited the Gbetagamma-mediated modulations of G protein-coupled inwardly rectifying potassium channel by baclofen (24 +/- 14% of control) and attenuated the GlyR potentiation by ethanol (51 +/- 10% versus 10 +/- 3%).
| S-EPMC2784719 | biostudies-literature
Project description:The novel ginkgolide analog ginkgolide X was characterized functionally at human glycine and gamma-aminobutyric acid type A receptors (GlyRs and GABA(A)Rs, respectively) in the fluorescence-based FLIPR(TM) Membrane Potential assay. The compound inhibited the signaling of all GABA(A)R subtypes included in the study with high nanomolar/low micromolar IC(50) values, except the rho 1 receptor at which it was a significantly weaker antagonist. Ginkgolide X also displayed high nanomolar/low micromolar IC(50) values at the homomeric alpha1 and alpha2 GlyRs, whereas it was inactive at the heteromeric alpha 1 beta and alpha 2 beta subtypes at concentrations up to 300 microm. Thus, the functional properties of the compound were significantly different from those of the naturally occurring ginkgolides A, B, C, J, and M but similar to those of picrotoxin. In a mutagenesis study the 6' M2 residues in the GlyR ion channel were identified as the primary molecular determinant of the selectivity profile of ginkgolide X, and a 6' M2 ring consisting of five Thr residues was found to be of key importance for its activity at the GABA(A)R. Conformational analysis and docking of low-energy conformations of the native ginkgolide A and ginkgolide X into a alpha1 GlyR homology model revealed two distinct putative binding sites formed by the 6' M2 residues together with the 2' residues and the 10' and 13' residues, respectively. Thus, we propose that the distinct functionalities of ginkgolide X compared with the other ginkgolides could arise from different flexibility and thus different binding modes to the ion channel of the anionic Cys-loop receptor.
| S-EPMC2843176 | biostudies-literature