Altered Cerebellar Short-Term Plasticity but No Change in Postsynaptic AMPA-Type Glutamate Receptors in a Mouse Model of Juvenile Batten Disease

Juvenile Batten disease is the most common progressive neurodegenerative disorder of childhood. It is associated with mutations in the CLN3 gene, causing loss of function of CLN3 protein and degeneration of cerebellar and retinal neurons. It has been proposed that changes in granule cell AMPA-type glutamate receptors (AMPARs) contribute to the cerebellar dysfunction. In this study we compared AMPAR properties and synaptic transmission in cerebellar granule cells from wild-type and Cln3 knockout mice. In Cln3Δex1–6 cells the amplitude of AMPA-evoked whole-cell currents was unchanged. Similarly, we found no change in the amplitude, kinetics, or rectification of synaptic currents evoked by individual quanta, or in their underlying single-channel conductance. We found no change in cerebellar expression of GluA2 or GluA4 protein. By contrast, we observed a reduced number of quantal events following mossy-fiber stimulation in Sr2+, altered short-term plasticity in conditions of reduced extracellular Ca2+, and reduced mossy fiber vesicle number. Thus, while our results suggest early presynaptic changes in the Cln3Δex1–6 mouse model of juvenile Batten disease, they reveal no evidence for altered postsynaptic AMPARs

Homomeric Q/R edited AMPA receptors conduct when desensitized

Desensitization is a canonical property of ligand-gated ion channels, causing progressive current decline in the continued presence of agonist. AMPA-type glutamate receptors, which mediate fast excitatory signaling throughout the brain, exhibit profound desensitization. Recent cryo-EM studies of AMPAR assemblies show their ion channels to be closed in the desensitized state. Here we report the surprising finding that homomeric Q/R edited AMPARs still allow ions to flow when the receptors are desensitized. GluA2(R) expressed alone, or with auxiliary subunits (γ-2, γ-8 or GSG1L), generates large steady-state currents and anomalous current-variance relationships. Using fluctuation analysis, single-channel recording, and kinetic modeling we demonstrate that the steady-state current is mediated predominantly by ‘conducting desensitized’ receptors. When combined with crystallography this unique functional readout of a hith-erto silent state enabled us to examine cross-linked cysteine mutants to probe the conformation of the desensitized ligand binding domain of functioning AMPAR complexes within the plasma membrane

Acid-sensing ion channel 1a drives AMPA receptor plasticity following ischemia and acidosis in hippocampal CA1 neurons

The CA1 region of the hippocampus is particularly vulnerable to ischemic damage. While NMDA receptors play a major role in excitotoxicity, it is thought to be exacerbated in this region by two forms of post-ischemic AMPA receptor (AMPAR) plasticity - namely, anoxic long-term potentiation (a-LTP), and a delayed increase in the prevalence of Ca2+ -permeable GluA2-lacking AMPARs (CP-AMPARs). The acid-sensing ion channel 1a (ASIC1a) which is expressed in CA1 pyramidal neurons, is also known to contribute to post-ischemic neuronal death and to physiologically induced LTP. This raises the question - does ASIC1a activation drive the post-ischemic forms of AMPAR plasticity in CA1 pyramidal neurons? We have tested this by examining organotypic hippocampal slice cultures (OHSCs) exposed to oxygen glucose deprivation (OGD), and dissociated cultures of hippocampal pyramidal neurons (HPN) exposed to low pH (acidosis). We find that both a-LTP and the delayed increase in the prevalence of CP-AMPARs are dependent on ASIC1a activation during ischemia. Indeed, acidosis alone is sufficient to induce the increase in CP-AMPARs. We also find that inhibition of ASIC1a channels circumvents any potential neuroprotective benefit arising from block of CP-AMPARs. By demonstrating that ASIC1a activation contributes to post-ischemic AMPAR plasticity, our results identify a functional interaction between acidotoxicity and excitotoxicity in hippocampal CA1 cells, and provide insight into the role of ASIC1a and CP-AMPARs as potential drug targets for neuroprotection. We thus propose that ASIC1a activation can drive certain forms of CP-AMPAR plasticity, and that inhibiting ASIC1a affords neuroprotection

Retrograde semaphorin-plexin signalling drives homeostatic synaptic plasticity.

Homeostatic signalling systems ensure stable but flexible neural activity and animal behaviour. Presynaptic homeostatic plasticity is a conserved form of neuronal homeostatic signalling that is observed in organisms ranging from Drosophila to human. Defining the underlying molecular mechanisms of neuronal homeostatic signalling will be essential in order to establish clear connections to the causes and progression of neurological disease. During neural development, semaphorin-plexin signalling instructs axon guidance and neuronal morphogenesis. However, semaphorins and plexins are also expressed in the adult brain. Here we show that semaphorin 2b (Sema2b) is a target-derived signal that acts upon presynaptic plexin B (PlexB) receptors to mediate the retrograde, homeostatic control of presynaptic neurotransmitter release at the neuromuscular junction in Drosophila. Further, we show that Sema2b-PlexB signalling regulates presynaptic homeostatic plasticity through the cytoplasmic protein Mical and the oxoreductase-dependent control of presynaptic actin. We propose that semaphorin-plexin signalling is an essential platform for the stabilization of synaptic transmission throughout the developing and mature nervous system. These findings may be relevant to the aetiology and treatment of diverse neurological and psychiatric diseases that are characterized by altered or inappropriate neural function and behaviour

TARP γ-7 selectively enhances synaptic expression of calcium-permeable AMPARs

Regulation of calcium-permeable AMPA receptors (CP-AMPARs) is crucial in normal synaptic function and neurological disease states. Although transmembrane AMPAR regulatory proteins (TARPs) such as stargazin (γ-2) modulate the properties of calcium-impermeable AMPARs (CI-AMPARs) and promote their synaptic targeting, the TARP-specific rules governing CP-AMPAR synaptic trafficking remain unclear. We used RNA interference to manipulate AMPAR-subunit and TARP expression in γ-2–lacking stargazer cerebellar granule cells—the classic model of TARP deficiency. We found that TARP γ-7 selectively enhanced the synaptic expression of CP-AMPARs and suppressed CI-AMPARs, identifying a pivotal role of γ-7 in regulating the prevalence of CP-AMPARs. In the absence of associated TARPs, both CP-AMPARs and CI-AMPARs were able to localize to synapses and mediate transmission, although their properties were altered. Our results also establish that TARPed synaptic receptors in granule cells require both γ-2 and γ-7 and reveal an unexpected basis for the loss of AMPAR-mediated transmission in stargazer mice

Reelin Secreted by GABAergic Neurons Regulates Glutamate Receptor Homeostasis

BACKGROUND: Reelin is a large secreted protein of the extracellular matrix that has been proposed to participate to the etiology of schizophrenia. During development, reelin is crucial for the correct cytoarchitecture of laminated brain structures and is produced by a subset of neurons named Cajal-Retzius. After birth, most of these cells degenerate and reelin expression persists in postnatal and adult brain. The phenotype of neurons that bind secreted reelin and whether the continuous secretion of reelin is required for physiological functions at postnatal stages remain unknown. METHODOLOGY/PRINCIPAL FINDINGS: Combining immunocytochemical and pharmacological approaches, we first report that two distinct patterns of reelin expression are present in cultured hippocampal neurons. We show that in hippocampal cultures, reelin is secreted by GABAergic neurons displaying an intense reelin immunoreactivity (IR). We demonstrate that secreted reelin binds to receptors of the lipoprotein family on neurons with a punctate reelin IR. Secondly, using calcium imaging techniques, we examined the physiological consequences of reelin secretion blockade. Blocking protein secretion rapidly and reversibly changes the subunit composition of N-methyl-D-aspartate glutamate receptors (NMDARs) to a predominance of NR2B-containing NMDARs. Addition of recombinant or endogenously secreted reelin rescues the effects of protein secretion blockade and reverts the fraction of NR2B-containing NMDARs to control levels. Therefore, the continuous secretion of reelin is necessary to control the subunit composition of NMDARs in hippocampal neurons. CONCLUSIONS/SIGNIFICANCE: Our data show that the heterogeneity of reelin immunoreactivity correlates with distinct functional populations: neurons synthesizing and secreting reelin and/or neurons binding reelin. Furthermore, we show that continuous reelin secretion is a strict requirement to maintain the composition of NMDARs. We propose that reelin is a trans-neuronal messenger secreted by GABAergic neurons that regulates NMDARs homeostasis in postnatal hippocampus. Defects in reelin secretion could play a major role in the development of neuropsychiatric disorders, particularly those associated with deregulation of NMDARs such as schizophrenia

Rare mutations in N-methyl-D-aspartate glutamate receptors in autism spectrum disorders and schizophrenia

Pharmacological, genetic and expression studies implicate N-methyl-D-aspartate (NMDA) receptor hypofunction in schizophrenia (SCZ). Similarly, several lines of evidence suggest that autism spectrum disorders (ASD) could be due to an imbalance between excitatory and inhibitory neurotransmission. As part of a project aimed at exploring rare and/or de novo mutations in neurodevelopmental disorders, we have sequenced the seven genes encoding for NMDA receptor subunits (NMDARs) in a large cohort of individuals affected with SCZ or ASD (n=429 and 428, respectively), parents of these subjects and controls (n=568). Here, we identified two de novo mutations in patients with sporadic SCZ in GRIN2A and one de novo mutation in GRIN2B in a patient with ASD. Truncating mutations in GRIN2C, GRIN3A and GRIN3B were identified in both subjects and controls, but no truncating mutations were found in the GRIN1, GRIN2A, GRIN2B and GRIN2D genes, both in patients and controls, suggesting that these subunits are critical for neurodevelopment. The present results support the hypothesis that rare de novo mutations in GRIN2A or GRIN2B can be associated with cases of sporadic SCZ or ASD, just as it has recently been described for the related neurodevelopmental disease intellectual disability. The influence of genetic variants appears different, depending on NMDAR subunits. Functional compensation could occur to counteract the loss of one allele in GRIN2C and GRIN3 family genes, whereas GRIN1, GRIN2A, GRIN2B and GRIN2D appear instrumental to normal brain development and function

Disease-associated missense mutations in GluN2B subunit alter NMDA receptor ligand binding and ion channel properties.

Genetic and bioinformatic analyses have identified missense mutations in GRIN2B encoding the NMDA receptor GluN2B subunit in autism, intellectual disability, Lennox Gastaut and West Syndromes. Here, we investigated several such mutations using a near-complete, hybrid 3D model of the human NMDAR and studied their consequences with kinetic modelling and electrophysiology. The mutants revealed reductions in glutamate potency; increased receptor desensitisation; and ablation of voltage-dependent Mg block. In addition, we provide new views on Mg and NMDA channel blocker binding sites. We demonstrate that these mutants have significant impact on excitatory transmission in developing neurons, revealing profound changes that could underlie their associated neurological disorders. Of note, the NMDAR channel mutant GluN2B unusually allowed Mg permeation, whereas nearby N615I reduced Ca permeability. By identifying the binding site for an NMDAR antagonist that is used in the clinic to rescue gain-of-function phenotypes, we show that drug binding may be modified by some GluN2B disease-causing mutations

Multiple roles of GluN2B-containing NMDA receptors in synaptic plasticity in juvenile hippocampus

AbstractIn the CA1 area of the hippocampus N-methyl-d-aspartate receptors (NMDARs) mediate the induction of long-term depression (LTD), short-term potentiation (STP) and long-term potentiation (LTP). All of these forms of synaptic plasticity can be readily studied in juvenile hippocampal slices but the involvement of particular NMDAR subunits in the induction of these different forms of synaptic plasticity is currently unclear. Here, using NVP-AAM077, Ro 25-6981 and UBP145 to target GluN2A-, 2B- and 2D-containing NMDARs respectively, we show that GluN2B-containing NMDARs (GluN2B) are involved in the induction of LTD, STP and LTP in slices prepared from P14 rat hippocampus. A concentration of Ro (1 μM) that selectively blocks GluN2B-containing diheteromers is able to block LTD. It also inhibits a component of STP without affecting LTP. A higher concentration of Ro (10 μM), that also inhibits GluN2A/B triheteromers, blocks LTP. UBP145 selectively inhibits the Ro-sensitive component of STP whereas NVP inhibits LTP. These data are consistent with a role of GluN2B diheretomers in LTD, a role of both GluN2B- and GluN2D- containing NMDARs in STP and a role of GluN2A/B triheteromers in LTP.This article is part of the Special Issue entitled ‘Ionotropic glutamate receptors’