These authors contributed equally to this work.Changes in the synaptic content of a-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)-type glutamate receptors lead to synaptic efficacy modifications, involved in synaptic plasticity mechanisms believed to underlie learning and memory formation. Early in development, GluR4 is highly expressed in the hippocampus, and GluR4-containing AMPA receptors are inserted into synapses. During synapse maturation, the number of AMPA receptors at the synapse is dynamically regulated, and both addition and removal of receptors from postsynaptic sites occur through regulated mechanisms. GluR4 delivery to synapses in rat hippocampal slices was shown to require protein kinase A (PKA)-mediated phosphorylation of GluR4 at serine 842 (Ser842). Protein kinase C (PKC) can also phosphorylate Ser842, and we have shown that PKCg can associate with GluR4. Here we show that activation of PKC in retina neurons, or in human embryonic kidney 293 cells cotransfected with GluR4 and PKCg, increases GluR4 surface expression and Ser842 phosphorylation. Moreover, mutation of amino acids R821A, K825A and R826A at the GluR4 C-terminal, within the interacting region of GluR4 with PKCg, abolishes the interaction between PKCg and GluR4 and prevents the stimulatory effect of PKCg on GluR4 Ser842 phosphorylation and surface expression. These data argue for a role of anchored PKCg in Ser842 phosphorylation and targeting to the plasma membrane. The triple GluR4 mutant is, however, phosphorylated by PKA, and it is targeted to the synapse in CA1 hippocampal neurons in organotypic rat hippocampal slices. The present findings show that the interaction between PKCg and GluR4 is specifically required to assure PKC-driven phosphorylation and surface membrane expression of GluR4. a-Amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)-type ionotropic glutamate receptors mediate the majority of fast excitatory synaptic transmission in the brain and are believed to be involved in learning and memory formation. It has been shown that these receptors can be added to and removed from the postsynaptic membrane, resulting in changes in synaptic efficacy (1). These changes in synaptic strength are involved in synaptic plasticity mechanisms [long-term potentiation (LTP) and long-term depression (LTD)], which are believed to be the molecular basis of learning and memory (1,2).The AMPA receptors are heterooligomeric structures formed by four subunits [GluR1-4; (3)]. The combination of different receptor subunits results in distinct trafficking properties of the AMPA receptors (4,5). GluR4-containing AMPA receptors exhibit fast currents and are expressed in several regions of the central nervous system (CNS) (6-11). In the hippocampus, GluR4 is expressed mainly in early postnatal development, and GluR4-containing AMPA receptors are delivered to the synapse by spontaneous activity (11). In this brain region, synaptic delivery of GluR4-containing AMPA receptors is dependent on protein kinase A (PKA) phosphorylation of the seri...

Spinocerebellar ataxia type 14 (SCA14) is an autosomal dominant neurodegenerative disorder, first described in a Japanese family, showing linkage to chromosome 19q13.4-qter. Recently, mutations have been identified in the PRKCG gene in families with SCA14. The PRKCG gene encodes the protein kinase Cc (PKCc), a member of a serine/threonine kinase family involved in signal transduction important for several cellular processes, including cell proliferation and synaptic transmission. To identify the disease-causing mutation in a large group of ataxia patients, we searched for mutations in the PRKCG gene. We ascertained 366 unrelated patients with spinocerebellar ataxia, either pure or with associated features such as epilepsy, mental retardation, seizures, paraplegia, and tremor. A C-to-G transversion in exon 4, resulting in a histidine-to-glutamine change at codon 101 of the PKCc protein, was identified in patients from a family with slowly progressive pure cerebellar ataxia. Functional studies performed in HEK293 cells transfected with normal or mutant construct showed that this mutation affects PKCc stability or solubility, verified by time-dependent decreased protein levels in cell culture. In conclusion, the H101Q mutation causes slowly progressive uncomplicated ataxia by interfering with PKCc stability or solubility, which consequently may cause in either case a decrease in the overall PKCc-dependent phosphorylation.

Glutamate receptor phosphorylation has been implicated in several forms of modulation of synaptic transmission. It has been reported that protein kinase A (PKA) can phosphorylate the a-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor subunit GluR4 on Ser842, both in vitro and in vivo. Here, we studied the regulation of GluR4 phosphorylation and intracellular trafficking by PKA and by metabotropic receptors coupled to adenylyl cyclase (AC), in cultured chick retinal amacrine-like neurones, which are enriched in GluR4. The regulation of AMPA receptor activity by PKA and by metabotropic AC-coupled receptors was also investigated by measuring the [Ca 2+ ] i response to kainate in Na + -free medium. Stimulation of AC with forskolin (FSK), or using the selective agonist of dopamine D 1 receptors (±)-1-phenyl-2,3,4,5-tetrahydro-(1H)-3-benzazepine-7,8-diol (SKF38393), increased the [Ca 2+ ] i response to kainate, GluR4 phosphorylation at Ser842 and GluR4 surface expression. Pre-incubation of the cells with (2S,2¢R,3¢R)-2-(2¢,3¢-dicarboxycyclopropyl)glycine (DCG-IV), an agonist of group II metabotropic glutamate receptors (mGluR), which are coupled to inhibition of AC, inhibited the effect of FSK and of SKF38393 on AMPA receptor activity, GluR4 phosphorylation and expression at the plasma membrane. These results indicate that there is a functional cross-talk between dopamine D 1 receptors and group II mGluR in the regulation of GluR4 phosphorylation and AMPA receptor activity. Our data show that GluR4 phosphorylation at Ser842 by PKA, and its recruitment to the plasma membrane upon phosphorylation, is regulated by metabotropic receptors.

Glutamate receptors of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type play an important role in synaptic plasticity and contribute to cell death under excitotoxic conditions. AMPA receptors form heterotetramers of four homologous subunits (GluR1-4), which exist in two functionally different isoforms, flip and flop, generated by alternative splicing. We identified transcripts for alternatively spliced isoforms of AMPA receptor subunits which lack both the flip and the flop exons, in hippocampal and retinal cultures. These transcripts originate AMPA receptor subunits lacking the flip/flop cassette, the fourth transmembrane domain and the intracellular C-terminus. Truncated GluR1 associates with full-length GluR1 and exerts a dominant negative effect, giving rise to non-functional receptors. Moreover, truncated GluR1 reaches the cell surface, but is not efficiently targeted to the synapse. Hippocampal neuronal transfection with truncated GluR1 resulted in a significant reduction in apoptotic neuronal death triggered by toxic concentrations of glutamate. Furthermore, mRNA coding for the truncated subunits is consistently detected in some regions of the brain in epileptic rats and in hippocampal neurons submitted to toxic concentrations of glutamate. The existence of truncated AMPA receptor subunits may constitute an intrinsic neuroprotective mechanism.

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The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors for the neurotransmitter glutamate are oligomeric structures responsible for most fast excitatory responses in the central nervous system. The activity of AMPA receptors can be directly regulated by protein phosphorylation, which may also affect the interaction with intracellular proteins and, consequently, their recycling and localization to defined postsynaptic sites. This review focuses on recent advances in understanding the dynamic regulation of AMPA receptors, on a short- and long-term basis, and its implications in synaptic plasticity.

A key distinction between today's and future networks is the appetite for reliable communication to support emerging critical-communication services. In this paper, we study multi-operator connectivity as a form of redundancy to support the design of reliable networks and investigate its trade-offs. This approach is motivated by 3GPP standardisation initiatives of dual-connectivity and similar techniques in industrial wired networks. We deploy a risk awareness performance metric to assess reliability: this superquantile metric accounts for periods of connectivity shortfalls. Our analysis shows that multi-operator connectivity brings significant reliability gains, in particular when network deployments by different operators exhibit high complementarity in coverage. We also explore the effects of multi-connectivity on spectral efficiency in times of high demand for bandwidth. Our study is based on a real-world dataset comprising signal strength indicators of three mobile operators in Dublin, Ireland.

The next generation of mobile networks will bring an appetite for reliable communication, which enables emerging critical-communication services. In this paper, we present network sharing between operators as a way to provide increased reliability while using the already existing mobile network infrastructure. Our results indicate that network sharing significantly improves network performance during periods of connectivity shortfalls, benefiting the design of reliable networks. Our conclusions are drawn from a real-world dataset of signal quality indicators for three mobile operators in Dublin, Ireland.