RESUMEN
N-methyl-D-aspartate (NMDA)-type ionotropic glutamate receptors have essential roles in neurotransmission and synaptic plasticity. Previously, we identified an evolutionarily conserved protein, NRAP-1, that is required for NMDA receptor (NMDAR) function in C. elegans. Here, we demonstrate that NRAP-1 was sufficient to gate NMDARs and greatly enhanced glutamate-mediated NMDAR gating, thus conferring coincident activation properties to the NMDAR. Intriguingly, vertebrate NMDARs-and chimeric NMDARs where the amino-terminal domain (ATD) of C. elegans NMDARs was replaced by the ATD from vertebrate receptors-were spontaneously active when ectopically expressed in C. elegans neurons. Thus, the ATD is a primary determinant of NRAP-1- and glutamate-mediated gating of NMDARs. We determined the crystal structure of NRAP-1 at 1.9-Å resolution, which revealed two distinct domains positioned around a central low-density lipoprotein receptor class A domain. The NRAP-1 structure, combined with chimeric and mutational analyses, suggests a model where the three NRAP-1 domains work cooperatively to modify the gating of NMDARs.
Asunto(s)
Caenorhabditis elegans , Receptores de N-Metil-D-Aspartato , Animales , Receptores de N-Metil-D-Aspartato/metabolismo , Caenorhabditis elegans/metabolismo , N-Metilaspartato , Transmisión Sináptica , Ácido GlutámicoRESUMEN
Synaptic plasticity depends on rapid experience-dependent changes in the number of neurotransmitter receptors. Previously, we demonstrated that motor-mediated transport of AMPA receptors (AMPARs) to and from synapses is a critical determinant of synaptic strength. Here, we describe two convergent signaling pathways that coordinate the loading of synaptic AMPARs onto scaffolds, and scaffolds onto motors, thus providing a mechanism for experience-dependent changes in synaptic strength. We find that an evolutionarily conserved JIP-protein scaffold complex and two classes of mitogen-activated protein kinase (MAPK) proteins mediate AMPAR transport by kinesin-1 motors. Genetic analysis combined with in vivo, real-time imaging in Caenorhabditis elegans revealed that CaMKII is required for loading AMPARs onto the scaffold, and MAPK signaling is required for loading the scaffold complex onto motors. Our data support a model where CaMKII signaling and a MAPK-signaling pathway cooperate to facilitate the rapid exchange of AMPARs required for early stages of synaptic plasticity.
Asunto(s)
Proteínas Quinasas Activadas por Mitógenos , Receptores AMPA , Animales , Caenorhabditis elegans , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Plasticidad Neuronal/fisiología , Receptores AMPA/metabolismo , Transducción de Señal , Sinapsis/metabolismoRESUMEN
NMDA receptors (NMDARs) are a subtype of postsynaptic ionotropic glutamate receptors that function as molecular coincidence detectors, have critical roles in models of learning, and are associated with a variety of neurological and psychiatric disorders. To date, no auxiliary proteins that modify NMDARs have been identified. Here, we report the identification of NRAP-1, an auxiliary protein in C. elegans that modulates NMDAR function. NMDAR-mediated currents were eliminated in nrap-1 mutants, as was NMDA-dependent behavior. We show that reconstitution of NMDA-gated current in Xenopus oocytes, or C. elegans muscle cells, depends on NRAP-1 and that recombinant NRAP-1 can convert silent NMDARs to functional channels. Our data indicate that NRAP-1, secreted from presynaptic neurons, localizes to glutamatergic synapses, where it associates with postsynaptic NMDARs to modify receptor gating. Thus, our studies reveal a novel mechanism for synaptic regulation via pre-synaptic control of NMDAR-mediated synaptic transmission.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Interneuronas/citología , Proteínas de la Membrana/genética , Movimiento/fisiología , Proteínas Nucleares/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Sinapsis/fisiología , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Regulación de la Expresión Génica/genética , Ácido Glutámico/farmacología , Interneuronas/efectos de los fármacos , Activación del Canal Iónico/efectos de los fármacos , Activación del Canal Iónico/genética , Proteínas de la Membrana/efectos de los fármacos , Movimiento/efectos de los fármacos , Células Musculares/citología , Células Musculares/efectos de los fármacos , Mutación/genética , N-Metilaspartato/farmacología , Proteínas Nucleares/genética , Oocitos , Proteínas de Unión al ARN , Receptores de N-Metil-D-Aspartato/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Sinapsis/efectos de los fármacos , Sinapsis/genética , XenopusRESUMEN
Excitatory glutamatergic synaptic transmission is critically dependent on maintaining an optimal number of postsynaptic AMPA receptors (AMPARs) at each synapse of a given neuron. Here, we show that presynaptic activity, postsynaptic potential, voltage-gated calcium channels (VGCCs) and UNC-43, the C. elegans homolog of CaMKII, control synaptic strength by regulating motor-driven AMPAR transport. Genetic mutations in unc-43, or spatially and temporally restricted inactivation of UNC-43/CaMKII, revealed its essential roles in the transport of AMPARs from the cell body and in the insertion and removal of synaptic AMPARs. We found that an essential target of UNC-43/CaMKII is kinesin light chain and that mouse CaMKII rescued unc-43 mutants, suggesting conservation of function. Transient expression of UNC-43/CaMKII in adults rescued the transport defects, while optogenetic stimulation of select synapses revealed CaMKII's role in activity-dependent plasticity. Our results demonstrate unanticipated, fundamentally important roles for UNC-43/CaMKII in the regulation of synaptic strength.
Asunto(s)
Caenorhabditis elegans/fisiología , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/genética , Cinesinas/metabolismo , Neuronas/metabolismo , Canales de Potasio con Entrada de Voltaje/fisiología , Receptores de Glutamato/metabolismo , Animales , Animales Modificados Genéticamente , Transporte Biológico/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Potenciación a Largo Plazo/fisiología , Ratones , Mutación , Plasticidad Neuronal/genética , Técnicas de Placa-Clamp , Sinapsis/fisiologíaRESUMEN
A primary determinant of the strength of neurotransmission is the number of AMPA-type glutamate receptors (AMPARs) at synapses. However, we still lack a mechanistic understanding of how the number of synaptic AMPARs is regulated. Here, we show that UNC-116, the C. elegans homolog of vertebrate kinesin-1 heavy chain (KIF5), modifies synaptic strength by mediating the rapid delivery, removal, and redistribution of synaptic AMPARs. Furthermore, by studying the real-time transport of C. elegans AMPAR subunits in vivo, we demonstrate that although homomeric GLR-1 AMPARs can diffuse to and accumulate at synapses in unc-116 mutants, glutamate-gated currents are diminished because heteromeric GLR-1/GLR-2 receptors do not reach synapses in the absence of UNC-116/KIF5-mediated transport. Our data support a model in which ongoing motor-driven delivery and removal of AMPARs controls not only the number but also the composition of synaptic AMPARs, and thus the strength of synaptic transmission.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/fisiología , Proteínas de Ciclo Celular/fisiología , Cinesinas/fisiología , Receptores AMPA/metabolismo , Transmisión Sináptica/fisiología , Animales , Proteínas de Caenorhabditis elegans/efectos de los fármacos , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/genética , Cicloheximida/farmacología , Ácido Glutámico/farmacología , Cinesinas/genética , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Mutación , Receptores AMPA/efectos de los fármacosRESUMEN
The strength of synaptic communication at central synapses depends on the number of ionotropic glutamate receptors, particularly the class gated by the agonist AMPA (AMPARs). Cornichon proteins, evolutionarily conserved endoplasmic reticulum cargo adaptors, modify the properties of vertebrate AMPARs when coexpressed in heterologous cells. However, the contribution of cornichons to behavior and in vivo nervous system function has yet to be determined. Here, we take a genetic approach to these questions by studying CNI-1--the sole cornichon homolog in C. elegans. cni-1 mutants hyperreverse, a phenotype associated with increased glutamatergic synaptic transmission. Consistent with this behavior, we find larger glutamate-gated currents in cni-1 mutants with a corresponding increase in AMPAR number. Furthermore, we observe opposite phenotypes in transgenic worms that overexpress CNI-1 or vertebrate homologs. In reconstitution studies, we provide support for an evolutionarily conserved role for cornichons in regulating the export of vertebrate and invertebrate AMPARs.
Asunto(s)
Caenorhabditis elegans/metabolismo , Retículo Endoplásmico/metabolismo , Proteínas de la Membrana/metabolismo , Receptores AMPA/metabolismo , Sinapsis/metabolismo , Transmisión Sináptica/fisiología , Animales , Células Cultivadas , Ácido Glutámico/metabolismo , Mutación/genética , Neuronas/citología , Neuronas/metabolismo , Transporte de Proteínas/fisiología , Receptores AMPA/agonistas , Receptores AMPA/genética , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiónico/metabolismoRESUMEN
The neurotransmitter glutamate mediates excitatory synaptic transmission by gating ionotropic glutamate receptors (iGluRs). AMPA receptors (AMPARs), a subtype of iGluR, are strongly implicated in synaptic plasticity, learning, and memory. We previously discovered two classes of AMPAR auxiliary proteins in C. elegans that modify receptor kinetics and thus change synaptic transmission. Here, we have identified another auxiliary protein, SOL-2, a CUB-domain protein that associates with both the related auxiliary subunit SOL-1 and with the GLR-1 AMPAR. In sol-2 mutants, behaviors dependent on glutamatergic transmission are disrupted, GLR-1-mediated currents are diminished, and GLR-1 desensitization and pharmacology are modified. Remarkably, a secreted variant of SOL-1 delivered in trans can rescue sol-1 mutants, and this rescue depends on in cis expression of SOL-2. Finally, we demonstrate that SOL-1 and SOL-2 have an ongoing role in the adult nervous system to control AMPAR-mediated currents.
Asunto(s)
Proteínas de Caenorhabditis elegans/fisiología , Caenorhabditis elegans/fisiología , Lipoproteínas LDL/fisiología , Proteínas de la Membrana/fisiología , Receptores AMPA/fisiología , Sinapsis/fisiología , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Femenino , Células HEK293 , Humanos , Proteínas Relacionadas con Receptor de LDL , Proteínas de la Membrana/genética , Proteínas de la Membrana/aislamiento & purificación , Datos de Secuencia Molecular , Oocitos , Estructura Terciaria de Proteína/fisiología , Receptores de N-Metil-D-Aspartato , Transmisión Sináptica/fisiología , Xenopus laevisAsunto(s)
Sistema Nervioso/metabolismo , Transducción de Señal , Membranas Sinápticas/metabolismo , Proteínas Wnt/metabolismo , Animales , Caenorhabditis elegans/crecimiento & desarrollo , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas Dishevelled , Drosophila/crecimiento & desarrollo , Drosophila/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Larva/metabolismo , Receptores Huérfanos Similares al Receptor Tirosina Quinasa/metabolismo , Receptores Acoplados a Proteínas G/metabolismoRESUMEN
The adult nervous system is plastic, allowing us to learn, remember, and forget. Experience-dependent plasticity occurs at synapses--the specialized points of contact between neurons where signaling occurs. However, the mechanisms that regulate the strength of synaptic signaling are not well understood. Here, we define a Wnt-signaling pathway that modifies synaptic strength in the adult nervous system by regulating the translocation of one class of acetylcholine receptors (AChRs) to synapses. In Caenorhabditis elegans, we show that mutations in CWN-2 (Wnt ligand), LIN-17 (Frizzled), CAM-1 (Ror receptor tyrosine kinase), or the downstream effector DSH-1 (disheveled) result in similar subsynaptic accumulations of ACR-16/α7 AChRs, a consequent reduction in synaptic current, and predictable behavioral defects. Photoconversion experiments revealed defective translocation of ACR-16/α7 to synapses in Wnt-signaling mutants. Using optogenetic nerve stimulation, we demonstrate activity-dependent synaptic plasticity and its dependence on ACR-16/α7 translocation mediated by Wnt signaling via LIN-17/CAM-1 heteromeric receptors.
Asunto(s)
Caenorhabditis elegans/fisiología , Receptores Colinérgicos/metabolismo , Vía de Señalización Wnt , Animales , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Emparejamiento Cromosómico , Mutación , Sistema Nervioso , Unión Neuromuscular , Plasticidad Neuronal , Proteínas Tirosina Quinasas Receptoras/genética , Proteínas Tirosina Quinasas Receptoras/metabolismo , Receptores Huérfanos Similares al Receptor Tirosina Quinasa , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Proteínas Wnt/metabolismoRESUMEN
The recent discovery that vertebrate homologs of Drosophila cornichon associate with AMPA receptors led to the unexpected notion that cornichons play a role in synaptic transmission. In this issue of Neuron, Kato et al. find that cornichons modulate the gating of TARP-associated AMPA receptors by preventing their resensitization to glutamate.
RESUMEN
BACKGROUND: Ionotropic glutamate receptors (iGluRs) are glutamate-gated ion channels that mediate excitatory neurotransmission in the central nervous system. Based on both molecular and pharmacological criteria, iGluRs have been divided into two major classes, the non-NMDA class, which includes both AMPA and kainate subtypes of receptors, and the NMDA class. One evolutionarily conserved feature of iGluRs is their desensitization in the continued presence of glutamate. Thus, when in a desensitized state, iGluRs can be bound to glutamate, yet the channel remains closed. However, the relevance of desensitization to nervous system function has remained enigmatic. RESULTS: Here, we report the identification and characterization of a novel polypeptide (con-ikot-ikot) from the venom of a predatory marine snail Conus striatus that specifically disrupts the desensitization of AMPA receptors (AMPARs). The stoichiometry of con-ikot-ikot appears reminiscent of the proposed subunit organization of AMPARs, i.e., a dimer of dimers, suggesting that it acts as a molecular four-legged clamp that holds the AMPAR channel open. Application of con-ikot-ikot to hippocampal slices caused a large and rapid increase in resting AMPAR-mediated current leading to neuronal death. CONCLUSIONS: Our findings provide insight into the mechanisms that regulate receptor desensitization and demonstrate that in the arms race between prey and predators, evolution has selected for a toxin that blocks AMPAR desensitization, thus revealing the fundamental importance of desensitization for regulating neural function.
Asunto(s)
Caracol Conus/metabolismo , Venenos de Moluscos/química , Neurotoxinas/farmacología , Péptidos/farmacología , Receptores AMPA/metabolismo , Animales , Benzotiadiazinas/farmacología , Sitios de Unión , Fraccionamiento Químico , Cromatografía Líquida de Alta Presión , Caracol Conus/química , Dimerización , Conductividad Eléctrica , Hipocampo/efectos de los fármacos , Neurotoxinas/química , Neurotoxinas/aislamiento & purificación , Técnicas de Placa-Clamp , Péptidos/química , Péptidos/aislamiento & purificación , Ratas , Receptores AMPA/química , Proteínas Recombinantes/química , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/farmacología , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , XenopusRESUMEN
Neurotransmission in the brain is critically dependent on excitatory synaptic signaling mediated by AMPA-class ionotropic glutamate receptors (AMPARs). AMPARs are known to be associated with Transmembrane AMPA receptor Regulatory Proteins (TARPs). In vertebrates, at least four TARPs appear to have redundant roles as obligate chaperones for AMPARs, thus greatly complicating analysis of TARP participation in synaptic function. We have overcome this limitation by identifying and mutating the essential set of TARPs in C. elegans (STG-1 and STG-2). In TARP mutants, AMPAR-mediated currents and worm behaviors are selectively disrupted despite apparently normal surface expression and clustering of the receptors. Reconstitution experiments indicate that both STG-1 and STG-2 can functionally substitute for vertebrate TARPs to modify receptor function. Thus, we show that TARPs are obligate auxiliary subunits for AMPARs with a primary, evolutionarily conserved functional role in the modification of current kinetics.
Asunto(s)
Reacción de Prevención/fisiología , Proteínas de Caenorhabditis elegans/metabolismo , Canales de Calcio/metabolismo , Potenciales de la Membrana/fisiología , Proteínas del Tejido Nervioso/metabolismo , Receptores AMPA/metabolismo , Animales , Animales Modificados Genéticamente , Secuencia de Bases , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Canales de Calcio/genética , Evolución Molecular , Proteínas de Transporte de Membrana/metabolismo , Datos de Secuencia Molecular , Mutación , Proteínas del Tejido Nervioso/genética , Isoformas de Proteínas/metabolismo , Homología de Secuencia de Ácido NucleicoRESUMEN
Small, high-impedance neurons with short processes, similar to those found in the soil nematode Caenorhabditis elegans, are predicted to transmit electrical signals by passive propagation. However, we have found that certain neurons in C. elegans fire regenerative action potentials. These neurons resembled Schmitt triggers, as their potential state appears to be bistable. Transitions between up and down states could be triggered by application of the neurotransmitter glutamate or brief current pulses.
Asunto(s)
Potenciales de Acción/fisiología , Caenorhabditis elegans/fisiología , Conducción Nerviosa/fisiología , Neuronas/clasificación , Neuronas/fisiología , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/genética , Animales , Animales Modificados Genéticamente , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Calcio/metabolismo , Estimulación Eléctrica , Retroalimentación/fisiología , Ácido Glutámico/metabolismo , Ácido Glutámico/farmacología , Proteínas Fluorescentes Verdes/genética , Canales Iónicos/genética , Canales Iónicos/metabolismo , Iones/metabolismo , Conducción Nerviosa/efectos de los fármacos , Neuronas/efectos de los fármacos , Técnicas de Placa-Clamp , Receptores AMPA/genética , Receptores AMPA/metabolismo , Sodio/metabolismo , Estimulación QuímicaRESUMEN
Learning and memory are essential processes of both vertebrate and invertebrate nervous systems that allow animals to survive and reproduce. The neurotransmitter glutamate signals via ionotropic glutamate receptors (iGluRs) that have been linked to learning and memory formation; however, the signaling pathways that contribute to these behaviors are still not well understood. We therefore undertook a genetic and electrophysiological analysis of learning and memory in the nematode Caenorhabditis elegans. Here, we show that two genes, nmr-1 and nmr-2, are predicted to encode the subunits of an NMDA-type (NMDAR) iGluR that is necessary for memory retention in C. elegans. We cloned nmr-2, generated a deletion mutation in the gene, and showed that like nmr-1, nmr-2 is required for in vivo NMDA-gated currents. Using an associative-learning paradigm that pairs starvation with the attractant NaCl, we also showed that the memory of a learned avoidance response is dependent on NMR-1 and NMR-2 and that expression of NMDARs in a single pair of interneurons is sufficient for normal memory. Our results provide new insights into the molecular and cellular mechanisms underlying the memory of a learned event.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Memoria , N-Metilaspartato/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Secuencia de Aminoácidos , Animales , Conducta Apetitiva/efectos de los fármacos , Aprendizaje por Asociación , Reacción de Prevención , Caenorhabditis elegans/efectos de los fármacos , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas Portadoras/metabolismo , Quimiotaxis , Alimentos , Interneuronas/metabolismo , Datos de Secuencia Molecular , Mutación , Receptores de N-Metil-D-Aspartato/genética , Homología de Secuencia de Aminoácido , Cloruro de Sodio/farmacología , InaniciónRESUMEN
The neurotransmitter glutamate mediates excitatory synaptic transmission by activating ionotropic glutamate receptors (iGluRs). In Caenorhabditis elegans, the GLR-1 receptor subunit is required for glutamate-gated current in a subset of interneurons that control avoidance behaviors. Current mediated by GLR-1-containing iGluRs depends on SOL-1, a transmembrane CUB-domain protein that immunoprecipitates with GLR-1. We have found that reconstitution of glutamate-gated current in heterologous cells depends on three proteins, STG-1 (a C. elegans stargazin-like protein), SOL-1, and GLR-1. Here, we use genetic and pharmacological perturbations along with rapid perfusion electrophysiological techniques to demonstrate that SOL-1 functions to slow the rate and limit the extent of receptor desensitization as well as to enhance the recovery from desensitization. We have also identified a SOL-1 homologue from Drosophila and show that Dro SOL1 has a conserved function in promoting C. elegans glutamate-gated currents. SOL-1 homologues may play critical roles in regulating glutamatergic neurotransmission in more complex nervous systems.
Asunto(s)
Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/fisiología , Secuencia Conservada , Proteínas de Drosophila/fisiología , Proteínas de la Membrana/fisiología , Receptores AMPA/metabolismo , Secuencia de Aminoácidos , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/química , Canales de Calcio/química , Canales de Calcio/genética , Células Cultivadas , Concanavalina A/metabolismo , Proteínas de Drosophila/genética , Proteínas de la Membrana/genética , Datos de Secuencia Molecular , Receptores AMPA/genética , Receptores AMPA/fisiologíaRESUMEN
alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors (AMPARs) are a major subtype of ionotropic glutamate receptors (iGluRs) that mediate rapid excitatory synaptic transmission in the vertebrate brain. Putative AMPARs are also expressed in the nervous system of invertebrates. In Caenorhabditis elegans, the GLR-1 receptor subunit is expressed in neural circuits that mediate avoidance behaviors and is required for glutamate-gated current in the AVA and AVD interneurons. Glutamate-gated currents can be recorded from heterologous cells that express vertebrate AMPARs; however, when C. elegans GLR-1 is expressed in heterologous cells, little or no glutamate-gated current is detected. This finding suggests that other receptor subunits or auxiliary proteins are required for function. Here, we identify Ce STG-1, a C. elegans stargazin-like protein, and show that expression of Ce STG-1 together with GLR-1 and the CUB-domain protein SOL-1 reconstitutes glutamate-gated currents in Xenopus oocytes. Ce STG-1 and homologues cloned from Drosophila (Dro STG1) and Apis mellifera (Apis STG1) have evolutionarily conserved functions and can partially substitute for one another to reconstitute glutamate-gated currents from rat, Drosophila, and C. elegans. Furthermore, we show that Ce STG-1 and Apis STG1 are primarily required for function independent of possible roles in promoting the surface expression of invertebrate AMPARs.
Asunto(s)
Proteínas de Caenorhabditis elegans/fisiología , Canales de Calcio/fisiología , Proteínas de Insectos/fisiología , Receptores AMPA/fisiología , Secuencia de Aminoácidos , Animales , Abejas/genética , Abejas/fisiología , Proteínas de Caenorhabditis elegans/biosíntesis , Proteínas de Caenorhabditis elegans/genética , Canales de Calcio/biosíntesis , Canales de Calcio/genética , Línea Celular , Humanos , Proteínas de Insectos/biosíntesis , Proteínas de Insectos/genética , Datos de Secuencia Molecular , Oocitos/metabolismo , XenopusRESUMEN
Most rapid excitatory synaptic signaling in the brain is mediated by postsynaptic ionotropic glutamate receptors (iGluRs) that are gated open by the neurotransmitter glutamate. In Caenorhabditis elegans, sol-1 encodes a CUB-domain transmembrane protein that is required for currents that are mediated by the GLR-1 iGluR. Mutations in sol-1 do not affect GLR-1 expression, localization, membrane insertion, or stabilization at synapses, suggesting that SOL-1 is required for iGluR function. Here, we provide evidence that SOL-1 is an auxiliary subunit that modulates the gating of GLR-1 receptors. We show that mutant variants of GLR-1 with altered gating partially restore glutamate-gated current and GLR-1-dependent behaviors in sol-1 mutants. Domain analysis of SOL-1 indicates that extracellular CUB domain 3 is required for function and that a secreted variant partially restores glutamate-gated currents and behavior. Also, we show that endogenous glutamatergic synaptic currents are absent in sol-1 mutants. Our data suggest that GLR-1 iGluRs are not simply stand-alone molecules and require the SOL-1 auxiliary protein to promote the open state of the receptor. Our analysis presents the possibility that glutamatergic signaling in other organisms may be similarly modified by SOL-1-like transmembrane proteins.
Asunto(s)
Proteínas de Caenorhabditis elegans/fisiología , Receptores AMPA/fisiología , Animales , Conducta Animal , Caenorhabditis elegans , Membrana Celular/metabolismo , Electrofisiología , Variación Genética , Ácido Glutámico/química , Proteínas Fluorescentes Verdes/metabolismo , Inmunoprecipitación , Cinética , Ligandos , Proteínas de la Membrana/química , Modelos Genéticos , Mutación , Oocitos/metabolismo , Plásmidos/metabolismo , Estructura Terciaria de Proteína , Receptores de Glutamato/metabolismo , Transducción de Señal , Transmisión Sináptica , Factores de Tiempo , XenopusRESUMEN
Most rapid excitatory synaptic signaling is mediated by glutamatergic neurotransmission. An important challenge in neurobiology is to understand the molecular architecture of functional glutamatergic synapses. By combining the techniques of genetics, molecular biology and electrophysiology in C. elegans we have the potential to identify and characterize the molecules that contribute to the function of glutamatergic synapses. In C. elegans both excitatory and inhibitory ionotropic glutamate receptors are linked to neural circuits and behavior. Genetic analysis has identified genes required for receptor expression, trafficking, localization, stabilization and function at synapses. Significantly, novel proteins required for glutamate receptor function have been discovered in the worm. These advances may also lead to a better understanding of glutamatergic signaling in vertebrates.
Asunto(s)
Caenorhabditis elegans/fisiología , Receptores de Glutamato/fisiología , Animales , Conducta Animal/fisiología , Canales de Cloruro/fisiología , Fenómenos Fisiológicos del Sistema Nervioso , Receptores de Glutamato/genética , Transducción de SeñalRESUMEN
Area-restricted search (ARS) is a foraging strategy used by many animals to locate resources. The behavior is characterized by a time-dependent reduction in turning frequency after the last resource encounter. This maximizes the time spent in areas in which resources are abundant and extends the search to a larger area when resources become scarce. We demonstrate that dopaminergic and glutamatergic signaling contribute to the neural circuit controlling ARS in the nematode Caenorhabditis elegans. Ablation of dopaminergic neurons eliminated ARS behavior, as did application of the dopamine receptor antagonist raclopride. Furthermore, ARS was affected by mutations in the glutamate receptor subunits GLR-1 and GLR-2 and the EAT-4 glutamate vesicular transporter. Interestingly, preincubation on dopamine restored the behavior in worms with defective dopaminergic signaling, but not in glr-1, glr-2, or eat-4 mutants. This suggests that dopaminergic and glutamatergic signaling function in the same pathway to regulate turn frequency. Both GLR-1 and GLR-2 are expressed in the locomotory control circuit that modulates the direction of locomotion in response to sensory stimuli and the duration of forward movement during foraging. We propose a mechanism for ARS in C. elegans in which dopamine, released in response to food, modulates glutamatergic signaling in the locomotory control circuit, thus resulting in an increased turn frequency.
Asunto(s)
Conducta Apetitiva/fisiología , Caenorhabditis elegans/fisiología , Dopamina/fisiología , Ácido Glutámico/fisiología , Animales , Conducta Apetitiva/efectos de los fármacos , Conducta Animal/fisiología , Caenorhabditis elegans/efectos de los fármacos , Dopamina/farmacología , Conducta Alimentaria/efectos de los fármacos , Conducta Alimentaria/fisiología , Ácido Glutámico/farmacología , Locomoción/fisiología , Receptores de Glutamato/genética , Receptores de Glutamato/fisiología , Transducción de Señal/fisiología , Tacto/fisiologíaRESUMEN
Ionotropic glutamate receptors (iGluRs) mediate most excitatory synaptic signalling between neurons. Binding of the neurotransmitter glutamate causes a conformational change in these receptors that gates open a transmembrane pore through which ions can pass. The gating of iGluRs is crucially dependent on a conserved amino acid that was first identified in the 'lurcher' ataxic mouse. Through a screen for modifiers of iGluR function in a transgenic strain of Caenorhabditis elegans expressing a GLR-1 subunit containing the lurcher mutation, we identify suppressor of lurcher (sol-1). This gene encodes a transmembrane protein that is predicted to contain four extracellular beta-barrel-forming domains known as CUB domains. SOL-1 and GLR-1 are colocalized at the cell surface and can be co-immunoprecipitated. By recording from neurons expressing GLR-1, we show that SOL-1 is an accessory protein that is selectively required for glutamate-gated currents. We propose that SOL-1 participates in the gating of non-NMDA (N-methyl-D-aspartate) iGluRs, thereby providing a previously unknown mechanism of regulation for this important class of neurotransmitter receptor.