Publications by authors named "Riad Seddik"

16 Publications

  • Page 1 of 1

Glial Endozepines Reverse High-Fat Diet-Induced Obesity by Enhancing Hypothalamic Response to Peripheral Leptin.

Mol Neurobiol 2020 Aug 9;57(8):3307-3333. Epub 2020 Jun 9.

CNRS UMR 7291, Laboratoire de Neurosciences Cognitives, Faculté des Sciences et Techniques de St Jérôme, Avenue Escadrille Normandie-Niemen, Université Aix-Marseille, 13397, Marseille cedex 20, France.

Research on energy homeostasis has focused on neuronal signaling; however, the role of glial cells has remained little explored. Glial endozepines exert anorexigenic actions by mechanisms which remain poorly understood. In this context, the present study was designed to decipher the mechanisms underlying the anorexigenic action of endozepines and to investigate their potential curative effect on high-fat diet-induced obesity. We carried out a combination of physiological, pharmacological, and molecular analyses together to dissect the underlying mechanisms of endozepine-induced hypophagia. To evaluate the potential anti-obesity effect of endozepines, different model of obesity were used, i.e., ob/ob and diet-induced obese mice. We show that the intracerebral administration of endozepines enhances satiety by targeting anorexigenic brain circuitry and induces STAT3 phosphorylation, a hallmark of leptin signaling. Strikingly, endozepines are entirely ineffective at reducing food intake in the presence of a circulating leptin antagonist and in leptin-deficient mice (ob/ob) but potentiate the reduced food intake and weight loss induced by exogenous leptin administration in these animals. Endozepines reversed high fat diet-induced obesity by reducing food intake and restored leptin-induced STAT3 phosphorylation in the hypothalamus. Interestingly, we observed that glucose and insulin synergistically enhance tanycytic endozepine expression and release. Finally, endozepines, which induce ERK activation necessary for leptin transport into the brain in cultured tanycytes, require tanycytic leptin receptor expression to promote STAT3 phosphorylation in the hypothalamus. Our data identify endozepines as potential anti-obesity compounds in part through the modulation of the LepR-ERK-dependent tanycytic leptin shuttle.
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http://dx.doi.org/10.1007/s12035-020-01944-zDOI Listing
August 2020

GABA receptors modulate Ca but not G protein-gated inwardly rectifying K channels in cerebrospinal-fluid contacting neurones of mouse brainstem.

J Physiol 2019 01 10;597(2):631-651. Epub 2018 Dec 10.

Aix Marseille Univ, CNRS, INT, Inst Neurosci Timone, Marseille, France.

Key Points: Medullo-spinal CSF contacting neurones (CSF-cNs) located around the central canal are conserved in all vertebrates and suggested to be a novel sensory system intrinsic to the CNS. CSF-cNs receive GABAergic inhibitory synaptic inputs involving ionotropic GABA receptors, but the contribution of metabotropic GABA receptors (GABA -Rs) has not yet been studied. Here, we indicate that CSF-cNs express functional GABA -Rs that inhibit postsynaptic calcium channels but fail to activate inhibitory potassium channel of the Kir3-type. We further show that GABA -Rs localise presynaptically on GABAergic and glutamatergic synaptic inputs contacting CSF-cNs, where they inhibit the release of GABA and glutamate. Our data are the first to address the function of GABA -Rs in CSF-cNs and show that on the presynaptic side they exert a classical synaptic modulation whereas at the postsynaptic level they have an atypical action by modulating calcium signalling without inducing potassium-dependent inhibition.

Abstract: Medullo-spinal neurones that contact the cerebrospinal fluid (CSF-cNs) are a population of evolutionary conserved cells located around the central canal. CSF-cN activity has been shown to be regulated by inhibitory synaptic inputs involving ionotropic GABA receptors, but the contribution of the G-protein coupled GABA receptors has not yet been studied. Here, we used a combination of immunofluorescence, electrophysiology and calcium imaging to investigate the expression and function of GABA -Rs in CSF-cNs of the mouse brainstem. We found that CSF-cNs express GABA -Rs, but their selective activation failed to induce G protein-coupled inwardly rectifying potassium (GIRK) currents. Instead, CSF-cNs express primarily N-type voltage-gated calcium (Ca 2.2) channels, and GABA -Rs recruit Gβγ subunits to inhibit Ca channel activity induced by membrane voltage steps or under physiological conditions by action potentials. Moreover, using electrical stimulation, we indicate that GABAergic inhibitory (IPSCs) and excitatory glutamatergic (EPSCs) synaptic currents can be evoked in CSF-cNs showing that mammalian CSF-cNs are also under excitatory control by glutamatergic synaptic inputs. We further demonstrate that baclofen reversibly reduced the amplitudes of both IPSCs and EPSCs evoked in CSF-cNs through a presynaptic mechanism of regulation. In summary, these results are the first to demonstrate the existence of functional postsynaptic GABA -Rs in medullar CSF-cNs, as well as presynaptic GABA auto- and heteroreceptors regulating the release of GABA and glutamate. Remarkably, postsynaptic GABA -Rs associate with Ca but not GIRK channels, indicating that GABA -Rs function as a calcium signalling modulator without GIRK-dependent inhibition in CSF-cNs.
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http://dx.doi.org/10.1113/JP277172DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6332714PMC
January 2019

KCTD Hetero-oligomers Confer Unique Kinetic Properties on Hippocampal GABAB Receptor-Induced K+ Currents.

J Neurosci 2017 02 21;37(5):1162-1175. Epub 2016 Dec 21.

Department of Biomedicine, Institute of Physiology, University of Basel, 4056 Basel, Switzerland,

GABA receptors are the G-protein coupled receptors for the main inhibitory neurotransmitter in the brain, GABA. GABA receptors were shown to associate with homo-oligomers of auxiliary KCTD8, KCTD12, KCTD12b, and KCTD16 subunits (named after their T1 K-channel tetramerization domain) that regulate G-protein signaling of the receptor. Here we provide evidence that GABA receptors also associate with hetero-oligomers of KCTD subunits. Coimmunoprecipitation experiments indicate that two-thirds of the KCTD16 proteins in the hippocampus of adult mice associate with KCTD12. We show that the KCTD proteins hetero-oligomerize through self-interacting T1 and H1 homology domains. Bioluminescence resonance energy transfer measurements in live cells reveal that KCTD12/KCTD16 hetero-oligomers associate with both the receptor and the G-protein. Electrophysiological experiments demonstrate that KCTD12/KCTD16 hetero-oligomers impart unique kinetic properties on G-protein-activated Kir3 currents. During prolonged receptor activation (one min) KCTD12/KCTD16 hetero-oligomers produce moderately desensitizing fast deactivating K currents, whereas KCTD12 and KCTD16 homo-oligomers produce strongly desensitizing fast deactivating currents and nondesensitizing slowly deactivating currents, respectively. During short activation (2 s) KCTD12/KCTD16 hetero-oligomers produce nondesensitizing slowly deactivating currents. Electrophysiological recordings from hippocampal neurons of KCTD knock-out mice are consistent with these findings and indicate that KCTD12/KCTD16 hetero-oligomers increase the duration of slow IPSCs. In summary, our data demonstrate that simultaneous assembly of distinct KCTDs at the receptor increases the molecular and functional repertoire of native GABA receptors and modulates physiologically induced K current responses in the hippocampus.

Significance Statement: The KCTD proteins 8, 12, and 16 are auxiliary subunits of GABA receptors that differentially regulate G-protein signaling of the receptor. The KCTD proteins are generally assumed to function as homo-oligomers. Here we show that the KCTD proteins also assemble hetero-oligomers in all possible dual combinations. Experiments in live cells demonstrate that KCTD hetero-oligomers form at least tetramers and that these tetramers directly interact with the receptor and the G-protein. KCTD12/KCTD16 hetero-oligomers impart unique kinetic properties to GABA receptor-induced Kir3 currents in heterologous cells. KCTD12/KCTD16 hetero-oligomers are abundant in the hippocampus, where they prolong the duration of slow IPSCs in pyramidal cells. Our data therefore support that KCTD hetero-oligomers modulate physiologically induced K current responses in the brain.
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http://dx.doi.org/10.1523/JNEUROSCI.2181-16.2016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6596860PMC
February 2017

A single polycystic kidney disease 2-like 1 channel opening acts as a spike generator in cerebrospinal fluid-contacting neurons of adult mouse brainstem.

Neuropharmacology 2016 Feb 26;101:549-65. Epub 2015 Jul 26.

Aix Marseille Université, PPSN EA 4674, 13397, Marseille, France. Electronic address:

Cerebrospinal fluid contacting neurons (CSF-cNs) are found around the central canal of all vertebrates. They present a typical morphology, with a single dendrite that projects into the cavity and ends in the CSF with a protuberance. These anatomical features have led to the suggestion that CSF-cNs might have sensory functions, either by sensing CSF movement or composition, but the physiological mechanisms for any such role are unknown. This hypothesis was recently supported by the demonstration that in several vertebrate species medullo-spinal CSF-cNs selectively express Polycystic Kidney Disease 2-Like 1 proteins (PKD2L1). PKD2L1 are members of the 'transient receptor potential (TRP)' superfamily, form non-selective cationic channels of high conductance, are regulated by various stimuli including protons and are therefore suggested to act as sensory receptors. Using patch-clamp whole-cell recordings of CSF-cNs in brainstem slices obtained from wild type and mutant PKD2L1 mice, we demonstrate that spontaneously active unitary currents in CSF-cNs are due to PKD2L1 channels that are capable, with a single opening, of triggering action potentials. Thus PKD2L1 might contribute to the setting of CSF-cN spiking activity. We also reveal that CSF-cNs have the capacity of discriminating between alkalinization and acidification following activation of specific conductances (PKD2L1 vs. ASIC) generating specific responses. Altogether, this study reinforces the idea that CSF-cNs represent sensory neurons intrinsic to the central nervous system and suggests a role for PKD2L1 channels as spike generators.
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http://dx.doi.org/10.1016/j.neuropharm.2015.07.030DOI Listing
February 2016

Up-regulation of GABA(B) receptor signaling by constitutive assembly with the K+ channel tetramerization domain-containing protein 12 (KCTD12).

J Biol Chem 2013 Aug 10;288(34):24848-56. Epub 2013 Jul 10.

Department of Biomedicine, University of Basel, CH-4056 Basel, Switzerland.

GABA(B) receptors are the G-protein coupled receptors (GPCRs) for GABA, the main inhibitory neurotransmitter in the central nervous system. Native GABA(B) receptors comprise principle and auxiliary subunits that regulate receptor properties in distinct ways. The principle subunits GABA(B1a), GABA(B1b), and GABA(B2) form fully functional heteromeric GABA(B(1a,2)) and GABA(B(1b,2)) receptors. Principal subunits regulate forward trafficking of the receptors from the endoplasmic reticulum to the plasma membrane and control receptor distribution to axons and dendrites. The auxiliary subunits KCTD8, -12, -12b, and -16 are cytosolic proteins that influence agonist potency and G-protein signaling of GABA(B(1a,2)) and GABA(B(1b,2)) receptors. Here, we used transfected cells to study assembly, surface trafficking, and internalization of GABA(B) receptors in the presence of the KCTD12 subunit. Using bimolecular fluorescence complementation and metabolic labeling, we show that GABA(B) receptors associate with KCTD12 while they reside in the endoplasmic reticulum. Glycosylation experiments support that association with KCTD12 does not influence maturation of the receptor complex. Immunoprecipitation and bioluminescence resonance energy transfer experiments demonstrate that KCTD12 remains associated with the receptor during receptor activity and receptor internalization from the cell surface. We further show that KCTD12 reduces constitutive receptor internalization and thereby increases the magnitude of receptor signaling at the cell surface. Accordingly, knock-out or knockdown of KCTD12 in cultured hippocampal neurons reduces the magnitude of the GABA(B) receptor-mediated K(+) current response. In summary, our experiments support that the up-regulation of functional GABA(B) receptors at the neuronal plasma membrane is an additional physiological role of the auxiliary subunit KCTD12.
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http://dx.doi.org/10.1074/jbc.M113.476770DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3750179PMC
August 2013

Opposite effects of KCTD subunit domains on GABA(B) receptor-mediated desensitization.

J Biol Chem 2012 Nov 3;287(47):39869-77. Epub 2012 Oct 3.

Department of Biomedicine, University of Basel, 4056 Basel, Switzerland.

GABA(B) receptors assemble from principle and auxiliary subunits. The principle subunits GABA(B1) and GABA(B2) form functional heteromeric GABA(B(1,2)) receptors that associate with homotetramers of auxiliary KCTD8, -12, -12b, or -16 (named after their K(+) channel tetramerization domain) subunits. These auxiliary subunits constitute receptor subtypes with distinct functional properties. KCTD12 and -12b generate desensitizing receptor responses while KCTD8 and -16 generate largely non-desensitizing receptor responses. The structural elements of the KCTDs underlying these differences in desensitization are unknown. KCTDs are modular proteins comprising a T1 tetramerization domain, which binds to GABA(B2), and a H1 homology domain. KCTD8 and -16 contain an additional C-terminal H2 homology domain that is not sequence-related to the H1 domains. No functions are known for the H1 and H2 domains. Here we addressed which domains and sequence motifs in KCTD proteins regulate desensitization of the receptor response. We found that the H1 domains in KCTD12 and -12b mediate desensitization through a particular sequence motif, T/NFLEQ, which is not present in the H1 domains of KCTD8 and -16. In addition, the H2 domains in KCTD8 and -16 inhibit desensitization when expressed C-terminal to the H1 domains but not when expressed as a separate protein in trans. Intriguingly, the inhibitory effect of the H2 domain is sequence-independent, suggesting that the H2 domain sterically hinders desensitization by the H1 domain. Evolutionary analysis supports that KCTD12 and -12b evolved desensitizing properties by liberating their H1 domains from antagonistic H2 domains and acquisition of the T/NFLEQ motif.
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http://dx.doi.org/10.1074/jbc.M112.412767DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3501043PMC
November 2012

GABAB receptors: physiological functions and mechanisms of diversity.

Adv Pharmacol 2010 ;58:231-55

Department of Biomedicine, Institute of Physiology, Pharmazentrum, University of Basel, Basel, Switzerland.

GABA(B) receptors are the G-protein-coupled receptors (GPCRs) for gamma-aminobutyric acid (GABA), the main inhibitory neurotransmitter in the central nervous system. GABA(B) receptors are implicated in the etiology of a variety of psychiatric disorders and are considered attractive drug targets. With the cloning of GABA(B) receptor subunits 13 years ago, substantial progress was made in the understanding of the molecular structure, physiology, and pharmacology of these receptors. However, it remained puzzling that native studies demonstrated a heterogeneity of GABA(B) responses that contrasted with a very limited diversity of cloned GABA(B) receptor subunits. Until recently, the only firmly established molecular diversity consisted of two GABA(B1) subunit isoforms, GABA(B1a) and GABA(B1b), which assemble with GABA(B2) subunits to generate heterodimeric GABA(B(1a,2)) and GABA(B(1b,2)) receptors. Using genetic, ultrastructural, biochemical, and electrophysiological approaches, it has been possible to identify functional properties that segregate with these two receptors. Moreover, receptor modifications and factors that can alter the receptor response have been identified. Most importantly, recent data reveal the existence of a family of auxiliary GABA(B) receptor subunits that assemble as tetramers with the C-terminal domain of GABA(B2) subunits and drastically alter pharmacology and kinetics of the receptor response. The data are most consistent with native GABA(B) receptors minimally forming dimeric assemblies of units composed of GABA(B1), GABA(B2), and a tetramer of auxiliary subunits. This represents a substantial departure from current structural concepts for GPCRs.
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http://dx.doi.org/10.1016/S1054-3589(10)58010-4DOI Listing
November 2010

NMDA receptor-dependent GABAB receptor internalization via CaMKII phosphorylation of serine 867 in GABAB1.

Proc Natl Acad Sci U S A 2010 Aug 19;107(31):13924-9. Epub 2010 Jul 19.

Department of Biomedicine, Institute of Physiology, Pharmazentrum, University of Basel, 4056 Basel, Switzerland.

GABAB receptors are the G-protein-coupled receptors for GABA, the main inhibitory neurotransmitter in the brain. GABAB receptors are abundant on dendritic spines, where they dampen postsynaptic excitability and inhibit Ca2+ influx through NMDA receptors when activated by spillover of GABA from neighboring GABAergic terminals. Here, we show that an excitatory signaling cascade enables spines to counteract this GABAB-mediated inhibition. We found that NMDA application to cultured hippocampal neurons promotes dynamin-dependent endocytosis of GABAB receptors. NMDA-dependent internalization of GABAB receptors requires activation of Ca2+/Calmodulin-dependent protein kinase II (CaMKII), which associates with GABAB receptors in vivo and phosphorylates serine 867 (S867) in the intracellular C terminus of the GABAB1 subunit. Blockade of either CaMKII or phosphorylation of S867 renders GABAB receptors refractory to NMDA-mediated internalization. Time-lapse two-photon imaging of organotypic hippocampal slices reveals that activation of NMDA receptors removes GABAB receptors within minutes from the surface of dendritic spines and shafts. NMDA-dependent S867 phosphorylation and internalization is predominantly detectable with the GABAB1b subunit isoform, which is the isoform that clusters with inhibitory effector K+ channels in the spines. Consistent with this, NMDA receptor activation in neurons impairs the ability of GABAB receptors to activate K+ channels. Thus, our data support that NMDA receptor activity endocytoses postsynaptic GABAB receptors through CaMKII-mediated phosphorylation of S867. This provides a means to spare NMDA receptors at individual glutamatergic synapses from reciprocal inhibition through GABAB receptors.
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http://dx.doi.org/10.1073/pnas.1000909107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2922270PMC
August 2010

Native GABA(B) receptors are heteromultimers with a family of auxiliary subunits.

Nature 2010 May 18;465(7295):231-5. Epub 2010 Apr 18.

Institute of Physiology II, University of Freiburg, Engesserstrasse 4, 79108 Freiburg, Germany.

GABA(B) receptors are the G-protein-coupled receptors for gamma-aminobutyric acid (GABA), the main inhibitory neurotransmitter in the brain. They are expressed in almost all neurons of the brain, where they regulate synaptic transmission and signal propagation by controlling the activity of voltage-gated calcium (Ca(v)) and inward-rectifier potassium (K(ir)) channels. Molecular cloning revealed that functional GABA(B) receptors are formed by the heteromeric assembly of GABA(B1) with GABA(B2) subunits. However, cloned GABA(B(1,2)) receptors failed to reproduce the functional diversity observed with native GABA(B) receptors. Here we show by functional proteomics that GABA(B) receptors in the brain are high-molecular-mass complexes of GABA(B1), GABA(B2) and members of a subfamily of the KCTD (potassium channel tetramerization domain-containing) proteins. KCTD proteins 8, 12, 12b and 16 show distinct expression profiles in the brain and associate tightly with the carboxy terminus of GABA(B2) as tetramers. This co-assembly changes the properties of the GABA(B(1,2)) core receptor: the KCTD proteins increase agonist potency and markedly alter the G-protein signalling of the receptors by accelerating onset and promoting desensitization in a KCTD-subtype-specific manner. Taken together, our results establish the KCTD proteins as auxiliary subunits of GABA(B) receptors that determine the pharmacology and kinetics of the receptor response.
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http://dx.doi.org/10.1038/nature08964DOI Listing
May 2010

A mouse model for visualization of GABA(B) receptors.

Genesis 2009 Sep;47(9):595-602

Department of Biomedicine, Institute of Physiology, University of Basel, Basel, Switzerland.

GABA(B) receptors are the G-protein-coupled receptors for the neurotransmitter gamma-aminobutyric acid (GABA). Receptor subtypes are based on the subunit isoforms GABA(B1a) and GABA(B1b), which combine with GABA(B2) subunits to form heteromeric receptors. Here, we used a modified bacterial artificial chromosome (BAC) containing the GABA(B1) gene to generate transgenic mice expressing GABA(B1a) and GABA(B1b) subunits fused to the enhanced green fluorescence protein (eGFP). We demonstrate that the GABA(B1)-eGFP fusion proteins reproduce the cellular expression patterns of endogenous GABA(B1) proteins in the brain and in peripheral tissue. Crossing the GABA(B1)-eGFP BAC transgene into the GABA(B1) (-/-) background restores pre and postsynaptic GABA(B) functions, showing that the GABA(B1)-eGFP fusion proteins substitute for the lack of endogenous GABA(B1) proteins. Finally, we demonstrate that the GABA(B1)-eGFP fusion proteins replicate the temporal expression patterns of native GABA(B) receptors in cultured neurons. These transgenic mice therefore provide a validated tool for direct visualization of native GABA(B) receptors.
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http://dx.doi.org/10.1002/dvg.20535DOI Listing
September 2009

The GABAB1a isoform mediates heterosynaptic depression at hippocampal mossy fiber synapses.

J Neurosci 2009 Feb;29(5):1414-23

Department of Biomedicine, Institute of Physiology, Pharmazentrum, University of Basel, 4056 Basel, Switzerland.

GABA(B) receptor subtypes are based on the subunit isoforms GABA(B1a) and GABA(B1b), which associate with GABA(B2) subunits to form pharmacologically indistinguishable GABA(B(1a,2)) and GABA(B(1b,2)) receptors. Studies with mice selectively expressing GABA(B1a) or GABA(B1b) subunits revealed that GABA(B(1a,2)) receptors are more abundant than GABA(B(1b,2)) receptors at glutamatergic terminals. Accordingly, it was found that GABA(B(1a,2)) receptors are more efficient than GABA(B(1b,2)) receptors in inhibiting glutamate release when maximally activated by exogenous application of the agonist baclofen. Here, we used a combination of genetic, ultrastructural and electrophysiological approaches to analyze to what extent GABA(B(1a,2)) and GABA(B(1b,2)) receptors inhibit glutamate release in response to physiological activation. We first show that at hippocampal mossy fiber (MF)-CA3 pyramidal neuron synapses more GABA(B1a) than GABA(B1b) protein is present at presynaptic sites, consistent with the findings at other glutamatergic synapses. In the presence of baclofen at concentrations >or=1 microm, both GABA(B(1a,2)) and GABA(B(1b,2)) receptors contribute to presynaptic inhibition of glutamate release. However, at lower concentrations of baclofen, selectively GABA(B(1a,2)) receptors contribute to presynaptic inhibition. Remarkably, exclusively GABA(B(1a,2)) receptors inhibit glutamate release in response to synaptically released GABA. Specifically, we demonstrate that selectively GABA(B(1a,2)) receptors mediate heterosynaptic depression of MF transmission, a physiological phenomenon involving transsynaptic inhibition of glutamate release via presynaptic GABA(B) receptors. Our data demonstrate that the difference in GABA(B1a) and GABA(B1b) protein levels at MF terminals is sufficient to produce a strictly GABA(B1a)-specific effect under physiological conditions. This consolidates that the differential subcellular localization of the GABA(B1a) and GABA(B1b) proteins is of regulatory relevance.
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http://dx.doi.org/10.1523/JNEUROSCI.3697-08.2009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6666076PMC
February 2009

Synapse loss in cortex of agrin-deficient mice after genetic rescue of perinatal death.

J Neurosci 2007 Jul;27(27):7183-95

Biozentrum, University of Basel, CH-4056 Basel, Switzerland.

Agrin-deficient mice die at birth because of aberrant development of the neuromuscular junctions. Here, we examined the role of agrin at brain synapses. We show that agrin is associated with excitatory but not inhibitory synapses in the cerebral cortex. Most importantly, we examined the brains of agrin-deficient mice whose perinatal death was prevented by the selective expression of agrin in motor neurons. We find that the number of presynaptic and postsynaptic specializations is strongly reduced in the cortex of 5- to 7-week-old mice. Consistent with a reduction in the number of synapses, the frequency of miniature postsynaptic currents was greatly decreased. In accordance with the synaptic localization of agrin to excitatory synapses, changes in the frequency were only detected for excitatory but not inhibitory synapses. Moreover, we find that the muscle-specific receptor tyrosine kinase MuSK, which is known to be an essential component of agrin-induced signaling at the neuromuscular junction, is also localized to a subset of excitatory synapses. Finally, some components of the mitogen-activated protein (MAP) kinase pathway, which has been shown to be activated by agrin in cultured neurons, are deregulated in agrin-deficient mice. In summary, our results provide strong evidence that agrin plays an important role in the formation and/or the maintenance of excitatory synapses in the brain, and we provide evidence that this function involves MAP kinase signaling.
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http://dx.doi.org/10.1523/JNEUROSCI.1609-07.2007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6794585PMC
July 2007

Corelease of GABA/glycine in lamina-X of the spinal cord of neonatal rats.

Neuroreport 2007 Jul;18(10):1025-9

Department of Physiology, University of Basel, Pharmazentrum, Klingelbergstrasse 50/70, CH-4056 Basel, Switzerland.

Spinal-cord slices from neonatal rats were used to record lamina-X neurons using the patch-clamp technique under whole cell recording configuration. Lamina-X surrounds the central canal of the spinal cord and contains sympathetic preganglionic neurons of the central autonomic nucleus. Miniature inhibitory postsynaptic currents were recorded in the presence of tetrodotoxin and kynurenic acid to block action potential-dependent transmitter release and glutamatergic transmissions, respectively. We recorded mixed gamma-amino-n-butyric acid/glycine miniature synaptic currents suggesting that gamma-amino-n-butyric acid and glycine can be coreleased from the same single synaptic vesicles, and that this corelease can be detected by the postsynaptic cell. In addition, acetylcholine can induce the release of gamma-amino-n-butyric acid/glycine by acting presynaptically at nicotinic receptors located on the gamma-amino-n-butyric acid ergic/glycinergic terminals.
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http://dx.doi.org/10.1097/WNR.0b013e3281667c0cDOI Listing
July 2007

Modulation of GABAergic synaptic transmission by terminal nicotinic acetylcholine receptors in the central autonomic nucleus of the neonatal rat spinal cord.

Neuropharmacology 2006 Jul 5;51(1):77-89. Epub 2006 May 5.

Department of Physiology, University of Basel, Pharmazentrum, Klingelbergstrasse 50/70, CH-4056 Basel, Switzerland.

Using patch clamp recordings from an in vitro spinal cord slice preparation of neonatal rats (9-15days old), we characterized the GABAergic synaptic transmission in sympathetic preganglionic neurones (SPN) of the central autonomic nucleus (CA) of lamina X. Local applications of isoguvacine (100microM), a selective agonist at GABA(A) receptors, induced in all cells tested a chloride current which was abolished by bicuculline, a competitive antagonist at GABA(A) receptors. In addition, 25% of the recorded cells displayed spontaneous tetrodotoxin-insensitive and bicuculline-sensitive chloride miniature inhibitory postsynaptic currents (mIPSCs). Acetylcholine (100microM) increased the frequency of GABAergic mIPSCs without affecting their amplitudes or their kinetic properties indicating a presynaptic site of action. The presynaptic effect of ACh was restricted to GABAergic neurones synapsing onto sympathetic preganglionic neurones. The facilitatory effect of ACh was abolished in the absence of external calcium or in the presence of 100microM cadmium added to the bath solution. Choline 10mM, an agonist at alpha7 nicotinic acetylcholine receptors (nAChRs) or muscarine (10microM), a muscarinic receptor agonist, did not reproduce the presynaptic effect of ACh. The presynaptic effect of ACh was blocked by 1microM of dihydro-beta-erythroidine (DHbetaE), an antagonist of non-alpha7 nAChRs but was insensitive to alpha7 nAChRs antagonists (strychnine, alpha-bungarotoxin and methyllycaconitine) or to the muscarinic receptor antagonist atropine (10microM). It was concluded that SPNs of the central autonomic nucleus displayed a functional GABAergic transmission which is facilitated by terminal non alpha7 nAChRs.
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http://dx.doi.org/10.1016/j.neuropharm.2006.03.007DOI Listing
July 2006

The rat spinal cord slice: Its use in generating pharmacological evidence for cholinergic transmission using the alpha7 subtype of nicotinic receptors in the central autonomic nucleus.

J Pharmacol Toxicol Methods 2005 May-Jun;51(3):243-52

Laboratoire de Neurophysiologie Cellulaire et Intégrée, UMR 7519 CNRS/ULP, 21 rue R.Descartes, 67084 Strasbourg Cedex, France.

Lamina X surrounds the central canal of the spinal cord and is an important site for the convergence of somatic and visceral afferent inputs relaying nociceptive information. Lamina X contains sympathetic preganglionic neurons (SPN) in the so-called central autonomic nucleus which may participate to viscero-autonomic reflexes. Here, we describe a transversal slice preparation of postnatal rat thoracolumbar spinal cord which allows the detailed characterization of the morphology, electrophysiological properties, synaptic activities and receptor pharmacology of neurons surrounding the central canal. By means of the patch clamp technique, in its whole cell configuration, and by the use of various pharmacological tools, we show here that lamina X neurons of the central autonomic nucleus express functional alpha7 nicotinic receptors which are located postsynaptically on SPNs where they are involved in a fast cholinergic transmission. Thus, this in vitro preparation is useful to study the mechanisms and the pharmacology of viscero-autonomic reflexes.
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http://dx.doi.org/10.1016/j.vascn.2004.08.013DOI Listing
August 2005

Choline induces Ca2+ entry in cultured sympathetic neurones isolated from rat superior cervical ganglion.

Eur J Pharmacol 2003 Jun;471(3):165-76

Laboratoire de Neurophysiologie Cellulaire et Intégrée, Institut de Physiologie et de Chimie Biologique, CNRS UMR 7519, Université Louis Pasteur, 21 rue R. Descartes, 67084 Cedex, Strasbourg, France.

Choline has been shown to be a specific agonist at alpha7 nicotinic acetylcholine receptors, which are the most Ca(2+) permeable of the ionotropic receptor channels. Whole-cell patch recording combined with the measurement of intracellular free Ca(2+) concentration ([Ca(2+)](i), using Indo1, in cultured rat superior cervical ganglion neurones demonstrated that application of choline induced a slowly desensitizing inward current and increased [Ca(2+)](i). The effect was dose dependent with an EC(50) of 1.6 mM and an n(H) of 1.19. The relationship between the elevation of [Ca(2+)](i) (Delta[Ca(2+)](i)) and charge transfer analysed under various recording conditions showed that the Delta[Ca(2+)](i) induced by choline resulted from an influx of Ca(2+) through nicotinic acetylcholine receptors. The effect of choline on the membrane current and Delta[Ca(2+)](i) was not affected by either short application or pretreatment with alpha-bungarotoxin (50 nM) and methyllycaconitine (1 nM), two alpha7 nicotinic receptors antagonists. These results indicate that activation of non-alpha7 nicotinic acetylcholine receptors by choline significantly increases the Ca(2+) concentration in rat superior cervical ganglion neurones.
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http://dx.doi.org/10.1016/s0014-2999(03)01860-0DOI Listing
June 2003