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Fabio Benfenati - One of the best experts on this subject based on the ideXlab platform.
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synapsin i controls synaptic maturation of long range projections in the lateral amygdala in a targeted selective fashion
Frontiers in Cellular Neuroscience, 2019Co-Authors: Eleonora Lugara, Fabio Benfenati, Antonio De Fusco, Gabriele Lignani, Yann HumeauAbstract:The amygdala, and more precisely its lateral nucleus, is thought to confer emotional valence to external stimuli by generating long-term plasticity changes at long-range projections to principal cells. Aversive experience has also been shown to modify pre- and post-synaptic markers in the amygdala, suggesting their possible role in the structural organization of adult amygdala networks. Here, we focused on the maturation of cortical and thalamic long-term projections on principal neurons and interneurons in the lateral amygdala (LA) and on how it is affected by deletion of the presynaptic protein synapsin I. We firstly performed dual electrophysiological recordings of identified cells in juvenile and adult GAD67-GFP mice after independent stimulation of cortical and thalamic afferent systems. The results demonstrate that synaptic strengthening occurs during development at synapses projecting to LA principal neurons, but not interneurons. As synaptic strengthening underlying fear conditioning depends on the presence and increasing expression of synapsin I, we successively tested if synapsin I contributes to synaptic strengthening during development. Interestingly, the physiological strengthening of cortical and thalamic synapses projecting to LA principal neurons was virtually abolished in synapsin I knockout mice, while no differences were observed in the excitatory projections to interneurons, suggesting that the synapse-specificity of synaptic strengthening by synapsin I is attributable to its neuronal target. Immunohistochemistry analysis demonstrated that the presence of synapsin I is restricted to excitatory contacts projecting to principal neurons in LA of adult mice. These results indicate that synapsin I is a key regulator of the maturation of synaptic connectivity in this brain region and that is expression is dependent on postsynaptic identity.
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synapsin i controls synaptic maturation of long range projections in the lateral amygdala in a targeted selective fashion
Frontiers in Cellular Neuroscience, 2019Co-Authors: Fabio Benfenati, Eleonora Lugara, Antonio De Fusco, Gabriele Lignani, Yann HumeauAbstract:The amygdala, and more precisely its lateral nucleus, is thought to attribute emotional valence to external stimuli by generating long-term plasticity changes at long-range projections to principal cells. Aversive experience has also been shown to modify pre- and post-synaptic markers in the amygdala, suggesting their possible role in the structural organization of adult amygdala networks. Here, we focused on how the maturation of cortical and thalamic long-range projections occurs on principal neurons and interneurons in the lateral amygdala (LA). We performed dual electrophysiological recordings of identified cells in juvenile and adult GAD67-GFP mice after independent stimulation of cortical and thalamic afferent systems. The results demonstrate that synaptic strengthening occurs during development at synapses projecting to LA principal neurons, but not interneurons. As synaptic strengthening underlies fear conditioning which depends, in turn, on presence and increasing expression of synapsin I, we tested if synapsin I contributes to synaptic strengthening during development. Interestingly, the physiological synaptic strengthening of cortical and thalamic synapses projecting to LA principal neurons was virtually abolished in synapsin I knockout mice, but not differences were observed in the excitatory projections to interneurons. Immunohistochemistry analysis showed that the presence of synapsin I is restricted to excitatory contacts projecting to principal neurons in LA of adult mice. These results indicate that synapsin I is a key regulator of the maturation of synaptic connectivity in this brain region and that is expression is dependent on postsynaptic identity.
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Data_Sheet_1_Synapsin I Controls Synaptic Maturation of Long-Range Projections in the Lateral Amygdala in a Targeted Selective Fashion.PDF
2019Co-Authors: Eleonora Lugara, Fabio Benfenati, Antonio De Fusco, Gabriele Lignani, Yann HumeauAbstract:The amygdala, and more precisely its lateral nucleus, is thought to attribute emotional valence to external stimuli by generating long-term plasticity changes at long-range projections to principal cells. Aversive experience has also been shown to modify pre- and post-synaptic markers in the amygdala, suggesting their possible role in the structural organization of adult amygdala networks. Here, we focused on how the maturation of cortical and thalamic long-range projections occurs on principal neurons and interneurons in the lateral amygdala (LA). We performed dual electrophysiological recordings of identified cells in juvenile and adult GAD67-GFP mice after independent stimulation of cortical and thalamic afferent systems. The results demonstrate that synaptic strengthening occurs during development at synapses projecting to LA principal neurons, but not interneurons. As synaptic strengthening underlies fear conditioning which depends, in turn, on presence and increasing expression of synapsin I, we tested if synapsin I contributes to synaptic strengthening during development. Interestingly, the physiological synaptic strengthening of cortical and thalamic synapses projecting to LA principal neurons was virtually abolished in synapsin I knockout mice, but not differences were observed in the excitatory projections to interneurons. Immunohistochemistry analysis showed that the presence of synapsin I is restricted to excitatory contacts projecting to principal neurons in LA of adult mice. These results indicate that synapsin I is a key regulator of the maturation of synaptic connectivity in this brain region and that is expression is dependent on postsynaptic identity.
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nonsense mediated mrna decay and loss of function of the protein underlie the x linked epilepsy associated with the w356 mutation in synapsin i
PLOS ONE, 2013Co-Authors: Maila Giannandrea, Fabio Benfenati, Fabrizia C Guarnieri, Niels H Gehring, Elena Monzani, Andreas E Kulozik, Flavia ValtortaAbstract:Synapsins are a family of neuronal phosphoproteins associated with the cytosolic surface of synaptic vesicles. Experimental evidence suggests a role for synapsins in synaptic vesicle clustering and recycling at the presynaptic terminal, as well as in neuronal development and synaptogenesis. Synapsin knock-out (Syn1−/−) mice display an epileptic phenotype and mutations in the SYN1 gene have been identified in individuals affected by epilepsy and/or autism spectrum disorder. We investigated the impact of the c.1067G>A nonsense transition, the first mutation described in a family affected by X-linked syndromic epilepsy, on the expression and functional properties of the synapsin I protein. We found that the presence of a premature termination codon in the human SYN1 transcript renders it susceptible to nonsense-mediated mRNA decay (NMD). Given that the NMD efficiency is highly variable among individuals and cell types, we investigated also the effects of expression of the mutant protein and found that it is expressed at lower levels compared to wild-type synapsin I, forms perinuclear aggregates and is unable to reach presynaptic terminals in mature hippocampal neurons grown in culture. Taken together, these data indicate that in patients carrying the W356× mutation the function of synapsin I is markedly impaired, due to both the strongly decreased translation and the altered function of the NMD-escaped protein, and support the value of Syn1−/− mice as an experimental model mimicking the human pathology.
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Tyrosine phosphorylation of synapsin I by Src regulates synaptic-vesicle trafficking.
Journal of Cell Science, 2010Co-Authors: Mirko Messa, Enrico Defranchi, Franco Onofri, Flavia Valtorta, Anna Fassio, Sonia Congia, Fabio BenfenatiAbstract:Synapsins are synaptic vesicle (SV)-associated phosphoproteins involved in the regulation of neurotransmitter release. Synapsins reversibly tether SVs to the cytoskeleton and their phosphorylation by serine/threonine kinases increases SV availability for exocytosis by impairing their association with SVs and/or actin. We recently showed that synapsin I, through SH3- or SH2-mediated interactions, activates Src and is phosphorylated by the same kinase at Tyr301. Here, we demonstrate that, in contrast to serine phosphorylation, Src-mediated tyrosine phosphorylation of synapsin I increases its binding to SVs and actin, and increases the formation of synapsin dimers, which are both potentially involved in SV clustering. Synapsin I phosphorylation by Src affected SV dynamics and was physiologically regulated in brain slices in response to depolarization. Expression of the non-phosphorylatable (Y301F) synapsin I mutant in synapsin-I-knockout neurons increased the sizes of the readily releasable and recycling pools of SVs with respect to the wild-type form, which is consistent with an increased availability of recycled SVs for exocytosis. The data provide a mechanism for the effects of Src on SV trafficking and indicate that tyrosine phosphorylation of synapsins, unlike serine phosphorylation, stimulates the reclustering of recycled SVs and their recruitment to the reserve pool.
Flavia Valtorta - One of the best experts on this subject based on the ideXlab platform.
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nonsense mediated mrna decay and loss of function of the protein underlie the x linked epilepsy associated with the w356 mutation in synapsin i
PLOS ONE, 2013Co-Authors: Maila Giannandrea, Fabio Benfenati, Fabrizia C Guarnieri, Niels H Gehring, Elena Monzani, Andreas E Kulozik, Flavia ValtortaAbstract:Synapsins are a family of neuronal phosphoproteins associated with the cytosolic surface of synaptic vesicles. Experimental evidence suggests a role for synapsins in synaptic vesicle clustering and recycling at the presynaptic terminal, as well as in neuronal development and synaptogenesis. Synapsin knock-out (Syn1−/−) mice display an epileptic phenotype and mutations in the SYN1 gene have been identified in individuals affected by epilepsy and/or autism spectrum disorder. We investigated the impact of the c.1067G>A nonsense transition, the first mutation described in a family affected by X-linked syndromic epilepsy, on the expression and functional properties of the synapsin I protein. We found that the presence of a premature termination codon in the human SYN1 transcript renders it susceptible to nonsense-mediated mRNA decay (NMD). Given that the NMD efficiency is highly variable among individuals and cell types, we investigated also the effects of expression of the mutant protein and found that it is expressed at lower levels compared to wild-type synapsin I, forms perinuclear aggregates and is unable to reach presynaptic terminals in mature hippocampal neurons grown in culture. Taken together, these data indicate that in patients carrying the W356× mutation the function of synapsin I is markedly impaired, due to both the strongly decreased translation and the altered function of the NMD-escaped protein, and support the value of Syn1−/− mice as an experimental model mimicking the human pathology.
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synaptic and extrasynaptic origin of the excitation inhibition imbalance in the hippocampus of synapsin i ii iii knockout mice
Cerebral Cortex, 2013Co-Authors: Flavia Valtorta, Fabrizia Cesca, Pasqualina Farisello, Davide Boido, Thierry Nieus, Lucian Medrihan, Pietro BaldelliAbstract:Synapsins (Syn I, Syn II, and Syn III) are a family of synaptic vesicle phosphoproteins regulating synaptic transmission and plasticity. SYN1/2 genes have been identified as major epilepsy susceptibility genes in humans and synapsin I/II/III triple knockout (TKO) mice are epileptic. However, excitatory and inhibitory synaptic transmission and short-term plasticity have never been analyzed in intact neuronal circuits of TKO mice. To clarify the generation and expression of the epileptic phenotype, we performed patch-clamp recordings in the CA1 region of acute hippocampal slices from 1-month-old presymptomatic and 6-month-old epileptic TKO mice and age-matched controls. We found a strong imbalance between basal glutamatergic and g-aminobutyric acid (GABA)ergic transmission with increased evoked excitatory postsynaptic current and impaired evoked inhibitory postsynaptic current amplitude. This imbalance was accompanied by a parallel derangement of shortterm plasticity paradigms, with enhanced facilitation of glutamatergic transmission in the presymptomatic phase and milder depression of inhibitory synapses in the symptomatic phase. Interestingly, a lower tonic GABAA current due to the impaired GABA release is responsible for the more depolarized resting potential found in TKO CA1 neurons, which makes them more susceptible to fire. All these changes preceded the appearance of epilepsy, indicating that the distinct changes in excitatory and inhibitory transmission due to the absence of Syns initiate the epileptogenic process.
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the synapsins key actors of synapse function and plasticity
Progress in Neurobiology, 2010Co-Authors: Fabrizia Cesca, Pietro Baldelli, Flavia ValtortaAbstract:The synapsins are a family of neuronal phosphoproteins evolutionarily conserved in invertebrate and vertebrate organisms. Their best-characterised function is to modulate neurotransmitter release at the pre-synaptic terminal, by reversibly tethering synaptic vesicles (SVs) to the actin cytoskeleton. However, many recent data have suggested novel functions for synapsins in other aspects of the pre-synaptic physiology, such as SV docking, fusion and recycling. Synapsin activity is tightly regulated by several protein kinases and phosphatases, which modulate the association of synapsins to SVs as well as their interaction with actin filaments and other synaptic proteins. In this context, synapsins act as a link between extracellular stimuli and the intracellular signalling events activated upon neuronal stimulation. Genetic manipulation of synapsins in various in vivo models has revealed that, although not essential for the basic development and functioning of neuronal networks, these proteins are extremely important in the fine-tuning of neuronal plasticity, as shown by the epileptic phenotype and behavioural abnormalities characterising mouse lines lacking one or more synapsin isoforms. In this review, we summarise the current knowledge about how the various members of the synapsin family are involved in the modulation of the pre-synaptic physiology. We give a comprehensive description of the molecular basis of synapsin function, as well as an overview of the more recent evidence linking mutations in the synapsin proteins to the onset of severe central nervous system diseases such as epilepsy and schizophrenia.
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Tyrosine phosphorylation of synapsin I by Src regulates synaptic-vesicle trafficking.
Journal of Cell Science, 2010Co-Authors: Mirko Messa, Enrico Defranchi, Franco Onofri, Flavia Valtorta, Anna Fassio, Sonia Congia, Fabio BenfenatiAbstract:Synapsins are synaptic vesicle (SV)-associated phosphoproteins involved in the regulation of neurotransmitter release. Synapsins reversibly tether SVs to the cytoskeleton and their phosphorylation by serine/threonine kinases increases SV availability for exocytosis by impairing their association with SVs and/or actin. We recently showed that synapsin I, through SH3- or SH2-mediated interactions, activates Src and is phosphorylated by the same kinase at Tyr301. Here, we demonstrate that, in contrast to serine phosphorylation, Src-mediated tyrosine phosphorylation of synapsin I increases its binding to SVs and actin, and increases the formation of synapsin dimers, which are both potentially involved in SV clustering. Synapsin I phosphorylation by Src affected SV dynamics and was physiologically regulated in brain slices in response to depolarization. Expression of the non-phosphorylatable (Y301F) synapsin I mutant in synapsin-I-knockout neurons increased the sizes of the readily releasable and recycling pools of SVs with respect to the wild-type form, which is consistent with an increased availability of recycled SVs for exocytosis. The data provide a mechanism for the effects of Src on SV trafficking and indicate that tyrosine phosphorylation of synapsins, unlike serine phosphorylation, stimulates the reclustering of recycled SVs and their recruitment to the reserve pool.
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The highly conserved synapsin domain E mediates synapsin dimerization and phospholipid vesicle clustering.
The Biochemical journal, 2010Co-Authors: Ilaria Monaldi, Silvia Giovedi, Flavia Valtorta, Massimo Vassalli, Angela Bachi, Enrico Millo, Roberto Raiteri, Anna FassioAbstract:Synapsins are abundant SV (synaptic vesicle)-associated phosphoproteins that regulate synapse formation and function. The highly conserved C-terminal domain E was shown to contribute to several synapsin functions, ranging from formation of the SV reserve pool to regulation of the kinetics of exocytosis and SV cycling, although the molecular mechanisms underlying these effects are unknown. In the present study, we used a synthetic 25-mer peptide encompassing the most conserved region of domain E (Pep-E) to analyse the role of domain E in regulating the interactions between synapsin I and liposomes mimicking the phospholipid composition of SVs (SV-liposomes) and other pre-synaptic protein partners. In affinity-chromatography and cross-linking assays, Pep-E bound to endogenous and purified exogenous synapsin I and strongly inhibited synapsin dimerization, indicating a role in synapsin oligomerization. Consistently, Pep-E (but not its scrambled version) counteracted the ability of holo-synapsin I to bind and coat phospholipid membranes, as analysed by AFM (atomic force microscopy) topographical scanning, and significantly decreased the clustering of SV-liposomes induced by holo-synapsin I in FRET (Forster resonance energy transfer) assays, suggesting a causal relationship between synapsin oligomerization and vesicle clustering. Either Pep-E or a peptide derived from domain C was necessary and sufficient to inhibit both dimerization and vesicle clustering, indicating the participation of both domains in these activities of synapsin I. The results provide a molecular explanation for the effects of domain E in nerve terminal physiology and suggest that its effects on the size and integrity of SV pools are contributed by the regulation of synapsin dimerization and SV clustering.
Paul Greengard - One of the best experts on this subject based on the ideXlab platform.
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Colocalization of synapsin and actin during synaptic vesicle recycling
The Journal of cell biology, 2003Co-Authors: Ona Bloom, Vincent A Pieribone, Lennart Brodin, Peter Löw, Emma Evergren, Nikolay Tomilin, Ole Kjaerulff, Paul Greengard, Oleg ShupliakovAbstract:It has been hypothesized that in the mature nerve terminal, interactions between synapsin and actin regulate the clustering of synaptic vesicles and the availability of vesicles for release during synaptic activity. Here, we have used immunogold electron microscopy to examine the subcellular localization of actin and synapsin in the giant synapse in lamprey at different states of synaptic activity. In agreement with earlier observations, in synapses at rest, synapsin immunoreactivity was preferentially localized to a portion of the vesicle cluster distal to the active zone. During synaptic activity, however, synapsin was detected in the pool of vesicles proximal to the active zone. In addition, actin and synapsin were found colocalized in a dynamic filamentous cytomatrix at the sites of synaptic vesicle recycling, endocytic zones. Synapsin immunolabeling was not associated with clathrin-coated intermediates but was found on vesicles that appeared to be recycling back to the cluster. Disruption of synapsin function by microinjection of antisynapsin antibodies resulted in a prominent reduction of the cytomatrix at endocytic zones of active synapses. Our data suggest that in addition to its known function in clustering of vesicles in the reserve pool, synapsin migrates from the synaptic vesicle cluster and participates in the organization of the actin-rich cytomatrix in the endocytic zone during synaptic activity.
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Identification of synapsin I peptides that insert into lipid membranes
Biochemical Journal, 2001Co-Authors: James J. Cheetham, Fabio Benfenati, Sabine Hilfiker, Thomas Weber, Paul Greengard, Andrew J CzernikAbstract:The synapsins constitute a family of synaptic vesicle-associated phosphoproteins essential for regulating neurotransmitter release and synaptogenesis. The molecular mechanisms underlying the selective targeting of synapsin I to synaptic vesicles are thought to involve specific protein-protein interactions, while the high-affinity binding to the synaptic vesicle membrane may involve both protein-protein and protein-lipid interactions. The highly hydrophobic N-terminal region of the protein has been shown to bind with high affinity to the acidic phospholipids phosphatidylserine and phosphatidylinositol and to penetrate the hydrophobic core of the lipid bilayer. To precisely identify the domains of synapsin I which mediate the interaction with lipids, synapsin I was bound to liposomes containing the membrane-directed carbene-generating reagent 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine and subjected to photolysis. Isolation and N-terminal amino acid sequencing of 125I-labelled synapsin I peptides derived from CNBr cleavage indicated that three distinct regions in the highly conserved domain C of synapsin I insert into the hydrophobic core of the phospholipid bilayer. The boundaries of the regions encompass residues 166-192, 233-258 and 278-327 of bovine synapsin I. These regions are surface-exposed in the crystal structure of domain C of bovine synapsin I and are evolutionarily conserved among isoforms across species. The present data offer a molecular explanation for the high-affinity binding of synapsin I to phospholipid bilayers and synaptic vesicles.
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Specificity of the binding of synapsin I to Src homology 3 domains.
The Journal of biological chemistry, 2000Co-Authors: Franco Onofri, Silvia Giovedi, Hung-teh Kao, Flavia Valtorta, P De Camilli, Lucilla Bongiorno Borbone, Paul Greengard, Fabio BenfenatiAbstract:Abstract Synapsins are synaptic vesicle-associated phosphoproteins involved in synapse formation and regulation of neurotransmitter release. Recently, synapsin I has been found to bind the Src homology 3 (SH3) domains of Grb2 and c-Src. In this work we have analyzed the interactions between synapsins and an array of SH3 domains belonging to proteins involved in signal transduction, cytoskeleton assembly, or endocytosis. The binding of synapsin I was specific for a subset of SH3 domains. The highest binding was observed with SH3 domains of c-Src, phospholipase C-γ, p85 subunit of phosphatidylinositol 3-kinase, full-length and NH2-terminal Grb2, whereas binding was moderate with the SH3 domains of amphiphysins I/II, Crk, α-spectrin, and NADPH oxidase factor p47phox and negligible with the SH3 domains of p21ras GTPase-activating protein and COOH-terminal Grb2. Distinct sites in the proline-rich COOH-terminal region of synapsin I were found to be involved in binding to the various SH3 domains. Synapsin II also interacted with SH3 domains with a partly distinct binding pattern. Phosphorylation of synapsin I in the COOH-terminal region by Ca2+/calmodulin-dependent protein kinase II or mitogen-activated protein kinase modulated the binding to the SH3 domains of amphiphysins I/II, Crk, and α-spectrin without affecting the high affinity interactions. The SH3-mediated interaction of synapsin I with amphiphysins affected the ability of synapsin I to interact with actin and synaptic vesicles, and pools of synapsin I and amphiphysin I were shown to associate in isolated nerve terminals. The ability to bind multiple SH3 domains further implicates the synapsins in signal transduction and protein-protein interactions at the nerve terminal level.
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Distinct Roles of Synapsin I and Synapsin II during Neuronal Development
Molecular Medicine, 1998Co-Authors: Adriana Ferreira, Lih-shen Chin, Lorene M. Lanier, Kenneth S Kosik, Paul GreengardAbstract:The synapsins are a family of neuron-specific proteins, associated with the cytoplasmic surface of synaptic vesicles, which have been shown to regulate neurotransmitter release in mature synapses and to accelerate development of the nervous system. Using neuronal cultures from mice lacking synapsin I, synapsin II, or both synapsins I and II, we have now found that synapsin I and synapsin II play distinct roles in neuronal development. Deletion of synapsin II, but not synapsin I, greatly retarded axon formation. Conversely, deletion of synapsin I, but not synapsin II, greatly retarded synapse formation. Remarkably, the deletion of both synapsins led to partial restoration of the wild phenotype. The results suggest that the synapsins play separate but coordinated developmental roles.
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Suppression of synapsin II inhibits the formation and maintenance of synapses in hippocampal culture
Proceedings of the National Academy of Sciences of the United States of America, 1995Co-Authors: Adriana Ferreira, Paul Greengard, Kenneth S KosikAbstract:Abstract Numerous synaptic proteins, including several integral membrane proteins, have been assigned roles in synaptic vesicle fusion with or retrieval from the presynaptic plasma membrane. In contrast, the synapsins, neuron-specific phosphoproteins associated with the cytoplasmic surface of synaptic vesicles, appear to play a much broader role, being involved in the regulation of neurotransmitter release and in the organization of the nerve terminal. Here we have administered antisense synapsin II oligonucleotides to dissociated hippocampal neurons, either before the onset of synaptogenesis or 1 week after the onset of synaptogenesis. In both cases, synapsin II was no longer detectable within 24-48 h of treatment. After 5 days of treatment, cultures were analyzed for the presence of synapses by synapsin I and synaptophysin antibody labeling and by electron microscopy. Cultures in which synapsin II was suppressed after axon elongation, but before synapse formation, did not develop synapses. Cultures in which synapsin II was suppressed after the development of synapses lost most of their synapses. Remarkably, with the removal of the antisense oligonucleotides, neurons and their synaptic connections recovered. These studies lead us to conclude that synapsin II is involved in the formation and maintenance of synapses in hippocampal neurons.
Andrew J Czernik - One of the best experts on this subject based on the ideXlab platform.
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structural domains involved in the regulation of transmitter release by synapsins
The Journal of Neuroscience, 2005Co-Authors: Sabine Hilfiker, Frederic Doussau, Angus C Nairn, Andrew J CzernikAbstract:Synapsins are a family of neuron-specific phosphoproteins that regulate neurotransmitter release by associating with synaptic vesicles. Synapsins consist of a series of conserved and variable structural domains of unknown function. We performed a systematic structure-function analysis of the various domains of synapsin by assessing the actions of synapsin fragments on neurotransmitter release, presynaptic ultrastructure, and the biochemical interactions of synapsin. Injecting a peptide derived from domain A into the squid giant presynaptic terminal inhibited neurotransmitter release in a phosphorylation-dependent manner. This peptide had no effect on vesicle pool size, synaptic depression, or transmitter release kinetics. In contrast, a peptide fragment from domain C reduced the number of synaptic vesicles in the periphery of the active zone and increased the rate and extent of synaptic depression. This peptide also slowed the kinetics of neurotransmitter release without affecting the number of docked vesicles. The domain C peptide, as well as another peptide from domain E that is known to have identical effects on vesicle pool size and release kinetics, both specifically interfered with the binding of synapsins to actin but not with the binding of synapsins to synaptic vesicles. This suggests that both peptides interfere with release by preventing interactions of synapsins with actin. Thus, interactions of domains C and E with the actin cytoskeleton may allow synapsins to perform two roles in regulating release, whereas domain A has an actin-independent function that regulates transmitter release in a phosphorylation-sensitive manner.
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Neuronal nitric-oxide synthase localization mediated by a ternary complex with synapsin and CAPON.
Proceedings of the National Academy of Sciences of the United States of America, 2002Co-Authors: Samie R. Jaffrey, Andrew J Czernik, Adele M. Snowman, Solomon H. SnyderAbstract:The specificity of the reactions of nitric oxide (NO) with its neuronal targets is determined in part by the precise localizations of neuronal NO synthase (nNOS) within the cell. The targeting of nNOS is mediated by adapter proteins that interact with its PDZ domain. Here, we show that the nNOS adapter protein, CAPON, interacts with synapsins I, II, and III through an N-terminal phosphotyrosine-binding domain interaction, which leads to a ternary complex comprising nNOS, CAPON, and synapsin I. The significance of this ternary complex is demonstrated by changes in subcellular localization of nNOS in mice harboring genomic deletions of both synapsin I and synapsin II. These results suggest a mechanism for specific actions of NO at presynaptic sites.
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opposing changes in phosphorylation of specific sites in synapsin i during ca2 dependent glutamate release in isolated nerve terminals
The Journal of Neuroscience, 2001Co-Authors: Jasmina N Jovanovic, Angus C Nairn, Talvinder S Sihra, Hugh C Hemmings, Andrew J CzernikAbstract:Synapsins are major neuronal phosphoproteins involved in regulation of neurotransmitter release. Synapsins are well established targets for multiple protein kinases within the nerve terminal, yet little is known about dephosphorylation processes involved in regulation of synapsin function. Here, we observed a reciprocal relationship in the phosphorylation–dephosphorylation of the established phosphorylation sites on synapsin I. We demonstrate that, in vitro , phosphorylation sites 1, 2, and 3 of synapsin I (P-site 1 phosphorylated by cAMP-dependent protein kinase; P-sites 2 and 3 phosphorylated by Ca2+–calmodulin-dependent protein kinase II) were excellent substrates for protein phosphatase 2A, whereas P-sites 4, 5, and 6 (phosphorylated by mitogen-activated protein kinase) were efficiently dephosphorylated only by Ca2+–calmodulin-dependent protein phosphatase 2B–calcineurin. In isolated nerve terminals, rapid changes in synapsin I phosphorylation were observed after Ca2+entry, namely, a Ca2+-dependent phosphorylation of P-sites 1, 2, and 3 and a Ca2+-dependent dephosphorylation of P-sites 4, 5, and 6. Inhibition of calcineurin activity by cyclosporin A resulted in a complete block of Ca2+-dependent dephosphorylation of P-sites 4, 5, and 6 and correlated with a prominent increase in ionomycin-evoked glutamate release. These two opposing, rapid, Ca2+-dependent processes may play a crucial role in the modulation of synaptic vesicle trafficking within the presynaptic terminal.
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Identification of synapsin I peptides that insert into lipid membranes
Biochemical Journal, 2001Co-Authors: James J. Cheetham, Fabio Benfenati, Sabine Hilfiker, Thomas Weber, Paul Greengard, Andrew J CzernikAbstract:The synapsins constitute a family of synaptic vesicle-associated phosphoproteins essential for regulating neurotransmitter release and synaptogenesis. The molecular mechanisms underlying the selective targeting of synapsin I to synaptic vesicles are thought to involve specific protein-protein interactions, while the high-affinity binding to the synaptic vesicle membrane may involve both protein-protein and protein-lipid interactions. The highly hydrophobic N-terminal region of the protein has been shown to bind with high affinity to the acidic phospholipids phosphatidylserine and phosphatidylinositol and to penetrate the hydrophobic core of the lipid bilayer. To precisely identify the domains of synapsin I which mediate the interaction with lipids, synapsin I was bound to liposomes containing the membrane-directed carbene-generating reagent 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine and subjected to photolysis. Isolation and N-terminal amino acid sequencing of 125I-labelled synapsin I peptides derived from CNBr cleavage indicated that three distinct regions in the highly conserved domain C of synapsin I insert into the hydrophobic core of the phospholipid bilayer. The boundaries of the regions encompass residues 166-192, 233-258 and 278-327 of bovine synapsin I. These regions are surface-exposed in the crystal structure of domain C of bovine synapsin I and are evolutionarily conserved among isoforms across species. The present data offer a molecular explanation for the high-affinity binding of synapsin I to phospholipid bilayers and synaptic vesicles.
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synapsins as regulators of neurotransmitter release
Philosophical Transactions of the Royal Society B, 1999Co-Authors: Sabine Hilfiker, Vincent A Pieribone, Andrew J CzernikAbstract:One of the crucial issues in understanding neuronal transmission is to define the role(s) of the numerous proteins that are localized within presynaptic terminals and are thought to participate in the regulation of the synaptic vesicle life cycle. Synapsins are a multigene family of neuron–specific phosphoproteins and are the most abundant proteins on synaptic vesicles. Synapsins are able to interact in vitro with lipid and protein components of synaptic vesicles and with various cytoskeletal proteins, including actin. These and other studies have led to a model in which synapsins, by tethering synaptic vesicles to each other and to an actin–based cytoskeletal meshwork, maintain a reserve pool of vesicles in the vicinity of the active zone. Perturbation of synapsin function in a variety of preparations led to a selective disruption of this reserve pool and to an increase in synaptic depression, suggesting that the synapsin–dependent cluster of vesicles is required to sustain release of neurotransmitter in response to high levels of neuronal activity. In a recent study performed at the squid giant synapse, perturbation of synapsin function resulted in a selective disruption of the reserve pool of vesicles and in addition, led to an inhibition and slowing of the kinetics of neurotransmitter release, indicating a second role for synapsins downstream from vesicle docking. These data suggest that synapsins are involved in two distinct reactions which are crucial for exocytosis in presynaptic nerve terminals. This review describes our current understanding of the molecular mechanisms by which synapsins modulate synaptic transmission, while the increasingly well–documented role of the synapsins in synapse formation and stabilization lies beyond the scope of this review.
Yann Humeau - One of the best experts on this subject based on the ideXlab platform.
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synapsin i controls synaptic maturation of long range projections in the lateral amygdala in a targeted selective fashion
Frontiers in Cellular Neuroscience, 2019Co-Authors: Eleonora Lugara, Fabio Benfenati, Antonio De Fusco, Gabriele Lignani, Yann HumeauAbstract:The amygdala, and more precisely its lateral nucleus, is thought to confer emotional valence to external stimuli by generating long-term plasticity changes at long-range projections to principal cells. Aversive experience has also been shown to modify pre- and post-synaptic markers in the amygdala, suggesting their possible role in the structural organization of adult amygdala networks. Here, we focused on the maturation of cortical and thalamic long-term projections on principal neurons and interneurons in the lateral amygdala (LA) and on how it is affected by deletion of the presynaptic protein synapsin I. We firstly performed dual electrophysiological recordings of identified cells in juvenile and adult GAD67-GFP mice after independent stimulation of cortical and thalamic afferent systems. The results demonstrate that synaptic strengthening occurs during development at synapses projecting to LA principal neurons, but not interneurons. As synaptic strengthening underlying fear conditioning depends on the presence and increasing expression of synapsin I, we successively tested if synapsin I contributes to synaptic strengthening during development. Interestingly, the physiological strengthening of cortical and thalamic synapses projecting to LA principal neurons was virtually abolished in synapsin I knockout mice, while no differences were observed in the excitatory projections to interneurons, suggesting that the synapse-specificity of synaptic strengthening by synapsin I is attributable to its neuronal target. Immunohistochemistry analysis demonstrated that the presence of synapsin I is restricted to excitatory contacts projecting to principal neurons in LA of adult mice. These results indicate that synapsin I is a key regulator of the maturation of synaptic connectivity in this brain region and that is expression is dependent on postsynaptic identity.
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synapsin i controls synaptic maturation of long range projections in the lateral amygdala in a targeted selective fashion
Frontiers in Cellular Neuroscience, 2019Co-Authors: Fabio Benfenati, Eleonora Lugara, Antonio De Fusco, Gabriele Lignani, Yann HumeauAbstract:The amygdala, and more precisely its lateral nucleus, is thought to attribute emotional valence to external stimuli by generating long-term plasticity changes at long-range projections to principal cells. Aversive experience has also been shown to modify pre- and post-synaptic markers in the amygdala, suggesting their possible role in the structural organization of adult amygdala networks. Here, we focused on how the maturation of cortical and thalamic long-range projections occurs on principal neurons and interneurons in the lateral amygdala (LA). We performed dual electrophysiological recordings of identified cells in juvenile and adult GAD67-GFP mice after independent stimulation of cortical and thalamic afferent systems. The results demonstrate that synaptic strengthening occurs during development at synapses projecting to LA principal neurons, but not interneurons. As synaptic strengthening underlies fear conditioning which depends, in turn, on presence and increasing expression of synapsin I, we tested if synapsin I contributes to synaptic strengthening during development. Interestingly, the physiological synaptic strengthening of cortical and thalamic synapses projecting to LA principal neurons was virtually abolished in synapsin I knockout mice, but not differences were observed in the excitatory projections to interneurons. Immunohistochemistry analysis showed that the presence of synapsin I is restricted to excitatory contacts projecting to principal neurons in LA of adult mice. These results indicate that synapsin I is a key regulator of the maturation of synaptic connectivity in this brain region and that is expression is dependent on postsynaptic identity.
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Data_Sheet_1_Synapsin I Controls Synaptic Maturation of Long-Range Projections in the Lateral Amygdala in a Targeted Selective Fashion.PDF
2019Co-Authors: Eleonora Lugara, Fabio Benfenati, Antonio De Fusco, Gabriele Lignani, Yann HumeauAbstract:The amygdala, and more precisely its lateral nucleus, is thought to attribute emotional valence to external stimuli by generating long-term plasticity changes at long-range projections to principal cells. Aversive experience has also been shown to modify pre- and post-synaptic markers in the amygdala, suggesting their possible role in the structural organization of adult amygdala networks. Here, we focused on how the maturation of cortical and thalamic long-range projections occurs on principal neurons and interneurons in the lateral amygdala (LA). We performed dual electrophysiological recordings of identified cells in juvenile and adult GAD67-GFP mice after independent stimulation of cortical and thalamic afferent systems. The results demonstrate that synaptic strengthening occurs during development at synapses projecting to LA principal neurons, but not interneurons. As synaptic strengthening underlies fear conditioning which depends, in turn, on presence and increasing expression of synapsin I, we tested if synapsin I contributes to synaptic strengthening during development. Interestingly, the physiological synaptic strengthening of cortical and thalamic synapses projecting to LA principal neurons was virtually abolished in synapsin I knockout mice, but not differences were observed in the excitatory projections to interneurons. Immunohistochemistry analysis showed that the presence of synapsin I is restricted to excitatory contacts projecting to principal neurons in LA of adult mice. These results indicate that synapsin I is a key regulator of the maturation of synaptic connectivity in this brain region and that is expression is dependent on postsynaptic identity.
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synapsin controls both reserve and releasable synaptic vesicle pools during neuronal activity and short term plasticity inaplysia
The Journal of Neuroscience, 2001Co-Authors: Yann Humeau, Frederic Doussau, Francesco Vitiello, Bernard PoulainAbstract:Neurotransmitter release is a highly efficient secretory process exhibiting resistance to fatigue and plasticity attributable to the existence of distinct pools of synaptic vesicles (SVs), namely a readily releasable pool and a reserve pool from which vesicles can be recruited after activity. Synaptic vesicles in the reserve pool are thought to be reversibly tethered to the actin-based cytoskeleton by the synapsins, a family of synaptic vesicle-associated phosphoproteins that have been shown to play a role in the formation, maintenance, and regulation of the reserve pool of synaptic vesicles and to operate during the post-docking step of the release process. In this paper, we have investigated the physiological effects of manipulating synapsin levels in identified cholinergic synapses of Aplysia californica. When endogenous synapsin was neutralized by the injection of specific anti-synapsin antibodies, the amount of neurotransmitter released per impulse was unaffected, but marked changes in the secretory response to high-frequency stimulation were observed, including the disappearance of post-tetanic potentiation (PTP) that was substituted by post-tetanic depression (PTD), and increased rate and extent of synaptic depression. Opposite changes on post-tetanic potentiation were observed when synapsin levels were increased by injecting exogenous synapsin I. Our data demonstrate that the presence of synapsin-dependent reserve vesicles allows the nerve terminal to release neurotransmitter at rates exceeding the synaptic vesicle recycling capacity and to dynamically change the efficiency of release in response to conditioning stimuli (e.g., post-tetanic potentiation). Moreover, synapsin-dependent regulation of the fusion competence of synaptic vesicles appears to be crucial for sustaining neurotransmitter release during short periods at rates faster than the replenishment kinetics and maintaining synchronization of quanta in evoked release.
