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Colloque organisé par Thierry Galli (Institut Jacques Monod, Paris) et Yann Humeau (Institut interdisciplinaire des Neurosciences, Bordeaux) du 10 au 15 septembre 2012ParticipantsAnnie Andrieux, Fabio Benfenati, Gerard Borst, Jamel Chelly, Lydia Danglot, Frédéric Darios, Pietro De Camilli, Thierry Galli, Frédéric Gambino, Etienne Herzog, Yann Humeau, Philippe Isope, David Perrais, Maja Petkovic, Bernard Poulain, Jakob B. Sorensen, David Tareste, Matthijs Verhage--Abstract The seminar entitled « Presynapse: function, plasticity, dysregulations » brought together experts working on functional and molecular presynaptic mechanisms, which are essential for neuronal communication. The purpose of the discussions was to understand the role played by the absence of important presynaptic factors in a wide range of nervous pathologies, including severe forms of epilepsy, schizophrenia, syndromic and non-syndromic cognitive deficits and familial migraines. Among the new concepts discussed during this seminar, we noted the emergence of fusogenic lipids as major players in the regulation of presynapses, their study becoming facilitated by the introduction of new optogenetic tools. Functional studies also helped to highlight the presence of vesicular pools in addition to those usually described for the fine tuning of synaptic efficacy during episodes of high frequency neuronal activity. From pathophysiological studies, it appeared fairly obvious that the presynapses are functional sites affected by genetic mutations associated with mental disturbances. In this respect, the role of many actors such as microtubules, monomeric G proteins, phosphoproteins and trans-synaptic signaling proteins was described. Interestingly, many large-scale programs characterizing these changes are underway, including examination of the co-responsibility of large networks of genes as a cause of cognitive disorders. Indeed, the accumulation of « weak » alleles combined with the appearance of a stochastic combination of mutations could explain the emergence of synaptic pathologies including genetic ones which are not strictly monogenic or polygenic. In a final round table, the importance of complementarity of research on large gene networks and systematic research on specific genes as well as the emergence of new methodological tools for the study of presynapses were addressed.--Compte-renduThe seminar entitled “Presynapse: function, plasticity, dysregulations” hosted experts of both functional and molecular aspects of the presynaptic compartment. The participants discussed the role of this highly specialized subcellular structure supporting neurotransmitter release. Synapse dysfunctions are associated with pathophysiological conditions named synaptopathies. It is more and more evident that some cognitive disorders are associated with the mutation, haplo-insufficiency or complete absence of key presynaptic proteins like synapsins or munc-18. Indeed, as reminded by J. Chelly, a growing number of genes encoding synaptic proteins are associated with cognitive disorders ranging from non syndromic (“pure”) intellectual disabilities (ID) and autism spectrum disorders to syndromic ID associated with autistic features, epilepsia or schizophrenia. The question of the coresponsibility of multiple genes to generate severe ID forms was discussed by M. Verhage. He showed that presynaptic ras/ERK signaling pathways may induce the degradation of synaptic proteins by the proteasome and play an important role in presynaptic regulations. He also presented the large European funded “Synaptologics” project to phenotype the behavior of a large number of mouse mutants which show synaptic vulnerability. The non-linear accumulation of weak polymorphisms and de novo mutations may be a mechanism through which complex genetic synaptopathies occur.Before the synapse is formed, axons and dendrites grow and connect. M. Petkovic presented new work suggesting an important role of axonal endoplasmic reticulum in the growth cone progression. Eventually, several membrane trafficking mechanisms may be involved in generating axons and dendrites.The notion that subtle deregulation of of synapses “destabilization” would be causal for ID spectrum pathologies was pushed forward by several participants: evidences first came from physiological studies by Y. Humeau’s group demonstrating that several unrelated ID mutations could generate very similar functional presynaptic phenotypes. Furthermore, a role for the accompanying partners of neurons, the so-called astroglial cells, was proposed by J. Chelly in the case of Mecp2 mutations, as some phenotypes could be generated by non cell-autonomous processes. From J. Chelly and A. Andrieux’s presentations, microtubules appeared as main regulators of presynaptic events and it is not yet clear if this is due to their structuring properties or their role axonal transport to the nerve terminal. This opens the perspective that microtubule-related drugs that are used to treat cancer could also be used against some intellectual disabilities when we are able to design versions that cross the blood brain barrier.Another emerging concept was the role of excitatory/inhibitory (E/I) imbalance in the pathophysiology of ID. As described by F. Benfenati and M. Verhage, a number of mutations (Synapsins, Munc-18, etc…) are associated with epileptic seizures in mouse models and human patients. Y. Humeau exposed results from a multi-scale analysis of Il1rapl1 mutant mice, in which perturbations of associative learning could be attributed to discrete E/I imbalance in neuronal microcircuits. This was thought to result from cell-specific impact of the mutation in particular cell populations. Thus, specific vulnerability of synapse to ID mutations might be meaningful to explain the difficulties in memory formation and expression in ID models and possibly in patients. In a final discussion, the crucial question of the long term consequences of epileptic phenotype, usually not persisting at adult ages, onto the later-on intellectual capabilities of individuals was discussed.Physiological studies (Patch clamp recordings and live cell imaging) presented by E. Herzog, P. Isope, G. Borst and B. Poulain on different synaptic preparations addressed the notion of vesicular pools. First, the strong link between the kinetic and availability of various synaptic pools was particularly evident, illustrated by the presence of some extra-pools of vesicles allowing invertebrate and vertebrate synapses to continuously release neurotransmitter during high frequency trains. New forms of presynaptic, synapsin-dependent forms of short-term plasticity were described, opening the question of the morpho/functional definition of one of the quantal parameters: the number of active releasing sites. G. Borst provided data on mutations in a calcium channel involved in neurotransmitter release, the mutation of which is associated with familial hemiplegic migraine. He demonstrated that some specific adaptations of presynaptic release by GABAergic metabotropic receptors might play a role in the developmental setting of presynaptic plasticity. E. Herzog also pointed the existence of a “super” pool of vesicles diffusing across several “en passant” presynaptic boutons of the same axon. He presented evidence for an unconventional role of the vesicular glutamate transporter in the mobility and clustering of synaptic vesicles. Thus, in addition to canonical vesicular pools, such as a the readily-releasable and the reserve pools, newly defined vesicular pools may be present at presynapses to allow for fine tuning of neurotransmitter release upon various ranges of neuronal activities.Several presentations addressed the question of the diversity of molecular mechanisms leading to presynaptic secretory mechanisms. J. Sorensen showed that detailed characterization of secretory pools lead to the idea of much larger number of states in which secretory granules can be before they release. These may be related to a combination of different molecular mechanisms involved in their secretion. T. Galli showed the molecular network and regulations that his group identified in the case of VAMP7-mediated secretion and he and L. Danglot discussed the possibility that synaptic vesicles might be equipped with several molecular mechanisms for their release. These molecular mechanisms could operate in parallel, generating distinct release routes for synaptic vesicles.D. Tareste presented biophysical approaches to study the force needed to fuse synaptic vesicles with the presynaptic plasma membrane. These approaches measure with great precision the thermodynamics of synaptic vesicle tethering, docking and priming steps leading to the exocytotic release of their content. D. Tareste could further explain how regulatory proteins influence the thermodynamics of snare complex formation.P. De Camilli presented the latest results of his laboratory towards the deciphering of the endocytic machinery. He showed that Endophilins are non essential to the clathrin coated endocytosis but that their removal slows down the process very significantly by limiting the uncoating synaptic vesicles similarly to synaptojanin. The presentation of D. Perrais also focused on endocytosis and more generally on the traffic of post-synaptic receptors using leading edge dynamic microscopy and discussed the importance of regulating the amount of these receptors at the neuronal surface in learning and memory.F. Darios and P. De Camilli presented their work on lipids and presynaptic regulations. F Darios showed the role of sphingosine, arachidonic acid and fatty acids in activating presynaptic exocytosis. He further showed the link between arachidonic acid and spastic paraplegia. P. De Camilli proposed how phosphatidylinositides could regulate exocytosis and endocytosis. He also showed sophisticated new optogenetics approaches to manipulate and monitor ex vivo phosphoinositides production in specific cell compartments and how these will allow asking yet unresolved questions in the field. With lengthy discussions and a final round-table, the workshop concluded on recent tremendous progress in the field, the genetic overlap for many forms of human intellectual disabilities, autism and ASD, epilepsy, and psychiatric disorders and convergence on presynaptic proteins as key regulators of neuronal signaling, the crucial but still to be studied role of lipids in presynaptic mechanisms and the unanticipated diversity of synaptic vesicles and secretory mechanisms. The participants went back to their lab cheered up with new ideas.