NeuroNex2: Enabling Identification and Impact of Synaptic Weight in Functional Networks; NSF reference 2014862

Synapses form trillions of connections between billions of neurons in the brain to establish neural circuits that allow us to sense, think, act, learn, and remember. Synaptic weight is a crucial concept to understand the nervous system, yet its clear definition remains elusive, despite more than a century of searching. This NeuroNex Network assembles world experts to study synapses from molecules to behavior, in order to answer this fundamental and ambitious question: What constitutes synaptic weight and what role does it play in shaping neural circuits?

How neuronal synapses control behaviour is a critical question in neuroscience. Behaviour is often a response to memories from prior experience. During learning, pre- and postsynaptic mechanisms of plasticity cooperate to elicit long-term changes in synaptic weight - broadly defined as the strength and influence of individual synapses or groups of synapses acting together to achieve a response. Revealing synaptic weight through a multi-modal analysis of synaptic molecules, subcellular composition, and morphology in the context of neural circuits is crucial to understand complex circuit function underlying behaviour. We need to study the diversity of synapses in different brain areas, across species, and in different functional states. For example, synaptic strength and other computational properties are affected by short-term or long-term changes in the probability of presynaptic transmitter release. Upon activation, presynaptic sites undergo dramatic changes as vesicles and other organelles appear, change morphology, relocate, and disappear on multiple time scales. The corresponding structural dynamics can now be captured with millisecond precision using a combination of optogenetics and electron microscopy. This exquisite timing allows characterization, at the spatial nanoscale, of dynamics of synapses undergoing plasticity, growth, or elimination in response to natural and experimental stimuli.

The ideal goal of determining the structure, composition, and circuitry of virtually every synapse in the brain has driven new imaging and cell biology techniques. Our proposed international NeuroNex Network will build upon existing investments in methodology and technology, including the enhanced resolution for 3DEM analysis of synapses based on tomoSEM, a new method with great promise for wide field and high-throughput studies and with sufficient resolution to assess ultrastructural features of synapses throughout tissue volumes great enough to contain local circuits. Advanced labelling and tissue fixation provide new understanding about the ultrastructural features that reflect synaptic states established by molecular and physiological criteria.

Our international NeuroNex Network brings expertise i) to integrate this transformative technology into the respective brain studies, ii) to combine and compare it with additional advanced methods, and to develop a more comprehensive, multi-scale understanding of the synapse across diverse brain circuits and species, and iii) to develop and combine computational tools to collect, analyze, and interpret the wealth of data. The complementary expertise covered by this international NeuroNex Network will generate unprecedented knowledge on the molecular and cellular mechanisms that establish synaptic weight and plasticity in individual synapses and how they translate to understanding synapses in circuits and behaviour.

Traditionally, synapses have been treated as on or off switches, that is, one-bit machines. Recent models, based on synapse size as a proxy for synaptic weight, show that this assumption is wrong. In fact, the information content can be much higher, for example, being >4 bits at hippocampal synapses. Synaptic weight is controlled over broad temporal and spatial scales that are dynamically regulated by activity in neural circuits. New evidence points to subcellular resources (endoplasmic reticulum, mitochondria, endosomes, ribosomes) that broker and drive synaptic efficacy and plasticity through mechanisms that regulate local protein synthesis. Thus, an understanding of synaptic weights needs to be addressed at both subcellular and circuit levels. We hypothesize that synaptic weight is defined by the differential composition and co-occurrence of key proteins and subcellular resources. We will use multidisciplinary approaches to assess these features in well-defined states. Comparisons will be made across neural circuits involving multiple cell types, brain regions, and diverse behaviors in several species. Mapping consistent predictors of synaptic state arising from these analyses onto neural connectomes will enhance tremendously our understanding of the roles of synaptic weight in circuit organization and function.
To achieve these goals, new technologies are needed to bridge multiple scales in image resolution and to collect sufficiently large tissue volumes to perform circuit level analyses. An important new approach involves conical tilt tomography on the scanning electron microscope operating in the transmission mode (tomoSEM). TomoSEM aims to fill the current resolution-to-volume gap between methods used for structural biology (high resolution, small volumes) and connectomics (relatively low resolution, large volumes). TomoSEM provides the axial (~15 nm) and in-plane (~2 nm) resolution needed to identify and quantify subcellular components in large volumes.

Our international NeuroNex Network builds upon the foundation of existing investments in neurotechnology and brings together the necessary cross-discipline experience to delve into previously unanswered questions about brain function. Through international collaboration, our Network will also pioneer new approaches to worldwide collaboration that maximizes the outcome from NSF investments. Progress in our understanding of the healthy brain, cognition, and the emergence of behavior will have a profound impact on society across a broad swath of domains and outcomes. Our international Network will focus on addressing unanswered questions in neuroscience at a scale that would not be possible without this collaborative discovery effort.

This NeuroNex Network includes mentoring activities across the educational spectrum from K-12 outreach programs to undergraduate, graduate and postdoctoral researchers. University courses, in-person workshops and conferences, inter-laboratory student and fellow exchanges, and online instruction will be used to train scientists and disseminate knowledge about brain ultrastructure and function as revealed by our international NeuroNex Network.
Bringing together a diverse set of investigators and infrastructure, this Network will create an enhanced international resource for research and education, which in turn will set the stage for answering future questions about brains. Sharing data enables theorists to quickly test their hypotheses and propose new experiments. Access to the variety of tools across the Network allows efficient use of resources to apply the optimal tool to the particular question. By leveraging the existing community portal for 3DEM sharing and outreach, we will provide tools and training for other neuroscientists to utilize the advances in technology. The 3DEM portal will serve as a highly accessible public resource of the Network's technology, data, and knowledge discovery, thus enhancing the infrastructure for research and education in the field.

Our NeuroNex Network is itself composed of networks of interacting investigators - a network of networks - and could serve as a model for future international collaborations, open sharing of new technologies, and sustainable software development, all enabling reproducible research. The personnel and trainees of this NeuroNex Network represent a diversity of backgrounds, including those traditionally underrepresented in the science, technology, engineering, and mathematics workforce. As an open public resource, we expect to play an important part in encouraging innovation and diversity in neuroscience and related fields.