Biology subjects

Metzbower, S. R.

Publications and source records attributed to Metzbower, S. R..

4 recordsLinked to original sources

bioRxiv2023

Trans-synaptic molecular context of NMDA receptor nanodomains

Tight coordination of the spatial relationships between protein complexes is required for cellular function. In neuronal synapses, many proteins responsible for neurotransmission organize into subsynaptic nanoclusters whose trans-cellular alignment modulates synaptic signal propagation. However, the spatial relationships between these proteins and NMDA receptors (NMDARs), which are required for learning and memory, remain undefined. Here, we mapped the relationship of key NMDAR subunits to reference proteins in the active zone and postsynaptic density using multiplexed super-resolution DNA-PAINT microscopy. GluN2A and GluN2B subunits formed nanoclusters with diverse configurations that, surprisingly, were not localized near presynaptic vesicle release sites marked by Munc13-1. Despite this, we found a subset of release sites was enriched with NMDARs, and modeling of glutamate release and receptor activation in measured synapses indicated this nanotopography promotes NMDAR activation. This subset of release sites was internally denser with Munc13-1, aligned with abundant PSD-95, and associated closely with specific NMDAR nanodomains. Further, NMDAR activation drove rapid reorganization of this release site/receptor relationship, suggesting a structural mechanism for tuning NMDAR-mediated synaptic transmission. This work reveals a new principle regulating NMDAR signaling and suggests that synaptic functional architecture depends on the assembly of and trans-cellular spatial relationships between multiprotein nanodomains.

neuroscience↗

bioRxiv2023

Molecular definition of distinct active zone protein machineries for Ca2+ channel clustering and synaptic vesicle priming

Action potentials trigger neurotransmitter release with minimal delay. Active zones mediate this temporal precision by co-organizing primed vesicles with CaV2 Ca2+ channels. The presumed model is that scaffolding proteins directly tether primed vesicles to CaV2s. We find that CaV2 clustering and vesicle priming are executed by separate machineries. At hippocampal synapses, CaV2 nanoclusters are positioned at variable distances from those of the priming protein Munc13. The active zone organizer RIM anchors both proteins, but distinct interaction motifs independently execute these functions. In heterologous cells, Liprin- and RIM from co- assemblies that are separate from CaV2-organizing complexes upon co-transfection. At synapses, Liprin-1-4 knockout impairs vesicle priming, but not CaV2 clustering. The cell adhesion protein PTP{sigma} recruits Liprin-, RIM and Munc13 into priming complexes without co- clustering of CaV2s. We conclude that active zones consist of distinct complexes to organize CaV2s and vesicle priming, and Liprin- and PTP{sigma} specifically support priming site assembly.

neuroscience↗

bioRxiv2023

Distinct SAP102 and PSD-95 nano-organization defines multiple types of synaptic scaffold protein domains at single synapses

The MAGUK family of scaffold proteins plays a central role in maintaining and modulating synaptic signaling, providing a framework to retain and position receptors, signaling molecules, and other synaptic components. Of these scaffold proteins, SAP102 and PSD-95 are essential for synaptic function at distinct developmental timepoints and perform overlapping as well as unique roles. While their similar structures allow for common binding partners, SAP102 is expressed earlier in synapse development and is required for synaptogenesis, whereas PSD-95 expression peaks later in development and is associated with synapse maturation. PSD-95 and other key synaptic proteins organize into subsynaptic nanodomains that have a significant impact on synaptic transmission, but the nanoscale organization of SAP102 is unknown. How SAP102 is organized within the synapse, and how it relates spatially to PSD-95 on a nanometer scale, could impact how SAP102 clusters synaptic proteins and underlie its ability to perform its unique functions. Here we used DNA-PAINT super-resolution microscopy to measure SAP102 nano-organization and its spatial relationship to PSD-95 at individual synapses. We found that like PSD-95, SAP102 accumulates in high-density subsynaptic nanoclusters. However, SAP102 nanoclusters were smaller and denser than PSD-95 nanoclusters across development. Additionally, only a subset of SAP102 nanoclusters co-organized with PSD-95, revealing that within individual synapses there are nanodomains that contain either one or both proteins. This organization into both shared and distinct subsynaptic nanodomains may underlie the ability of SAP102 and PSD-95 to perform both common and unique synaptic functions. Significance statementSAP102 and PSD-95 are two key members of the MAGUK family of synaptic scaffold proteins that are critical for synapse development, maintenance, and modification during plasticity. Because PSD-95 has a highly complex subsynaptic nanostructure that impacts synaptic function, we asked if SAP102 is similarly organized into nanoclusters at individual synapses and how it relates to PSD-95 within synapses. We found that SAP102 forms subsynaptic nanoclusters with unique properties, and that within individual synapses proteins both concentrate into overlapping nanodomains, as well as form MAGUK-specific nanodomains. This demonstrates that organization of synaptic proteins into nanoclusters is likely to be maintained within the family of MAGUK proteins and reveals potential mechanism for specializing functions within individual synapses based on scaffold protein nanodomains.

neuroscience↗

bioRxiv2019

Properties of individual hippocampal synapses influencing NMDA-receptor activation by spontaneous neurotransmission.

NMDA receptor (NMDAR) activation is critical for maintenance and modification of synapse strength. Specifically, NMDAR activation by spontaneous glutamate release has been shown to mediate forms of synaptic plasticity as well as synaptic development. Interestingly, there is evidence that within individual synapses each release mode may be segregated such that postsynaptically there are distinct pools of responsive receptors. In order to examine potential regulators of NMDAR activation due to spontaneous glutamate release in cultured rat hippocampal neurons, we utilized GCaMP6f imaging at single synapses in concert with confocal and super-resolution imaging. Using these single spine approaches, we found that Ca2+ entry activated by spontaneous release tends to be carried by GluN2B-NMDARs. Additionally, the amount of NMDAR activation varies greatly both between synapses and within synapses, and is unrelated to spine and synapse size, but does correlate loosely with synapse distance from the soma. Despite the critical role of spontaneous activation of NMDARs in maintaining synaptic function, their activation seems to be controlled factors other than synapse size or synapse distance from the soma. It is most likely that NMDAR activation by spontaneous release influenced variability in subsynaptic receptor position, release site position, vesicle content, and channel properties. Therefore, spontaneous activation of NMDARs appears to be regulated distinctly from other receptor types, notably AMPARs, within individual synapses.\n\nSignificance StatementUnderstanding the underlying synaptic mechanisms for learning and memory is critically important to the field of neuroscience and for human health. A key neurotransmitter receptor type involved in learning is the NMDA receptor, and exploration of its regulation is vital. In this study, we optimized optical tools to allow detailed characterization of NMDA receptor activity at single synapses, along with analysis of structural features of the imaged synapses. The amount of receptor activation is independent of the size of the synapse, but weakly dependent on synapse position within the dendritic tree. Notably, we found that NMDA receptors activated following spontaneous neurotransmitter release tend be GluN2B-containing receptors. Thus, the unique mechanisms that regulate the number and positioning of these receptors within synapses will have important consequences for control of synaptic development and signaling.

neuroscience↗