News and Publications

Patzke C, Dai J, Brockmann MM, Sun Z, Fenske P, Rosenmund C, Südhof TC. (2021) Cannabinoid receptor activation acutely increases synaptic vesicle numbers by activating synapsins in human synapses. Molecular Psychiatry

Cannabis and cannabinoid drugs are central agents that are used widely recreationally and are employed broadly for treating psychiatric conditions. Cannabinoids primarily act by stimulating presynaptic CB1 receptors (CB1Rs), the most abundant Gprotein-coupled receptors in brain. CB1R activation decreases neurotransmitter release by inhibiting presynaptic Ca2+ channels and induces long-term plasticity by decreasing cellular cAMP levels. Here we identified an unanticipated additional mechanism of acute cannabinoid signaling in presynaptic terminals that regulates the size of synaptic vesicle pools available for neurotransmitter release. Specifically, we show that activation of CB1Rs in human and mouse neurons rapidly recruits vesicles to nerve terminals by suppressing the cAMP-dependent phosphorylation of synapsins. We confirmed this unanticipated mechanism using conditional deletion of synapsin-1, the predominant synapsin isoform in human neurons, demonstrating that synapsin-1 significantly contributes to the CB1R-dependent regulation of neurotransmission. Interestingly, acute activation of the Gi-DREADD hM4D mimics the effect of CB1R activation in a synapsin-1- dependent manner, suggesting that the control of synaptic vesicle numbers by synapsin-1 phosphorylation is a general presynaptic mechanism of neuromodulation. Thus, we uncovered a CB1R-dependent presynaptic mechanism that rapidly regulates the organization and neurotransmitter release properties of synapses.

Figure caption: CB1 cannabinoid receptor activation rapidly increases synaptic vesicle numbers in human synapses.

  1. Schematic rationale of experiments
  2. Human neurons before and after acute treatment for activation of CB1 receptors with WIN55 or inhibition with AM251 stained for presynaptic markers synapsin (‘Pan-Syn’), synaptophysin (‘Syph’), and MAP2.
  3. EM micrographs of high-pressure frozen human neurons before and after acute treatment with WIN55 or AM251

Luo F, Sclip A, Merrill S, Südhof TC. (2021) Neurexins regulate presynaptic GABAB-receptors at central synapses.

Diverse signaling complexes are precisely assembled at the presynaptic active zone for dynamic modulation of synaptic transmission and synaptic plasticity. Presynaptic GABAB-receptors nucleate critical signaling complexes regulating neurotransmitter release at most synapses. However, the molecular mechanisms underlying assembly of GABAB-receptor signaling complexes remain unclear. Here we show that neurexins are required for the localization and function of presynaptic GABAB-receptor signaling complexes. At four model synapses, excitatory calyx of Held synapses in the brainstem, excitatory and inhibitory synapses on hippocampal CA1-region pyramidal neurons, and inhibitory basket cell synapses in the cerebellum, deletion of neurexins rendered neurotransmitter release significantly less sensitive to GABAB-receptor activation. Moreover, deletion of neurexins caused a loss of GABAB-receptors from the presynaptic active zone of the calyx synapse. These findings extend the role of neurexins at the presynaptic active zone to enabling GABAB-receptor signaling, supporting the notion that neurexins function as central organizers of active zone signaling complexes.

Figure caption: Neurexins are required for intact functional organization of GABAB-receptors at the calyx of Held. a, The diagram of the calyx of Held synapse. b, Strategy for selective deletions of all neurexins at the calyx of Held by crossing PV-Cre mice with triple Nrxn123 cKO mice. c, Representative traces of EPSC before and after application of 20 µM SKF-97541 (SKF), a potent and selective GABAB-receptor agonist, recorded in acute slices from littermate control and neurexin123 TKO mice at P12–P14. The normalized EPSCs before and after SKF are shown in inset. d, Summary graphs of EPSC1 amplitudes before and after SKF for control and Nrxn123 TKO mice. e, Summary graphs of EPSC1 remaining unblocked by SKF application. f, Summary graphs of the paired-pulse ratio (PPR) before and after SKF in control and Nrxn123 TKO mice. g, Representative confocal microscopy images of MNTB-containing brainstem slice with specific labeling of VGluT1 (green) and GABAB-receptor subunit 2 (GABAB2, red) from both littermate control and Nrxn123 TKO mice at P12. h, Summary of GABAB2 immunostaining intensity (normalized to control). I & j, Same as g & h except for specific labeling of VGluT1 (green) and GABAB-receptor subunit 1 (GABAB1, red).

Data are means ± SEM. Number of cells (from at least three mice per group) analyzed are indicated in the bars; Statistical differences were assessed by Student’s t test or two way ANOVA with Bonferroni post hoc test (*P < 0.05; **P < 0.01; ***P < 0.001)

Luo, F., Sclip, A., Jiang, M., and Südhof, T.C. (2020) Neurexins Cluster Ca2+ Channels within presynaptic Active Zone. EMBO J. 39, e103208.

Luo et al. demonstrate that deletion of all neurexins from the calyx of Held synapse in the brainstem suppresses neurotransmitter release evoked by action potentials because the influx of calcium into the presynaptic terminal is impaired. The release machinery itself is intact, and the total amount of depolarization-evoked calcium influx into the nerve terminal is not decreased. Instead, the loss of neurexins suppresses neurotransmitter release because the calcium channels are more distant to the release sites, causing a spatial uncoupling of calcium influx and neurotransmitter release sites.

Figure caption: Deletion of neurexins impairs neurotransmitter release at an ambient 1 mM calcium concentration by suppressing the release probability, as indicated by the rise in paired-pulse ratio. The same suppression of release by the neurexin deletion is observed at a higher 2 mM ambient calcium concentration when the terminal contains the slow calcium buffer EGTA, which has no effect on release in wild-type synapses.

Sando R, Südhof TC. (2021) Latrophilin GPCR signaling mediates synapse formation. eLife. 2021; 10: e65717.

Neural circuit assembly in the brain requires precise establishment of synaptic connections, but the mechanisms of synapse assembly remain incompletely understood. Latrophilins are postsynaptic adhesion-GPCRs that engage in trans-synaptic complexes with presynaptic teneurins and FLRTs. In mouse CA1-region neurons, Latrophilin-2 and Latrophilin-3 are essential for formation of entorhinal-cortex-derived and Schaffer-collateral-derived synapses, respectively. However, it is unknown whether latrophilins function as GPCRs in synapse formation. Here, we show that Latrophilin-2 and Latrophilin-3 exhibit constitutive GPCR activity that increases cAMP levels, which was blocked by a mutation interfering with G-protein and arrestin interactions of GPCRs. The same mutation impaired the ability of Latrophilin-2 and Latrophilin-3 to rescue the synapse-loss phenotype in Latrophilin-2 and Latrophilin-3 knockout neurons in vivo. Our results suggest that Latrophilin-2 and Latrophilin-3 require GPCR signaling in synapse formation, indicating that latrophilins promote synapse formation in the hippocampus by activating a classical GPCR-signaling pathway.

Figure caption: Latrophilins (Lphns) function as postsynaptic GPCRs that mediate excitatory hippocampal synapse formation and specificity. Mutations that disrupt Lphn signal transduction abolish their ability to rescue synapse formation deficits in Lphn conditional KO cultured
hippocampal neurons.

Chen, MB, Jiang X, Quake QR, Südhof TC. (2020) Persistent transcriptional programmes are associated with remote memory. Nature, 2020 Nov;587(7834):437-442.

The role of gene expression during learning and in short-term memories has been studied extensively, but less is known about remote memories, which can persist for a lifetime. Here we used long-term contextual fear memory as a paradigm to probe the single-cell gene expression landscape that underlies remote memory storage in the medial prefrontal cortex. We found persistent activity-specific transcriptional alterations in diverse populations of neurons that lasted for weeks after fear learning. Out of a vast plasticity-coding space, we identified genes associated with membrane fusion that could have important roles in the maintenance of remote memory. Unexpectedly, astrocytes and microglia also acquired persistent gene expression signatures that were associated with remote memory, suggesting that they actively contribute to memory circuits. The discovery of gene expression programmes associated with remote memory engrams adds an important dimension of activity-dependent cellular states to existing brain taxonomy atlases and sheds light on the elusive mechanisms of remote memory storage.

Figure Caption: a, Volcano plots of non-neuronal cell types when comparing cells in FR over NF nice. DEGs (FDR >0.01, log2FC >1) are labelled in red, and exemplary DEGs (high log2FC and log10FDR) are labelled in black. b, Number of non-neuronal cells collected in this study, for each cell type and experimental condition. c, Heat map of a subset of neuronal ligands and glial receptors that are found to be differentially perturbed upon memory consolidation. Only receptors and ligands which were found to be (differentially) expressed are shown. d, Left, heat map of the log2FC of DEGs (FR over NF) in neurons that are classified as ligands. Middle and right, Sankey plot of known ligand-receptor pairs and heat map of the average scaled expression level of the corresponding receptors in each cell type. e, Left, heat map of the log2FC of DEGs (FR over NF) in neurons that are classified as receptors. Middle and right, Sankey plot of known ligand-receptor pairs and heat map of the average scaled expression level of the corresponding ligands in each cell type.