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ELife May 2024Vesicles within presynaptic terminals are thought to be segregated into a variety of readily releasable and reserve pools. The nature of the pools and trafficking...
Vesicles within presynaptic terminals are thought to be segregated into a variety of readily releasable and reserve pools. The nature of the pools and trafficking between them is not well understood, but pools that are slow to mobilize when synapses are active are often assumed to feed pools that are mobilized more quickly, in a series. However, electrophysiological studies of synaptic transmission have suggested instead a parallel organization where vesicles within slowly and quickly mobilized reserve pools would separately feed independent reluctant- and fast-releasing subdivisions of the readily releasable pool. Here, we use FM-dyes to confirm the existence of multiple reserve pools at hippocampal synapses and a parallel organization that prevents intermixing between the pools, even when stimulation is intense enough to drive exocytosis at the maximum rate. The experiments additionally demonstrate extensive heterogeneity among synapses in the relative sizes of the slowly and quickly mobilized reserve pools, which suggests equivalent heterogeneity in the numbers of reluctant and fast-releasing readily releasable vesicles that may be relevant for understanding information processing and storage.
Topics: Animals; Hippocampus; Synaptic Vesicles; Synapses; Synaptic Transmission; Rats; Exocytosis; Presynaptic Terminals
PubMed: 38727712
DOI: 10.7554/eLife.88212 -
The Journal of Neuroscience : the... Jun 2024Understanding the function of the human brain requires determining basic properties of synaptic transmission in human neurons. One of the most fundamental parameters...
Understanding the function of the human brain requires determining basic properties of synaptic transmission in human neurons. One of the most fundamental parameters controlling neurotransmitter release is the presynaptic action potential, but its amplitude and duration remain controversial. Presynaptic action potentials have so far been measured with high temporal resolution only in a limited number of vertebrate but not in human neurons. To uncover properties of human presynaptic action potentials, we exploited recently developed tools to generate human glutamatergic neurons by transient expression of Neurogenin 2 (Ngn2) in pluripotent stem cells. During maturation for 3 to 9 weeks of culturing in different established media, the proportion of cells with multiple axon initial segments decreased, while the amount of axonal tau protein and neuronal excitability increased. Super-resolution microscopy revealed the alignment of the pre- and postsynaptic proteins, Bassoon and Homer. Synaptic transmission was surprisingly reliable at frequencies of 20, 50, and 100 Hz. The synchronicity of synaptic transmission during high-frequency transmission increased during 9 weeks of neuronal maturation. To analyze the mechanisms of synchronous high-frequency glutamate release, we developed direct presynaptic patch-clamp recordings from human neurons. The presynaptic action potentials had large overshoots to ∼25 mV and short durations of ∼0.5 ms. Our findings show that Ngn2-induced neurons represent an elegant model system allowing for functional, structural, and molecular analyses of glutamatergic synaptic transmission with high spatiotemporal resolution in human neurons. Furthermore, our data predict that glutamatergic transmission is mediated by large and rapid presynaptic action potentials in the human brain.
Topics: Humans; Induced Pluripotent Stem Cells; Action Potentials; Synapses; Neurons; Presynaptic Terminals; Nerve Tissue Proteins; Synaptic Transmission; Cells, Cultured; Basic Helix-Loop-Helix Transcription Factors; Cell Differentiation
PubMed: 38724283
DOI: 10.1523/JNEUROSCI.0971-23.2024 -
The Journal of Physiology Jun 2024Neurons in the central nervous system communicate with each other by activating billions of tiny synaptic boutons distributed along their fine axons. These presynaptic...
Neurons in the central nervous system communicate with each other by activating billions of tiny synaptic boutons distributed along their fine axons. These presynaptic varicosities are very crowded environments, comprising hundreds of synaptic vesicles. Only a fraction of these vesicles can be recruited in a single release episode, either spontaneous or evoked by action potentials. Since the seminal work by Fatt and Katz, spontaneous release has been modelled as a memoryless process. Nevertheless, at central synapses, experimental evidence indicates more complex features, including non-exponential distributions of release intervals and power-law behaviour in their rate. To describe these features, we developed a probabilistic model of spontaneous release based on Brownian motion of synaptic vesicles in the presynaptic environment. To account for different diffusion regimes, we based our simulations on fractional Brownian motion. We show that this model can predict both deviation from the Poisson hypothesis and power-law features in experimental quantal release series, thus suggesting that the vesicular motion by diffusion could per se explain the emergence of these properties. We demonstrate the efficacy of our modelling approach using electrophysiological recordings at single synaptic boutons and ultrastructural data. When this approach was used to simulate evoked responses, we found that the replenishment of the readily releasable pool driven by Brownian motion of vesicles can reproduce the characteristic binomial release distributions seen experimentally. We believe that our modelling approach supports the idea that vesicle diffusion and readily releasable pool dynamics are crucial factors for the physiological functioning of neuronal communication. KEY POINTS: We developed a new probabilistic model of spontaneous and evoked vesicle fusion based on simple biophysical assumptions, including the motion of vesicles before they dock to the release site. We provide closed-form equations for the interval distribution of spontaneous releases in the special case of Brownian diffusion of vesicles, showing that a power-law heavy tail is generated. Fractional Brownian motion (fBm) was exploited to simulate anomalous vesicle diffusion, including directed and non-directed motion, by varying the Hurst exponent. We show that our model predicts non-linear features observed in experimental spontaneous quantal release series as well as ultrastructural data of synaptic vesicles spatial distribution. Evoked exocytosis based on a diffusion-replenished readily releasable pool might explain the emergence of power-law behaviour in neuronal activity.
Topics: Synaptic Vesicles; Animals; Synaptic Transmission; Models, Neurological; Presynaptic Terminals; Rats; Diffusion
PubMed: 38723211
DOI: 10.1113/JP284926 -
The Journal of Neuroscience : the... Jun 2024Acetylcholine is a robust neuromodulator of the limbic system and a critical regulator of arousal and emotions. The anterior cingulate cortex (ACC) and the amygdala...
Acetylcholine is a robust neuromodulator of the limbic system and a critical regulator of arousal and emotions. The anterior cingulate cortex (ACC) and the amygdala (AMY) are key limbic structures that are both densely innervated by cholinergic afferents and interact with each other for emotional regulation. The ACC is composed of functionally distinct dorsal (A24), rostral (A32), and ventral (A25) areas that differ in their connections with the AMY. The structural substrates of cholinergic modulation of distinct ACC microcircuits and outputs to AMY are thought to depend on the laminar and subcellular localization of cholinergic receptors. The present study examines the distribution of muscarinic acetylcholine receptors, m1 and m2, on distinct excitatory and inhibitory neurons and on AMY-targeting projection neurons within ACC areas, via immunohistochemistry and injections of neural tracers into the basolateral AMY in adult rhesus monkeys of both sexes. We found that laminar densities of m1+ and m2+ expressing excitatory and inhibitory neurons depended on area and cell type. Among the ACC areas, ventral subgenual ACC A25 exhibited greater m2+ localization on presynaptic inhibitory axon terminals and greater density of m1+ and m2+ expressing AMY-targeting (tracer+) pyramidal neurons. These patterns suggest robust cholinergic disinhibition and potentiation of amygdalar outputs from the limbic ventral ACC, which may be linked to the hyperexcitability of this subgenual ACC area in depression. These findings reveal the anatomical substrate of diverse cholinergic modulation of specific ACC microcircuits and amygdalar outputs that mediate cognitive-emotional integration and dysfunctions underlying stress and affective disorders.
Topics: Animals; Gyrus Cinguli; Macaca mulatta; Male; Female; Receptor, Muscarinic M2; Receptor, Muscarinic M1; Nerve Net; Acetylcholine; Neural Pathways; Neurons
PubMed: 38719447
DOI: 10.1523/JNEUROSCI.0953-23.2024 -
Brain Research Sep 2024Dynamin is a microtubule (MT) binding protein playing a key role in vesicle endocytosis. In a brain slice model, tau loaded in presynaptic terminals assembles MTs,...
Dynamin is a microtubule (MT) binding protein playing a key role in vesicle endocytosis. In a brain slice model, tau loaded in presynaptic terminals assembles MTs, thereby impairing vesicle endocytosis via depletion of cytosolic dynamin. The peptide PHDP5, derived from the pleckstrin homology domain of dynamin 1, inhibits dynamin-MT interaction and rescues endocytosis and synaptic transmission impaired by tau when co-loaded in presynaptic terminals. We tested whether in vivo administration of PHDP5 could rescue the learning/memory deficits observed in Alzheimer's disease (AD) model mice. A modified PHDP5 incorporating a cell-penetrating peptide (CPP) and a FITC fluorescent marker was delivered intranasally to Tau609 transgenic (Tg) and 3xTg-AD mice. FITC-positive puncta were observed in the hippocampus of mice infused with PHDP5 or scrambled (SPHDP5) peptide, but not in saline-infused controls. In the Morris water maze (MWM) test for spatial learning/memory, AD model mice treated with FITC-PHDP5-CPP showed prominent improvements in learning and memory, performing close to the level of saline-infused WT mice control. In contrast, mice treated with a scrambled construct (FITC-SPHDP5-CPP) showed no significant improvement. We conclude that PHDP5 can be a candidate for human AD therapy.
Topics: Animals; Alzheimer Disease; Mice; Mice, Transgenic; Memory Disorders; Disease Models, Animal; Spatial Learning; Microtubules; Hippocampus; Maze Learning; Dynamins; Male; tau Proteins
PubMed: 38718851
DOI: 10.1016/j.brainres.2024.148987 -
Frontiers in Neural Circuits 2024A morphologically present but non-functioning synapse is termed a silent synapse. Silent synapses are categorized into "postsynaptically silent synapses," where AMPA...
A morphologically present but non-functioning synapse is termed a silent synapse. Silent synapses are categorized into "postsynaptically silent synapses," where AMPA receptors are either absent or non-functional, and "presynaptically silent synapses," where neurotransmitters cannot be released from nerve terminals. The presence of presynaptically silent synapses remains enigmatic, and their physiological significance is highly intriguing. In this study, we examined the distribution and developmental changes of presynaptically active and silent synapses in individual neurons. Our findings show a gradual increase in the number of excitatory synapses, along with a corresponding decrease in the percentage of presynaptically silent synapses during neuronal development. To pinpoint the distribution of presynaptically active and silent synapses, i.e., their positional information, we employed Sholl analysis. Our results indicate that the distribution of presynaptically silent synapses within a single neuron does not exhibit a distinct pattern during synapse development in different distance from the cell body. However, irrespective of neuronal development, the proportion of presynaptically silent synapses tends to rise as the projection site moves farther from the cell body, suggesting that synapses near the cell body may exhibit higher synaptic transmission efficiency. This study represents the first observation of changes in the distribution of presynaptically active and silent synapses within a single neuron.
Topics: Animals; Hippocampus; Neurons; Synapses; Cells, Cultured; Presynaptic Terminals; Excitatory Postsynaptic Potentials; Rats; Synaptic Transmission
PubMed: 38715983
DOI: 10.3389/fncir.2024.1358570 -
ELife May 2024The cytosolic proteins synucleins and synapsins are thought to play cooperative roles in regulating synaptic vesicle (SV) recycling, but mechanistic insight is lacking....
The cytosolic proteins synucleins and synapsins are thought to play cooperative roles in regulating synaptic vesicle (SV) recycling, but mechanistic insight is lacking. Here, we identify the synapsin E-domain as an essential functional binding-partner of α-synuclein (α-syn). Synapsin E-domain allows α-syn functionality, binds to α-syn, and is necessary and sufficient for enabling effects of α-syn at synapses of cultured mouse hippocampal neurons. Together with previous studies implicating the E-domain in clustering SVs, our experiments advocate a cooperative role for these two proteins in maintaining physiologic SV clusters.
Topics: Animals; Humans; Mice; alpha-Synuclein; Cells, Cultured; Hippocampus; Neurons; Protein Binding; Protein Domains; Synapses; Synapsins; Synaptic Vesicles
PubMed: 38713200
DOI: 10.7554/eLife.89687 -
BioRxiv : the Preprint Server For... Apr 2024Memory engrams are formed through experience-dependent remodeling of neural circuits, but their detailed architectures have remained unresolved. Using 3D electron...
Memory engrams are formed through experience-dependent remodeling of neural circuits, but their detailed architectures have remained unresolved. Using 3D electron microscopy, we performed nanoscale reconstructions of the hippocampal CA3-CA1 pathway following chemogenetic labeling of cellular ensembles with a remote history of correlated excitation during associative learning. Projection neurons involved in memory acquisition expanded their connectomes via multi-synaptic boutons without altering the numbers and spatial arrangements of individual axonal terminals and dendritic spines. This expansion was driven by presynaptic activity elicited by specific negative valence stimuli, regardless of the co-activation state of postsynaptic partners. The rewiring of initial ensembles representing an engram coincided with local, input-specific changes in the shapes and organelle composition of glutamatergic synapses, reflecting their weights and potential for further modifications. Our findings challenge the view that the connectivity among neuronal substrates of memory traces is governed by Hebbian mechanisms, and offer a structural basis for representational drifts.
PubMed: 38712256
DOI: 10.1101/2024.04.23.590812 -
Biochemical and Biophysical Research... Jul 2024Calcium (Ca) in mitochondria plays crucial roles in neurons including modulating metabolic processes. Moreover, excessive Ca in mitochondria can lead to cell death....
Calcium (Ca) in mitochondria plays crucial roles in neurons including modulating metabolic processes. Moreover, excessive Ca in mitochondria can lead to cell death. Thus, altered mitochondrial Ca regulation has been implicated in several neurodegenerative diseases including Huntington's disease (HD). HD is a progressive hereditary neurodegenerative disorder that results from abnormally expanded cytosine-adenine-guanine trinucleotide repeats in the huntingtin gene. One neuropathological hallmark of HD is neuronal loss in the striatum and cortex. However, mechanisms underlying selective loss of striatal and cortical neurons in HD remain elusive. Here, we measured the basal Ca levels and Ca uptake in single presynaptic mitochondria during 100 external electrical stimuli using highly sensitive mitochondria-targeted Ca indicators in cultured cortical and striatal neurons of a knock-in mouse model of HD (zQ175 mice). We observed elevated presynaptic mitochondrial Ca uptake during 100 electrical stimuli in HD cortical neurons compared with wild-type (WT) cortical neurons. We also found the highly elevated presynaptic mitochondrial basal Ca level and Ca uptake during 100 stimuli in HD striatal neurons. The elevated presynaptic mitochondrial basal Ca level in HD striatal neurons and Ca uptake during stimulation in HD striatal and cortical neurons can disrupt neurotransmission and induce mitochondrial Ca overload, eventually leading to neuronal death in the striatum and cortex of HD.
Topics: Animals; Huntington Disease; Calcium; Mitochondria; Mice; Corpus Striatum; Cerebral Cortex; Presynaptic Terminals; Cells, Cultured; Disease Models, Animal; Gene Knock-In Techniques; Neurons; Mice, Transgenic
PubMed: 38704892
DOI: 10.1016/j.bbrc.2024.150010 -
EMBO Reports Jun 2024GABA receptors (GBRs), the G protein-coupled receptors for GABA, regulate synaptic transmission throughout the brain. A main synaptic function of GBRs is the gating of...
GABA receptors (GBRs), the G protein-coupled receptors for GABA, regulate synaptic transmission throughout the brain. A main synaptic function of GBRs is the gating of Cav2.2-type Ca channels. However, the cellular compartment where stable GBR/Cav2.2 signaling complexes form remains unknown. In this study, we demonstrate that the vesicular protein synaptotagmin-11 (Syt11) binds to both the auxiliary GBR subunit KCTD16 and Cav2.2 channels. Through these dual interactions, Syt11 recruits GBRs and Cav2.2 channels to post-Golgi vesicles, thus facilitating assembly of GBR/Cav2.2 signaling complexes. In addition, Syt11 stabilizes GBRs and Cav2.2 channels at the neuronal plasma membrane by inhibiting constitutive internalization. Neurons of Syt11 knockout mice exhibit deficits in presynaptic GBRs and Cav2.2 channels, reduced neurotransmitter release, and decreased GBR-mediated presynaptic inhibition, highlighting the critical role of Syt11 in the assembly and stable expression of GBR/Cav2.2 complexes. These findings support that Syt11 acts as a vesicular scaffold protein, aiding in the assembly of signaling complexes from low-abundance components within transport vesicles. This mechanism enables insertion of pre-assembled functional signaling units into the synaptic membrane.
Topics: Animals; Synaptotagmins; Mice; Signal Transduction; Mice, Knockout; Humans; Neurons; Synaptic Transmission; Receptors, GABA-B; Presynaptic Terminals; Calcium Channels, N-Type; Golgi Apparatus; Protein Binding; HEK293 Cells
PubMed: 38698221
DOI: 10.1038/s44319-024-00147-0