Direct and indirect pathways for heterosynaptic interaction underlying developmental synapse elimination in the mouse cerebellum.

Developmental synapse elimination is crucial for shaping mature neural circuits. In the neonatal mouse cerebellum, Purkinje cells (PCs) receive excitatory synaptic inputs from multiple climbing fibers (CFs) and synapses from all but one CF are eliminated by around postnatal day 20. Heterosynaptic interaction between CFs and parallel fibers (PFs), the axons of cerebellar granule cells (GCs) forming excitatory synapses onto PCs and molecular layer interneurons (MLIs), is crucial for CF synapse elimination. However, mechanisms for this heterosynaptic interaction are largely unknown. Here we show that deletion of AMPA-type glutamate receptor functions in GCs impairs CF synapse elimination mediated by metabotropic glutamate receptor 1 (mGlu1) signaling in PCs. Furthermore, CF synapse elimination is impaired by deleting NMDA-type glutamate receptors from MLIs. We propose that PF activity is crucial for CF synapse elimination by directly activating mGlu1 in PCs and indirectly enhancing the inhibition of PCs through activating NMDA receptors in MLIs.

A nanofluidic spiking synapse.

Currently, the nanofluidic synapse can only perform basic neuromorphic pulse patterns. One immediate problem that needs to be addressed to further its capability of brain-like computing is the realization of a nanofluidic spiking device. Here, we report the use of a poly(3,4-ethylenedioxythiophene) polystyrene sulfonate membrane to achieve bionic ionic current-induced spiking. In addition to the simulation of various electrical pulse patterns, our synapse could produce transmembrane ionic current-induced spiking, which is highly analogous to biological action potentials with similar phases and excitability. Moreover, the spiking properties could be modulated by ions and neurochemicals. We expect that this work could contribute to biomimetic spiking computing in solution.

Structured connectivity in the output of the cerebellar cortex.

The spatial organization of a neuronal circuit is critically important for its function since the location of neurons is often associated with function. In the cerebellum, the major output of the cerebellar cortex are synapses made from Purkinje cells onto neurons in the cerebellar nuclei, yet little has been known about the spatial organization of these synapses. We explored this question using whole-cell electrophysiology and optogenetics in acute sagittal cerebellar slices to produce spatial connectivity maps of cerebellar cortical output in mice. We observed non-random connectivity where Purkinje cell inputs clustered in cerebellar transverse zones: while many nuclear neurons received inputs from a single zone, several multi-zonal connectivity motifs were also observed. Single neurons receiving input from all four zones were overrepresented in our data. These findings reveal that the output of the cerebellar cortex is spatially structured and represents a locus for multimodal integration in the cerebellum.

Induction of LTP mechanisms in dually innervated dendritic spines.

Dendritic spines are the postsynaptic compartments of excitatory synapses, however, a substantial subset of spines additionally receives inhibitory input. In such dually innervated spines (DiSs), excitatory long-term potentiation (LTP) mechanisms are suppressed, but can be enabled by blocking tonic inhibitory GABAB receptor signaling. Here we show that LTP mechanisms at DiSs are also enabled by two other excitatory LTP stimuli. In hippocampal neurons, these chemical LTP (cLTP) stimuli induced robust movement of the Ca2+/calmodulin-dependent protein kinase II (CaMKII) to DiSs. Such synaptic CaMKII accumulation is an essential LTP mechanism at singly innervated spines (SiSs). Indeed, CaMKII accumulation at DiSs was also accompanied by other readouts for successful LTP induction: spine growth and surface insertion of GluA1. Thus, DiSs are capable of the same LTP mechanisms as SiSs, although induction of these mechanism additionally requires either reduced inhibitory signaling or increased excitatory stimulation. This additional regulation may provide further computational control.

Repeated stimulation of feeding mechanoafferents in Aplysia generates responses consistent with the release of food.

How does repeated stimulation of mechanoafferents affect feeding motor neurons? Monosynaptic connections from a mechanoafferent population in the Aplysia buccal ganglia to five motor followers with different functions were examined during repeated stimulus trains. The mechanoafferents produced both fast and slow synaptic outputs, which could be excitatory or inhibitory. In contrast, other Aplysia mechanoafferents produce only fast excitation on their followers. In addition, patterns of synaptic connections were different to the different motor followers. Some followers received both fast excitation and fast inhibition, whereas others received exclusively fast excitation. All followers showed strong decreases in fast postsynaptic potential (PSP) amplitude within a stimulus train. Fast and slow synaptic connections were of net opposite signs in some followers but not in others. For one follower, synaptic contacts were not uniform from all subareas of the mechanoafferent cluster. Differences in properties of the buccal ganglia mechanoafferents and other Aplysia mechanoafferents may arise because the buccal ganglia neurons innervate the interior of the feeding apparatus, rather than an external surface, and connect to motor neurons for muscles with different motor functions. Fast connection patterns suggest that these synapses may be activated when food slips, biasing the musculature to release food. The largest slow inhibitory synaptic PSPs may contribute to a delay in the onset of the next behavior. Additional functions are also possible.

Synaptic Engram.

The concept of the engram refers to structural and/or physiological changes that underlie memory associations during learning. However, the precise biological basis of the engram remains elusive, with ongoing controversy regarding whether it resides at the cellular level or within the synaptic connections between activated cells. Here, we briefly review the studies investigating the cellular engram and the challenges they encounter. Subsequently, we delve into the biological basis of the engram within synaptic connections. In this regard, we introduce the history of synaptic engrams and discuss recent findings suggesting that synaptic plasticity serves as a substrate for memory. Additionally, we provide an overview of key technologies utilized in the study of synaptic plasticity.

Clustered synapses develop in distinct dendritic domains in visual cortex before eye opening.

Synaptic inputs to cortical neurons are highly structured in adult sensory systems, such that neighboring synapses along dendrites are activated by similar stimuli. This organization of synaptic inputs, called synaptic clustering, is required for high-fidelity signal processing, and clustered synapses can already be observed before eye opening. However, how clustered inputs emerge during development is unknown. Here, we employed concurrent in vivo whole-cell patch-clamp and dendritic calcium imaging to map spontaneous synaptic inputs to dendrites of layer 2/3 neurons in the mouse primary visual cortex during the second postnatal week until eye opening. We found that the number of functional synapses and the frequency of transmission events increase several fold during this developmental period. At the beginning of the second postnatal week, synapses assemble specifically in confined dendritic segments, whereas other segments are devoid of synapses. By the end of the second postnatal week, just before eye opening, dendrites are almost entirely covered by domains of co-active synapses. Finally, co-activity with their neighbor synapses correlates with synaptic stabilization and potentiation. Thus, clustered synapses form in distinct functional domains presumably to equip dendrites with computational modules for high-capacity sensory processing when the eyes open.

Rem2 interacts with CaMKII at synapses and restricts long-term potentiation in hippocampus.

Synaptic plasticity, the process whereby neuronal connections are either strengthened or weakened in response to stereotyped forms of stimulation, is widely believed to represent the molecular mechanism that underlies learning and memory. The holoenzyme calcium/calmodulin-dependent protein kinase II (CaMKII) plays a well-established and critical role in the induction of a variety of forms of synaptic plasticity such as long-term potentiation (LTP), long-term depression (LTD) and depotentiation. Previously, we identified the GTPase Rem2 as a potent, endogenous inhibitor of CaMKII. Here, we report that knock out of Rem2 enhances LTP at the Schaffer collateral to CA1 synapse in hippocampus, consistent with an inhibitory action of Rem2 on CaMKII in vivo. Further, re-expression of WT Rem2 rescues the enhanced LTP observed in slices obtained from Rem2 conditional knock out (cKO) mice, while expression of a mutant Rem2 construct that is unable to inhibit CaMKII in vitro fails to rescue increased LTP. In addition, we demonstrate that CaMKII and Rem2 interact in dendritic spines using a 2pFLIM-FRET approach. Taken together, our data lead us to propose that Rem2 serves as a brake on synaptic potentiation via inhibition of CaMKII activity. Further, the enhanced LTP phenotype we observe in Rem2 cKO slices reveals a previously unknown role for Rem2 in the negative regulation of CaMKII function.

Co-existence of synaptic plasticity and metastable dynamics in a spiking model of cortical circuits.

Evidence for metastable dynamics and its role in brain function is emerging at a fast pace and is changing our understanding of neural coding by putting an emphasis on hidden states of transient activity. Clustered networks of spiking neurons have enhanced synaptic connections among groups of neurons forming structures called cell assemblies; such networks are capable of producing metastable dynamics that is in agreement with many experimental results. However, it is unclear how a clustered network structure producing metastable dynamics may emerge from a fully local plasticity rule, i.e., a plasticity rule where each synapse has only access to the activity of the neurons it connects (as opposed to the activity of other neurons or other synapses). Here, we propose a local plasticity rule producing ongoing metastable dynamics in a deterministic, recurrent network of spiking neurons. The metastable dynamics co-exists with ongoing plasticity and is the consequence of a self-tuning mechanism that keeps the synaptic weights close to the instability line where memories are spontaneously reactivated. In turn, the synaptic structure is stable to ongoing dynamics and random perturbations, yet it remains sufficiently plastic to remap sensory representations to encode new sets of stimuli. Both the plasticity rule and the metastable dynamics scale well with network size, with synaptic stability increasing with the number of neurons. Overall, our results show that it is possible to generate metastable dynamics over meaningful hidden states using a simple but biologically plausible plasticity rule which co-exists with ongoing neural dynamics.

Synaptic Mechanisms of Ethanol Tolerance and Neuroplasticity: Insights from Invertebrate Models.

Alcohol tolerance is a neuroadaptive response that leads to a reduction in the effects of alcohol caused by previous exposure. Tolerance plays a critical role in the development of alcohol use disorder (AUD) because it leads to the escalation of drinking and dependence. Understanding the molecular mechanisms underlying alcohol tolerance is therefore important for the development of effective therapeutics and for understanding addiction in general. This review explores the molecular basis of alcohol tolerance in invertebrate models, Drosophila and C. elegans, focusing on synaptic transmission. Both organisms exhibit biphasic responses to ethanol and develop tolerance similar to that of mammals. Furthermore, the availability of several genetic tools makes them a great candidate to study the molecular basis of ethanol response. Studies in invertebrate models show that tolerance involves conserved changes in the neurotransmitter systems, ion channels, and synaptic proteins. These neuroadaptive changes lead to a change in neuronal excitability, most likely to compensate for the enhanced inhibition by ethanol.

Specific and comprehensive genetic targeting reveals brain-wide distribution and synaptic input patterns of GABAergic axo-axonic interneurons.

Axo-axonic cells (AACs), also called chandelier cells (ChCs) in the cerebral cortex, are the most distinctive type of GABAergic interneurons described in the neocortex, hippocampus, and basolateral amygdala (BLA). AACs selectively innervate glutamatergic projection neurons (PNs) at their axon initial segment (AIS), thus may exert decisive control over PN spiking and regulate PN functional ensembles. However, the brain-wide distribution, synaptic connectivity, and circuit function of AACs remain poorly understood, largely due to the lack of specific and reliable experimental tools. Here, we have established an intersectional genetic strategy that achieves specific and comprehensive targeting of AACs throughout the mouse brain based on their lineage (Nkx2.1) and molecular (Unc5b, Pthlh) markers. We discovered that AACs are deployed across essentially all the pallium-derived brain structures, including not only the dorsal pallium-derived neocortex and medial pallium-derived hippocampal formation, but also the lateral pallium-derived claustrum-insular complex, and the ventral pallium-derived extended amygdaloid complex and olfactory centers. AACs are also abundant in anterior olfactory nucleus, taenia tecta, and lateral septum. AACs show characteristic variations in density across neocortical areas and layers and across subregions of the hippocampal formation. Neocortical AACs comprise multiple laminar subtypes with distinct dendritic and axonal arborization patterns. Retrograde monosynaptic tracing from AACs across neocortical, hippocampal, and BLA regions reveal shared as well as distinct patterns of synaptic input. Specific and comprehensive targeting of AACs facilitates the study of their developmental genetic program and circuit function across brain structures, providing a ground truth platform for understanding the conservation and variation of a bona fide cell type across brain regions and species.

Whether we are memorising facts or reacting to a loud noise, nerve cells in different brain areas must be able to communicate with one another through precise, meaningful signals. Specialized nerve cells known as interneurons act as "traffic lights" to precisely regulate when and where this information flows in neural circuits. Axo-axonic cells are a rare type of inhibitory interneuron that are thought to be particularly important for controlling the passage of information between different groups of excitatory neurons. This is because they only connect to one key part of their target cell ­ the axon-initial segment ­ where the electrical signals needed for brain communication (known as action potentials) are initiated. Since axo-axonic cells are inhibitory interneurons, this connection effectively allows them to 'veto' the generation of these signals at their source. Although axo-axonic cells have been identified in three brain regions using traditional anatomical methods, there were no 'tags' readily available that can reliably identify them. Therefore, much about these cells remained unknown, including how widespread they are in the mammalian brain. To solve this problem, Raudales et al. investigated which genes are switched on in axo-axonic cells but not in other cells, identifying a unique molecular signature that could be used to mark, record, and manipulate these cells. Microscopy imaging of brain tissue from mice in which axo-axonic cells had been identified revealed that they are present in many more brain areas than previously thought, including nearly all regions of the broadly defined cerebral cortex and even the hypothalamus, which controls many innate behaviors. Axo-axonic cells were also 'wired up' differently, depending on where they were located; for example, those in brain areas associated with memory and emotions had wider-ranging input connections than other areas. The finding of Raudales et al. provide, for the first time, a method to directly track and manipulate axo-axonic cells in the brain. Since dysfunction in axo-axonic cells is also associated with neurological disorders like epilepsy and schizophrenia, gaining an insight into their distribution and connectivity could help to develop better treatments for these conditions.

Toward human-like adaptability in robotics through a retention-engineered synaptic control system.

Although advanced robots can adeptly mimic human movement and aesthetics, they are still unable to adapt or evolve in response to external experiences. To address this limitation, we propose an innovative approach that uses parallel-processable retention-engineered synaptic devices in the control system. This approach aims to simulate a human-like learning system without necessitating complex computational systems. The retention properties of the synaptic devices were modulated by adjusting the amount of Ag/AgCl ink sprayed. This changed the voltage drop across the interface between the gate electrode and the electrolyte. Furthermore, the unrestricted movement of ions in the electrolyte enhanced the signal multiplexing capability of the ion gel, enabling device-level parallel processing. By integrating the unique characteristics of the synaptic devices with actuators, we successfully emulated a human-like workout process that includes feedback between acute and chronic responses. The proposed control system offers an innovative approach to reducing system complexity and achieving a human-like learning system in the field of biomimicry.

From hidden hearing loss to supranormal auditory processing by neurotrophin 3-mediated modulation of inner hair cell synapse density.

Loss of synapses between spiral ganglion neurons and inner hair cells (IHC synaptopathy) leads to an auditory neuropathy called hidden hearing loss (HHL) characterized by normal auditory thresholds but reduced amplitude of sound-evoked auditory potentials. It has been proposed that synaptopathy and HHL result in poor performance in challenging hearing tasks despite a normal audiogram. However, this has only been tested in animals after exposure to noise or ototoxic drugs, which can cause deficits beyond synaptopathy. Furthermore, the impact of supernumerary synapses on auditory processing has not been evaluated. Here, we studied mice in which IHC synapse counts were increased or decreased by altering neurotrophin 3 (Ntf3) expression in IHC supporting cells. As we previously showed, postnatal Ntf3 knockdown or overexpression reduces or increases, respectively, IHC synapse density and suprathreshold amplitude of sound-evoked auditory potentials without changing cochlear thresholds. We now show that IHC synapse density does not influence the magnitude of the acoustic startle reflex or its prepulse inhibition. In contrast, gap-prepulse inhibition, a behavioral test for auditory temporal processing, is reduced or enhanced according to Ntf3 expression levels. These results indicate that IHC synaptopathy causes temporal processing deficits predicted in HHL. Furthermore, the improvement in temporal acuity achieved by increasing Ntf3 expression and synapse density suggests a therapeutic strategy for improving hearing in noise for individuals with synaptopathy of various etiologies.

Functional Dynamics and Selectivity of Two Parallel Corticocortical Pathways from Motor Cortex to Layer 5 Circuits in Somatosensory Cortex.

In the rodent whisker system, active sensing and sensorimotor integration are mediated in part by the dynamic interactions between the motor cortex (M1) and somatosensory cortex (S1). However, understanding these dynamic interactions requires knowledge about the synapses and how specific neurons respond to their input. Here, we combined optogenetics, retrograde labeling, and electrophysiology to characterize the synaptic connections between M1 and layer 5 (L5) intratelencephalic (IT) and pyramidal tract (PT) neurons in S1 of mice (both sexes). We found that M1 synapses onto IT cells displayed modest short-term depression, whereas synapses onto PT neurons showed robust short-term facilitation. Despite M1 inputs to IT cells depressing, their slower kinetics resulted in summation and a response that increased during short trains. In contrast, summation was minimal in PT neurons due to the fast time course of their M1 responses. The functional consequences of this reduced summation, however, were outweighed by the strong facilitation at these M1 synapses, resulting in larger response amplitudes in PT neurons than IT cells during repetitive stimulation. To understand the impact of facilitating M1 inputs on PT output, we paired trains of inputs with single backpropagating action potentials, finding that repetitive M1 activation increased the probability of bursts in PT cells without impacting the time dependence of this coupling. Thus, there are two parallel but dynamically distinct systems of M1 synaptic excitation in L5 of S1, each defined by the short-term dynamics of its synapses, the class of postsynaptic neurons, and how the neurons respond to those inputs.

Skewing information flow through pre- and postsynaptic plasticity in the mushroom bodies of Drosophila.

Animal brains need to store information to construct a representation of their environment. Knowledge of what happened in the past allows both vertebrates and invertebrates to predict future outcomes by recalling previous experience. Although invertebrate and vertebrate brains share common principles at the molecular, cellular, and circuit-architectural levels, there are also obvious differences as exemplified by the use of acetylcholine versus glutamate as the considered main excitatory neurotransmitters in the respective central nervous systems. Nonetheless, across central nervous systems, synaptic plasticity is thought to be a main substrate for memory storage. Therefore, how brain circuits and synaptic contacts change following learning is of fundamental interest for understanding brain computations tied to behavior in any animal. Recent progress has been made in understanding such plastic changes following olfactory associative learning in the mushroom bodies (MBs) of Drosophila A current framework of memory-guided behavioral selection is based on the MB skew model, in which antagonistic synaptic pathways are selectively changed in strength. Here, we review insights into plasticity at dedicated Drosophila MB output pathways and update what is known about the plasticity of both pre- and postsynaptic compartments of Drosophila MB neurons.

Physiological roles of endocytosis and presynaptic scaffold in vesicle replenishment at fast and slow central synapses.

After exocytosis, release sites are cleared of vesicular residues to replenish with transmitter-filled vesicles. Endocytic and scaffold proteins are thought to underlie this site-clearance mechanism. However, the physiological significance of this mechanism at diverse mammalian central synapses remains unknown. Here, we tested this in a physiologically optimized condition using action potential evoked EPSCs at fast calyx synapse and relatively slow hippocampal CA1 synapse, in post-hearing mice brain slices at 37°C and in 1.3 mM [Ca2+]. Pharmacological block of endocytosis enhanced synaptic depression at the calyx synapse, whereas it attenuated synaptic facilitation at the hippocampal synapse. Block of scaffold protein activity likewise enhanced synaptic depression at the calyx but had no effect at the hippocampal synapse. At the fast calyx synapse, block of endocytosis or scaffold protein activity significantly enhanced synaptic depression as early as 10 ms after the stimulation onset. Unlike previous reports, neither endocytic blockers nor scaffold protein inhibitors prolonged the recovery from short-term depression. We conclude that the release-site clearance by endocytosis can be a universal phenomenon supporting vesicle replenishment at both fast and slow synapses, whereas the presynaptic scaffold mechanism likely plays a specialized role in vesicle replenishment predominantly at fast synapses.

Hebbian priming of human motor learning.

Motor learning relies on experience-dependent plasticity in relevant neural circuits. In four experiments, we provide initial evidence and a double-blinded, sham-controlled replication (Experiment I-II) demonstrating that motor learning involving ballistic index finger movements is improved by preceding paired corticospinal-motoneuronal stimulation (PCMS), a human model for exogenous induction of spike-timing-dependent plasticity. Behavioral effects of PCMS targeting corticomotoneuronal (CM) synapses are order- and timing-specific and partially bidirectional (Experiment III). PCMS with a 2 ms inter-arrival interval at CM-synapses enhances learning and increases corticospinal excitability compared to control protocols. Unpaired stimulations did not increase corticospinal excitability (Experiment IV). Our findings demonstrate that non-invasively induced plasticity interacts positively with experience-dependent plasticity to promote motor learning. The effects of PCMS on motor learning approximate Hebbian learning rules, while the effects on corticospinal excitability demonstrate timing-specificity but not bidirectionality. These findings offer a mechanistic rationale to enhance motor practice effects by priming sensorimotor training with individualized PCMS.

Persistent spiking activity in neuromorphic circuits incorporating post-inhibitory rebound excitation.

Objective. This study introduces a novel approach for integrating the post-inhibitory rebound excitation (PIRE) phenomenon into a neuronal circuit. Excitatory and inhibitory synapses are designed to establish a connection between two hardware neurons, effectively forming a network. The model demonstrates the occurrence of PIRE under strong inhibitory input. Emphasizing the significance of incorporating PIRE in neuromorphic circuits, the study showcases generation of persistent activity within cyclic and recurrent spiking neuronal networks.Approach. The neuronal and synaptic circuits are designed and simulated in Cadence Virtuoso using TSMC 180 nm technology. The operating mechanism of the PIRE phenomenon integrated into a hardware neuron is discussed. The proposed circuit encompasses several parameters for effectively controlling multiple electrophysiological features of a neuron.Main results. The neuronal circuit has been tuned to match the response of a biological neuron. The efficiency of this circuit is evaluated by computing the average power dissipation and energy consumption per spike through simulation. The sustained firing of neural spikes is observed till 1.7 s using the two neuronal networks.Significance. Persistent activity has significant implications for various cognitive functions such as working memory, decision-making, and attention. Therefore, hardware implementation of these functions will require our PIRE-integrated model. Energy-efficient neuromorphic systems are useful in many artificial intelligence applications, including human-machine interaction, IoT devices, autonomous systems, and brain-computer interfaces.

Quantification of Microglial Engulfment of Synaptic Material Using Flow Cytometry.

Microglia play a pivotal role in synaptic refinement in the brain. Analysis of microglial engulfment of synapses is essential for comprehending this process; however, currently available methods for identifying microglial engulfment of synapses, such as immunohistochemistry (IHC) and imaging, are laborious and time-intensive. To address this challenge, herein we present in vitro and in vivo* assays that allow fast and high-throughput quantification of microglial engulfment of synapses using flow cytometry. In the in vivo* approach, we performed intracellular vGLUT1 staining following fresh cell isolation from adult mouse brains to quantify engulfment of vGLUT1+ synapses by microglia. In the in vitro synaptosome engulfment assay, we used freshly isolated cells from the adult mouse brain to quantify the engulfment of pHrodo Red-labeled synaptosomes by microglia. These protocols together provide a time-efficient approach to quantifying microglial engulfment of synapses and represent promising alternatives to labor-intensive image analysis-based methods. By streamlining the analysis, these assays can contribute to a better understanding of the role of microglia in synaptic refinement in different disease models.