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Cellular and circuit hyperexcitability are core features of fragile X syndrome and related autism spectrum disorder models. However, the cellular and synaptic bases of this hyperexcitability have proved elusive. We report in a mouse model of fragile X syndrome, glutamate uncaging onto individual dendritic spines yields stronger single-spine excitation than wild-type, with more silent spines. Furthermore, fewer spines are required to trigger an action potential with near-simultaneous uncaging at multiple spines. This is, in part, from increased dendritic gain due to increased intrinsic excitability, resulting from reduced hyperpolarization-activated currents, and increased NMDA receptor signaling. Using super-resolution microscopy we detect no change in dendritic spine morphology, indicating no structure-function relationship at this age. However, ultrastructural analysis shows a 3-fold increase in multiply-innervated spines, accounting for the increased single-spine glutamate currents. Thus, loss of FMRP causes abnormal synaptogenesis, leading to large numbers of poly-synaptic spines despite normal spine morphology, thus explaining the synaptic perturbations underlying circuit hyperexcitability., Fragile X syndrome and autism spectrum disorders are associated with circuit hyperexcitability, however, its cellular and synaptic bases are not well understood. Here, the authors report abnormal synaptogenesis with an increased prevalence of polysynaptic spines with normal morphology in a mouse model of fragile X.

Sensory hypersensitivity is a common and debilitating feature of neurodevelopmental disorders such as Fragile X Syndrome (FXS). How developmental changes in neuronal function culminate in network dysfunction that underlies sensory hypersensitivities is unknown. By systematically studying cellular and synaptic properties of layer 4 neurons combined with cellular and network simulations, we explored how the array of phenotypes in Fmr1-knockout (KO) mice produce circuit pathology during development. We show that many of the cellular and synaptic pathologies in Fmr1-KO mice are antagonistic, mitigating circuit dysfunction, and hence may be compensatory to the primary pathology. Overall, the layer 4 network in the Fmr1-KO exhibits significant alterations in spike output in response to thalamocortical input and distorted sensory encoding. This developmental loss of layer 4 sensory encoding precision would contribute to subsequent developmental alterations in layer 4-to-layer 2/3 connectivity and plasticity observed in Fmr1-KO mice, and circuit dysfunction underlying sensory hypersensitivity., Somatosensory hypersensitivity in Fmr-1 knockout mice is thought to arise from an increase in cortical circuit excitability. Here, the authors report that the loss of precision of sensory encoding in the Layer 4 of barrel cortex is the primary developmental circuit alteration that drives the other compensatory circuit dysfunction.

Injury induces synaptic, circuit, and systems reorganization. After unilateral amputation or stroke, this functional loss disrupts the interhemispheric interaction between intact and deprived somatomotor cortices to recruit deprived cortex in response to intact limb stimulation. This recruitment has been implicated in enhanced intact sensory function. In other patients, maladaptive consequences such as phantom limb pain can occur. We used unilateral whisker denervation in male and female mice to detect circuitry alterations underlying interhemispheric cortical reorganization. Enhanced synaptic strength from the intact cortex via the corpus callosum (CC) onto deep neurons in deprived primary somatosensory barrel cortex (S1BC) has previously been detected. It was hypothesized that specificity in this plasticity may depend on to which area these neurons projected. Increased connectivity to somatomotor areas such as contralateral S1BC, primary motor cortex (M1) and secondary somatosensory cortex (S2) may underlie beneficial adaptations, while increased connectivity to pain areas like anterior cingulate cortex (ACC) might underlie maladaptive pain phenotypes. Neurons from the deprived S1BC that project to intact S1BC were hyperexcitable, had stronger responses and reduced inhibitory input to CC stimulation. M1-projecting neurons also showed increases in excitability and CC input strength that was offset with enhanced inhibition. S2 and ACC-projecting neurons showed no changes in excitability or CC input. These results demonstrate that subgroups of output neurons undergo dramatic and specific plasticity after peripheral injury. The changes in S1BC-projecting neurons likely underlie enhanced reciprocal connectivity of S1BC after unilateral deprivation consistent with the model that interhemispheric takeover supports intact whisker processing.SIGNIFICANCE STATEMENTAmputation, peripheral injury, and stroke patients experience widespread alterations in neural activity after sensory loss. A hallmark of this reorganization is the recruitment of deprived cortical space which likely aids processing and thus enhances performance on intact sensory systems. Conversely, this recruitment of deprived cortical space has been hypothesized to underlie phenotypes like phantom limb pain and hinder recovery. A mouse model of unilateral denervation detected remarkable specificity in alterations in the somatomotor circuit. These changes underlie increased reciprocal connectivity between intact and deprived cortical hemispheres. This increased connectivity may help explain the enhanced intact sensory processing detected in humans.

Neurodegeneration driven by aberrant tau is a key feature of many dementias. Pathological stages of tauopathy are characterised by reduced synapse density and altered synapse function. Furthermore, changes in synaptic plasticity have been documented in the early stages of tauopathy suggesting that they may be a driver of later pathology. However, it remains unclear if synapse plasticity is specifically linked to the degeneration of neurons. This is partly because, in progressive dementias, pathology can vary widely from cell-to-cell along the prolonged disease time-course. To overcome this variability, we have taken a longitudinal experimental approach to track individual neurons through the progression of neurodegenerative tauopathy. Using repeated in vivo 2-photon imaging in rTg4510 transgenic mice, we have measured structural plasticity of presynaptic terminaux boutons and postsynaptic spines on individual axons and dendrites over long periods of time. By following individual neurons, we have measured synapse density across the neuronal population and tracked changes in synapse turnover in each neuron. We found that tauopathy drives a reduction in density of both presynaptic and postsynaptic structures and that this is partially driven by degeneration of individual axons and dendrites that are spread widely across the disease time-course. Both synaptic loss and neuronal degeneration was ameliorated by reduction in expression of the aberrant P301L transgene, but only if that reduction was initiated early in disease progression. Notably, neurite degeneration was preceded by alterations in synapse turnover that contrasted in axons and dendrites. In dendrites destined to die, there was a dramatic loss of spines in the week immediately before degeneration. In contrast, axonal degeneration was preceded by a progressive attenuation of presynaptic turnover that started many weeks before axon disappearance. Therefore, changes in synapse plasticity are harbingers of degeneration of individual neurites that occur at differing stages of tau-driven neurodegenerative disease, suggesting a cell or neurite autonomous process. Furthermore, the links between synapse plasticity and degeneration are distinct in axonal and dendritic compartments.Key findingsTauopathy driven by tau P301L in rTg4510 mice causes a progressive decrease in density of presynaptic terminaux boutons and postsynaptic dendritic spines in cortical excitatory neurons.Longitudinal imaging of individual axons and dendrites shows that there is a huge diversity of effects at varying times in different cells.Decreases in overall synapse density are driven partly, but not exclusively, by degeneration of dendrites and axons that are distributed widely across the time-course of disease.Suppression of pathological P301L tau expression can ameliorate accumulation of tau pathology, synapse loss and neurodegeneration, but only if administered early in disease progression.Neurite degeneration is preceded by aberrant structural synaptic plasticity in a cell-specific way that is markedly different in dendrites and axons.Degeneration of dendrites is immediately preceded by dramatic loss of dendritic spines.Axonal loss is characterised by a progressive attenuation of presynaptic bouton plasticity that starts months before degeneration.

Significance The corpus callosum is a large fiber bundle which connects contralateral brain regions. After unilateral perturbations such as stroke or amputation, interhemispheric connectivity is altered and often leads to bilateral somatomotor cortical hyperactivity in patients with poor recovery. This study reports that callosal targeting of deprived layer 5 neurons is maximally potentiated in mouse primary somatosensory barrel cortex after unilateral whisker denervation. These neurons also experience an increase in excitability and spontaneous excitatory amplitudes. These results should be relevant to the cortical responses observed in human patients after unilateral nerve transection, amputation, or stroke., Central or peripheral injury causes reorganization of the brain’s connections and functions. A striking change observed after unilateral stroke or amputation is a recruitment of bilateral cortical responses to sensation or movement of the unaffected peripheral area. The mechanisms underlying this phenomenon are described in a mouse model of unilateral whisker deprivation. Stimulation of intact whiskers yields a bilateral blood-oxygen-level−dependent fMRI response in somatosensory barrel cortex. Whole-cell electrophysiology demonstrated that the intact barrel cortex selectively strengthens callosal synapses to layer 5 neurons in the deprived cortex. These synapses have larger AMPA receptor- and NMDA receptor-mediated events. These factors contribute to a maximally potentiated callosal synapse. This potentiation occludes long-term potentiation, which could be rescued, to some extent, with prior long-term depression induction. Excitability and excitation/inhibition balance were altered in a manner consistent with cell-specific callosal changes and support a shift in the overall state of the cortex. This is a demonstration of a cell-specific, synaptic mechanism underlying interhemispheric cortical reorganization.

Pharmacological manipulation of specific neural circuits to optimize therapeutic index is an unrealized goal in neurology and psychiatry. AMPA receptors are important for excitatory synaptic transmission, and their antagonists are antiepileptic. Although efficacious, AMPA-receptor antagonists, including perampanel (Fycompa), the only approved antagonist for epilepsy, induce dizziness and motor impairment. We hypothesized that blockade of forebrain AMPA receptors without blocking cerebellar AMPA receptors would be antiepileptic and devoid of motor impairment. Taking advantage of an AMPA receptor auxiliary protein, TARP γ-8, which is selectively expressed in the forebrain and modulates the pharmacological properties of AMPA receptors, we discovered that LY3130481 selectively antagonized recombinant and native AMPA receptors containing γ-8, but not γ-2 (cerebellum) or other TARP members. Two amino acid residues unique to γ-8 determined this selectivity. We also observed antagonism of AMPA receptors expressed in hippocampal, but not cerebellar, tissue from an patient with epilepsy. Corresponding to this selective activity, LY3130481 prevented multiple seizure types in rats and mice and without motor side effects. These findings demonstrate the first rationally discovered molecule targeting specific neural circuitries for therapeutic advantage.

Sensory hypersensitivity is a common and debilitating feature of neurodevelopmental disorders such as Fragile X Syndrome (FXS). However, how developmental changes in neuronal function ultimately culminate in the network dysfunction that underlies sensory hypersensitivities is not known. To address this, we studied the layer 4 barrel cortex circuit inFmr1knockout mice, a critical sensory processing circuit in this mouse model of FXS. By systematically studying cellular and synaptic properties of layer 4 neurons and combining with cellular and network simulations, we explored how the array of phenotypes inFmr1knockout produce circuit pathology during development that result in sensory processing dysfunction. We show that many of the cellular and synaptic pathologies inFmr1knockout mice are antagonistic, mitigating circuit dysfunction, and hence can be regarded as compensatory to the primary pathology. Despite this compensation, the layer 4 network in theFmr1knockout exhibits significant alterations in spike output in response to ascending thalamocortical input that we show results in impaired sensory encoding. We suggest that it is this developmental loss of layer 4 sensory encoding precision that drives subsequent developmental alterations in layer 4 – layer 2/3 connectivity and plasticity observed in theFmr1knockout, and is a critical process producing sensory hypersensitivity.

Summary Synapse loss is a key feature of dementia, but it is unclear whether synaptic dysfunction precedes degenerative phases of the disease. Here, we show that even before any decrease in synapse density, there is abnormal turnover of cortical axonal boutons and dendritic spines in a mouse model of tauopathy-associated dementia. Strikingly, tauopathy drives a mismatch in synapse turnover; postsynaptic spines turn over more rapidly, whereas presynaptic boutons are stabilized. This imbalance between pre- and post-synaptic stability coincides with reduced synaptically driven neuronal activity in pre-degenerative stages of the disease., Graphical Abstract, Highlights • Density of cortical axonal boutons and dendritic spines is reduced early in tauopathy • Abnormalities in synaptic stability and size exist before decreases in synapse density • Turnover of dendritic spines is elevated, whereas presynaptic boutons are stabilized • Neuronal activity is reduced at stages associated with mismatched synaptic turnover, Using in vivo two-photon imaging in the rTg4510 tauopathy mouse model, Jackson et al. find that synapse stability is altered during the pre-degenerative stages of tauopathy. Mismatched abnormalities in pre- and post-synaptic turnover coincide with disrupted neuronal activity.

Traditionally, neurotransmitters are associated with a fast, or phasic, type of action on neurons in the central nervous system (CNS). However, accumulating evidence indicates that γ-aminobutyric acid (GABA) and glutamate can also have a continual, or tonic, influence on these cells. Here, in voltage- and current-clamp recordings in rat brain slices, we identify three types of tonically active receptors in a single CNS structure, the thalamic reticular nucleus (TRN). Thus, TRN contains constitutively active GABAA receptors (GABAA Rs), which are located on TRN neurons and generate a persistent outward Cl(-) current. When TRN neurons are depolarized, blockade of this current increases their action potential output in response to current injection. Furthermore, TRN contains tonically active GluN2B-containing N-methyl-D-aspartate receptors (NMDARs). These are located on reticuloreticular GABAergic terminals in TRN and generate a persistent facilitation of vesicular GABA release from these terminals. In addition, TRN contains tonically active metabotropic glutamate type 2 receptors (mGlu2Rs). These are located on glutamatergic cortical terminals in TRN and generate a persistent reduction of vesicular glutamate release from these terminals. Although tonically active GABAA Rs, NMDARs and mGlu2Rs operate through different mechanisms, we propose that the continual and combined activity of these three receptor types ultimately serves to hyperpolarize TRN neurons, which will differentially affect the output of these cells depending upon the current state of their membrane potential. Thus, when TRN cells are relatively depolarized, their firing in single-spike tonic mode will be reduced, whereas when these cells are relatively hyperpolarized, their ability to fire in multispike burst mode will be facilitated.

Feedforward GABAergic inhibition sets the dendritic integration window, thereby controlling timing and output in cortical circuits. However, the manner in which feedforward inhibitory circuits emerge is unclear, despite this being a critical step for neocortical development and function. We found that sensory experience drove plasticity of the feedforward inhibitory circuit in mouse layer 4 somatosensory barrel cortex in the second postnatal week via two distinct mechanisms. First, sensory experience selectively strengthened thalamocortical-to-feedforward interneuron inputs via a presynaptic mechanism but did not regulate other inhibitory circuit components. Second, experience drove a postsynaptic mechanism in which a downregulation of a prominent thalamocortical NMDA excitatory postsynaptic potential in stellate cells regulated the final expression of functional feedforward inhibitory input. Thus, experience is required for specific, coordinated changes at thalamocortical synapses onto both inhibitory and excitatory neurons, producing a circuit plasticity that results in maturation of functional feedforward inhibition in layer 4.

Summary Background Ionotropic glutamate receptors (iGluRs) are glutamate-gated ion channels that mediate excitatory neurotransmission in the central nervous system. Based on both molecular and pharmacological criteria, iGluRs have been divided into two major classes, the non-NMDA class, which includes both AMPA and kainate subtypes of receptors, and the NMDA class. One evolutionarily conserved feature of iGluRs is their desensitization in the continued presence of glutamate. Thus, when in a desensitized state, iGluRs can be bound to glutamate, yet the channel remains closed. However, the relevance of desensitization to nervous system function has remained enigmatic. Results Here, we report the identification and characterization of a novel polypeptide (con-ikot-ikot) from the venom of a predatory marine snail Conus striatus that specifically disrupts the desensitization of AMPA receptors (AMPARs). The stoichiometry of con-ikot-ikot appears reminiscent of the proposed subunit organization of AMPARs, i.e., a dimer of dimers, suggesting that it acts as a molecular four-legged clamp that holds the AMPAR channel open. Application of con-ikot-ikot to hippocampal slices caused a large and rapid increase in resting AMPAR-mediated current leading to neuronal death. Conclusions Our findings provide insight into the mechanisms that regulate receptor desensitization and demonstrate that in the arms race between prey and predators, evolution has selected for a toxin that blocks AMPAR desensitization, thus revealing the fundamental importance of desensitization for regulating neural function.

Transmission through the thalamus activates circuits involving the GABAergic neurons of the thalamic reticular nucleus (TRN). TRN cells receive excitatory inputs from thalamocortical and corticothalamic cells and send inhibitory projections to thalamocortical cells. The inhibitory output of TRN neurons largely depends on the level of excitatory drive to these cells but may also be partly under the control of mechanisms intrinsic to the TRN. We examined two such possible mechanisms, short-term plasticity at glutamatergic synapses in the TRN and intra-TRN inhibition. In rat brain slices, responses of TRN neurons to brief trains of stimuli applied to glutamatergic inputs were recorded in voltage- or current-clamp mode. In voltage clamp, TRN cells showed no change in alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor-mediated excitatory postsynaptic current amplitudes to stimulation at non-gamma frequencies (30 Hz), simulating background activity, but exhibited short-term depression in these amplitudes to stimulation at gamma frequencies (30 Hz), simulating sensory transmission. In current clamp, TRN cells increased their spike outputs in burst and tonic firing modes to increasing stimulus-train frequencies. These increases in spike output were most likely due to temporal summation of excitatory postsynaptic potentials. However, the frequency-dependent increase in tonic firing was attenuated at gamma stimulus frequencies, indicating that the synaptic depression selectively observed in this frequency range acts to suppress TRN cell output. In contrast, intra-TRN inhibition reduced spike output selectively at non-gamma stimulus frequencies. Thus, our data indicate that two intrinsic mechanisms play a role in controlling the tonic spike output of TRN neurons and these mechanisms are differentially related to two physiologically meaningful stimulus frequency ranges.

Long-term synaptic plasticity is an important mechanism underlying the development of cortical circuits in a number of brain regions. In barrel cortex NMDA receptor (NMDAR)-dependent long-term potentiation (LTP) and long-term depression (LTD) play a critical role in the development and experience-dependent plasticity of the topographical map of the rodent whiskers. However, the mechanisms underlying the induction and expression of these forms of plasticity are poorly characterised. Here we investigate the role of PKC in the regulation of synaptic strength in neonatal barrel cortex using patch-clamp recordings in brain slices. We demonstrate that PKC activity tonically maintains AMPA receptor-mediated transmission at thalamocortical synapses, and that basal transmission can be potentiated by PKC activation using postsynaptic infusion of phorbol ester. Furthermore, we show that induction of NMDAR-dependent LTP requires PKC activity. These findings demonstrate that PKC is required for the regulation of transmission at thalamocortical synapses, the major ascending sensory input to barrel cortex. Thalamocortical inputs in barrel cortex only express LTP during the first postnatal week during a critical period for experience-dependent plasticity in layer IV. Therefore, the requirement for PKC in LTP suggests an important role for this kinase in the development of the barrel cortex sensory map.

Developing neuronal networks acquire the ability to precisely time events, a key feature required for information processing. In the barrel cortex, encoding of information requires a high-precision temporal code with a resolution of ∼5 ms; however, it is not known what process drives the maturation in timing precision. Here, we report that long-term potentiation (LTP) at thalamocortical synapses in the neonatal layer IV barrel cortex produces a dramatic improvement in the timing of neuronal output and synaptic input. LTP strongly reduces the latency and variability of synaptically evoked action potentials, improving the fidelity of timing to within that predicted to be required for adult sensory processing. Such changes in timing also occur during development in the neonate. LTP also reduces the summation of EPSPs shortening the window for coincidence detection for synaptic input. In contrast to these reliable effects, LTP produced only a modest and variable change in synaptic efficacy. Thus, our findings suggest that the primary role of this form of neonatal LTP is for the acquisition of timing precision and the refinement of coincidence detection, rather than an increase in synaptic strength. Therefore, neonatal thalamocortical LTP may be a critical prerequisite for the maturation of information processing in the barrel cortex.

Neonatal hippocampus exhibits distinct patterns of network activity that are dependent on the interaction between inhibitory and excitatory transmission. Kainate receptors are ideally positioned to regulate this activity by virtue of their ability to regulate presynaptic function in GABAergic interneurones. Indeed, kainate receptors are highly expressed in neonatal hippocampal interneurones, yet the role and mechanisms by which they might regulate neonatal circuitry are unexplored. To address this we investigated the kainate receptor-dependent regulation of GABAergic transmission onto neonatal CA1 pyramidal neurones. Kainate receptor activation produced two distinct opposing effects, a very large increase in the frequency of spontaneous IPSCs, and a robust depression of evoked GABAergic transmission. The up-regulation of spontaneous transmission was due to activation of somatodendritic and axonal receptors while the depression of evoked transmission could be fully accounted for by a direct regulation of GABA release by kainate receptors located at the terminals. None of the effects of kainate receptor agonists were sensitive to GABAB receptor antagonists, nor was there any postsynaptic kainate receptor-dependent effects observed in CA1 pyramidal cells that could account for our findings. Our data demonstrate that kainate receptors profoundly regulate neonatal CA1 GABAergic circuitry by two distinct opposing mechanisms, and indicate that these two effects are mediated by functionally distinct populations of receptors. Thus kainate receptors are strategically located to play a critical role in shaping early hippocampal network activity and by virtue of this have a key role in hippocampal development.

The main input to primary sensory cortex is via thalamocortical (TC) axons that form the greatest number of synapses in layer 4, but also synapse onto neurons in layer 6. The development of the TC input to layer 4 has been widely studied, but less is known about the development of the layer 6 input. Here, we show that, in neonates, the input to layer 6 is as strong as that to layer 4. Throughout the first postnatal week, there is an experience-dependent strengthening specific to layer 4, which correlates with the ability of synapses in layer 4, but not in layer 6, to undergo long-term potentiation (LTP). This strengthening consists of an increase in axon branching and the divergence of connectivity in layer 4 without a change in the strength of individual connections. We propose that experience-driven LTP stabilizes transient TC synapses in layer 4 to increase strength and divergence specifically in layer 4 over layer 6.

NMDAR-dependent long-term depression involves the activation of protein phosphatase 1 (PP1) and 2B (calcineurin) and the subsequent dephosphorylation of synaptic proteins. In this issue of Neuron, Morishita et al. (2001) provide evidence that precise targeting of PP1 to synaptic substrates is critical for the expression of LTD.

High affinity glutamate transport plays an important role in maintaining a low extracellular glutamate concentration in the CNS. Excitotoxicity due to a loss of glutamate transporter function has been implicated in disease processes such as stroke and amyotrophic lateral sclerosis (ALS). We studied the effects of glutamate transport inhibitors on thalamocortical synapses at developing (postnatal day 3–8) layer IV neurons in the barrel cortex using the thalamocortical slice preparation and whole-cell recordings. Inhibition of glutamate transport by D,L-threo-β-hydroxyaspartate (THA), a combination of THA and dihydrokainate (DHK), or by L-trans-pyrrolidine-2,4-dicarboxylate (tPDC), caused a reversible blockade of AMPA and kainate receptor-mediated dual component excitatory postsynaptic currents (AMPA/KA EPSCs). This effect was not blocked by cyclothiazide (CTZ) indicating that is was not due to desensitisation of AMPARs. Under conditions in which NMDA receptors were unblocked the transport inhibitors caused the massive activation of NMDA receptors leading to the rapid loss of recordings. Previous studies using these transport inhibitors on brain slices from older animals reported no or only modest effects on synaptic transmission. Therefore the data in the present study suggest that neurons in the developing neocortex are particularly sensitive to glutamate transporter function. Furthermore the effects of transport inhibition are dependent upon whether neurons are sufficiently depolarised to relieve the voltage-dependent block of NMDA receptors.

Synapses are anatomical specializations at the interface of two neurons. They have fascinated researchers ever since Sherrington coined the term ‘synapse’ in 1879. Studies of synapses have provided amazing insight into their function and were boosted by the discovery, now more than 30 years ago, that synapses can undergo long-term activity-dependent plasticity. Over the last two decades much effort has been devoted to the study of long-term synaptic plasticity as a candidate mechanism underlying learning and memory. More recently dysfunctional synaptic plasticity has been implicated in several brain diseases, such as depression, addiction, dementia and anxiety disorders. Here we report on an international meeting that took place in Geneva in July 2008 that brought together scientists who study synapses and disease mechanisms. Brain diseases constitute a major burden to society. In Europe, alone, it is estimated that direct and indirect costs of brain diseases represent more than 350 billion euros per year. When broken down into individual diseases the most costly are depression (105 billion euros, a sum equal to the cost of all cardiovascular diseases), addiction (56 billion even without including nicotine abuse), dementia (54 billion) and anxiety disorders (40 billion), followed by autism and schizophrenia. The Synaptic Basis of Disease meeting, which was organized as a satellite meeting to the July 2008 FENS meeting in Geneva (Fig. 1), was convened to bring together leading researchers working on basic synaptic mechanisms with those working on neurological disorders. Indeed, many labs at the meeting have programs working on both aspects. The main focus of the meeting was to generate discussion amongst some of the leading experimentalists working on synaptic and disease mechanisms. Figure 1 The poster for the symposium

Transmission of sensory information through the dorsal thalamus involves two types of modality-related nuclei, first order and higher order, between which there are thought to be no intrathalamic interactions. We now show that within the somatosensory thalamus, cells in one nucleus, the ventrobasal complex, can influence activity in another nucleus, the medial division of the posterior complex. Stimulation of ventrobasal complex cells evoked inhibitory postsynaptic currents in cells of the medial division of the posterior complex. These currents exhibited the reversal potential and pharmacology of a GABAA receptor-mediated chloride conductance, indicating that they result from the activation of a disynaptic pathway involving the GABAergic cells of the thalamic reticular nucleus. These findings provide the first direct evidence for intrathalamic interactions between dorsal thalamic nuclei.

The subcellular locations of synapses on pyramidal neurons strongly influences dendritic integration and synaptic plasticity. Despite this, there is little quantitative data on spatial distributions of specific types of synaptic input. Here we use array tomography (AT), a high-resolution optical microscopy method, to examine thalamocortical (TC) input onto layer 5 pyramidal neurons. We first verified the ability of AT to identify synapses using parallel electron microscopic analysis of TC synapses in layer 4. We then use large-scale array tomography (LSAT) to measure TC synapse distribution on L5 pyramidal neurons in a 1.00 × 0.83 × 0.21 mm(3) volume of mouse somatosensory cortex. We found that TC synapses primarily target basal dendrites in layer 5, but also make a considerable input to proximal apical dendrites in L4, consistent with previous work. Our analysis further suggests that TC inputs are biased toward certain branches and, within branches, synapses show significant clustering with an excess of TC synapse nearest neighbors within 5-15 μm compared to a random distribution. Thus, we show that AT is a sensitive and quantitative method to map specific types of synaptic input on the dendrites of entire neurons. We anticipate that this technique will be of wide utility for mapping functionally-relevant anatomical connectivity in neural circuits.

Genetic causes, or predisposition, are increasingly accepted to be part of the ethiopathogenesis of many neuropsychiatric diseases. While genes can be studied in any type of cells, their physiological function in human brain cells is difficult to evaluate, particularly in living subjects. As a first step towards the characterisation of human inducible pluripotent stem cell (iPSC)-derived neurons from autism spectrum disorder (ASD) patients, we used gene expression and functional studies to define the regional identity of the typical forebrain differentiation, demonstrate expression patterns of genes of interest in ASD and understand the properties of ‘control’ iPSC-derived neurons (iCell-Neurons™), with a focus on receptors and ion channels that play a central role in synaptic physio-pathology. The gene expression profile of the iCell-Neurons™ closely resembled that observed in neonatal prefrontal cortex tissues. Functional studies, performed mainly using calcium flux assays, demonstrated the presence of ionotropic glutamate (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid and N-methyl-d-aspartate) and gamma-aminobutyric acid type A receptors. Voltage-gated sodium and calcium channels were also identified using similar techniques. Overall, the results reported here suggest that iCell-Neurons™ are a good cellular model of a relatively immature forebrain human neuron population that can be used both as a control in comparison to patients cells, and as host cells in which mutations, insertions and deletions can be used in order to study the molecular mechanisms of ASD and other neurological disorders in an isogenic cellular background.

Despite extensive investigation, it remains unclear whether presynaptic and/or postsynaptic modifications are primarily responsible for the expression of long-term potentiation (LTP) in the CA1 region of the hippocampus. Here we address this issue by using techniques that maximize the likelihood of stimulating a single axon and thereby presumably a single synapse before and after the induction of LTP. Several basic properties of synaptic transmission were examined including the probability of neurotransmitter release (Pr), the quantal size (q), and the so-called potency, which is defined as the average size of the synaptic response when release of transmitter does occur. LTP was routinely associated with an increase in potency, whereas increases in Pr alone were not observed. LTP was also reliably induced when baseline Pr was high, indicating that synapses with high Pr can express LTP. These results suggest that the mechanism for the expression of LTP involves an increase in q and is difficult to explain by an increase in Pr alone.

In cerebral cortex there is a developmental switch from NR2B- to NR2A-containing NMDA receptors (NMDARs) driven by activity and sensory experience. This subunit switch alters NMDAR function, influences synaptic plasticity, and its dysregulation is associated with neurological disorders. However, the mechanisms driving the subunit switch are not known. Here, we show in hippocampal CA1 pyramidal neurons that the NR2B to NR2A switch driven acutely by activity requires activation of NMDARs and mGluR5, involves PLC, Ca2+ release from IP3R-dependent stores, and PKC activity. In mGluR5 knockout mice the developmental NR2B-NR2A switch in CA1 is deficient. Moreover, in visual cortex of mGluR5 knockout mice, the NR2B-NR2A switch evoked in vivo by visual experience is absent. Thus, we establish that mGluR5 and NMDARs are required for the activity-dependent NR2B-NR2A switch and play a critical role in experience-dependent regulation of NMDAR subunit composition in vivo. In cerebral cortex there is a developmental switch from NR2B- to NR2A-containing NMDA receptors (NMDARs) driven by activity and sensory experience. This subunit switch alters NMDAR function, influences synaptic plasticity, and its dysregulation is associated with neurological disorders. However, the mechanisms driving the subunit switch are not known. Here, we show in hippocampal CA1 pyramidal neurons that the NR2B to NR2A switch driven acutely by activity requires activation of NMDARs and mGluR5, involves PLC, Ca2+ release from IP3R-dependent stores, and PKC activity. In mGluR5 knockout mice the developmental NR2B-NR2A switch in CA1 is deficient. Moreover, in visual cortex of mGluR5 knockout mice, the NR2B-NR2A switch evoked in vivo by visual experience is absent. Thus, we establish that mGluR5 and NMDARs are required for the activity-dependent NR2B-NR2A switch and play a critical role in experience-dependent regulation of NMDAR subunit composition in vivo.

Presynaptic kainate receptors play an important role in synaptic transmission and short-term plasticity to profoundly regulate network activity in many parts of the mammalian brain. In primary sensory neocortex, where short-term synaptic plasticity is important for receptive field structure and information processing, kainate receptors are highly expressed and regulate thalamocortical inputs, particularly during development. However, the mechanisms of the kainate receptor-dependent presynaptic regulation of thalamocortical transmission are unclear. We therefore investigated this issue using electrophysiology in neonatal thalamocortical slices of barrel cortex combined with pharmacology and biochemical analyses. We show that presynaptic kainate receptors can both facilitate or depress synaptic transmission depending on the extent of their activation. This bi-directional regulation is mediated in part by kainate receptors that directly influence thalamocortical axonal excitability, but also likely involves receptors acting at thalamocortical terminals to regulate transmitter release. The efficacy of kainate in regulating thalamocortical transmission is low compared to that reported for other inputs. Consistent with this low efficacy, our biochemical analyses indicate that the presynaptic kainate receptors regulating neonatal thalamocortical inputs likely lack the high kainate affinity GluK4 and 5 subunits. Thus thalamocortical transmission can be bi-directionally regulated by low affinity kainate receptors through two mechanisms. Such presynaptic regulation provides a potentially powerful mechanism to influence sensory processing during development of barrel cortex.

Summary Over-inhibition is thought to be one of the underlying causes of the cognitive deficits in Ts65Dn mice, the most widely used model of Down syndrome (DS). Here we demonstrate a direct link between gene triplication and defects in neuron production during embryonic development. These neurogenesis defects lead to an imbalance between excitatory and inhibitory neurons and to increased inhibitory drive in the Ts65Dn forebrain. We discovered that Olig1 and Olig2, two genes triplicated in DS and Ts65Dn, are over-expressed in the Ts65Dn forebrain. To test the hypothesis that Olig triplication is causative for the neurological phenotype, we used a genetic approach to normalize the dosage of these two genes and thereby rescued the inhibitory neuron phenotype in the Ts65Dn brain. These data identify seminal alterations during brain development and demonstrate a mechanistic relationship between triplicated genes and these brain abnormalities in the Ts65Dn mouse.

The mammalian neocortex is a six-layered structure containing a large number of different cell types arranged in a stereotypical pattern. This complex structure additionally contains a vast array of neuronal circuits, both local and long range. The basic repeated unit of the neocortex is thought to be the ‘cortical column’, a vertical association of cells and synaptic connections across the six laminae that form a functional processing unit. Each column alone contains thousands of neurons and millions of synapses. Thus, the mature neocortex is highly complex in structure, connectivity and function, and as such, is poorly understood. An additional challenge to neurobiologists is to understand how neocortex develops to produce its stereotyped yet complex organization. Neocortical development is a major focus in neurobiology, not only because it is of interest from a purely developmental standpoint, but also because understanding how neocortex develops provides important insight into mature neocortical organization and function. Mammalian neocortical development involves many mechanisms and stages. Early embryonic development involves cell specification and cell migration, which leads to an ‘inside-out’ development of the six neocortical layers. The deepest layer, layer 6, appears first and is followed sequentially by development of the progressively more superficial layers. During this time transcription factors and axon guidance mechanisms play a central role in the patterning of neocortex. This initial patterning is largely completed by birth. Postnatal development during the first few weeks of life is characterized by additional sets of processes. Neurons elaborate their dendritic and axonal arbors, which can then be followed by axonal pruning. This facilitates rapid changes in the connectivity of neocortical circuits as neuronal processes intermix. Neurons also exhibit developmental changes in their ion channel complement producing changes in excitability and firing properties. Overlaid on these changes in connectivity and intrinsic excitability are dramatic alterations in synaptic function. Ionotropic neurotransmitter receptors play a prominent role in this process as there are developmental changes in the number, distribution and subtype of GABAA receptors, and AMPA, NMDA and kainate subtype glutamate receptors. In addition, there is a change in the reversal potential for chloride ions such that GABAA receptor activation is depolarizing during the first postnatal week, but becomes hyperpolarizing and inhibitory by the second postnatal week. These changes in receptor function produce profound alterations in synaptic transmission during cortical development. The postnatal development of neocortical circuits is driven in large part by changes in synaptic function produced by long-term synaptic plasticity. Neuronal activity causes long-lasting changes in synaptic strength at excitatory (glutamatergic) and inhibitory (GABAergic) synapses. Such plasticity not only changes synaptic strength but also produces changes in receptor subtype expression, neuronal intrinsic excitability and structural elements such as dendritic spines. In addition, short-term dynamics at synaptic transmission are also regulated during development, altering the information transfer properties of specific circuits in neocortex. These postnatal synaptic changes combine with alterations in connectivity to produce functional cortical microcircuits that are the building blocks of large scale circuits at the level of columns and cortical areas. The current focus on the mechanisms of neocortical development has led to major advances in our understanding of many of the processes described above. Such advances have been driven by the recent proliferation of new technologies that allow an unprecedented insight into neocortical development across the many stages. To highlight the recent advances in this fast changing field, The Journal of Physiology convened a symposium organized by John Isaac and Dirk Feldmeyer entitled: ‘Mechanisms of Neocortical Development’ as an official satellite to the Society for Neuroscience Annual meeting held in Washington, DC in November 2008. The symposium brought together leading figures from the field spanning embryonic development, structural and functional development and the role of plasticity. The presentations can be grouped into four themes. The first theme was the embryonic development of neocortex. Zoltan Molnar (University of Oxford, UK) presented work on mechanisms involved in the production of the earliest cortical circuits in late embryonic and early postnatal development of neocortex (see Pinon et al. 2009). In the second theme, the development and role of GABAergic signalling in neocortex was discussed. Arnold Kriegstein (University of California, San Francisco, CA, USA) described studies in which the role of ‘depolarizing GABA’ in development of synaptic function and structure was investigated (Wang & Kriegstein, 2009). Josh Huang (Cold Spring Harbor Laboratory, NY, USA) described molecular mechanisms regulating the number and distribution of GABAergic synapses in cortex focusing on a role for GAD67 (Huang, 2009). John Isaac described synaptic mechanisms underlying the development and fine tuning of feed-forward GABAergic circuits in layer 4 barrel cortex. A third major theme was mechanisms contributing to development of excitatory circuits. Dirk Feldmeyer (Research Centre Julich and RWTH Aachen University, Germany) described the development of excitatory connections within layers 4 and 5A of the barrel cortex (Feldmeyer & Radnikow, 2009). Ingrid Bureau (University of France) presented her work on the development of layer 4–layer 2/3 excitatory connections in barrel cortex and the impact of fragile X mental retardation protein (FMRP) knockout on this process (Bureau, 2009). In a final theme the roles of activity-dependent plasticity was considered. Gina Turrigiano (Brandeis University, Boston, MA, USA) described work showing a prominent role of plasticity of neuronal excitability as a mechanism underlying experience-dependent plasticity in primary visual cortex. Mark Bear (Massachusetts Institute of Technology, Cambridge, MA, USA) discussed the role of long-term synaptic plasticity in mediating ocular dominance plasticity in visual cortex. This issue of The Journal contains a series of reviews written by Symposium speakers that provide an in-depth discussion of the major themes covered in the Symposium concerning the development of mammalian neocortex. These reviews, written by the experts in the field, highlight some of the most significant current thinking about neocortical development and thus provide an excellent resource for readers of The Journal to get the ‘scoop’ on what's ‘hot’ in neocortical development.

Light exposure changes the subunit composition and kinetics of NMDA receptors in the developing visual cortex. Quinlan and colleagues suggest that this may be due to rapid synaptic insertion of receptors containing newly synthesized NR2A subunits.

Many brain processes and the expression of many neuronal proteins are developmentally regulated; therefore, studying cellular and molecular processes in tissue from very young animals is vital to understanding the development of the brain. Working on tissue from very young animals (first postnatal week) presents a number of specific problems. This unit describes techniques to overcome these problems to produce in vitro slices from neonatal rodents and to reliably achieve high-quality patch-clamp recordings from neurons in neonatal brain slices.

Understanding basic neuronal mechanisms hold the hope for future treatment of brain disease. The 1st international conference on synapse, memory, drug addiction and pain was held in beautiful downtown Toronto, Canada on August 21-23, 2006. Unlike other traditional conferences, this new meeting focused on three major aims: (1) to promote new and cutting edge research in neuroscience; (2) to encourage international information exchange and scientific collaborations; and (3) to provide a platform for active scientists to discuss new findings. Up to 64 investigators presented their recent discoveries, from basic synaptic mechanisms to genes related to human brain disease. This meeting was in part sponsored by Molecular Pain, together with University of Toronto (Faculty of Medicine, Department of Physiology as well as Center for the Study of Pain). Our goal for this meeting is to promote future active scientific collaborations and improve human health through fundamental basic neuroscience researches. The second international meeting on Neurons and Brain Disease will be held in Toronto (August 29-31, 2007). © 2006 Bi et al; licensee BioMed Central Ltd.

Postnatal glutamatergic principal neuron synapses are typically presumed to express only calcium-impermeable (CI), GluR2-containing AMPARs under physiological conditions. Here, however, we demonstrate that long-term potentiation (LTP) in CA1 hippocampal pyramidal neurons causes rapid incorporation of GluR2-lacking calcium-permeable (CP)-AMPARs: CP-AMPARs are present transiently, being replaced by GluR2-containing AMPARs approximately 25 min after LTP induction. Thus, CP-AMPARs are physiologically expressed at CA1 pyramidal cell synapses during LTP, and may be required for LTP consolidation.

During the first week of life, there is a shift from kainate to AMPA receptor-mediated thalamocortical transmission in layer IV barrel cortex. However, the mechanisms underlying this change and the differential properties of AMPA and kainate receptor-mediated transmission remain essentially unexplored. To investigate this, we studied the quantal properties of AMPA and kainate receptor-mediated transmission using strontium-evoked miniature EPSCs. AMPA and kainate receptor-mediated transmission exhibited very different quantal properties but were never coactivated by a single quantum of transmitter, indicating complete segregation to different synapses within the thalamocortical input. Nonstationary fluctuation analysis showed that synaptic AMPA receptors exhibited a range of single-channel conductance (γ) and a strong negative correlation between γ and functional channel number, indicating that these two parameters are reciprocally regulated at thalamocortical synapses. We obtained the first estimate of γ for synaptic kainate receptors (

Kainate receptors (KARs) are highly expressed throughout the neonatal brain, but their function during development is unclear. Here, we show that the maturation of the hippocampus is associated with a switch in the functional role of presynaptic KARs. In a developmental period restricted to the first postnatal week, endogenousl-glutamate tonically activates KARs at CA3 glutamatergic synapses to regulate release in an action potential-independent manner. At synapses onto pyramidal cells, KARs inhibit glutamate release via a G-protein and PKC-dependent mechanism. In contrast, at glutamatergic terminals onto CA3 interneurons, presynaptic KARs can facilitate release in a G-protein-independent mechanism. In both cell types, however, KAR activation strongly upregulates inhibitory transmission. We show that, through the interplay of these novel diverse mechanisms, KARs strongly regulate the characteristic synchronous network activity observed in the neonatal hippocampus. By virtue of this, KARs are likely to play a central role in the development of hippocampal synaptic circuits.

Transmission through the dorsal thalamus involves nuclei that convey different aspects of sensory or motor information. Cells in the dorsal thalamus are strongly inhibited by the GABAergic cells of the thalamic reticular nucleus (TRN). Here we show that stimulation of cells in specific dorsal thalamic nuclei evokes robust IPSCs or IPSPs in other specific dorsal thalamic nuclei and vice versa. These IPSCs are GABA A receptor-mediated currents and are consistent with the activation of disynaptic intrathalamic pathways mediated by TRN. Thus, cells engaged in sensory analyses in the ventrobasal complex or the medial division of the posterior complex can interact with cells responsive to sensory events in the caudal intralaminar nuclei, whereas cells engaged in motor analyses in the ventrolateral nucleus can interact with cells responsive to motor events in the rostral intralaminar nuclei. Furthermore, sensory event-related cells in the caudal intralaminar nuclei can interact with motor event-related cells in the rostral intralaminar nuclei. In addition, single cells in one dorsal thalamic nucleus can receive convergent inhibitory inputs after stimulation of cells in two or more other dorsal thalamic nuclei, and TRN-mediated inhibitory inputs can momentarily switch off tonic firing of action potentials in dorsal thalamic cells. Our findings provide the first direct evidence for a rich network of intrathalamic pathways that allows modality-related and cross-modality inhibitory modulation between dorsal thalamic nuclei. Moreover, TRN-mediated switching between dorsal thalamic nuclei could provide a mechanism for the selection of competing transmissions of sensory and/or motor information through the dorsal thalamus.

Local recurrent excitatory circuits are ubiquitous in neocortex, yet little is known about their development or architecture. Here we introduce a quantitative technique for efficient single-cell resolution circuit mapping using 2-photon (2P) glutamate uncaging and analyze experience-dependent neonatal development of the layer 4 barrel cortex local excitatory circuit. We show that sensory experience specifically drives a 3-fold increase in connectivity at postnatal day (P) 9, producing a highly recurrent network. A profound dendritic spinogenesis occurs concurrent with the connectivity increase, but this is not experience dependent. However, in experience-deprived cortex, a much greater proportion of spines lack postsynaptic AMPA receptors (AMPARs) and synaptic connectivity via NMDA receptors (NMDARs) is the same as in normally developing cortex. Thus we describe a approach for quantitative circuit mapping and show that sensory experience sculpts an intrinsically developing template network, which is based on NMDAR-only synapses, by driving AMPARs into newly formed silent spines. Local recurrent excitatory circuits are ubiquitous in neocortex, yet little is known about their development or architecture. Here we introduce a quantitative technique for efficient single-cell resolution circuit mapping using 2-photon (2P) glutamate uncaging and analyze experience-dependent neonatal development of the layer 4 barrel cortex local excitatory circuit. We show that sensory experience specifically drives a 3-fold increase in connectivity at postnatal day (P) 9, producing a highly recurrent network. A profound dendritic spinogenesis occurs concurrent with the connectivity increase, but this is not experience dependent. However, in experience-deprived cortex, a much greater proportion of spines lack postsynaptic AMPA receptors (AMPARs) and synaptic connectivity via NMDA receptors (NMDARs) is the same as in normally developing cortex. Thus we describe a approach for quantitative circuit mapping and show that sensory experience sculpts an intrinsically developing template network, which is based on NMDAR-only synapses, by driving AMPARs into newly formed silent spines.

Previous studies have shown that pharmacological activation of presynaptic kainate receptors at glutamatergic synapses facilitates or depresses transmission in a dose-dependent manner. However, the only synaptically activated kainate autoreceptor described to date is facilitatory. Here, we describe a kainate autoreceptor that depresses synaptic transmission. This autoreceptor is present at developing thalamocortical synapses in the barrel cortex, specifically regulates transmission at frequencies corresponding to those observed in vivo during whisker activation, and is developmentally down regulated during the first postnatal week. This receptor may, therefore, limit the transfer of high-frequency activity to the developing cortex, the loss of which mechanism may be important for the maturation of sensory processing.