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Summary Segregation of retinal ganglion cell (RGC) axons by type and eye of origin is considered a hallmark of dorsal lateral geniculate nucleus (dLGN) structure. However, recent anatomical studies have shown that neurons in mouse dLGN receive input from multiple RGC types of both retinae. Whether convergent input leads to relevant functional interactions is unclear. We studied functional eye-specific retinogeniculate convergence using dual-color optogenetics in vitro. dLGN neurons were strongly dominated by input from one eye. Most neurons received detectable input from the non-dominant eye, but this input was weak, with a prominently reduced AMPAR:NMDAR ratio. Consistent with this, only a small fraction of thalamocortical neurons was binocular in vivo across visual stimuli and cortical projection layers. Anatomical overlap between RGC axons and dLGN neuron dendrites alone did not explain the strong bias toward monocularity. We conclude that functional eye-specific input selection and refinement limit convergent interactions in dLGN, favoring monocularity.

Eye-specific segregation of retinal ganglion cell (RGC) axons in the dorsal lateral geniculate nucleus (dLGN) is considered a hallmark of visual system development. However, a recent anatomical study showed that nearly half of the neurons in dLGN of adult mice still receive input from both retinae, but functional data about binocularity in mature dLGN is conflicting. Here, we found that a variable but small fraction of thalamocortical neurons is binocular in vivo. Using dual-channel optogenetics in vitro we correspondingly found that dLGN neurons are dominated by retinogeniculate input from one eye only, although most neurons also received small but detectable input from the non-dominant eye. Anatomical overlap between RGC axons and dLGN neuron dendrites did not explain this strong bias towards monocularity. Our data rather suggest that functional input selection and refinement, leaving the remaining non-dominant eye inputs in a juvenile-like state, underlies the prevalent monocularity of neurons in dLGN.

Summary Astrocytes are particularly promising candidates for reprogramming into neurons, as they maintain some of the original patterning information from their radial glial ancestors. However, to which extent the position of astrocytes influences the fate of reprogrammed neurons remains unknown. To elucidate this, we performed stab wound injury covering an entire neocortical column, including the gray matter (GM) and white matter (WM), and targeted local reactive astrocytes via injecting FLEx switch (Cre-On) adeno-associated viral (AAV) vectors into mGFAP-Cre mice. Single proneural factors were not sufficient for adequate reprogramming, although their combination with the nuclear receptor-related 1 protein (Nurr1) improved reprogramming efficiency. Nurr1 and Neurogenin 2 (Ngn2) resulted in high-efficiency reprogramming of targeted astrocytes into neurons that develop lamina-specific hallmarks, including the appropriate long-distance axonal projections. Surprisingly, in the WM, we did not observe any reprogrammed neurons, thereby unveiling a crucial role of region- and layer-specific differences in astrocyte reprogramming., Graphical Abstract, Highlights • AAV can be targeted to reactive astrocytes upon stab wound injury • Expression of Ngn2 and Nurr1 in these astrocytes induces pyramidal neurons • Induced pyramidal neurons acquire correct layer identity and axonal projections • Neurons cannot be induced in the white matter, Neurons dying after brain injury cannot be replaced. Mattugini, Bocchi, et al. show that local astrocytes can be converted into functional neurons acquiring appropriate layer identity and connectivity by expression of neurogenic factors in a mouse model of traumatic brain injury.

The majority of γ-aminobutyric acid (GABA)ergic interneurons have smooth dendrites with no or only few dendritic spines, but certain types of spiny GABAergic interneurons do actually contain substantial numbers of spines. The explanation for such spines has so far been purely structural: They increase the dendritic surface area and thus provide the opportunity to accommodate larger numbers of synapses. We reasoned that there may be specific functional properties for these spines and therefore, undertook to characterize interneuron spines functionally. We find a remarkable similarity to pyramidal cell spines: They receive excitatory synapses with calcium impermeable α-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptors, compartmentalize biochemical signals, and display activity-dependent morphological plasticity. Nevertheless, notable differences in spine density, neck length, and spine-dendrite coupling exist. Thus, dendritic spines on inhibitory interneurons have a number of important functional properties that go substantially beyond simply expanding the dendritic surface area. It therefore seems likely that spiny and aspiny interneurons may have very different roles in neural circuit function and plasticity.

The spatial organization of synaptic inputs on the dendritic tree of cortical neurons plays a major role for dendritic integration and neural computations, yet, remarkably little is known about it. We mapped the spatial organization of glutamatergic synapses between layer 5 pyramidal cells by combining optogenetics and 2-photon calcium imaging in mouse neocortical slices. To mathematically characterize the organization of inputs we developed an approach based on combinatorial analysis of the likelihoods of specific synapse arrangements. We found that the synapses of intralaminar inputs form clusters on the basal dendrites of layer 5 pyramidal cells. These clusters contain 4 to 14 synapses within ≤30 µm of dendrite. According to the spatiotemporal characteristics of synaptic summation, these numbers suggest that there will be non-linear dendritic integration of synaptic inputs during synchronous activation. DOI: http://dx.doi.org/10.7554/eLife.09222.001, eLife digest Neurons in the brain exchange information through points of contact called synapses. If electrical activity arriving at a number of synapses exceeds a certain threshold, it can trigger an electrical impulse, which is transmitted to the next neuron. Synapses generally connect with branch-like structures called dendrites on the receiving neuron. However, little is known about how synapses are arranged on dendrites. Gökçe et al. have now used a technique called optogenetics to work out the exact arrangement of a type of synapse on neurons in a part of the mouse brain that is devoted to vision. Optogenetics takes advantage of light-activated proteins that can trigger electrical activity. Gökçe et al. used mice that had been genetically engineered to produce these proteins in specific neurons, and then deliberately triggered electrical activity simply by shining light on these neurons. The experiments also used another technique called two-photon calcium imaging to monitor the activity of single synapses in response to the electrical activity triggered by optogenetics. Gökçe et al. found that these neurons have clusters of four to fourteen synapses within a space of 30 micrometers along a dendrite. Synapses in clusters that are active at the same time can interact and thereby generate electrical signals more effectively than synapses spread across the dendrites. Further experiments are now needed to map the synapses between other kinds of neurons, and to map synapses from two different inputs at the same time. DOI: http://dx.doi.org/10.7554/eLife.09222.002

Ca2+influx through synaptic NMDA receptors (NMDA-Rs) triggers a variety of adaptive cellular processes. To probe NMDA-R-mediated [Ca2+] signaling, we used two-photon glutamate uncaging to stimulate NMDA-Rs on individual dendritic spines of CA1 pyramidal neurons in rat brain slices. We measured NMDA-R currents at the soma and NMDA-R-mediated [Ca2+] transients in stimulated spines (Δ[Ca2+]). Uncaging-evoked NMDA-R current amplitudes were independent of the size of the stimulated spine, implying that smaller spines contain higher densities of functional NMDA-Rs. The ratio of Δ[Ca2+] over NMDA-R current was highly variable (factor of 10) across spines, especially for small spines. These differences were not explained by heterogeneity in spine sizes or diffusional coupling between spines and their parent dendrites. In addition, we find that small spines have NMDA-R currents that are sensitive to NMDA-R NR2B subunit-specific antagonists. With block of NR2B-containing receptors, the range of Δ[Ca2+]/NMDA-R current ratios and their average value were much reduced. Our data suggest that individual spines can regulate the subunit composition of their NMDA-Rs and the effective fractional Ca2+current through these receptors.

SummarySubsynaptic structures such as bouton, active zone, postsynaptic density (PSD) and dendritic spine, are highly correlated in their dimensions and also correlate with synapse strength. Why this is so and how such correlations are maintained during synaptic plasticity remains poorly understood. We induced spine enlargement by two-photon glutamate uncaging and examined the relationship between spine, PSD, and bouton size by two-photon time-lapse imaging and electron microscopy. In enlarged spines the PSD-associated protein Homer1c increased rapidly, whereas the PSD protein PSD-95 increased with a delay and only in cases of persistent spine enlargement. In the case of nonpersistent spine enlargement, the PSD proteins remained unchanged or returned to their original level. The ultrastructure at persistently enlarged spines displayed matching dimensions of spine, PSD, and bouton, indicating their correlated enlargement. This supports a model in which balancing of synaptic structures is a hallmark for the stabilization of structural modifications during synaptic plasticity.

Synaptic neurotransmitter release is restricted to active zones, where the processes of synaptic vesicle tethering, priming to fusion competence, and Ca2+-triggered fusion are taking place in a highly coordinated manner. We show that the active zone components Munc13-1, an essential vesicle priming protein, and RIM1, a Rab3 effector with a putative role in vesicle tethering, interact functionally. Disruption of this interaction causes a loss of fusion-competent synaptic vesicles, creating a phenocopy of Munc13-1-deficient neurons. RIM1 binding and vesicle priming are mediated by two distinct structural modules of Munc13-1. The Munc13-1/RIM1 interaction may create a functional link between synaptic vesicle tethering and priming, or it may regulate the priming reaction itself, thereby determining the number of fusion-competent vesicles.

Syntaxin-1 is a key component of the synaptic vesicle docking/fusion machinery that forms the SNARE complex with VAMP/synaptobrevin and SNAP-25. Identifying proteins that modulate SNARE complex formation is critical for understanding the molecular mechanisms underlying neurotransmitter release and its modulation. We have cloned and characterized a protein called syntaphilin that is selectively expressed in brain. Syntaphilin competes with SNAP-25 for binding to syntaxin-1 and inhibits SNARE complex formation by absorbing free syntaxin-1. Transient overexpression of syntaphilin in cultured hippocampal neurons significantly reduces neurotransmitter release. Furthermore, introduction of syntaphilin into presynaptic superior cervical ganglion neurons in culture inhibits synaptic transmission. These findings suggest that syntaphilin may function as a molecular clamp that controls free syntaxin-1 availability for the assembly of the SNARE complex, and thereby regulates synaptic vesicle exocytosis.

ADP ribosylation factors (ARFs) represent a family of small monomeric G proteins that switch from an inactive, GDP-bound state to an active, GTP-bound state. One member of this family, ARF6, translocates on activation from intracellular compartments to the plasma membrane and has been implicated in regulated exocytosis in neuroendocrine cells. Because GDP release in vivo is rather slow, ARF activation is facilitated by specific guanine nucleotide exchange factors like cytohesin-1 or ARNO. Here we show that msec7-1, a rat homologue of cytohesin-1, translocates ARF6 to the plasma membrane in living cells. Overexpression of msec7-1 leads to an increase in basal synaptic transmission at the Xenopus neuromuscular junction. msec7-1-containing synapses have a 5-fold higher frequency of spontaneous synaptic currents than control synapses. On stimulation, the amplitudes of the resulting evoked postsynaptic currents of msec7-1-overexpressing neurons are increased as well. However, further stimulation leads to a decline in amplitudes approaching the values of control synapses. This transient effect on amplitude is strongly reduced on overexpression of msec7-1E157K, a mutant incapable of translocating ARFs. Our results provide evidence that small G proteins of the ARF family and activating factors like msec7-1 play an important role in synaptic transmission, most likely by making more vesicles available for fusion at the plasma membrane.

SummaryA fundamental property of neuronal circuits is the ability to adapt to altered sensory inputs. It is well established that the functional synaptic changes underlying this adaptation are reflected by structural modifications in excitatory neurons. In contrast, the degree to which structural plasticity in inhibitory neurons accompanies functional changes is less clear. Here, we use two-photon imaging to monitor the fine structure of inhibitory neurons in mouse visual cortex after deprivation induced by retinal lesions. We find that a subset of inhibitory neurons carry dendritic spines, which form glutamatergic synapses. Removal of visual input correlates with a rapid and lasting reduction in the number of inhibitory cell spines. Similar to the effects seen for dendritic spines, the number of inhibitory neuron boutons dropped sharply after retinal lesions. Together, these data suggest that structural changes in inhibitory neurons may precede structural changes in excitatory circuitry, which ultimately result in functional adaptation following sensory deprivation.

With key roles in essential brain functions ranging from the basics of excitatory neurotransmission to the complexities of learning and memory, the N-methyl-d-aspartate (NMDA) receptor can be considered one of the fundamental neurotransmitter receptors in the brain. The NMDA receptor belongs to a family of ionotropic receptors for the excitatory amino acid glutamate and is characterized by high affinity for glutamate, high calcium permeability, and a voltage-dependent block by magnesium ions. This article focuses on the basic biophysical properties and physiological functions of NMDA receptors and how these are modulated by various signaling molecules and biochemical cascades under physiological conditions.

Spine growth and retraction with synapse formation and elimination plays an important role in shaping brain circuits during development and in the adult brain. Yet the temporal relationship between spine morphogenesis and the formation of functional synapses remains poorly defined. We imaged hippocampal pyramidal neurons to identify spines of different ages. We then used two-photon glutamate uncaging, whole-cell recording, and Ca2+ imaging to analyze the properties of nascent spines and their older neighbors. We found that new spines expressed glutamate sensitive currents that were indistinguishable from mature spines of comparable volumes. Some spines exhibited negligible AMPA receptor-mediated responses, but the occurrence of these ‘silent’ spines was uncorrelated with spine age. In contrast, NMDA receptor-mediated Ca2+ accumulations were significantly lower in new spines. New spines reconstructed using electron microscopy made synapses. Our data support a model in which outgrowth and enlargement of nascent spines is tightly coupled to formation and maturation of glutamatergic synapses.