Cortical Integration of Vestibular and Visual Cues for Navigation, Visual Processing, and Perception.

Despite increasing evidence of its involvement in several key functions of the cerebral cortex, the vestibular sense rarely enters our consciousness. Indeed, the extent to which these internal signals are incorporated within cortical sensory representation and how they might be relied upon for sensory-driven decision-making, during, for example, spatial navigation, is yet to be understood. Recent novel experimental approaches in rodents have probed both the physiological and behavioral significance of vestibular signals and indicate that their widespread integration with vision improves both the cortical representation and perceptual accuracy of self-motion and orientation. Here, we summarize these recent findings with a focus on cortical circuits involved in visual perception and spatial navigation and highlight the major remaining knowledge gaps. We suggest that vestibulo-visual integration reflects a process of constant updating regarding the status of self-motion, and access to such information by the cortex is used for sensory perception and predictions that may be implemented for rapid, navigation-related decision-making.

Accurate determination of marker location within whole-brain microscopy images.

High-resolution whole-brain microscopy provides a means for post hoc determination of the location of implanted devices and labelled cell populations that are necessary to interpret in vivo experiments designed to understand brain function. Here we have developed two plugins (brainreg and brainreg-segment) for the Python-based image viewer napari, to accurately map any object in a common coordinate space. We analysed the position of dye-labelled electrode tracks and two-photon imaged cell populations expressing fluorescent proteins. The precise location of probes and cells were physiologically interrogated and revealed accurate segmentation with near-cellular resolution.

Analysis of segmentation ontology reveals the similarities and differences in connectivity onto L2/3 neurons in mouse V1.

Quantitatively comparing brain-wide connectivity of different types of neuron is of vital importance in understanding the function of the mammalian cortex. Here we have designed an analytical approach to examine and compare datasets from hierarchical segmentation ontologies, and applied it to long-range presynaptic connectivity onto excitatory and inhibitory neurons, mainly located in layer 2/3 (L2/3), of mouse primary visual cortex (V1). We find that the origins of long-range connections onto these two general cell classes-as well as their proportions-are quite similar, in contrast to the inputs on to a cell type in L6. These anatomical data suggest that distal inputs received by the general excitatory and inhibitory classes of neuron in L2/3 overlap considerably.

A Circuit for Integration of Head- and Visual-Motion Signals in Layer 6 of Mouse Primary Visual Cortex.

To interpret visual-motion events, the underlying computation must involve internal reference to the motion status of the observer's head. We show here that layer 6 (L6) principal neurons in mouse primary visual cortex (V1) receive a diffuse, vestibular-mediated synaptic input that signals the angular velocity of horizontal rotation. Behavioral and theoretical experiments indicate that these inputs, distributed over a network of 100 L6 neurons, provide both a reliable estimate and, therefore, physiological separation of head-velocity signals. During head rotation in the presence of visual stimuli, L6 neurons exhibit postsynaptic responses that approximate the arithmetic sum of the vestibular and visual-motion response. Functional input mapping reveals that these internal motion signals arrive into L6 via a direct projection from the retrosplenial cortex. We therefore propose that visual-motion processing in V1 L6 is multisensory and contextually dependent on the motion status of the animal's head.

Interneurons and oligodendrocyte progenitors form a structured synaptic network in the developing neocortex.

NG2 cells, oligodendrocyte progenitors, receive a major synaptic input from interneurons in the developing neocortex. It is presumed that these precursors integrate cortical networks where they act as sensors of neuronal activity. We show that NG2 cells of the developing somatosensory cortex form a transient and structured synaptic network with interneurons that follows its own rules of connectivity. Fast-spiking interneurons, highly connected to NG2 cells, target proximal subcellular domains containing GABAA receptors with γ2 subunits. Conversely, non-fast-spiking interneurons, poorly connected with these progenitors, target distal sites lacking this subunit. In the network, interneuron-NG2 cell connectivity maps exhibit a local spatial arrangement reflecting innervation only by the nearest interneurons. This microcircuit architecture shows a connectivity peak at PN10, coinciding with a switch to massive oligodendrocyte differentiation. Hence, GABAergic innervation of NG2 cells is temporally and spatially regulated from the subcellular to the network level in coordination with the onset of oligodendrogenesis.

Postnatal down-regulation of the GABAA receptor γ2 subunit in neocortical NG2 cells accompanies synaptic-to-extrasynaptic switch in the GABAergic transmission mode.

NG2 cells, a main pool of glial progenitors, express γ-aminobutyric acid A (GABA(A)) receptors (GABA(A)Rs), the functional and molecular properties of which are largely unknown. We recently reported that transmission between GABAergic interneurons and NG2 cells drastically changes during development of the somatosensory cortex, switching from synaptic to extrasynaptic communication. Since synaptic and extrasynaptic GABA(A)Rs of neurons differ in their subunit composition, we hypothesize that GABA(A)Rs of NG2 cells undergo molecular changes during cortical development accompanying the switch of transmission modes. Single-cell RT-PCR and the effects of zolpidem and α5IA on evoked GABAergic currents reveal the predominance of functional α1- and α5-containing GABA(A)Rs at interneuron-NG2 cell synapses in the second postnatal week, while the α5 expression declines later in development when responses are exclusively extrasynaptic. Importantly, pharmacological and molecular analyses demonstrate that γ2, a subunit contributing to the clustering of GABA(A)Rs at postsynaptic sites in neurons, is down-regulated in NG2 cells in a cell type-specific manner in concomitance with the decline of synaptic activity and the switch of transmission mode. In keeping with the synaptic nature of γ2 in neurons, the down-regulation of this subunit is an important molecular hallmark of the change of transmission modes between interneurons and NG2 cells during development.

The stimulus selectivity and connectivity of layer six principal cells reveals cortical microcircuits underlying visual processing.

Sensory computations performed in the neocortex involve layer six (L6) cortico-cortical (CC) and cortico-thalamic (CT) signaling pathways. Developing an understanding of the physiological role of these circuits requires dissection of the functional specificity and connectivity of the underlying individual projection neurons. By combining whole-cell recording from identified L6 principal cells in the mouse primary visual cortex (V1) with modified rabies virus-based input mapping, we have determined the sensory response properties and upstream monosynaptic connectivity of cells mediating the CC or CT pathway. We show that CC-projecting cells encompass a broad spectrum of selectivity to stimulus orientation and are predominantly innervated by deep layer V1 neurons. In contrast, CT-projecting cells are ultrasparse firing, exquisitely tuned to orientation and direction information, and receive long-range input from higher cortical areas. This segregation in function and connectivity indicates that L6 microcircuits route specific contextual and stimulus-related information within and outside the cortical network.

Oligodendrocyte precursor cells are accurate sensors of local K+ in mature gray matter.

Oligodendrocyte precursor cells (OPCs) are the major source of myelinating oligodendrocytes during development. These progenitors are highly abundant at birth and persist in the adult where they are distributed throughout the brain. The large abundance of OPCs after completion of myelination challenges their unique role as progenitors in the healthy adult brain. Here we show that adult OPCs of the barrel cortex sense fine extracellular K(+) increases generated by neuronal activity, a property commonly assigned to differentiated astrocytes rather than to progenitors. Biophysical, pharmacological, and single-cell RT-PCR analyses demonstrate that this ability of OPCs establishes itself progressively through the postnatal upregulation of Kir4.1 K(+) channels. In animals with advanced cortical myelination, extracellular stimulation of layer V axons induces slow K(+) currents in OPCs, which amplitude correlates with presynaptic action potential rate. Moreover, using paired recordings, we demonstrate that the discharge of a single neuron can be detected by nearby adult OPCs, indicating that these cells are strategically located to detect local changes in extracellular K(+) concentration during physiological neuronal activity. These results identify a novel unitary neuron-OPC connection, which transmission does not rely on neurotransmitter release and appears late in development. Beyond their abundance in the mature brain, the postnatal emergence of a physiological response of OPCs to neuronal network activity supports the view that in the adult these cells are not progenitors only.

Cortical circuits: layer 6 is a gain changer.

Stimulation of excitatory cells in layer six of mouse visual cortex results in net inhibition of the spiking of neurons in upper cortical layers; this ascending intra-cortical drive provides a mechanism for gain modulation of sensory-evoked responses.

Central role of GABA in neuron-glia interactions.

The major types of glial cells-astrocytes, microglia, and cells of the oligodendroglial lineage-are known to express functional metabotropic and ionotropic GABA receptors. Neuronal signaling mechanisms allowing for the activation of these receptors in glia are probably as complex as those described among neurons and involve synaptic and extrasynaptic transmission modes. In addition, astrocytes can signal back to neurons by releasing GABA, probably through unconventional nonvesicular mechanisms. The decryption of the roles played by GABAergic signaling in neuron-glia interactions is only beginning, but it has been suggested that activation of glial cells by GABA influences important functions of the brain such as neuronal activity, differentiation, myelination, and neuroprotection. This review discusses the cellular mechanisms allowing the major types of glial cells to sense and transmit GABAergic signals and gives an overview of potential roles of this signaling pathway in developing and mature brains.

Is neuronal communication with NG2 cells synaptic or extrasynaptic?

NG2-expressing glial cells (NG2 cells) represent a major pool of progenitors able to generate myelinating oligodendrocytes, and perhaps astrocytes and neurones, in the postnatal brain. In the last decade, it has been demonstrated that NG2 cells receive functional glutamatergic and GABAergic synapses mediating fast synaptic transmission in different brain regions. However, several controversies exist in this field. While two classes of NG2 cells have been defined by the presence or absence of Na(+) channels, action potential firing and neuronal input, other studies suggest that all NG2 cells possess Na(+) conductances and are the target of quantal neuronal release, but are unable to trigger action potential firing. Here we bring new evidence supporting the idea that the level of expression of Na(+) conductances is not a criterion to discriminate NG2 cell subpopulations in the somatosensory cortex. Surprisingly, recent reports demonstrated that NG2 cells detect quantal glutamate release from unmyelinated axons in white matter regions. Yet, it is difficult from these studies to establish whether axonal vesicular release in white matter occurs at genuine synaptic junctions or at ectopic release sites. In addition, we recently reported a new mode of extrasynaptic communication between neurones and NG2 cells that relies on pure GABA spillover and does not require GABAergic synaptic input. This review discusses the properties of quantal neuronal release onto NG2 cells and gives an extended overview of potential extrasynaptic modes of transmission, from ectopic to diffuse volume transmission, between neurones and NG2 cells in the brain.

Postnatal switch from synaptic to extrasynaptic transmission between interneurons and NG2 cells.

NG2 cells, oligodendrocyte precursors, play a critical role in myelination during postnatal brain maturation, but a pool of these precursors is maintained in the adult and recruited to lesions in demyelinating diseases. NG2 cells in immature animals have recently been shown to receive synaptic inputs from neurons, and these have been assumed to persist in the adult. Here, we investigated the GABAergic synaptic activity of NG2 cells in acute slices of the barrel cortex of NG2-DsRed transgenic mice during the first postnatal month, which corresponds to the period of active myelination in the neocortex. Our data demonstrated that the frequency of spontaneous and miniature GABAergic synaptic activity of cortical NG2 cells dramatically decreases after the second postnatal week, indicating a decrease in the number of synaptic inputs onto NG2 cells during development. However, NG2 cells still receive GABAergic inputs from interneurons in the adult cortex. These inputs do not rely on the presence of functional synapses but involve a form of GABA spillover. This GABA volume transmission allows interneurons to induce phasic responses in target NG2 cells through the activation of extrasynaptic GABA(A) receptors. Hence, after development is complete, volume transmission allows NG2 cells to integrate neuronal activity patterns at frequencies occurring during in vivo sensory stimulation.

Holographic photolysis for multiple cell stimulation in mouse hippocampal slices.

BACKGROUND: Advanced light microscopy offers sensitive and non-invasive means to image neural activity and to control signaling with photolysable molecules and, recently, light-gated channels. These approaches require precise and yet flexible light excitation patterns. For synchronous stimulation of subsets of cells, they also require large excitation areas with millisecond and micrometric resolution. We have recently developed a new method for such optical control using a phase holographic modulation of optical wave-fronts, which minimizes power loss, enables rapid switching between excitation patterns, and allows a true 3D sculpting of the excitation volumes. In previous studies we have used holographic photololysis to control glutamate uncaging on single neuronal cells. Here, we extend the use of holographic photolysis for the excitation of multiple neurons and of glial cells. METHODS/PRINCIPAL FINDINGS: The system combines a liquid crystal device for holographic patterned photostimulation, high-resolution optical imaging, the HiLo microscopy, to define the stimulated regions and a conventional Ca(2+) imaging system to detect neural activity. By means of electrophysiological recordings and calcium imaging in acute hippocampal slices, we show that the use of excitation patterns precisely tailored to the shape of multiple neuronal somata represents a very efficient way for the simultaneous excitation of a group of neurons. In addition, we demonstrate that fast shaped illumination patterns also induce reliable responses in single glial cells. CONCLUSIONS/SIGNIFICANCE: We show that the main advantage of holographic illumination is that it allows for an efficient excitation of multiple cells with a spatiotemporal resolution unachievable with other existing approaches. Although this paper focuses on the photoactivation of caged molecules, our approach will surely prove very efficient for other probes, such as light-gated channels, genetically encoded photoactivatable proteins, photoactivatable fluorescent proteins, and voltage-sensitive dyes.

Functional alpha 7-containing nicotinic receptors of NG2-expressing cells in the hippocampus.

In the postnatal central nervous system, glial cells expressing the chondroitin sulfate proteoglycan NG2 (NG2-cells) constitute a cell population exhibiting several properties of oligodendrocyte precursors such as the ability to proliferate. One particular feature of NG2-cells is that they express several glutamatergic and GABAergic ionotropic receptors activated by synaptic neurotransmitter release. Here, we used patch-clamp recordings, immunostaining, calcium imaging, and intracellular labeling to test for the presence of ionotropic nicotinic acetylcholine receptors (nAChRs) in NG2-cells identified in acute hippocampal slices of mice. We demonstrated that these cells express functional nAChRs during the second postnatal week, i.e., the period in which they become the most abundant proliferative cell type of CA1 stratum radiatum. Pharmacological experiments showed that NG2-cells express alpha 7-containing nAChRs. In particular, the powerful positive allosteric modulator of these receptors PNU-120596 induced a 20-fold increase of agonist-induced currents and revealed rises in intracellular calcium concentration upon agonist applications. In addition, nanomolar concentrations of nicotine, which did not induce any response in these cells, largely desensitized nAChR-mediated currents. These data indicate that the functional expression of Ca(2+)-permeable alpha 7-containing nAChRs in hippocampal slices is not restricted to neurons and that the receptors of NG2-cells can be desensitized by low concentrations of nicotine.