All-optical reporting of inhibitory receptor driving force in the nervous system.

Ionic driving forces provide the net electromotive force for ion movement across receptors, channels, and transporters, and are a fundamental property of all cells. In the nervous system, fast synaptic inhibition is mediated by chloride permeable GABA and glycine receptors, and single-cell intracellular recordings have been the only method for estimating driving forces across these receptors (DF). Here we present a tool for quantifying inhibitory receptor driving force named ORCHID: all-Optical Reporting of CHloride Ion Driving force.

sUPRa is a dual-color reporter for unbiased quantification of the unfolded protein response with cellular resolution.

The unfolded protein response (UPR) maintains proteostasis upon endoplasmic reticulum (ER) stress, and is initiated by a range of physiological and pathological processes. While there have been advances in developing fluorescent reporters for monitoring individual signaling pathways of the UPR, this approach may not capture a cell's overall UPR activity. Here we describe a novel sensor of UPR activity, sUPRa, which is designed to report the global UPR.

Higher-order thalamocortical circuits are specified by embryonic cortical progenitor types in the mouse brain.

The sensory cortex receives synaptic inputs from both first-order and higher-order thalamic nuclei. First-order inputs relay simple stimulus properties from the periphery, whereas higher-order inputs relay more complex response properties, provide contextual feedback, and modulate plasticity. Here, we reveal that a cortical neuron's higher-order input is determined by the type of progenitor from which it is derived during embryonic development.

Optogenetic Determination of Dynamic and Cell-Type-Specific Inhibitory Reversal Potentials.

The reversal potential refers to the membrane potential at which the net current flow through a channel reverses direction. The reversal potential is determined by transmembrane ion gradients and, in turn, determines how the channel's activity will affect the membrane potential. Traditional investigation into the reversal potential of inhibitory ligand-gated ion channels (E) has relied upon the activation of endogenous receptors, such as the GABA-A receptor (GABAR).

Somnotate: A probabilistic sleep stage classifier for studying vigilance state transitions.

Electrophysiological recordings from freely behaving animals are a widespread and powerful mode of investigation in sleep research. These recordings generate large amounts of data that require sleep stage annotation (polysomnography), in which the data is parcellated according to three vigilance states: awake, rapid eye movement (REM) sleep, and non-REM (NREM) sleep. Manual and current computational annotation methods ignore intermediate states because the classification features become ambiguous, even though intermediate states contain important information regarding vigilance state dynamics.

Active cortical networks promote shunting fast synaptic inhibition in vivo.

Fast synaptic inhibition determines neuronal response properties in the mammalian brain and is mediated by chloride-permeable ionotropic GABA-A receptors (GABARs). Despite their fundamental role, it is still not known how GABARs signal in the intact brain. Here, we use in vivo gramicidin recordings to investigate synaptic GABAR signaling in mouse cortical pyramidal neurons under conditions that preserve native transmembrane chloride gradients.

A genetically targeted ion sensor reveals distinct seizure-related chloride and pH dynamics in GABAergic interneuron populations.

Intracellular chloride and pH play fundamental roles in determining a neuron's synaptic inhibition and excitability. Yet it has been difficult to measure changes in these ions during periods of heightened network activity, such as occur in epilepsy. Here we develop a version of the fluorescent reporter, ClopHensorN, to enable simultaneous quantification of chloride and pH in genetically defined neurons during epileptiform activity.

Effects of clozapine-N-oxide and compound 21 on sleep in laboratory mice.

Designer receptors exclusively activated by designer drugs (DREADDs) are chemogenetic tools for remote control of targeted cell populations using chemical actuators that bind to modified receptors. Despite the popularity of DREADDs in neuroscience and sleep research, potential effects of the DREADD actuator clozapine-N-oxide (CNO) on sleep have never been systematically tested. Here, we show that intraperitoneal injections of commonly used CNO doses (1, 5, and 10 mg/kg) alter sleep in wild-type male laboratory mice.

Intracellular chloride regulation mediates local sleep pressure in the cortex.

Extended wakefulness is associated with reduced performance and the build-up of sleep pressure. In the cortex, this manifests as changes in network activity. These changes show local variation depending on the waking experience, and their underlying mechanisms represent targets for overcoming the effects of tiredness.

Why won't it stop? The dynamics of benzodiazepine resistance in status epilepticus.

Status epilepticus is a life-threatening neurological emergency that affects both adults and children. Approximately 36% of episodes of status epilepticus do not respond to the current preferred first-line treatment, benzodiazepines. The proportion of episodes that are refractory to benzodiazepines is higher in low-income and middle-income countries (LMICs) than in high-income countries (HICs).

Disrupting Epileptiform Activity by Preventing Parvalbumin Interneuron Depolarization Block.

Inhibitory synaptic mechanisms oppose epileptic network activity in the brain. The breakdown in this inhibitory restraint and propagation of seizure activity has been linked to the overwhelming of feedforward inhibition, which is provided in large part by parvalbumin-expressing (PV) interneurons in the cortex. The underlying cellular processes therefore represent potential targets for understanding and preventing the propagation of seizure activity.

A role for the cortex in sleep-wake regulation.

Cortical and subcortical circuitry are thought to play distinct roles in the generation of sleep oscillations and global state control, respectively. Here we silenced a subset of neocortical layer 5 pyramidal and archicortical dentate gyrus granule cells in male mice by ablating SNAP25. This markedly increased wakefulness and reduced rebound of electroencephalographic slow-wave activity after sleep deprivation, suggesting a role for the cortex in both vigilance state control and sleep homeostasis.

Targeted single-cell RNA sequencing of transcription factors enhances the identification of cell types and trajectories.

Single-cell RNA sequencing (scRNA-seq) is a widely used method for identifying cell types and trajectories in biologically heterogeneous samples, but it is limited in its detection and quantification of lowly expressed genes. This results in missing important biological signals, such as the expression of key transcription factors (TFs) driving cellular differentiation. We show that targeted sequencing of ∼1000 TFs (scCapture-seq) in iPSC-derived neuronal cultures greatly improves the biological information garnered from scRNA-seq.

Pro-maturational Effects of Human iPSC-Derived Cortical Astrocytes upon iPSC-Derived Cortical Neurons.

Astrocytes influence neuronal maturation and function by providing trophic support, regulating the extracellular environment, and modulating signaling at synapses. The emergence of induced pluripotent stem cell (iPSC) technology offers a human system with which to validate and re-evaluate insights from animal studies. Here, we set out to examine interactions between human astrocytes and neurons derived from a common cortical progenitor pool, thereby recapitulating aspects of in vivo cortical development.

Chloride dynamics alter the input-output properties of neurons.

Fast synaptic inhibition is a critical determinant of neuronal output, with subcellular targeting of synaptic inhibition able to exert different transformations of the neuronal input-output function. At the receptor level, synaptic inhibition is primarily mediated by chloride-permeable Type A GABA receptors. Consequently, dynamics in the neuronal chloride concentration can alter the functional properties of inhibitory synapses.

Backpropagation and the brain.

During learning, the brain modifies synapses to improve behaviour. In the cortex, synapses are embedded within multilayered networks, making it difficult to determine the effect of an individual synaptic modification on the behaviour of the system. The backpropagation algorithm solves this problem in deep artificial neural networks, but historically it has been viewed as biologically problematic.

The transition to status epilepticus: how the brain meets the demands of perpetual seizure activity.

The pathophysiology leading to the development of status epilepticus (SE) remains a topic of significant scientific interest and clinical relevance. The use of multiple experimental and computational models has shown that SE relies on a complex interaction between mechanisms that operate at both a cellular and network level. In this narrative review, we will summarise the current knowledge on the factors that play a key role in allowing SE to develop and persist.

Embryonic progenitor pools generate diversity in fine-scale excitatory cortical subnetworks.

The mammalian neocortex is characterized by a variety of neuronal cell types and precise arrangements of synaptic connections, but the processes that generate this diversity are poorly understood. Here we examine how a pool of embryonic progenitor cells consisting of apical intermediate progenitors (aIPs) contribute to diversity within the upper layers of mouse cortex. In utero labeling combined with single-cell RNA-sequencing reveals that aIPs can generate transcriptionally defined glutamatergic cell types, when compared to neighboring neurons born from other embryonic progenitor pools.

Excitatory GABAergic signalling is associated with benzodiazepine resistance in status epilepticus.

Status epilepticus is defined as a state of unrelenting seizure activity. Generalized convulsive status epilepticus is associated with a rapidly rising mortality rate, and thus constitutes a medical emergency. Benzodiazepines, which act as positive modulators of chloride (Cl-) permeable GABAA receptors, are indicated as first-line treatment, but this is ineffective in many cases.

The Role of TRESK in Discrete Sensory Neuron Populations and Somatosensory Processing.

Two-pore domain K (K) channels generate K leak current, which serves a vital role in controlling and modulating neuronal excitability. This diverse family of K channels exhibit distinct expression and function across neuronal tissues. TWIK-related spinal cord K channel (TRESK) is a K channel with a particularly enriched role in sensory neurons and pain pathways.

Reproducibility of Molecular Phenotypes after Long-Term Differentiation to Human iPSC-Derived Neurons: A Multi-Site Omics Study.

Reproducibility in molecular and cellular studies is fundamental to scientific discovery. To establish the reproducibility of a well-defined long-term neuronal differentiation protocol, we repeated the cellular and molecular comparison of the same two iPSC lines across five distinct laboratories. Despite uncovering acceptable variability within individual laboratories, we detect poor cross-site reproducibility of the differential gene expression signature between these two lines.

Chemogenetic Recruitment of Specific Interneurons Suppresses Seizure Activity.

Current anti-epileptic medications that boost synaptic inhibition are effective in reducing several types of epileptic seizure activity. Nevertheless, these drugs can generate significant side-effects and even paradoxical responses due to the broad nature of their action. Recently developed chemogenetic techniques provide the opportunity to pharmacologically recruit endogenous inhibitory mechanisms in a selective and circuit-specific manner.

Neuronal Chloride Regulation via KCC2 Is Modulated through a GABA Receptor Protein Complex.

GABA receptors are G-protein-coupled receptors that mediate inhibitory synaptic actions through a series of downstream target proteins. It is increasingly appreciated that the GABA receptor forms part of larger signaling complexes, which enable the receptor to mediate multiple different effects within neurons. Here we report that GABA receptors can physically associate with the potassium-chloride cotransporter protein, KCC2, which sets the driving force for the chloride-permeable ionotropic GABA receptor in mature neurons.