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Increased tonic activity of locus coeruleus noradrenergic (LC-NE) neurons induces anxiety-like and aversive behavior. While some information is known about the afferent circuitry that endogenously drives this neural activity and behavior, the downstream receptors and anatomical projections that mediate these acute risk aversive behavioral states via the LC-NE system remain unresolved. Here we use a combination of retrograde tracing, fast-scan cyclic voltammetry, electrophysiology, and in vivo optogenetics with localized pharmacology to identify neural substrates downstream of increased tonic LC-NE activity in mice. We demonstrate that photostimulation of LC-NE fibers in the BLA evokes norepinephrine release in the basolateral amygdala (BLA), alters BLA neuronal activity, conditions aversion, and increases anxiety-like behavior. Additionally, we report that β-adrenergic receptors mediate the anxiety-like phenotype of increased NE release in the BLA. These studies begin to illustrate how the complex efferent system of the LC-NE system selectively mediates behavior through distinct receptor and projection-selective mechanisms.

Nicotinic acetylcholine receptors (nAChRs) mediate and modulate synaptic transmission throughout the brain, and contribute to learning, memory, and behavior. Dysregulation of α7-type nAChRs in neuropsychiatric as well as immunological and oncological diseases makes them attractive targets for pharmaceutical development. Recently, we identified NACHO as an essential chaperone for α7 nAChRs. Leveraging the robust recombinant expression of α7 nAChRs with NACHO, we utilized genome-wide cDNA library screening and discovered that several anti-apoptotic Bcl-2 family proteins further upregulate receptor assembly and cell surface expression. These effects are mediated by an intracellular motif on α7 that resembles the BH3 binding domain of pro-apoptotic Bcl-2 proteins, and can be blocked by BH3 mimetic Bcl-2 inhibitors. Overexpression of Bcl-2 member Mcl-1 in neurons enhanced surface expression of endogenous α7 nAChRs, while a combination of chemotherapeutic Bcl2-inhibitors suppressed neuronal α7 receptor assembly. These results demonstrate that Bcl-2 proteins link α7 nAChR assembly to cell survival pathways. The α7 nicotinic acetylcholine receptor (nAChR) plays a major role in shaping the activity of neuronal circuits and contributes to the pathophysiology of several neurological disorders. Following cDNA library screening, the authors identify anti-apoptotic, Bcl-2 family proteins as enhancers of α7 nAChR assembly, acting through an intracellular BH3-like domain during receptor biogenesis in the endoplasmic reticulum.

Successful integration of advanced semiconductor devices with biological systems will accelerate basic scientific discoveries and their translation into clinical technologies. In neuroscience generally, and in optogenetics in particular, the ability to insert light sources, detectors, sensors, and other components into precise locations of the deep brain yields versatile and important capabilities. Here, we introduce an injectable class of cellular-scale optoelectronics that offers such features, with examples of unmatched operational modes in optogenetics, including completely wireless and programmed complex behavioral control over freely moving animals. The ability of these ultrathin, mechanically compliant, biocompatible devices to afford minimally invasive operation in the soft tissues of the mammalian brain foreshadow applications in other organ systems, with potential for broad utility in biomédical science and engineering.

Anxiety disorders are debilitating psychiatric illnesses with detrimental effects on human health. These heightened states of arousal are often in the absence of obvious threatening cues and are difficult to treat owing to a lack of understanding of the neural circuitry and cellular machinery mediating these conditions. Activation of noradrenergic circuitry in the basolateral amygdala is thought to have a role in stress, fear, and anxiety, and the specific cell and receptor types responsible is an active area of investigation. Here we take advantage of two novel cellular approaches to dissect the contributions of G-protein signaling in acute and social anxiety-like states. We used a chemogenetic approach utilizing the Gαs DREADD (rM3Ds) receptor and show that selective activation of generic Gαs signaling is sufficient to induce acute and social anxiety-like behavioral states in mice. Second, we use a recently characterized chimeric receptor composed of rhodopsin and the β2-adrenergic receptor (Opto-β2AR) with in vivo optogenetic techniques to selectively activate Gαs β-adrenergic signaling exclusively within excitatory neurons of the basolateral amygdala. We found that optogenetic induction of β-adrenergic signaling in the basolateral amygdala is sufficient to induce acute and social anxiety-like behavior. These findings support the conclusion that activation of Gαs signaling in the basolateral amygdala has a role in anxiety. These data also suggest that acute and social anxiety-like states may be mediated through signaling pathways identical to β-adrenergic receptors, thus providing support that inhibition of this system may be an effective anxiolytic therapy.

Optogenetics has provided a revolutionary approach to dissecting biological phenomena. However, the generation and use of optically active GPCRs in these contexts is limited and it is unclear how well an opsin-chimera GPCR might mimic endogenous receptor activity. Here we show that a chimeric rhodopsin/β 2 adrenergic receptor (opto-β 2 AR) is similar in dynamics to endogenous β 2 AR in terms of: cAMP generation, MAP kinase activation and receptor internalization. In addition, we develop and characterize a novel toolset of optically active, functionally selective GPCRs that can bias intracellular signalling cascades towards either G-protein or arrestin-mediated cAMP and MAP kinase pathways. Finally, we show how photoactivation of opto-β 2 AR in vivo modulates neuronal activity and induces anxiety-like behavioural states in both fiber-tethered and wireless, freely moving animals when expressed in brain regions known to contain β 2 ARs. These new GPCR approaches enhance the utility of optogenetics and allow for discrete spatiotemporal control of GPCR signalling in vitro and in vivo . Optogenetic activation of β2-adrenergic receptors (β2-AR) has been achieved, but not characterized in detail. Here, Siuda et al . show that light-controlled opto-β2AR mimics endogenous β2AR activity in vitro and in vivo , and develop novel, optically active, functionally selective receptors to bias β2AR intracellular signaling mechanisms.

Anxiety disorders are debilitating psychiatric illnesses with detrimental effects on human health. These heightened states of arousal are often in the absence of obvious threatening cues and are difficult to treat owing to a lack of understanding of the neural circuitry and cellular machinery mediating these conditions. Activation of noradrenergic circuitry in the basolateral amygdala is thought to have a role in stress, fear, and anxiety, and the specific cell and receptor types responsible is an active area of investigation. Here we take advantage of two novel cellular approaches to dissect the contributions of G-protein signaling in acute and social anxiety-like states. We used a chemogenetic approach utilizing the Gαs DREADD (rM3Ds) receptor and show that selective activation of generic Gαs signaling is sufficient to induce acute and social anxiety-like behavioral states in mice. Second, we use a recently characterized chimeric receptor composed of rhodopsin and the β2 -adrenergic receptor (Opto-β2 AR) with in vivo optogenetic techniques to selectively activate Gαs β-adrenergic signaling exclusively within excitatory neurons of the basolateral amygdala. We found that optogenetic induction of β-adrenergic signaling in the basolateral amygdala is sufficient to induce acute and social anxiety-like behavior. These findings support the conclusion that activation of Gαs signaling in the basolateral amygdala has a role in anxiety. These data also suggest that acute and social anxiety-like states may be mediated through signaling pathways identical to β-adrenergic receptors, thus providing support that inhibition of this system may be an effective anxiolytic therapy.

Optogenetics has provided a revolutionary approach to dissecting biological phenomena. Traditional techniques however use binary control schemes to selectively turn on/off neurons in the presence of light. While useful in helping to understand aspects of the cell biology and neural circuitry in healthy and diseased states, these approaches are limited in their ability to mirror endogenous neuromodulator receptor signaling. For this reason, the development of optically active G-protein coupled receptors (GPCRs) allows for more fine tuned modulation of cellular activity. This, in combination with the spatiotemporal control offered through optogenetics, provides more refined in vitro and in vivo GPCR toolkits. Here we generated, and characterized two optically active GPCRs. A Gαs- coupled chimera of rhodopsin and the β2-adrenergic receptor (opto-β2AR) and a Gαi- coupled chimera of rhodopsin and the mu-opioid receptor (opto-MOR). We first fully compare these receptors to their wild type counterparts using canonical in vitro readouts of GPCR activity and demonstrate that our chimeric receptors indeed behave as their biological complements. We then package these receptors into viral hosts and transfect them into various brain regions of interest. Utilizing electrophysiological and behavioral measurements we demonstrate that optical activation of opto-β2AR mimics noradrenergic activity, producing an anxiety-like behavioral phenotype, while optical activation of opto-MOR shows robust effects on motivational behavioral, similar to endogenous mu-opioid receptor activity. Taken together, we show that these optically active GPCR approaches enhance the utility of optogenetics and allow for discrete spatiotemporal control of GPCR signaling in vitro and in vivo, thus expanding the toolbox for understanding both neurotransmitter and neuropeptide signaling within neural circuits.

Optogenetics has provided a revolutionary approach to dissecting biological phenomena. However, the generation and use of optically active GPCRs in these contexts is limited and it is unclear how well an opsin-chimera GPCR might mimic endogenous receptor activity. Here we show that a chimeric rhodopsin/β2 adrenergic receptor (opto-β2 AR) is similar in dynamics to endogenous β2 AR in terms of: cAMP generation, MAP kinase activation and receptor internalization. In addition, we develop and characterize a novel toolset of optically active, functionally selective GPCRs that can bias intracellular signalling cascades towards either G-protein or arrestin-mediated cAMP and MAP kinase pathways. Finally, we show how photoactivation of opto-β2 AR in vivo modulates neuronal activity and induces anxiety-like behavioural states in both fiber-tethered and wireless, freely moving animals when expressed in brain regions known to contain β2 ARs. These new GPCR approaches enhance the utility of optogenetics and allow for discrete spatiotemporal control of GPCR signalling in vitro and in vivo.