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1.
Nature ; 620(7972): 154-162, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37495689

RESUMO

Fasting initiates a multitude of adaptations to allow survival. Activation of the hypothalamic-pituitary-adrenal (HPA) axis and subsequent release of glucocorticoid hormones is a key response that mobilizes fuel stores to meet energy demands1-5. Despite the importance of the HPA axis response, the neural mechanisms that drive its activation during energy deficit are unknown. Here, we show that fasting-activated hypothalamic agouti-related peptide (AgRP)-expressing neurons trigger and are essential for fasting-induced HPA axis activation. AgRP neurons do so through projections to the paraventricular hypothalamus (PVH), where, in a mechanism not previously described for AgRP neurons, they presynaptically inhibit the terminals of tonically active GABAergic afferents from the bed nucleus of the stria terminalis (BNST) that otherwise restrain activity of corticotrophin-releasing hormone (CRH)-expressing neurons. This disinhibition of PVHCrh neurons requires γ-aminobutyric acid (GABA)/GABA-B receptor signalling and potently activates the HPA axis. Notably, stimulation of the HPA axis by AgRP neurons is independent of their induction of hunger, showing that these canonical 'hunger neurons' drive many distinctly different adaptations to the fasted state. Together, our findings identify the neural basis for fasting-induced HPA axis activation and uncover a unique means by which AgRP neurons activate downstream neurons: through presynaptic inhibition of GABAergic afferents. Given the potency of this disinhibition of tonically active BNST afferents, other activators of the HPA axis, such as psychological stress, may also work by reducing BNST inhibitory tone onto PVHCrh neurons.


Assuntos
Jejum , Sistema Hipotálamo-Hipofisário , Neurônios , Sistema Hipófise-Suprarrenal , Proteína Relacionada com Agouti/metabolismo , Hormônio Liberador da Corticotropina/metabolismo , Jejum/fisiologia , Neurônios GABAérgicos/metabolismo , Ácido gama-Aminobutírico/metabolismo , Sistema Hipotálamo-Hipofisário/citologia , Sistema Hipotálamo-Hipofisário/metabolismo , Neurônios/metabolismo , Núcleo Hipotalâmico Paraventricular/citologia , Núcleo Hipotalâmico Paraventricular/metabolismo , Sistema Hipófise-Suprarrenal/citologia , Sistema Hipófise-Suprarrenal/inervação , Sistema Hipófise-Suprarrenal/metabolismo , Terminações Pré-Sinápticas/metabolismo , Núcleos Septais/citologia , Núcleos Septais/metabolismo
2.
Nat Methods ; 21(7): 1275-1287, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38811857

RESUMO

Information is transmitted between brain regions through the release of neurotransmitters from long-range projecting axons. Understanding how the activity of such long-range connections contributes to behavior requires efficient methods for reversibly manipulating their function. Chemogenetic and optogenetic tools, acting through endogenous G-protein-coupled receptor pathways, can be used to modulate synaptic transmission, but existing tools are limited in sensitivity, spatiotemporal precision or spectral multiplexing capabilities. Here we systematically evaluated multiple bistable opsins for optogenetic applications and found that the Platynereis dumerilii ciliary opsin (PdCO) is an efficient, versatile, light-activated bistable G-protein-coupled receptor that can suppress synaptic transmission in mammalian neurons with high temporal precision in vivo. PdCO has useful biophysical properties that enable spectral multiplexing with other optogenetic actuators and reporters. We demonstrate that PdCO can be used to conduct reversible loss-of-function experiments in long-range projections of behaving animals, thereby enabling detailed synapse-specific functional circuit mapping.


Assuntos
Neurônios , Optogenética , Optogenética/métodos , Animais , Neurônios/fisiologia , Neurônios/metabolismo , Transmissão Sináptica , Opsinas/genética , Opsinas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Camundongos , Humanos , Sinapses/fisiologia , Sinapses/metabolismo
3.
Cell ; 147(7): 1446-57, 2011 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-22196724

RESUMO

The capture and utilization of light is an exquisitely evolved process. The single-component microbial opsins, although more limited than multicomponent cascades in processing, display unparalleled compactness and speed. Recent advances in understanding microbial opsins have been driven by molecular engineering for optogenetics and by comparative genomics. Here we provide a Primer on these light-activated ion channels and pumps, describe a group of opsins bridging prior categories, and explore the convergence of molecular engineering and genomic discovery for the utilization and understanding of these remarkable molecular machines.


Assuntos
Opsinas/genética , Opsinas/metabolismo , Sequência de Aminoácidos , Animais , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Opsinas/química , Filogenia , Engenharia de Proteínas
4.
Cereb Cortex ; 33(6): 2838-2856, 2023 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-35788286

RESUMO

Focal cortical epilepsies are frequently refractory to available anticonvulsant drug therapies. One key factor contributing to this state is the limited availability of animal models that allow to reliably study focal cortical seizures and how they recruit surrounding brain areas in vivo. In this study, we selectively expressed the inhibitory chemogenetic receptor, hM4D, in GABAergic neurons in focal cortical areas using viral gene transfer. GABAergic silencing using Clozapine-N-Oxide (CNO) demonstrated reliable induction of local epileptiform events in the electroencephalogram signal of awake freely moving mice. Anesthetized mice experiments showed consistent induction of focal epileptiform-events in both the barrel cortex (BC) and the medial prefrontal cortex (mPFC), accompanied by high-frequency oscillations, a known characteristic of human seizures. Epileptiform-events showed propagation indication with favored propagation pathways: from the BC on 1 hemisphere to its counterpart and from the BC to the mPFC, but not vice-versa. Lastly, sensory whisker-pad stimulation evoked BC epileptiform events post-CNO, highlighting the potential use of this model in studying sensory-evoked seizures. Combined, our results show that targeted chemogenetic inhibition of GABAergic neurons using hM4D can serve as a novel, versatile, and reliable model of focal cortical epileptic activity suitable for systematically studying cortical ictogenesis in different cortical areas.


Assuntos
Clozapina , Epilepsias Parciais , Neurônios GABAérgicos , Neurônios , Regulação Viral da Expressão Gênica , Clozapina/análogos & derivados , Eletroencefalografia , Convulsões , Animais
5.
Nature ; 525(7570): 519-22, 2015 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-26375004

RESUMO

It is commonly assumed, but has rarely been demonstrated, that sex differences in behaviour arise from sexual dimorphism in the underlying neural circuits. Parental care is a complex stereotypic behaviour towards offspring that is shared by numerous species. Mice display profound sex differences in offspring-directed behaviours. At their first encounter, virgin females behave maternally towards alien pups while males will usually ignore the pups or attack them. Here we show that tyrosine hydroxylase (TH)-expressing neurons in the anteroventral periventricular nucleus (AVPV) of the mouse hypothalamus are more numerous in mothers than in virgin females and males, and govern parental behaviours in a sex-specific manner. In females, ablating the AVPV TH(+) neurons impairs maternal behaviour whereas optogenetic stimulation or increased TH expression in these cells enhance maternal care. In males, however, this same neuronal cluster has no effect on parental care but rather suppresses inter-male aggression. Furthermore, optogenetic activation or increased TH expression in the AVPV TH(+) neurons of female mice increases circulating oxytocin, whereas their ablation reduces oxytocin levels. Finally, we show that AVPV TH(+) neurons relay a monosynaptic input to oxytocin-expressing neurons in the paraventricular nucleus. Our findings uncover a previously unknown role for this neuronal population in the control of maternal care and oxytocin secretion, and provide evidence for a causal relationship between sexual dimorphism in the adult brain and sex differences in parental behaviour.


Assuntos
Hipotálamo/citologia , Hipotálamo/fisiologia , Comportamento Materno/fisiologia , Ocitocina/metabolismo , Caracteres Sexuais , Agressão , Animais , Núcleo Hipotalâmico Anterior/citologia , Núcleo Hipotalâmico Anterior/enzimologia , Núcleo Hipotalâmico Anterior/fisiologia , Neurônios Dopaminérgicos/enzimologia , Neurônios Dopaminérgicos/metabolismo , Feminino , Hipotálamo/enzimologia , Masculino , Camundongos , Ocitocina/sangue , Núcleo Hipotalâmico Paraventricular/citologia , Núcleo Hipotalâmico Paraventricular/enzimologia , Núcleo Hipotalâmico Paraventricular/fisiologia , Período Pós-Parto , Sinapses/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo
6.
Proc Natl Acad Sci U S A ; 114(26): E5167-E5176, 2017 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-28611221

RESUMO

Key mitochondrial functions such as ATP production, Ca2+ uptake and release, and substrate accumulation depend on the proton electrochemical gradient (ΔµH+) across the inner membrane. Although several drugs can modulate ΔµH+, their effects are hardly reversible, and lack cellular specificity and spatial resolution. Although channelrhodopsins are widely used to modulate the plasma membrane potential of excitable cells, mitochondria have thus far eluded optogenetic control. Here we describe a toolkit of optometabolic constructs based on selective targeting of channelrhodopsins with distinct functional properties to the inner mitochondrial membrane of intact cells. We show that our strategy enables a light-dependent control of the mitochondrial membrane potential (Δψm) and coupled mitochondrial functions such as ATP synthesis by oxidative phosphorylation, Ca2+ dynamics, and respiratory metabolism. By directly modulating Δψm, the mitochondria-targeted opsins were used to control complex physiological processes such as spontaneous beats in cardiac myocytes and glucose-dependent ATP increase in pancreatic ß-cells. Furthermore, our optometabolic tools allow modulation of mitochondrial functions in single cells and defined cell regions.


Assuntos
Sinalização do Cálcio/fisiologia , Channelrhodopsins/metabolismo , Células Secretoras de Insulina/metabolismo , Potencial da Membrana Mitocondrial/fisiologia , Mitocôndrias Cardíacas/metabolismo , Miócitos Cardíacos/metabolismo , Optogenética , Animais , Células HEK293 , Células HeLa , Humanos , Células Secretoras de Insulina/citologia , Consumo de Oxigênio/fisiologia , Ratos , Ratos Sprague-Dawley
7.
Annu Rev Neurosci ; 34: 389-412, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21692661

RESUMO

Genetically encoded, single-component optogenetic tools have made a significant impact on neuroscience, enabling specific modulation of selected cells within complex neural tissues. As the optogenetic toolbox contents grow and diversify, the opportunities for neuroscience continue to grow. In this review, we outline the development of currently available single-component optogenetic tools and summarize the application of various optogenetic tools in diverse model organisms.


Assuntos
Engenharia Genética/métodos , Neurônios/fisiologia , Animais , Animais Geneticamente Modificados , Humanos , Transdução de Sinal Luminoso/genética , Modelos Genéticos , Rede Nervosa/fisiologia , Inibição Neural/genética , Neurociências , Opsinas/genética
8.
Nature ; 482(7385): 369-74, 2012 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-22266941

RESUMO

Channelrhodopsins (ChRs) are light-gated cation channels derived from algae that have shown experimental utility in optogenetics; for example, neurons expressing ChRs can be optically controlled with high temporal precision within systems as complex as freely moving mammals. Although ChRs have been broadly applied to neuroscience research, little is known about the molecular mechanisms by which these unusual and powerful proteins operate. Here we present the crystal structure of a ChR (a C1C2 chimaera between ChR1 and ChR2 from Chlamydomonas reinhardtii) at 2.3 Å resolution. The structure reveals the essential molecular architecture of ChRs, including the retinal-binding pocket and cation conduction pathway. This integration of structural and electrophysiological analyses provides insight into the molecular basis for the remarkable function of ChRs, and paves the way for the precise and principled design of ChR variants with novel properties.


Assuntos
Cátions/metabolismo , Chlamydomonas reinhardtii/química , Ativação do Canal Iônico/efeitos da radiação , Canais Iônicos/química , Luz , Rodopsina/química , Animais , Bacteriorodopsinas/química , Sítios de Ligação , Bovinos , Chlamydomonas reinhardtii/genética , Cristalografia por Raios X , Canais Iônicos/genética , Canais Iônicos/efeitos da radiação , Modelos Moleculares , Mutação , Conformação Proteica , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/efeitos da radiação , Retinaldeído/metabolismo , Rodopsina/genética , Rodopsina/efeitos da radiação , Bases de Schiff/química , Eletricidade Estática
9.
Nature ; 477(7363): 171-8, 2011 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-21796121

RESUMO

Severe behavioural deficits in psychiatric diseases such as autism and schizophrenia have been hypothesized to arise from elevations in the cellular balance of excitation and inhibition (E/I balance) within neural microcircuitry. This hypothesis could unify diverse streams of pathophysiological and genetic evidence, but has not been susceptible to direct testing. Here we design and use several novel optogenetic tools to causally investigate the cellular E/I balance hypothesis in freely moving mammals, and explore the associated circuit physiology. Elevation, but not reduction, of cellular E/I balance within the mouse medial prefrontal cortex was found to elicit a profound impairment in cellular information processing, associated with specific behavioural impairments and increased high-frequency power in the 30-80 Hz range, which have both been observed in clinical conditions in humans. Consistent with the E/I balance hypothesis, compensatory elevation of inhibitory cell excitability partially rescued social deficits caused by E/I balance elevation. These results provide support for the elevated cellular E/I balance hypothesis of severe neuropsychiatric disease-related symptoms.


Assuntos
Modelos Neurológicos , Inibição Neural/fisiologia , Neurônios/metabolismo , Córtex Pré-Frontal/fisiologia , Córtex Pré-Frontal/fisiopatologia , Comportamento Social , Animais , Transtorno Autístico/fisiopatologia , Modelos Animais de Doenças , Células HEK293 , Hipocampo/citologia , Humanos , Aprendizagem , Transtornos Mentais/fisiopatologia , Camundongos , Atividade Motora , Opsinas/metabolismo , Esquizofrenia/fisiopatologia
10.
Nat Methods ; 9(12): 1171-9, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23169303

RESUMO

Optogenetics with microbial opsin genes has enabled high-speed control of genetically specified cell populations in intact tissue. However, it remains a challenge to independently control subsets of cells within the genetically targeted population. Although spatially precise excitation of target molecules can be achieved using two-photon laser-scanning microscopy (TPLSM) hardware, the integration of two-photon excitation with optogenetics has thus far required specialized equipment or scanning and has not yet been widely adopted. Here we take a complementary approach, developing opsins with custom kinetic, expression and spectral properties uniquely suited to scan times typical of the raster approach that is ubiquitous in TPLSMlaboratories. We use a range of culture, slice and mammalian in vivo preparations to demonstrate the versatility of this toolbox, and we quantitatively map parameter space for fast excitation, inhibition and bistable control. Together these advances may help enable broad adoption of integrated optogenetic and TPLSMtechnologies across experimental fields and systems.


Assuntos
Microscopia Confocal/instrumentação , Neurônios/fisiologia , Opsinas/genética , Optogenética , Animais , Células Cultivadas , Desenho de Equipamento , Masculino , Potenciais da Membrana/fisiologia , Camundongos , Fótons , Transfecção
11.
Nature ; 459(7247): 698-702, 2009 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-19396159

RESUMO

Synchronized oscillations and inhibitory interneurons have important and interconnected roles within cortical microcircuits. In particular, interneurons defined by the fast-spiking phenotype and expression of the calcium-binding protein parvalbumin have been suggested to be involved in gamma (30-80 Hz) oscillations, which are hypothesized to enhance information processing. However, because parvalbumin interneurons cannot be selectively controlled, definitive tests of their functional significance in gamma oscillations, and quantitative assessment of the impact of parvalbumin interneurons and gamma oscillations on cortical circuits, have been lacking despite potentially enormous significance (for example, abnormalities in parvalbumin interneurons may underlie altered gamma-frequency synchronization and cognition in schizophrenia and autism). Here we use a panel of optogenetic technologies in mice to selectively modulate multiple distinct circuit elements in neocortex, alone or in combination. We find that inhibiting parvalbumin interneurons suppresses gamma oscillations in vivo, whereas driving these interneurons (even by means of non-rhythmic principal cell activity) is sufficient to generate emergent gamma-frequency rhythmicity. Moreover, gamma-frequency modulation of excitatory input in turn was found to enhance signal transmission in neocortex by reducing circuit noise and amplifying circuit signals, including inputs to parvalbumin interneurons. As demonstrated here, optogenetics opens the door to a new kind of informational analysis of brain function, permitting quantitative delineation of the functional significance of individual elements in the emergent operation and function of intact neural circuitry.


Assuntos
Córtex Cerebral/fisiologia , Interneurônios/fisiologia , Parvalbuminas/fisiologia , Animais , Linhagem Celular , Sincronização Cortical , Eletrofisiologia , Potenciais Evocados , Interneurônios/citologia , Camundongos , Neocórtex/fisiologia , Periodicidade , Estimulação Luminosa , Córtex Pré-Frontal/fisiologia , Transdução de Sinais
12.
Nat Methods ; 9(2): 159-72, 2011 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-22179551

RESUMO

Diverse optogenetic tools have allowed versatile control over neural activity. Many depolarizing and hyperpolarizing tools have now been developed in multiple laboratories and tested across different preparations, presenting opportunities but also making it difficult to draw direct comparisons. This challenge has been compounded by the dependence of performance on parameters such as vector, promoter, expression time, illumination, cell type and many other variables. As a result, it has become increasingly complicated for end users to select the optimal reagents for their experimental needs. For a rapidly growing field, critical figures of merit should be formalized both to establish a framework for further development and so that end users can readily understand how these standardized parameters translate into performance. Here we systematically compared microbial opsins under matched experimental conditions to extract essential principles and identify key parameters for the conduct, design and interpretation of experiments involving optogenetic techniques.


Assuntos
Opsinas/metabolismo , Potenciais de Ação , Animais , Cinética , Luz , Células Piramidais/fisiologia
13.
Nat Commun ; 15(1): 5883, 2024 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-39003286

RESUMO

Rodents continuously move their heads and whiskers in a coordinated manner while perceiving objects through whisker-touch. Studies in head-fixed rodents showed that the ventroposterior medial (VPM) and posterior medial (POm) thalamic nuclei code for whisker kinematics, with POm involvement reduced in awake animals. To examine VPM and POm involvement in coding head and whisker kinematics in awake, head-free conditions, we recorded thalamic neuronal activity and tracked head and whisker movements in male mice exploring an open arena. Using optogenetic tagging, we found that in freely moving mice, both nuclei equally coded whisker kinematics and robustly coded head kinematics. The fraction of neurons coding head kinematics increased after whisker trimming, ruling out whisker-mediated coding. Optogenetic activation of thalamic neurons evoked overt kinematic changes and increased the fraction of neurons leading changes in head kinematics. Our data suggest that VPM and POm integrate head and whisker information and can influence head kinematics during tactile perception.


Assuntos
Neurônios , Optogenética , Vibrissas , Animais , Vibrissas/fisiologia , Masculino , Neurônios/fisiologia , Camundongos , Fenômenos Biomecânicos , Movimentos da Cabeça/fisiologia , Cabeça/fisiologia , Camundongos Endogâmicos C57BL , Percepção do Tato/fisiologia , Tálamo/fisiologia , Tálamo/citologia
14.
Curr Opin Neurobiol ; 86: 102879, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38692167

RESUMO

Although aggression is associated with several psychiatric disorders, there is no effective treatment nor a rigorous definition for "pathological aggression". Mice make a valuable model for studying aggression. They have a dynamic social structure that depends on the habitat and includes reciprocal interactions between the mice's aggression levels, social dominance hierarchy (SDH), and resource allocation. Nevertheless, the classical behavioral tests for territorial aggression and SDH in mice are reductive and have limited ethological and translational relevance. Recent work has explored the use of semi-natural environments to simultaneously study dominance-related behaviors, resource allocation, and aggressive behavior. Semi-natural setups allow experimental control of the environment combined with manipulations of neural activity. We argue that these setups can help bridge the translational gap in aggression research toward discovering neuronal mechanisms underlying maladaptive aggression.


Assuntos
Agressão , Predomínio Social , Animais , Agressão/fisiologia , Camundongos , Comportamento Animal/fisiologia , Humanos , Etologia/métodos
15.
Nat Commun ; 14(1): 1667, 2023 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-36966143

RESUMO

The medial prefrontal cortex (mPFC) mediates a variety of complex cognitive functions via its vast and diverse connections with cortical and subcortical structures. Understanding the patterns of synaptic connectivity that comprise the mPFC local network is crucial for deciphering how this circuit processes information and relays it to downstream structures. To elucidate the synaptic organization of the mPFC, we developed a high-throughput optogenetic method for mapping large-scale functional synaptic connectivity in acute brain slices. We show that in male mice, mPFC neurons that project to the basolateral amygdala (BLA) display unique spatial patterns of local-circuit synaptic connectivity, which distinguish them from the general mPFC cell population. When considering synaptic connections between pairs of mPFC neurons, the intrinsic properties of the postsynaptic cell and the anatomical positions of both cells jointly account for ~7.5% of the variation in the probability of connection. Moreover, anatomical distance and laminar position explain most of this fraction in variation. Our findings reveal the factors determining connectivity in the mPFC and delineate the architecture of synaptic connections in the BLA-projecting subnetwork.


Assuntos
Tonsila do Cerebelo , Complexo Nuclear Basolateral da Amígdala , Camundongos , Masculino , Animais , Vias Neurais/fisiologia , Tonsila do Cerebelo/fisiologia , Córtex Pré-Frontal/fisiologia , Neurônios/fisiologia
16.
Res Sq ; 2023 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-36798372

RESUMO

In the classical model of the basal ganglia, direct pathway striatal projection neurons (dSPNs) send projections to the substantia nigra (SNr) and entopeduncular nucleus to regulate motor function. Recent studies have re-established that dSPNs also possess "bridging" collaterals within the globus pallidus (GPe), yet the significance of these collaterals for behavior is unknown. Here we use in vivo optical and chemogenetic tools combined with deep learning approaches to dissect the roles of bridging collaterals in motor function. We find that dSPNs projecting to the SNr send synchronous motor-related information to the GPe via axon collaterals. Inhibition of native activity in dSPN GPe terminals impairs motor activity and function via regulation of pallidostriatal Npas1 neurons. We propose a model by which dSPN GPe collaterals ("striatopallidal Go pathway") act in concert with the canonical terminals in the SNr to support motor control by inhibiting Npas1 signals going back to the striatum.

17.
Nat Commun ; 14(1): 6712, 2023 10 23.
Artigo em Inglês | MEDLINE | ID: mdl-37872145

RESUMO

In the classical model of the basal ganglia, direct pathway striatal projection neurons (dSPNs) send projections to the substantia nigra (SNr) and entopeduncular nucleus to regulate motor function. Recent studies have re-established that dSPNs also possess axon collaterals within the globus pallidus (GPe) (bridging collaterals), yet the significance of these collaterals for behavior is unknown. Here we use in vivo optical and chemogenetic tools combined with deep learning approaches in mice to dissect the roles of dSPN GPe collaterals in motor function. We find that dSPNs projecting to the SNr send synchronous motor-related information to the GPe via axon collaterals. Inhibition of native activity in dSPN GPe terminals impairs motor activity and function via regulation of Npas1 neurons. We propose a model by which dSPN GPe axon collaterals (striatopallidal Go pathway) act in concert with the canonical terminals in the SNr to support motor control by inhibiting Npas1 neurons.


Assuntos
Axônios , Neurônios , Camundongos , Animais , Neurônios/metabolismo , Axônios/metabolismo , Globo Pálido/fisiologia , Corpo Estriado/metabolismo , Gânglios da Base/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo
18.
bioRxiv ; 2023 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-37425961

RESUMO

Information is transmitted between brain regions through the release of neurotransmitters from long-range projecting axons. Understanding how the activity of such long-range connections contributes to behavior requires efficient methods for reversibly manipulating their function. Chemogenetic and optogenetic tools, acting through endogenous G-protein coupled receptor (GPCRs) pathways, can be used to modulate synaptic transmission, but existing tools are limited in sensitivity, spatiotemporal precision, or spectral multiplexing capabilities. Here we systematically evaluated multiple bistable opsins for optogenetic applications and found that the Platynereis dumerilii ciliary opsin (PdCO) is an efficient, versatile, light-activated bistable GPCR that can suppress synaptic transmission in mammalian neurons with high temporal precision in-vivo. PdCO has superior biophysical properties that enable spectral multiplexing with other optogenetic actuators and reporters. We demonstrate that PdCO can be used to conduct reversible loss-of-function experiments in long-range projections of behaving animals, thereby enabling detailed synapse-specific functional circuit mapping.

19.
J Cell Sci ; 123(Pt 11): 1940-7, 2010 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-20484665

RESUMO

Regulation of exocytosis by voltage-gated K(+) channels has classically been viewed as inhibition mediated by K(+) fluxes. We recently identified a new role for Kv2.1 in facilitating vesicle release from neuroendocrine cells, which is independent of K(+) flux. Here, we show that Kv2.1-induced facilitation of release is not restricted to neuroendocrine cells, but also occurs in the somatic-vesicle release from dorsal-root-ganglion neurons and is mediated by direct association of Kv2.1 with syntaxin. We further show in adrenal chromaffin cells that facilitation induced by both wild-type and non-conducting mutant Kv2.1 channels in response to long stimulation persists during successive stimulation, and can be attributed to an increased number of exocytotic events and not to changes in single-spike kinetics. Moreover, rigorous analysis of the pools of released vesicles reveals that Kv2.1 enhances the rate of vesicle recruitment during stimulation with high Ca(2+), without affecting the size of the readily releasable vesicle pool. These findings place a voltage-gated K(+) channel among the syntaxin-binding proteins that directly regulate pre-fusion steps in exocytosis.


Assuntos
Células Cromafins/metabolismo , Exocitose , Gânglios Espinais/patologia , Neurônios/metabolismo , Vesículas Secretórias/metabolismo , Canais de Potássio Shab/metabolismo , Animais , Animais Recém-Nascidos , Sinalização do Cálcio , Células Cultivadas , Células Cromafins/patologia , Eletrofisiologia , Neurônios/patologia , Proteínas Qa-SNARE/metabolismo , Ratos , Ratos Wistar , Canais de Potássio Shab/genética
20.
Nat Neurosci ; 11(6): 631-3, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18432196

RESUMO

The introduction of two microbial opsin-based tools, channelrhodopsin-2 (ChR2) and halorhodopsin (NpHR), to neuroscience has generated interest in fast, multimodal, cell type-specific neural circuit control. Here we describe a cation-conducting channelrhodopsin (VChR1) from Volvox carteri that can drive spiking at 589 nm, with excitation maximum red-shifted approximately 70 nm compared with ChR2. These results demonstrate fast photostimulation with yellow light, thereby defining a functionally distinct third category of microbial rhodopsin proteins.


Assuntos
Proteínas de Transporte/fisiologia , Cor , Neurônios/fisiologia , Estimulação Luminosa/métodos , Volvox/química , Animais , Animais Recém-Nascidos , Proteínas de Transporte/genética , Células Cultivadas , Relação Dose-Resposta a Droga , Estimulação Elétrica , Halorrodopsinas/fisiologia , Hipocampo/citologia , Humanos , Canais Iônicos , Luz , Potenciais da Membrana/fisiologia , Potenciais da Membrana/efeitos da radiação , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley , Transfecção , Xenopus laevis
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