Reversible Suppression of Glutamatergic Neurotransmission of Cerebellar Granule Cells<i>In Vivo</i>by Genetically Manipulated Expression of Tetanus Neurotoxin Light Chain

Yamamoto, Misaki; Wada, Norio; Kitabatake, Yasuji; Watanabe, Dai; Anzai, Masayuki; Yokoyama, Minesuke; Teranishi, Yutaka; Nakanishi, Shigetada

doi:10.1523/jneurosci.23-17-06759.2003

Cited by 139 publications

(122 citation statements)

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“…In the RNB transgenic mice, the tetanus toxin light chain is restrictedly expressed in granule cells under the control of a tetracycline-controlled reverse transactivator (rtTA) (29). Tetanus toxin specifically cleaves synaptic vesicle VAMP2 (30), resulting in blockade of transmitter release from synaptic vesicles (Fig.…”

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“…Tetanus toxin specifically cleaves synaptic vesicle VAMP2 (30), resulting in blockade of transmitter release from synaptic vesicles (Fig. 1b) (29). Consequently, granule cell inputs to Purkinje cells are blocked and reversibly recovered by administration and omission, respectively, of doxycycline (DOX) (29).…”

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“…1b) (29). Consequently, granule cell inputs to Purkinje cells are blocked and reversibly recovered by administration and omission, respectively, of doxycycline (DOX) (29). These RNB mice were thus used based on the following rationale: when granule cell inputs to Purkinje cells are selectively blocked, the CS signal is not transmitted to Purkinje cells but still conveyed to the interpositus nucleus via the direct mossy fiber pathway (Fig.…”

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Conditioned eyeblink learning is formed and stored without cerebellar granule cell transmission

Wada

Kishimoto

Watanabe

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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), the authors note that in Fig. 2a, the same trace was inadvertently used for the electrophysiological records of RNB (DOXuntreated) and RNB (DOX-withdrawn). This error does not affect the conclusions of the article. The corrected figure and its legend appear below. (e and f ) Mean Ϯ SEM of firing rates of simple spikes (e) and complex spikes ( f) are shown with columns and bars, respectively. *** , P Ͻ 0.001 (comparison between DOX-treated RNB mice and all five other animals by the Scheffé test). (g) The extracellular recording site (red) was confirmed at the Purkinje cell layer by fluorescent Nissl staining. ML, molecular layer; PL, Purkinje cell layer; GL, granule cell layer. eyeblink conditioning ͉ interpositus nucleus ͉ Purkinje cell ͉ reversible neurotransmission blockade ͉ synaptic plasticity C lassical conditioning of the eyeblink reflex is elicited by paired presentation of conditioned stimulus (CS) with reinforcing unconditioned stimulus (US) (1-4). This associative motor learning is a basic form of cerebellum-dependent leaning. In eyeblink conditioning, the CS pathway includes the pontine nucleus and mossy fibers, whereas the US pathway includes the inferior olive and the climbing fibers (1-5). The mossy fibers project directly to the interpositus nucleus and indirectly to the cerebellar cortex via granule cells (1-5). The CS and US signals are thus conveyed to both the cerebellar cortex and the interpositus nucleus (Fig. 1a). The localization and underlying mechanisms of eyeblink conditioning have been extensively studied by different approaches including gene targeting (6-11), lesioning (12-15), mutant analysis (16), and pharmacological inactivation analyses (17-23). On the basis of these studies, one model proposes an essential role of the cerebellar Purkinje cell circuit in memory traces (24)(25)(26). According to this model, convergence of the CS and US signals induces long-term depression at the parallel fiber-Purkinje cell synapses. Because Purkinje cells are inhibitory, long-term depression causes a disinhibition of the interpositus nucleus and an increased input in the downstream motor pathways. The other model proposes that the memory trace is located in the interpositus nucleus and that conditioned responses (CRs) are properly evoked in response to CS by a timing signal from Purkinje cells (1,3,27,28). Eyeblink conditioning involves multiple learning processes, at least acquisition, expression, and storage of motor learning. Once the expression process of memory traces is impaired, it becomes difficult to define the key neural circuits responsible for expression, formation, and storage of memory traces.In this investigation, we adopted a novel reversible neurotransmission blocking (RNB) technique to explore the role of Purkinje cells and the interpositus nucleus in associative eyeblink conditioning. In the RNB transgenic mice, the tetanus toxin light chain is restrictedly expressed in granule cells under the control of a tetracycline-controlled reverse transactivator (rtTA) (29). Tetanus t...

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Conditioned eyeblink learning is formed and stored without cerebellar granule cell transmission

Wada

Kishimoto

Watanabe

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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“…The tetanus toxin light chain (TeTxLC) gene has been shown to inhibit the neuronal function in transgenic f lies and mice (3,18). To examine whether TeTxLC can inhibit neuronal function in zebrafish, we constructed a transgenic fish line that harbored a single-copy insertion of the TeTxLC gene downstream of 5xUAS (UAS:TeTxLC) (Fig.…”

Section: Gal4ff-mediated Tetxlc Expression Can Inhibit Neuronal Functmentioning

confidence: 99%

Genetic dissection of neural circuits by Tol2 transposon-mediated Gal4 gene and enhancer trapping in zebrafish

Asakawa

Suster

Mizusawa

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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Targeted gene expression is a powerful approach to study the function of genes and cells in vivo. In Drosophila, the P elementmediated Gal4-UAS method has been successfully used for this purpose. However, similar methods have not been established in vertebrates. Here we report the development of a targeted gene expression methodology in zebrafish based on the Tol2 transposable element and its application to the functional study of neural circuits. First, we developed gene trap and enhancer trap constructs carrying an engineered yeast Gal4 transcription activator (Gal4FF) and transgenic reporter fish carrying the GFP or the RFP gene downstream of the Gal4 recognition sequence (UAS) and showed that the Gal4FF can activate transcription through UAS in zebrafish. Second, by using this Gal4FF-UAS system, we performed large-scale screens and generated a large collection of fish lines that expressed Gal4FF in specific tissues, cells, and organs. Finally, we developed transgenic effector fish carrying the tetanus toxin light chain (TeTxLC) gene downstream of UAS, which is known to block synaptic transmission. We crossed the Gal4FF fish with the UAS:TeTxLC fish and analyzed double transgenic embryos for defects in touch response. From this analysis, we discovered that targeted expression of TeTxLC in distinct populations of neurons in the brain and the spinal cord caused distinct abnormalities in the touch response behavior. These studies illustrate that our Gal4FF gene trap and enhancer trap methods should be an important resource for genetic analysis of neuronal functions and behavior in vertebrates.targeted gene expression ͉ Gal4-UAS ͉ tetanus toxin ͉ touch response ͉ interneuron

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“…The circuit is inactivated through blocking neurotransmission. This system has been used successfully to inactivate neurons and circuits selectively and reversibly [41,42,163,164]. For instance, in order to test the roles of the circuit between propriospinal neurons and spinal motor neurons in dexterous hand movement of primates, AAV encoding reverse tetracycline transactivator was injected into the origin of the circuit where the propriospinal neurons are located, and retrograde gene transfer lentivirus carrying TeNT under the control of tetracycline responsive element was injected into the termination of the circuit where the spinal motor neurons reside.…”

Section: Inducible Tetanus Neurotoxinmentioning

confidence: 99%

Selective Manipulation of Neural Circuits

Park

Carmel

2016

Neurotherapeutics

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Unraveling the complex network of neural circuits that form the nervous system demands tools that can manipulate specific circuits. The recent evolution of genetic tools to target neural circuits allows an unprecedented precision in elucidating their function. Here we describe two general approaches for achieving circuit specificity. The first uses the genetic identity of a cell, such as a transcription factor unique to a circuit, to drive expression of a molecule that can manipulate cell function. The second uses the spatial connectivity of a circuit to achieve specificity: one genetic element is introduced at the origin of a circuit and the other at its termination. When the two genetic elements combine within a neuron, they can alter its function. These two general approaches can be combined to allow manipulation of neurons with a specific genetic identity by introducing a regulatory gene into the origin or termination of the circuit. We consider the advantages and disadvantages of both these general approaches with regard to specificity and efficacy of the manipulations. We also review the genetic techniques that allow gain-and loss-of-function within specific neural circuits. These approaches introduce light-sensitive channels (optogenetic) or drug sensitive channels (chemogenetic) into neurons that form specific circuits. We compare these tools with others developed for circuit-specific manipulation and describe the advantages of each. Finally, we discuss how these tools might be applied for identification of the neural circuits that mediate behavior and for repair of neural connections.

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Reversible Suppression of Glutamatergic Neurotransmission of Cerebellar Granule CellsIn Vivoby Genetically Manipulated Expression of Tetanus Neurotoxin Light Chain

Cited by 139 publications

References 62 publications

Conditioned eyeblink learning is formed and stored without cerebellar granule cell transmission

Conditioned eyeblink learning is formed and stored without cerebellar granule cell transmission

Genetic dissection of neural circuits by Tol2 transposon-mediated Gal4 gene and enhancer trapping in zebrafish

Selective Manipulation of Neural Circuits

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