Dataset Information

Efficient gene transfers into neocortical neurons connected by NMDA NR1-containing synapses.

ABSTRACT: BACKGROUND:Within a circuit, specific neurons and synapses are hypothesized to have essential roles in circuit physiology and learning, and dysfunction in these neurons and synapses causes specific disorders. These critical neurons and synapses are embedded in complex circuits containing many neuron and synapse types. NEW METHOD:We established technology that can deliver different genes into pre- and post-synaptic neurons connected by a specific synapse type. The first, presynaptic gene transfer employs standard gene transfer technology to express a synthetic peptide neurotransmitter which has three domains, a dense core vesicle sorting domain for processing the protein as a peptide neurotransmitter, a receptor-binding domain, here a small peptide that binds to NMDA NR1 subunits, and the His tag. Upon release, this peptide neurotransmitter binds to its cognate receptor on postsynaptic neurons. Gene transfer selectively into these postsynaptic neurons employs antibody-mediated, targeted gene transfer and anti-His tag antibodies, which recognize the His tag domain in the synthetic peptide neurotransmitter. RESULTS:For the model system, we studied the connection from projection neurons in postrhinal cortex to specific neurons in perirhinal cortex. In our initial report, gene transfer to connected neurons was 20+1% specific. Here, we optimized the technology; we improved the transfection for packaging by using a modern using a modern lipid, Lipofectamine 3000, and used a modern confocal microscope to collect data. We now report 80+2% specific gene transfer to connected neurons. COMPARISON WITH EXISTING METHODS:There is no existing method with this capability. CONCLUSIONS:This technology may enable studies on the roles of specific neurons and synapses in circuit physiology and learning, and support gene therapy treatments for specific disorders.

SUBMITTER: Nagayach A

PROVIDER: S-EPMC6760849 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Efficient gene transfers into neocortical neurons connected by NMDA NR1-containing synapses.

Nagayach Aarti A Singh Anshuman A Geller Alfred I AI

Journal of neuroscience methods 20190809

<h4>Background</h4>Within a circuit, specific neurons and synapses are hypothesized to have essential roles in circuit physiology and learning, and dysfunction in these neurons and synapses causes specific disorders. These critical neurons and synapses are embedded in complex circuits containing many neuron and synapse types.<h4>New method</h4>We established technology that can deliver different genes into pre- and post-synaptic neurons connected by a specific synapse type. The first, presynapti ...[more]

PMID: 31404560

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Project description:Local neocortical circuits play critical roles in information processing, including synaptic plasticity, circuit physiology, and learning, and GABAergic inhibitory interneurons have key roles in these circuits. Moreover, specific neurological disorders, including schizophrenia and autism, are associated with deficits in GABAergic transmission in these circuits. GABAergic synapses represent a small fraction of neocortical synapses, and are embedded in complex local circuits that contain many neuron and synapse types. Thus, it is challenging to study the physiological roles of GABAergic inhibitory interneurons and their synapses, and to develop treatments for the specific disorders caused by dysfunction at these GABAergic synapses. To these ends, we report a novel technology that can deliver different genes into pre- and post-synaptic neocortical interneurons connected by a GABAergic synapse: First, standard gene transfer into the presynaptic neurons delivers a synthetic peptide neurotransmitter, containing three domains, a dense core vesicle sorting domain, a GABAA receptor-binding domain, a single-chain variable fragment anti-GABAA ß2 or ß3, and the His tag. Second, upon release, this synthetic peptide neurotransmitter binds to GABAA receptors on the postsynaptic neurons. Third, as the synthetic peptide neurotransmitter contains the His tag, antibody-mediated, targeted gene transfer using anti-His tag antibodies is selective for these neurons. We established this technology by expressing the synthetic peptide neurotransmitter in GABAergic neurons in the middle layers of postrhinal cortex, and the delivering the postsynaptic vector into connected GABAergic neurons in the upper neocortical layers. Targeted gene transfer was 61% specific for the connected neurons, but untargeted gene transfer was only 21% specific for these neurons. This technology may support studies on the roles of GABAergic inhibitory interneurons in circuit physiology and learning, and support gene therapy treatments for specific disorders associated with deficits at GABAergic synapses.

Dataset Information

Efficient gene transfers into neocortical neurons connected by NMDA NR1-containing synapses.

Publications

Efficient gene transfers into neocortical neurons connected by NMDA NR1-containing synapses.

Similar Datasets

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