Neural circuit-specific gene manipulation in mouse brain in vivo using split-intein-mediated split-Cre system

Kim, Yong-Eun; Kim, Sunwhi; Kim, Il Hwan

doi:10.1016/j.xpro.2022.101807

Cited by 5 publications

(6 citation statements)

References 20 publications

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“…These include rAAV2/1-hSyn- Cre , rAAV2.retro-hSyn- Cre , rAAV2/1-Ef1α- CreN-InteinN , rAAV2.retro-Ef1α- InteinC-CreC , rAAV2/8-Ef1α- CreN-InteinN , and rAAV2/9-Syn -Flex-jGCaMP8m . The production and subsequent titer measurement of these rAAVs were based on protocols from our previous studies 38, 60 , with necessary adjustments. Notably, to produce rAAV2/1-Ef1α- CreN-InteinN and rAAV2/1-hSyn- Cre , we utilized the pAAV2/1 packaging plasmid (Addgene #112862), diverging from our previous use of the pAAV 2/8 or 2/9 plasmids.…”

Section: Methodsmentioning

confidence: 99%

“…Stereotaxic surgeries for viral injections were conducted in accordance with the protocols established in our previous studies 37, 38 . Briefly, mice aged 8-10 weeks were anesthetized using an isoflurane vaporizer (VetEquip).…”

Section: Methodsmentioning

confidence: 99%

“…1e). This technique is an improvement of our previously established strategy for identifying single long-range neural circuits [36][37][38][39] (see Extended Fig. 3 and Video 3).…”

Section: Comparison Of Mnm Approach With Conventional Raav Tracing Me...mentioning

confidence: 99%

“…Here we introduce the Monosynaptically-interconnected Neural Network Module (MNM) approach, a novel technique developed from our previously established strategy for selecting single long-range neural circuits [36][37][38][39] . The 'MNM' concept refines the broader term 'module' in network neuroscience, offering more precise identification of monosynaptically-interconnected distant brain regions via axons.…”

Section: Introductionmentioning

confidence: 99%

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Monosynaptically-interconnected Network Module (MNM) Approach for High-Resolution Brain Sub-Network Analysis

Kim,

Ujihara

et al. 2024

Preprint

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We introduce the Monosynaptically-interconnected Network Module (MNM) approach, an innovative method designed for efficiently analyzing the anatomical structure and functional dynamics of specific brain network modulesin vivo. Utilizing an Intein-mediated split-Cre system combined with bidirectional adeno-associated viruses, this technique precisely targets and manipulates monosynaptically interconnected modular subnetworks in freely moving animals. We demonstrate its utility through anatomical and functional mapping of a specific MNM encompassing the prefrontal cortex (PFC), basolateral amygdala (BLA), and intermediary hub regions. Specifically, the MNM approach with Cre-reporter mice visualizes detailed network architecture and enables the tracing of axonal connections among the nodes in the network. Furthermore, integration of the MNM approach with Cre-dependent Ca2+indicator and multi-fiber photometry in freely moving mice reveals enhanced correlative network activities in social contexts. This versatile technique offers significant potential for advancing our understanding of network functions that underlie complex behaviors, providing a modular network perspective.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…1e). This technique is an improvement of our previously established strategy for identifying single long-range neural circuits [36][37][38][39] (see Extended Fig. 3 and Video 3).…”

Section: Comparison Of Mnm Approach With Conventional Raav Tracing Me...mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Monosynaptically-interconnected Network Module (MNM) Approach for High-Resolution Brain Sub-Network Analysis

Kim,

Ujihara

et al. 2024

Preprint

Self Cite

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“…To produce AAV-PHP.v1-CLDN5-GSK3β S9A -HA and AAV-PHP.v1-CLDN5-HA, HEK293T cells were co-transfected with pAdDeltaF6, pUCmini-iCAP-PHP.V1 plasmid, and AAV-CLDN5-GSK3β S9A -HA or AAV-CLDN5-HA plasmid expressing GSK3β S9A -HA or HA under control of CLDN5 promoter. AAV puri cation was performed as previously described 23,24,60,61 . After 72 h incubation, HEK293T cells were collected and lysed with lysis buffer [20 mM Tris-HCl (pH8.5), 150 mM NaCl] through four times of freeze-thaw process in dry ice/ethanol bath and 37 °C water bath.…”

Section: Aav Production and Titrationmentioning

confidence: 99%

Endothelial SHANK3 Controls Tight Junction of Neonatal Blood-Brain Barrier

Kim,

Kim

et al. 2024

Preprint

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Autism spectrum disorder (ASD) is a neurodevelopmental disability condition arising from the combination of genetic and environmental factors. Despite the blood-brain barrier (BBB) serving as a crucial gatekeeper, conveying environmental influences into the brain parenchyma, the contributions of BBB in ASD pathogenesis remain largely uncharted. Here we report that a novel form of SHANK3, a gene strongly associated with Phelan-McDermid syndrome and ASD2, expresses in the BBB-forming brain endothelial cells (BECs) and regulates tight junctional (TJ) integrity essential for maintaining the BBB’s barrier function. Loss of Shank3 disrupts the TJ of BECs, leading to increased paracellular passage through endothelial linings. Neonates with endothelium-specific SHANK3 knockout exhibit increased BBB permeability, reduced neuronal excitability, and impaired ultra-sonic communications. Surprisingly, while BBB permeability is restored during adult age, the mutant mice continue to display reduced neuronal excitability, impaired sociability, and repetitive responses. Further investigation reveals that the hyperpermeability resulting from TJ disruption is attributed to the β-Catenin imbalance triggered by SHANK3 knockout, which is rescued by normalizing β-Catenin signaling in vitro and in vivo. These findings highlight a previously unknown pathogenic mechanism stemming from the ASD-risk SHANK3, emphasizing the significance of neonatal BECs in the BBB as a potential therapeutic target of ASD

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Adamtsl3 mediates DCC signaling to selectively promote GABAergic synapse function

Cramer,

Pinan-Lucarre,

Cavaccini

et al. 2023

Cell Reports

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Neural circuit-specific gene manipulation in mouse brain in vivo using split-intein-mediated split-Cre system

Cited by 5 publications

References 20 publications

Monosynaptically-interconnected Network Module (MNM) Approach for High-Resolution Brain Sub-Network Analysis

Monosynaptically-interconnected Network Module (MNM) Approach for High-Resolution Brain Sub-Network Analysis

Endothelial SHANK3 Controls Tight Junction of Neonatal Blood-Brain Barrier

Adamtsl3 mediates DCC signaling to selectively promote GABAergic synapse function

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