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iGABASnFR2
Improved GABA (γ-aminobutyric acid) Indicators for Synaptic Imaging
iGABASnFR2 is the current (2022) generation of GABA-Sensing Fluorescent Reporters developed at HHMI's Janelia Research Campus. Other sensor technologies such as SF-iGluSnFR, jGCaMP8, and jRGECO are optimized for excitatory synaptic transmission and action potentials. iGABASnFr complements these technologies with the ability to detect inhibitory synaptic transmission and inhibitory post-synaptic currents. iGABASnFr2 has the best performance among GABA indicators and is completely genetically coded, which is optimal for in vivo use. See the post for additional data.
Fig. 1(a) Schematic representation of iGABASnFR2 function.
Aberrations in GABAergic signaling occur in Alzheimer’s disease, Parkinson’s, Huntington’s, schizophrenia, autism and Rett syndrome. The imaging of iGABASnFR in animal models will lead to a greater understanding of these mechanisms. iGABASnFR could also allow easy screening of candidate transporters such as the vertebrate mitochondrial GABA transporter.
The GENIE Project Team at HHMI Janelia developed these second-generation genetically encoded GABA indicators in collaboration with Jonathan Marvin and Loren Looger. The sensor is based on iGABASnFR and has the following extensively-tested features:
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Positive-going version (iGABASnFR2) and negative-going version (iGABASnFR2n)
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iGABASnFR2: 4x higher sensitivity to APs than iGABASnFR, iGABASnFR2n: 2x higher sensitivity to APs than iGABASnFR.
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Both enable single-AP high-speed imaging
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Higher affinity for GABA (iGABASnFR2: EC50: 6.4±0.2 μM, ~7x tighter than iGABASnFR, ΔF/Fmax=0.5±0.05, ~2x higher than iGABASnFR) -
Imaged with standard GFP filter set
Fig. 2. iGABASnFR2 and 2n performance against previous generation iGABASnFR
Availability
Plasmids/AAVs available from Addgene:
Positive-going (dim to bright with GABA binding)
| Name | ID | Plasmid | AAV |
|---|---|---|---|
| pGP-CAG-iGABASnFR2-WPRE-bGH-polyA | 514.4409 | 218868 | - |
| pGP-AAV-GFAP-iGABASnFR2-WPRE | 608.4409 | 218871 | AAV1/AAV5 |
| pGP-AAV-syn-iGABASnFR2-WPRE | 609.4409 | 218874 | AAV1/AAV5 |
| pGP-AAV-syn-flex-iGABASnFR2-WPRE | 610.4409 | 218877 | AAV1/AAV5 |
| pGP-AAV-CAG-flex-iGABASnFR2-WPRE | 616.4409 | 218880 | * |
| pGP-AAV-CAG-iGABASnFR2-WPRE | 640.4409 | 218883 | * |
| pGP-RSET-iGABASnFR2-6xHis | 602.10 | 218885 | - |
Negative-going (bright to dim with GABA binding)
| Name | ID | Plasmid | AAV |
|---|---|---|---|
| pGP-CAG-iGABASnFR2n-WPRE-bGH-polyA | 514.4445 | 218869 | - |
| pGP-AAV-GFAP-iGABASnFR2n-WPRE | 608.4445 | 218872 | * |
| pGP-AAV-syn-iGABASnFR2n-WPRE | 609.4445 | 218875 | * |
| pGP-AAV-syn-flex-iGABASnFR2n-WPRE | 610.4445 | 218878 | * |
| pGP-AAV-CAG-flex-iGABASnFR2n-WPRE | 616.4445 | 218881 | * |
| pGP-AAV-CAG-iGABASnFR2n-WPRE | 640.4445 | 218884 | * |
| pGP-RSET-iGABASnFR2n-6xHis | 602.4445 | 218886 | - |
Non-binding controls (no fluorescence change with GABA)
| Name | ID | Plasmid | AAV |
|---|---|---|---|
| pGP-CAG-iGABASnFR2(no bind)-WPRE-bGH-polyA | 514.4444 | 218870 | - |
| pGP-AAV-GFAP-iGABASnFR2(no bind)-WPRE | 608.4444 | 218873 | AAV1/AAV5 |
| pGP-AAV-syn-iGABASnFR2(no bind)-WPRE | 609.4444 | 218876 | AAV1/AAV5 |
| pGP-AAV-syn-flex-iGABASnFR2(no bind)-WPRE | 610.4444 | 218879 | AAV1/AAV5 |
| pGP-AAV-CAG-flex-iGABASnFR2(no bind)-WPRE | 616.4444 | 218882 | * |
| pGP-RSET-iGABASnFR2(no bind)-6xHis | 602.4444 | 218887 | - |
* Request Addgene to make AAV
Opportunity
For non-profit/academic reagent requests and more information, please contact GENIEreagents@janelia.hhmi.org. For commercial licenses (internal research or commercial sale), please contact innovation@janelia.hhmi.org.
Janelia Tech ID
2021-020
Patent Protection
US Patent Application Publication US20220050103
