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Abstract The leucine‐rich repeat‐containing family 8 member A (LRRC8A) is an essential subunit of the volume‐regulated anion channel (VRAC). VRAC is critical for cell volume control, but its broader physiological functions remain under investigation. Recent studies in the field indicate thatLrrc8adisruption in the brain astrocytes reduces neuronal excitability, impairs synaptic plasticity and memory, and protects against cerebral ischemia. In the present work, we generated brain‐wide conditional LRRC8A knockout mice (LRRC8A bKO) usingNestinCre‐drivenLrrc8aflox/floxexcision in neurons, astrocytes, and oligodendroglia. LRRC8A bKO animals were born close to the expected Mendelian ratio and developed without overt histological abnormalities, but, surprisingly, all died between 5 and 9 weeks of age with a seizure phenotype, which was confirmed by video and EEG recordings. Brain slice electrophysiology detected changes in the excitability of pyramidal cells and modified GABAergic inputs in the hippocampal CA1 region of LRRC8A bKO. LRRC8A‐null hippocampi showed increased immunoreactivity of the astrocytic marker GFAP, indicating reactive astrogliosis. We also found decreased whole‐brain protein levels of the GABA transporter GAT‐1, the glutamate transporter GLT‐1, and the astrocytic enzyme glutamine synthetase. Complementary HPLC assays identified reduction in the tissue levels of the glutamate and GABA precursor glutamine. Together, these findings suggest that VRAC provides vital control of brain excitability in mouse adolescence. VRAC deletion leads to a lethal phenotype involving progressive astrogliosis and dysregulation of astrocytic uptake and supply of amino acid neurotransmitters and their precursors.
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This content will become publicly available on January 28, 2026
Light-activated tetanus neurotoxin for conditional proteolysis and inducible synaptic inhibition in vivo
Abstract The light chain of tetanus neurotoxin (TeNT) is a 52 kD metalloprotease that potently inhibits synaptic transmission by cleaving the endogenous vesicle fusion protein VAMP2. To mitigate the toxicity of TeNT and harness it as a conditional tool for neuroscience, we engineered Light-Activated TeNT (LATeNT) via insertion of the light-sensitive LOV domain into an allosteric site. LATeNT was optimized by directed evolution and shown to have undetectable activity in the dark mammalian brain. Following 30 seconds of weak blue light exposure, however, LATeNT potently inhibited synaptic transmission in multiple brain regions. The effect could be reversed over 24 hours. We used LATeNT to discover an interneuron population in hippocampus that controls anxiety-like behaviors in mouse, and to control the secretion of endogenous insulin from pancreatic beta cells. Synthetic circuits incorporating LATeNT converted drug, Ca2+, or receptor activation into transgene expression or reporter protein secretion. Due to its large dynamic range, rapid kinetics, and highly specific mechanism of action, LATeNT should be a robust tool for conditional proteolysis and spatiotemporal control of synaptic transmissionin vivo.
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- Award ID(s):
- 2014862
- PAR ID:
- 10623742
- Publisher / Repository:
- bioRxiv
- Date Published:
- Format(s):
- Medium: X
- Institution:
- bioRxiv
- Sponsoring Org:
- National Science Foundation
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