Selective and Quickly Reversible Inactivation of Mammalian Neurons In Vivo Using the Drosophila Allatostatin Receptor

Abstract

Genetic strategies for perturbing activity of selected neurons hold great promise for understanding circuitry and behavior. Several such strategies exist, but there has been no direct demonstration of reversible inactivation of mammalian neurons in vivo. We previously reported quickly reversible inactivation of neurons in vitro using expression of the Drosophila allatostatin receptor (AlstR). Here, adeno-associated viral vectors are used to express AlstR in vivo in cortical and thalamic neurons of rats, ferrets, and monkeys. Application of the receptor's ligand, allatostatin (AL), leads to a dramatic reduction in neural activity, including responses of visual neurons to optimized visual stimuli. Additionally, AL eliminates activity in spinal cords of transgenic mice conditionally expressing AlstR. This reduction occurs selectively in AlstR-expressing neurons. Inactivation can be reversed within minutes upon washout of the ligand and is repeatable, demonstrating that the AlstR/AL system is effective for selective, quick, and reversible silencing of mammalian neurons in vivo.

A Caenorhabditis elegans allatostatin/galanin-like receptor NPR-9 inhibits local search behavior in response to feeding cues

“…In this system the allatostatin receptor (9) and glutamate-gated chloride channels (28) function as inhibitors and the ionotropic glutamate receptors function as stimulators (29). In addition, the Drosophila allatostatin receptor can reversibly inactivate mammalian neurons in vivo (30). In the absence of food, npr-9 mutants mimic the locomotory behavior of wild-type N2 animals.…”

Inducible and titratable silencing ofCaenorhabditis elegansneurons in vivo with histamine-gated chloride channels

“…Several chemical genetic systems have been developed based on G protein-coupled receptor (GPCR) transgenes that open endogenous potassium channels in response to synthetic or biologically nonnative ligands (9,10). Inhibitory receptor−ligand pairs have also been developed to imitate GABA, which opens ligand-gated chloride channels.…”

Rapidly inducible, genetically targeted inactivation of neural and synaptic activity in vivo

“…Similarly, in slices of the spinal cord from mice expressing AlstR in a subpopulation of spinal cord inhibitory neurons, reversible suppression of excitability was observed in the presence of allatostatin (10 nM) with a time course of approximately 20 min for both induction and reversal [15 ]. Tan et al used this system successfully in rats, ferrets, and monkeys to selectively suppress neuronal activity in genetically defined visual cortical and thalamic neurons [12 ]. Local application of allatostatin (100 nM) to the relevant neuronal tissue (within 1 mm) was necessary because of restricted diffusion of the peptide, and its effect on neuronal excitability was inducible and reversible with the similar time course of 10-20 min.…”

“…Allatostatin needs to be applied within 1 mm of the targeted neurons. If cells in the cortex are being silenced, the peptide can be applied to the surface of the brain; however, it needs to be injected with a glass pipette if deeper brain areas are targeted [12 ]. The light source for the activation of halorhodopsin may need to be close to the targeted neurons as well given the power required for stimulation and the light-scattering and absorption properties of the brain tissue and is often done with an optical fiber for stimulating deep in the brain.…”

“…The light source for the activation of halorhodopsin may need to be close to the targeted neurons as well given the power required for stimulation and the light-scattering and absorption properties of the brain tissue and is often done with an optical fiber for stimulating deep in the brain. Although both glass pipettes for AL delivery and optical fibers can be quite small in diameter (30 mm for the pipette [12 ] and 100 mm for the fiber [34]) and thus may not cause much damage, care may need to be taken in some cases to ensure that an important fiber tract has not been disrupted. Dimerizers that are used for the induction of MIST-dependent silencing do not impose such strict restrictions on delivery.…”