DESCRIPTION: Conditionally silencing the activity of specific neural ensembles is a powerful approach for mapping the circuits responsible for specific behaviors. While microbial opsin tools currently exist to silence neural activity with light, these tools have significant limitations tha make them unsuitable for applications that require persistent silencing over the course of minutes or hours. Longer term neuronal silencing has been classically achieved by expressing the catalytic light chain of Clostridium neurotoxins, which disrupt neurotransmitter release, however this approach suffers from poor temporal and spatial control. Recent chemogenetic approaches, including designer receptors exclusively activated by designer drugs (DREADDs) have been developed for persistent silencing, but lack the spatiotemporal benefits of optogenetic approaches. Thus, there remains an unmet need for silencing tools that combine the robust and persistent silencing qualities of Clostridium neurotoxin and chemogenetic approaches, with the spatiotemporal control of optogenetics. Here we will develop a completely new toolkit for rapid, extended neural silencing with a single, brief light pulse that can be readiy activated by either single or multiphoton excitation for in vivo systems. Our tools will not only b applicable to neurotransmitter systems, but also to neuromodulatory systems and glial transmission. These powerful tools will complement and extend the existing opto- and chemogenetic silencing toolset by allowing rapid, robust and persistent silencing with a single light pulse. Because our strategy operates on a completely different principle than current tools, it could be multiplexed with existing approaches for complex remote control of circuit activity during behavior.