Chemogenetics revealed: DREADD occupancy and activation via converted clozapine

TLDR

DREADDs are chemogenetic tools that enable remote control of neuronal activity in freely moving animals by activating engineered receptors with the designer drug clozapine N‑oxide. The study aimed to confirm the in vivo mechanism of action of CNO at DREADDs. CNO fails to cross the blood–brain barrier and binds poorly to DREADDs, but is rapidly converted to clozapine, which penetrates the brain, binds DREADDs with high affinity, and elicits DREADD‑mediated behaviors even at subthreshold doses.

Abstract

The chemogenetic technology DREADD (designer receptors exclusively activated by designer drugs) is widely used for remote manipulation of neuronal activity in freely moving animals. DREADD technology posits the use of "designer receptors," which are exclusively activated by the "designer drug" clozapine N-oxide (CNO). Nevertheless, the in vivo mechanism of action of CNO at DREADDs has never been confirmed. CNO does not enter the brain after systemic drug injections and shows low affinity for DREADDs. Clozapine, to which CNO rapidly converts in vivo, shows high DREADD affinity and potency. Upon systemic CNO injections, converted clozapine readily enters the brain and occupies central nervous system-expressed DREADDs, whereas systemic subthreshold clozapine injections induce preferential DREADD-mediated behaviors.