Optogenetic manipulation of second messengers in neurons and cardiomyocytes with microbial rhodopsins and adenylyl cyclase

Abstract

Even though microbial photosensitive proteins have been used for optogenetics, their use should be optimized to precisely control second messengersin vivo. We exploitedGtCCR4 andKnChR, cation channelrhodopsins from algae,BeGC1, a guanylyl cyclase rhodopsin from a fungus, and photoactivated adenylyl cyclases (PACs) from cyanobacteria (OaPAC) or bacteria (bPAC), to control cell functions in zebrafish. Optical activation ofGtCCR4 andKnChR in the hindbrain reticulospinal V2a neurons, which are involved in locomotion, immediately induced swimming behavior, whereas activation ofBeGC1 or PACs was achieved at a short latency.KnChR had the highest locomotion-inducing activity of all the channelrhodopsins examined. Activation ofGtCCR4 andKnChR in cardiomyocytes induced cardiac arrest, whereas activation ofbPAC gradually induced bradycardia.KnChR activation led to an increase in intracellular Ca2+in the heart, suggesting that depolarization caused cardiac arrest. These data suggest that these optogenetic tools can be used to reveal the roles of second messengers in various cell types in vertebrates.

Neuropeptidergic regulation of neuromuscular signaling in larval zebrafish alters swimming behavior and synaptic transmission

“…One class of tools for optogenetic manipulation are photoactivated adenylyl cyclases (PACs) such as bPAC, mediating light-dependent synthesis of the second messenger cAMP for precise temporal and spatial control and acute tuning of intracellular cAMP concentrations [31][32][33][34]. bPAC was shown to be enzymatically active in early zebrafish embryos [35] and has been applied to analyze the role of cAMP signaling in axonal regeneration [36] or to induce swimming behavior by activating hindbrain reticulospinal V2a neurons [37].…”