The dynamics of an infrared light‐activated adenylyl cyclase during the manipulation of behavior in Caenhorabditis elegans
Authors: Szurgot Mary , Janton Francis Matt , Nelson Matthew 
Published: November 2023

Optogenetic tools that can induce cAMP(cyclic adenosine monophosphate), a secondary messenger which amplifies signals received by a cell to regulate physiological responses, provide an exciting new approach for determining where and how cAMP functions in living animals. One such tool is a red light‐activated adenylyl cyclase called IlaC. When red light is absorbed by the phytochrome domain of IlaC the adenylyl cyclase domain is activated and converts ATP to cAMP. To quantify the dynamics of IlaC in vivo and as a means for studying sleep behaviors, we have engineered various Caenorhabditis elegans strains expressing IlaC in different subsets of neurons.

First, we have been measuring the biochemical dynamics of the IlaC tool in vivo. To do this, we have been using a cAMP detection assay on lysates of pan‐neuronal IlaC‐expressing worms exposed to either red or green light. After exposing these worms to red or green light for different lengths of time, we quantify cAMP levels within the worms to see how quickly IlaC acts and how significantly it increases intracellular cAMP. Our preliminary results show that when exposed to just one minute of red light, IlaC‐expressing worms have approximately 70% more cAMP than those exposed to one minute of green light. Thus, IlaC is acting relatively quickly to induce cAMP in the nervous system of C. elegans.

Second, we are using the IlaC tool to study C. elegans sleep behaviors. Although all animals demonstrate sleep, the reason for this behavior and the mechanisms of its regulation are still largely unknown. By using a model system like C. elegans, pathways that regulate sleep and wakefulness can quickly be identified and studied with single‐cell resolution. The cAMP pathway regulates sleep in mice, fruit flies, and worms. Increased levels of cAMP are associated with wakefulness, while decreased levels regulate sleep. The exact details of how and where cAMP regulates sleep are unknown. Our preliminary results show that when animals expressing IlaC in all neurons are monitored under red light (active IlaC) but not green light (inactive IlaC), animals display drastically less sleep. Thus, cAMP acts in the nervous system and IlaC will be a useful tool for determining the exact site of action during sleep in C. elegans.

In order to pinpoint which neurons regulate sleep within the nervous system, we constructed a number of C. elegans strains that express transgenes composed of tissue‐specific promoters. These various promoters drive IlaC expression in certain sets of neurons such as motor neurons, command interneurons, and other neurons believed to be associated with quiescent behavior. One such neuron is the DVA neuron, a neuron thought to be involved in the sleep pathway. Preliminary results show a decrease in sleep following an exogenous stressor compared to wild‐type worms. Further experiments will help determine the role the DVA neuron plays in regulating sleep, as well as elucidate other specific neurons responsible for sleep regulation in C. elegans.