Neuronal control of sleep in Caenorhabditis elegans
von Inka Busack
Datum der mündl. Prüfung:2020-10-27
Betreuer:Prof. Dr. Henrik Bringmann
Gutachter:Prof. Dr. Henrik Bringmann
Gutachter:Prof. Dr. Tim Gollisch
EnglischSleep is crucial for all organisms with a nervous system. Amongst other functions, it is required for energy allocation, higher brain functions and the control of physiological processes. Sleep-active neurons have previously been identified in many species. These neurons act as the motor of sleep as their depolarization causes inhibition of wakefulness circuits and leads to sleep induction. However, how these sleep-active neurons get regulated and how exactly they are involved in molecular pathways for the benefits of sleep remains unclear. In this study I focused on the neuronal component of sleep regulation in the model organism Caenorhabditis elegans. In C. elegans the ring interneuron RIS functions as single sleep active neuron. First, I aimed to identify a neuronal circuit that regulates RIS activity. I found that RIS is controlled by the command interneuron PVC through a positive feedback loop. The interneurons PVC and RIM act together to activate RIS and sleep is most likely induced at the transition from forward to reverse locomotion. While RIS activity and hence sleep gets regulated by the nervous system, I could also show through pan-neuronal imaging that the control is reciprocal and RIS depolarization directly inhibits nervous system activity. Next, I intended to design a stand-alone device for optogenetic long-term experiments: the OptoGenBox. Optogenetics is a method in which through genetically knocked-in actuators and light, for instance, individual neurons can get de- or hyperpolarized. Implementation of the OptoGenBox was successful and I could show that long-term optogenetic sleep inhibition by hyperpolarization of RIS leads to a reduced longevity of arrested first larval stage worms. Lastly, I investigated the functions of sleep in C. elegans. Selected health span assays and investigation of synaptic changes did not reveal further functions of sleep. To better assess sleep benefits, strains, in which RIS was either constantly de- or hyperpolarized through genetically knocked-in ion channels, were generated and characterized. Constant de- as well as hyperpolarization of RIS led to a reduction in sleep but diverging longevity effects in the arrested first larval stage. In conclusion, sleep in C. elegans is highly controlled by the nervous system and sleep induction is not only dependent on sleep-active neurons but furthermore wake-active circuits that activate sleep neurons. As sleep is evolutionary conserved, these circuits are most likely also existent in organisms with more complex nervous systems such as mammals. The OptoGenBox as well as the here presented new RIS manipulated worm strains present potent tools to further investigate neuronal circuits and protective pathways downstream of the sleep neuron RIS.
Keywords: C. elegans, sleep, optogenetics, RIS, PVC, RIM, OptoGenBox