Electrophysiological characterisation of Drosophila central clock neurons in health and disease

The importance of normal circadian clock function in maintaining good overall health has become widely accepted. Daily rhythms in physiology and behaviour are driven by a complex circuit of circadian neurons, of which in Drosophila remains an elusive part of circadian control. A study published in 2011 revealed the ability of CRY to behave as a neuronal activator in light conditions, eliciting an increase in firing frequency when activated; possibly via the modulation of potassium channels in the cell membrane using whole cell patch clamp. The use of this technique in Drosophila CNS neurons is limited due to difficulties in obtaining and interpreting the data generated. This project aims to contribute to the field via exploring its use and limitations, as well as dissect the role of blue light photoreceptor CRYPTOCHROME within the circadian clock neuron circuit by manipulating normal levels of CRY. This was achieved by using a constitutively active from of CRY, CRYΔ, a cry knockout, cry0 and compared these to controls with undisturbed levels of CRY. We also considered the impact of these changes depending on the time of day by recording l-LNv physiology in the morning and the evening. This study revealed that CRY modulates potassium channels throughout the day to support the circadian-regulated changes in l-LNv physiology, of which some are already known such as firing frequency and resting membrane potential. These changes have suggested that along with CRY’s instant ability to influence potassium channels in response to light input in the morning, a delayed signalling mechanism that impacts potassium channels in the evening could occur. It is well established that clock degradation occurs as an early symptom in many neurodegenerative diseases, such as Parkinson’s disease. We therefore studied the effect of α-Synuclein expression in l-LNv neurons throughout the process of aging. Interestingly, we have revealed that the process of aging influences aspects of normal l-LNv physiology. In conjunction, α-Synuclein expression disrupted these normal aging processes as well as affect firing frequency and potassium channels outside of this. Although more work is needed to consolidate these findings, this study has provided an insight into clock cell disruption that occurs prior to the loss of neurons. We hope this early signalling disruption could be further investigated to aid earlier diagnosis in the future.