Dissertations, Theses, and Capstone Projects

Date of Degree

6-2022

Document Type

Thesis

Degree Name

M.S.

Program

Cognitive Neuroscience

Advisor

Orie Shafer

Subject Categories

Animal Studies | Cognitive Neuroscience | Molecular and Cellular Neuroscience

Keywords

Drosophila, Chronobiology, Genetics

Abstract

Circadian rhythms are physiological and behavioral changes which follow a 24-hour cycle. Drosophila Melanogaster’s circadian clock neuronal network (CCNN) has been identified and several subpopulations have been characterized based on previous studies; the classifications of subpopulations of neurons within the CCNN are based on a return of anticipatory locomotor activity preceding the two daily light transitions (Lights on = dawn/Lights off = dusk). The neurons responsible for the return of anticipatory morning locomotor activity have been referred to as the M-cells, the group of neurons known to rescue evening anticipatory locomotor activity have been termed the E-cells. In this study I have selectively reinstated the Period (PER) protein using the Gal4-UAS system in specific subsets of the circadian clock neuron network using the per01mutant fly as the base. This mutant fly lacks the PER protein, a critical protein essential for driving the negative feedback loop which drives endogenous circadian timekeeping rendering this fly arrhythmic in environments with constant conditions (DD). In more natural conditions with a light-dark cycle, the mutant per01 fly will lack anticipatory locomotor activity preceding light transitions, the recovery of this anticipatory activity has been classically used as a proxy for the recovery for circadian rhythm. Using the GAL4-UAS system, I will isolate the contributions of specific circadian clock neurons to sleep in a standard light-dark (LD) environment, break-down the sleep architecture for day (siesta) and nighttime sleep. Previous research using this design has focused on the anticipatory activity mentioned earlier, my study will use this phenotype to confirm rescue of circadian rhythm in the genetic mosaics but will focus on sleep and sleep consolidation changes accompanying the rescue. I have also broken-down sleep-in Drosophila Melanogaster into the day which has been referred to in the field as siesta sleep, and night sleep due to the models’ diurnal rhythms.

My results indicate the M-neurons classically associated with anticipatory activity preceding lights on exert influence on nighttime sleep. With several night sleep parameters trending towards wildtype levels when PER was reinstated in the M-neurons. However, Siesta sleep remained similar to our parental Gal-4 control. Similarly, when both the M-neurons and E-neurons have had per reinstated confirmed by restoration of the morning and evening anticipatory activity, both siesta and nighttime sleep architecture recovers in the direction of wildtype levels. The consolidation of sleep recovers to wildtype levels. These results suggest the M-neurons exert influence over nighttime sleep and the E-neurons contributes to siesta sleep.

I will also discuss longer sleep bouts which have been correlated with deeper sleep by increased arousal threshold and metabolic changes after 30 minutes of inactivity. When looking at sleep parameters using a longer definition of sleep we see similar trends as the standard definition with the M-neurons having more influence over night sleep and the E-neurons having a profound impact on siesta sleep. Finally, I’ll discuss sleep in a constant environment (DD), with the lack of light transitions to guide behaviour the flies will be free running, which refers to their endogenous circadian clock and the period it sets. The difficulty with free-running analysis stemming from differing endogenous periods of day prohibited DD sleep parameter comparisons.

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