Drosophila melanogster are diurnal, meaning they are awake during the day and sleep at night (Dubowy and Seghal, 2017). Although, they do like to take a daytime siesta (much like some people I know). Flies are an amazing model to study circadian rhythms. Indeed Jeffrey Hall, Michael Rosbash and Michael Young won the 2017 Nobel Prize in Physiology or Medicine for their groundbreaking work on the molecular mechanisms of circadian rhythms using Drosophila as a model system. The Light/Dark (L/D) cycle can affect many facets of fly behavior.
First, the locomotor activity of flies is dependent on the L/D cycle, and nearly all behaviors that we measure in the lab depend on the ability of the flies to move. Flies are most active near dawn (when lights turn on) and dusk (when lights turn off) (Allada & Chung, 2010). Second, flies sleep at night so if you have an abnormal L/D cycle, you are probably messing with the flies’ sleep. Flies, like humans, need sleep for peak behavior performance. Sleep affects how much a fly learns and remembers, and benefits different memory stages (Dissel et al, 2015; Marquand et al, 2023)
What type of Light/Dark cycle should you use for optimal behavior? We used to use a 12/12 cycle on our incubator but for the past 10 years, the lab has shifted to a 14/10 cycle to allow the larks and owls of the lab (people not flies) to use the same incubator. We haven’t noticed any real effects on the behaviors we routinely run in the lab, but we are always cognisant of when the peak activity and peak rest times are and take that into account when running any assays that require consistent fly activity. This doesn’t mean that our 14/10 cycle doesn’t affect behavior - maybe it does, but the variation in the behavior assay itself supercedes variation that might be caused by our L/D cycle. I am very reluctant to decrease the amount of night to less than 10 hours, but we haven’t tested this rigorously. In any case, always write down the L/D cycle of your incubator, if there were any openings during dark hours, and the time of day the behavior experiment was performed.
What times of the day are best for fly behavior? The dilemma if you are measuring any behavior other than general locomotor activity, is that you don’t know how high activity or low activity is going to affect whatever behavior you want to measure. My recommendation is to do some pilot tests to see where you get the most consistent behavioral response (ie least variable between flies or groups of flies). Note that I didn’t write ‘strongest behavioral response’ and that is because sometimes the highest effect size can occur at a time where the lowest effect size is found in a different fly or group of flies. This might result in too much variability to get a consistent pattern in behavior.
Some general trends that we have found occur in our lab:
For most of our induced group locomotor activity and single-fly locomotor activity behaviors we find that morning works best (1 or more hours after lights on).
Olfactory responses tend to do well in the dark and I have found early afternoon works well for that.
Feeding behaviors are also rhythmic (Krupp & Levine, 2010). We have found that responses to food-reward stimuli seem to be most consistent in the afternoon. However, I recommend finishing up the experiments before activity ramps up near lights-out.
Responses to aversive stimuli seem pretty consistent no matter the time of day.
That all being said, for planned classroom and laboratory class type activities, I don’t worry about the L/D cycle because the behaviors are typically robust enough that you’ll see them regardless of the L/D cycle the flies have been kept at.
What time of day works best for the behaviors you are measuring?