effect-of-choline-on-drosophila

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Sorry one more questions: (4) How do you know that epigenetic changes will only affect "bad" genes in the brain and not "good" ones? What defines a "bad" gene from a "good" one?

4) I truly do not know what effect the choline fully has on the fly brain overall, only that from my experiment, it seems that choline has a positive effect on memory when administered in a proper dosage. The long term effects are still unknown, although no physical abnormalities were observed between groups of flies who were produced, born, and raised in environments with choline. In the future, I would like to gain access to more advanced tools to possibly map out neurons within choline-treated fly brain, or even use fluorescent stains to discover the amount and distribution of certain proteins and neurotransmitters throughout fly brains as well, as well as produce tens or even hundreds of generations of choline treated flies to see what evolutionary effects the addition of choline can have on fly populations as well as ew properties tat may emerge after several generations. I only used the terms “good” and “bad” to designate genes positive to fly health and function, and to designate genes potentially detrimental to fly health and function. In reality, these terms are quite subjective, as no gene can be purely defined as good or bad. Thank you again for your interest in my research! I you have any more questions, feel free to leave them above!

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Hi Lauren, great work that you have done here! I have a few questions for you. (1) Because choline is a nutrient, how effectively do you think it gets absorbed and if its bio-availability is low, are the differences in your concentrations significant? (2) What literature makes you believe that choline leads to epigenetic changes, specifically methylation of histone proteins? (3) Are changes in memory after choline addition through epigenetic mechanistic? How could you prove this?

Thank you!

1) Choline is a polar molecule and also functions as an essential vitamin in humans that is needed for many bodily functions, so I would assume it is absorbed relatively effectively by the human body. According to food site.org, choline is found in animal products as well as in cruciferous veggies and legumes, so it is already quite prevalent in the diet of many. The differences in concentrationwere significant in terms of bioavailability, as three concentrations were used in addition to a control group (no choline at all). The first concentration was set by scaling down the suggested dosage on the choline supplements (capsules with pure choline powder sold on amazon as vitamins) from the recommended dosage for humans, which was two capsules, into grams and then adjusted from the average human weight down to the mass of a fly, and then calculated by the ratio of choline/mL of food solution i terms of approximate number of flies per vial so that theoretically each fly in this group (0.015g choline) would be getting a dosage roughly equal to that of the reccomended daily dosage. The next group, (0.03g choline) was intended to be a double dose, as it was calculated by doubling the recommended dosage to see the effects. Lastly, a “super dose” was calculated to see what the effects of putting multiple times the recommended dosage into the flies’ diet would have on their cognitive function.

2) There wasn’t a concrete article for which this was based on, but rather information learned in an AP biology class. Choline, by nature, contains lots of negatively charged methyl groups. The negative charges on these methyl groups then interact with positively charged histones tailed in DNA, which caused the DNA to become more tightly wound around the protein tails and thus causes some genes to be completely silenced as a result. In other words, the interactions between the histones tails and methyl groups of the choline molecules interact to silence certain genes, which was the basis of this experimental design.

3) From the results of my experiment, it seems that changes in memory after choline addition were not mechanistic, as the flies in the regularly dosed groups performed much better on the t-test and went to the bleach (which should have triggered their memory that bleach was formerly associated with a reward, and gone to the scent hoping for another reward) while the control group flies with no choline performed significantly worse, which indicates that the addition of an accurate choline dosage to food can help boost memory power in flies and brain function. I suppose to validate this, more t-test trials could be run, an extra vial with no beach or ammonia could be added to the t-test to act as a control with the behavioral assay, or other methods could be used to test brain function in flies. But, I found through research that a t-test was the easiest and most efficient way to see fly brain function, although there were a few other methods that could have been used (although I didn’t have access to these resources becasue I did this project in my high school science lab and thus didn’t have huge budget, advanced enough appliances, or even a age enough time frame to test more methods tha could have provided more concrete results, although I would love to explore these more advanced methods in the future to fisher validate the results and continue with my research. Thank you again for your interest in my research!

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Hi Lauren, can you explain how the training was done for the behavioral assay? Also, could you discuss the sample sizes? Why were fewer flies used for the control group? How does this affect the strength of your conclusion? Thanks!

Training the flies was done by exposing them to sucrose, a positive reinforcer, along with bleach as the odor stimuli in the hopes that the flies would learn to associate the scent of bleach with positive (sucrose reward). The same was done with NaCl, and ammonia. I did this specifically by placing a solution of either salt or of sucrose in vials with flies, then adding a cotton pad to seal it onto which I placed 10 drops of either ammonia or bleach. The samples sizes simply resulted from how many flies survived. Whether coincidental or not, the flies in the control group mostly died, whereas the flies in the other groups tended to have lower rates of death. There were approximately 30 flies in each vial to begin with, its just that several died while in the vials. This certainly could call other problems into question with my conclusion, but I believe the percentages of the results from the t-test speak well enough to describe the effects of choline on the flies, because the effects could still clearly be seen between the groups. However, this would certainly be a factor I would want to look into, and potentially rectify through further experimentation. Thank you for you interest in my project, please reach out if you have anymore questions. Also, I apologize for the delayed response. Please stay safe during these unprecedented times in our world.

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Thank you for your interest in my project! If you have any questions, please use the “Ask Me Anything” button at the top of the page. I don’t think this widget works on the tumblr app, but it does work when accessing my tumblr site through a browser on a mobile device, or on a desktop. This presentation mostly discusses the background and the purpose of this project. Thank you again!

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Hello, my name is Lauren McDonough, I’m currently a sophomore at American Heritage High School Boca-Delray in Derry Beach Florida, and this is my project!

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What is Choline?

Choline is a neurotransmitter, nutrient, and a metabolite in many species, including humans. It is the parent compound of the choline class, and has the molecular formula of C5H14NO+. Choline is important because it is a precursor of acetylcholine, which acts as a donor of methyl groups in various metabolic processes. Choline is also an important component in cell membranes, as the hydrophilic head of a phospholipid is made of choline. Choline is considered to be an essential vitamin, and can be oxidized to form betaine, which is a source of methyl groups for a variety of reactions. In humans, acetylcholine is an important neurotransmitter for mental focus and learning. Choline also plays an important role in cholinergic neurotransmission. This process is responsible for memory, mental clarity, concentration, and focus.

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References:

BrainMD. (2016, August 23). Focus on choline: Improve focus and memory. Retrieved October 12, 2019, from BrainMD Life website: https://brainmd.com/blog/focus-on-choline-improve-focus-memory/

Choline. (n.d.). Retrieved October 7, 2019, from WebMD website: https://www.webmd.com/vitamins/ai/ingredientmono-436/choline

Epigenetics: Fundamentals. (n.d.). Retrieved October 12, 2019, from What is Epigentics? website: https://www.whatisepigenetics.com/fundamentals/

Flies' intelligence shocks in study. (2014, May 23). SBS News. Retrieved from https://www.sbs.com.au/news/flies-intelligence-surprises-in-study

HMDB. (2005, November 16). Choline metabocard. Retrieved October 12, 2019, from Human Metabolome Database (HMDB) website: http://www.hmdb.ca/metabolites/HMDB0000097

Jeibmann, A., & Paulus, W. (n.d.). Drosophila melanogaster as a Model Organism of Brain Diseases. Int J Mol Sci. 2009 Feb; 10(2): 407–440. Retrieved from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2660653/

Martin, C. A., & Krantz, D. E. (2014). Drosophila melanogaster as a genetic model system to study neurotransmitter transporters. Neurochemistry international, 73, 71–88. doi:10.1016/j.neuint.2014.03.015

National Center for Biotechnology Information. (n.d.). Choline: Compound Summary. Retrieved October 12, 2019, from PubChem Database: https://pubchem.ncbi.nlm.nih.gov/compound/Choline

Phosphate buffered saline. (2016, October 6). Retrieved October 9, 2019, from Protocols online website: https://www.protocolsonline.com/recipes/phosphate-buffered-saline-pbs/

Scherer, S., Stocker, R. F., & Gerber, B. (n.d.). Olfactory Learning in Individually Assayed Drosophila Larvae. Learn. Mem. 2003. 10: 217-225. Retrieved from http://learnmem.cshlp.org/content/10/3/217.full.html#sec-2

Society for Science and the Public (2016-17). International Science and Engineering Fair 2016-17: International Rules & Guidelines. Washington, DC: Society for Science and the Public.

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Abstract:

The Epigenetic Effects of Choline on Memory and the Development of the Brain in Drosophila Melanogaster

This experiment desired to test if choline could alter the memory complex in flies. Choline is a neurotransmitter and large source of methyl groups, which act in epigenetic gene expression. It was hypothesized that the group with 0.015g of choline would perform best, followed by 0.03g, 0.1g, then, the control. Accurate choline dosages were calculated and added to food. The flies were transferred to vials and left for fourteen days. The flies were then placed into vials with fructose and a bleach solution and in separate vials, NaCl with an ammonia solution. Then, flies were added to a t-contraption to test their memory. After this, flies were dissected and imaged under a microscope to analyze differences between groups. For the t-test, in the control group, 20% went to bleach, 20% ammonia, and 60% chose neither. In the 0.015 group, 65% chose bleach, 10% ammonia, and 25% neither. In the 0.03 group, 56.25% chose bleach, 18.75% ammonia, and 25% neither. In the 0.1 group, 42.9% chose bleach, 14.3% ammonia, and 42.9% neither. In the dissected images of the flies, there seemed to be no visible differences between the groups, but this could have been due to the lack of more in-depth technology. The results of this project supported the hypothesis, as the 0.015 group did the best. In the future, this could be applied to humans, to see how choline alters human brain function.

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This project is titled: “The Epigenetic Effects of Choline on Memory and the Development of the Brain in Drosophila Melanogaster”

*Full title wouldn’t fit on header