Scanned with CamScanner Scanned with CamScanner Scanned with CamScanner By Dr. Sascha McKeon, Science Department, Blue Mountain Community College, CCBY 2016. Effects of alcohol and caffeine on Daphnia metabolism - A Sample Formal Lab Report – Group Number Your Name Experiment Date By Dr. Sascha McKeon, Science Department, Blue Mountain Community College, CCBY 2016. Introduction Metabolism comprises all chemical reactions involved in the cellular maintenance of a living organism. Central to these chemical reactions is the transformation of ingested nutrients into energy. This energy, in the form of Adenosine Triphosphate (ATP)i, is needed for both voluntary actions (like walking, running, or biking) and involuntary actions (like breathing, blood circulation, and cell growth). The resting metabolic rate (RMR) utilizes roughly 70% of the daily caloric intake to maintain vital body functions including heart rateii. During homeostasis, the body secretes many chemicals that act to increase or decrease heart rate, as needed. One of the main hormones that increase heart rate is epinephrine (adrenaline), which acts on the body's cells to cause a cascade of signaling eventsiii. In contrast, those which decrease the heart rate include: natriuretic peptides, substance P, neurokinin A, oxytocin, angiotensin 1-7iv. Caffeine mimics some of the effects of adrenaline by stimulating the adrenal glands fright or flight reflexE. This survival response is triggered when a perceived threat is experienced, initiating a cascade of hormones to increase the metabolic rate of the circulatory and respiratory systems. Caffeine also increases the amount of cAMP in the sinoatrial node, making it depolarize and 'beat' fasterv. Alcohol, on the other hand, reduces the activity of the central nervous system thereby reducing heart rate. Alcohol causes an imbalance of electrolytes which affect potassium levels that are critical in the role of nerve functionvi. By interfering with the communication pathways in the brain, alcohol can cause sedation and act as a depressant. The microcrustacean Daphnia, commonly referred to as the water flea, has been subject to intense biological investigations for over a centuryvii. Distant relatives of aquatic animals like crabs and shrimps, there are about twenty-five British species in the family Daphniidae, with the largest being Daphnia Magnavi. The size and transparent carapace make it possible to watch the internal organs at work, particularly the heart, which is usually red as it is full of oxygen-rich bloodv. Daphnia are sensitive sentinel species in freshwater ecosystems establishing the species as a versatile model system to investigate physiology and ecotoxicologyvii. Changes in metabolism are reflected in cardiac frequencyv. There are many inborn errors of metabolism that are treated through supplementationviii. Furthermore, substances like caffeine and alcohol have often become common place in the American diet, while facilitating downstream effects that have not been fully characterized. The objective of this study was to quantify the effects of alcohol and caffeine on the heart and assess the duration of each effect. Understanding the impact and extent on heart rate will provide context to their relevance and safety in dieting and substance abuse. Methods: By Dr. Sascha McKeon, Science Department, Blue Mountain Community College, CCBY 2016. A wet mount slide was prepared, and allowed to stand in fresh water for a minute, to let their heart rate return to normal. The heart rate was calculated by counting the beats of the Daphnia for fifteen seconds and then times that by four, Figure 1 diagrams the general anatomy of the crustacean. This process was normalized over a period of eight trials, to establish a threshold, using water as a negative control. A drop of solution was removed before the addition of a new to maintain a standard volume in the mount. This process was repeated with alcohol, using four concentrations varying from 2-8%, and again with caffeine and three levels (1-3%). Between experimental trials, the Daphnia were given a period of 5 minutes in pure water to return to a resting heart rate. To determine the lasting effects of either solution, daphnia were submerged in 3% solutions of either alcohol or caffeine, with the pulse recorded every minute until the number crossed the threshold. Each experiment was subjected to 3 replicates. The average value across replicates was assessed for precision and accuracy. Figure 1 Diagram of Basic Daphnia Anatomy Results: The average heart rate was approximately 68 bpm (calculated 67.9), using the data from Table 1 to normalize this resting rate, a SD of 138.5 was calculated when comparing the upper and lower variance to the mean, indicating low precision. The SEM indicated a variation of approximately 28 heart beats below or above the norm. Both solutions alter the heart rate of the daphnia, with impact increasing with relative concentration (Table 1 and 2). Overall, alcohol depressed the heart rate of the daphnia by thirty percent, while caffeine had a greater affect by increasing heart rate by four hundred percent (Figure 2). Daphnia exposed to caffeine exhibited an average increase of 3 bpm with each increment of solution strength (R2 = 0.9652, p = 0.002), while alcohol decreased the heart rate at by an average 6 bpm per increment (R² = 0.9441, p = 0.0057). When examining duration, the effects of caffeine and alcohol response occurred within a minute of exposure (Table 3). However, the minimization of their reaction occurs more readily with caffeine than alcohol (Figure 3). Caffeine effect wore off within a minute, while alcohol took four times that span to return to the baseline. By Dr. Sascha McKeon, Science Department, Blue Mountain Community College, CCBY 2016. Table 1 Normalization of Basal Heart Rate Table 2 Effect of Substances on Heart Rate Table 3 Lasting Effects on Heart Rate using 3% solutions Discussion: The experimental set up includes both systematic and random error. The pure water readings were more inconsistent, as evidenced by the high SD and SEM scores, which may indicate stress throughout the experiment. Daphnia are poikilothermic, which means that its body temperature and therefore its metabolic rate are affected directly by the temperature of the environmentv. When the daphnia are observed under the microscope it is in close proximity to the lamp. Given the small quantity of water in which the daphnia is immersed in, only a small amount of heat is required to raise the temperaturevi. It is likely that the temperature increased by several degrees, possibly increasing the daphnia’s heart rate, this may underestimate the effect By Dr. Sascha McKeon, Science Department, Blue Mountain Community College, CCBY 2016. of alcohol and overestimate the effect of caffeine, this constant bias, however, would not affect the precision of the dataset. Figure 2 Effect of Concentration on Heart rate Both Caffeine and alcohol affect the heart in more ways than just pulse rate. Caffeine can affect the main pumping chambers (ventricles), leading to an increase in the rate of contraction and relaxation of each heartbeatv. This means that, as well as beating faster, the heart's individual beats are associated with an increased volume of blood ejected into the circulation per unit time, thereby increasing outputv. Excessive alcohol consumption can damage the heart by weakening the heart muscle and decreasing blood pressureix. Damage to the cardiac nerves will also impact heart function by disrupting the internal pacemaker causing arrythmiasix. By Dr. Sascha McKeon, Science Department, Blue Mountain Community College, CCBY 2016. Figure 3 Duration of substance effect on heart rate Extrapolating the effects on Daphnia to the human condition would require a large-scale medical study. As humans are much larger than daphnia, one would expect a much larger quantity of alcohol or caffeine would be required to produce the same degree of change. Previous studies indicate two or three cups of strong coffee or tea contain enough caffeine to cause an increase in human heart rate of 5-20 beats/miniii. The effects of alcohol on the heart are determined based upon regularity and volume. In small amounts alcohol can increase heart ratevi,ix, can be depressive in moderationv, and dangerously increase heart rate in excessix. For people with underlying conditions, such as heart disease, fluctuations in chemicals that control the heart could prove fatal. Conclusion The heart of Daphnia, and so the heart of a human, is effected by both alcohol and caffeine correlating to concentration strength. Alcohol causes the heartrate to decrease significantly, while caffeine significantly increases the heartrate. The effects of caffeine are exhibited more readily and are metabolized faster by the body than that of alcohol. Both chemicals have far-reaching effects and should be used in moderation. Further studies on similar stimulants and depressants in addition to effects on human body systems will expand our understanding of human metabolic manipulation. By Dr. Sascha McKeon, Science Department, Blue Mountain Community College, CCBY 2016. i Coenzyme Q10 - The Energy Maker - ProHealth.com. (n.d.). Retrieved fromhttp://www.prohealth.com/library/showarticle.cfm?libid=15078 ii Unknown (2013) Importance of metabolism. Retrieved from http://www.journal-advocate.com/sterlingcommunity/ci_24434166/importance-metabolism-sterling-regional-medcenter iii Patel M (2015) Chemicals That Will Increase Your Heart Rate. Retrieved from http://www.livestrong.com/article/226152-norepinephrine-vs-epinephrine/ iv Lacka K and Czyzyk A (2008) Hormones and the cardiovascular system. Endokrynol POl, 59 (5): 420-32. v Handy R (2012) Investigator factors affecting the heartrate of Daphnia. Retrieved from http://www.nuffieldfoundation.org/practical-biology/investigating-factors-affecting-heart-rate-daphnia vi StudyZones.com | Activity Zone StudyZones.com | Activity ...(n.d.). Retrieved from http://www.studyzones.com/ActivityZone/ArticlePrint.cfm?SelectedObjectUUID=97C84 vii Pfrender M (2016) Model Organisms for Biomedical Research. Retrieved from http://modelorganisms.nih.gov/daphnia/ viii Unknown (2013) Importance of metabolism. Retrieved from http://www.journal-advocate.com/sterlingcommunity/ci_24434166/importance-metabolism-sterling-regional-medcenter ix Unknown (2010) Beyond Hangovers: Understanding alcohol’s impact on your health Retrieved from http://pubs.niaaa.nih.gov/publications/Hangovers/beyondHangovers.htm Scanned with CamScanner Scanned with CamScanner Scanned with CamScanner
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