15 Daphnia magna were used in total for this experiment. Five Daphnia for each treatment to be tested were transferred by a wide pipette onto a petri dish and put on the Daphnia cooling chamber; the cooling chamber brought down the heart rate of the Daphnia. There was enough water added with the Daphnia in order to ensure survival. To allow slight movement, yet restrict the Daphnia from swimming cotton wool put on the animal. To maintain a temperature conducive to the heart rate measurements, the temperature was maintained at a range of 15-18˚C. Both the petri dish with Daphnia to be tested and the petri dish with Daphnia being tested were on the cooling chamber to ensure constant consistent temperatures for each treatment. Once the Daphnia had reached the set temperature, it was allowed time to acclimate to the new temperature for approximately 2 minutes.
Research Protocol
The experiments focused on the three treatments of nicotine (100 μM), caffeine (50 mM), and ethanol (10% v/v). These solutions were made by 20 μL of each treatment; aliquots were made in order to dismiss any bias. Application of different treatments to different individuals would reduce any stress the Daphnia may have. Treatment solutions were added by using a P200 pipettor and directly placed on the animal.…show more content… Once the treatment was added the heart rate was observed immediately. The sequences were randomized since looking down the microscope affect the measurements negatively. Each observer measured the Daphnia twice prior to the addition of the treatment (control), and twice after treatment. In order to remove the treatment solution, the Daphnia was returned to rehabilitation and the petri dish was washed out for the new Daphnia to be