The Conditioning of Respiration Rate in Goldfish (Carassius auratus)Using Water Temperature and Visual StimuliDonna Tehrani and Patrick SchaferDepartment of Biological SciencesSaddleback CollegeMission Viejo, California 92692Goldfish (Carassius auratus) is a domesticated species of tropical fish that is often used in psychological and physiological experiments. They have been demonstrated to respond to classical (Pavlonian) conditioning and even alter their homeostatic processes based on various stimuli, including color. Twenty goldfish were trained to associate the color red with a decrease in water temperature and the color green with no change in water temperature. An experiment was run to see whether goldfish exposed to the color red would change their respiration rates even without achange in temperature by filming and recording their operculum movements. The change in operculum movements for one minute before and after exposure to the stimuli was compared to a control group which was shown the color green instead. Atwo-tailed t-test was performed to ascertain the difference between the changes in respiration rates between the two groups. There was no significant difference (p=0.084), which suggests that some fish do not alter their respiration rates in anticipation of temperature changes. Project SummaryMost species of tropical fish, including carp and goldfish, have superior color vision to humans or other mammals due to the structure of their retina. Therefore, it is unclear to what extent fish use visual cues to anticipate environmental changes, since they can respond to stimuli that are too subtle for humans to notice. They also possess theability to be psychologically and physiologically conditioned as a response to visual stimuli. Previous research suggests that it may be possible that goldfish can anticipate changes in their environment and modify their homeostatic processes. Goldfish possess color vision, and have demonstrated color constancy in a laboratory setting (Ingle 1985). It has also been shown that because of the structure of the brain and eye, goldfish are more adept at recognizing colors than patterns, especially different shades of green and red (Ingle 1965). In laboratory studies, goldfish have been shown to lower their respiration rates in response to lower temperatures (Schmidt-Nelson 1997). Also, Japanese carp, a closely related species, have demonstrated the possibility of respiratory conditioning using electrodes (Woodard 1971, Otis 1957). However, no research was found regarding the possibility of conditioning goldfish using temperature and color as a visual stimuli. We hypothesized that when goldfish were trained to associate a specific color with changes in water temperature, they would alter their respiration rates when shown the stimuli but without a change in temperature. This research could shed light on the extent that aquatic creatures regulate their homeostatic processes based solely on sensory information.Methods and MaterialsTwenty feeder goldfish were purchased from Petsmart and were trained over a period of two weeks. Training consisted of placing an individual goldfish with 0.5 L of water at 18-23 C into a cylindrical plastic container that was 18 centimeters across 46 centimeters high. The goldfish was given five minutes to acclimate itself to its surroundings, after which the container was completely surrounded by a red letter-size folder and 0.5 L of water at 0 C were poured in to decrease the respiration rate. The goldfish was kept in this condition for another five minutes, before being returned to one of two common tanks. After a couple hours, the same goldfish was placed in the same container with the same amount of water. After five minutes the container was surrounded with a green letter-sized folder instead and 0.5 L of room temperature water was poured inside the container. Each fish was exposed to both temperatures and colors daily, and the order in which they were placed in each condition was varied. After 14 days of conditioning, an experiment was run where 10 of the goldfish were randomly placed into the glass container and filmed by a Canon SX 150IX camera. Randomization was achieved by removing the goldfish from their two common tanks one at a time and alternating between placing them in the control and experimental group. After a four minute acclimation period, the number of operculum movements in one minute was recorded. The red folder was then displayed but 0.5 L of room temperature water were poured instead, and the number of operculum movements in the minute after the water was added was recorded. The procedure was repeated with the other 10 fish, except the green folder was displayed instead. The change in respiration rates between the two groups was recorded and an unpaired two-tailed t-test was performed between the groups.ResultsThe mean change in respiration rates in the experimental group was 2.9 ± 3.2 (±SEM, n=10), and mean change in respiration rates in the control group was -1.4 ± 6.7 (±SEM, n=10). The two groups were graphed against their mean change and we obtained a p-value of 0.084, using a two tailed t-test, which corresponds to no significant difference in the number of operculum movements between the two groups (Figure 1). Therefore, our data suggest the null hypothesis. Figure 1. This bar graph displays the average change in respiration rates between the experimental (n=10) and control group (n=10). A two-tailed unpaired t-test was performed on the difference in operculum movements, and there was no significant difference between the two groups (p=0.084). Standard error bars are shown.DiscussionOur results suggest that goldfish do not make immediate changes to their respiration rates even when they anticipate changes in water temperature. However, we cannot rule out the possibility that competing trends may have affected out results, since goldfish, like terrestrial animals have been shown to increase heart rate and respiration rate in anticipation of stress (Woodward 1971). Further experimentation might be performed by raising the temperature of the water instead of cooling it down, to see if this was the case. Also, more extreme changes in temperature might be employed to maximize the physiological stress and make a possible response more visible. Finally we could measure heart rate instead of respiration rate.In conclusion, it does not appear that goldfish physiologically prepare