11In order to track changes in personality as a function of mating, the behavior and excreted 12 hormone levels of adult female threespine sticklebacks was recorded both before and after 13 mating, and compared relative to females that did not have an opportunity to mate (control) and 14 to females that had an opportunity to mate and therefore experienced courtship socially but did 15 not mate (courtship control). Individual behavior was repeatedly recorded in three separate 16 behavioral assays designed to measure activity, social behavior and risk taking behavior. The 17 behavior of individuals in all three treatment groups was measured in each assay three times 18 both before and after females had an opportunity to mate, for a total of six trials per assay per 19 individual. Repeated samples of steroids excreted in water were collected and were measured 20 using GC/MS. 21 22 Study population 23 Threespine sticklebacks were collected from Putah Creek, California, U.S.A. in April 2017 prior to 24 the onset of the breeding season. Neither males nor females showed signs of reproductive 25 maturity when they were collected; therefore, we assume that females were virgins at the time 26 of the experiment. Fish were shipped to the University of Illinois at Urbana-Champaign 27 (Champaign, IL, U.S.A.) and maintained in the laboratory in mixed-sex group tanks (108 L × 33 W 28 × 24 H cm). Fish were housed in tanks (60.75 L × 30 W × 20 H cm, 20 fish/tank) for approximately 29 three weeks before the experiment began. 30 One week before the start of the experiment, females were housed in 'home' tanks (35.5 31 L × 33 W × 25 H cm, 10 fish/tank) with a gravel bottom, plastic plants, and a shelter (opaque white 32 PVC pot of 10 cm in diameter) and stayed in these home tanks for the duration of the experiment, 33 except when individuals were transferred to separate 'observation' tanks (60.75 L × 30 W × 20 H 34 cm) for behavior trials. The observation tanks were set up the same as the home tanks, i.e. gravel 35 at the bottom, plastic plants, and a shelter (white PVC pot). All trials were conducted from mid-36 June to early October 2017 i.e. during the reproduction period of this population. None of the 37 females in this experiment were infected with Schistocephalus solidus, a tapeworm known to 38 influence risk-taking behavior [1,2]. 39 Fish were maintained at 20°C on a summer photoperiod (16: 8 h light: dark cycle). A 40 recirculating flow-through system consisting of a series of particulate, biological, and UV filters 41 (Aquaneering, San Diego, USA) was used to clean the water. 10% of the water volume in the tanks 42 was replaced each day. The fish were daily fed a mixed diet consisting of frozen bloodworm, 43 frozen brine shrimp, and frozen Mysis shrimp ad lib each day. During behavioral assays, fish were 44 fed after assays ended. 45 46 Experimental design 47 157 All females were measured for length and weight on the last day of the After trials.158 159 Measuring excreted steroids 160After the third trial of both the Before ...
