Pheromonal Induction of Spatial Learning in Mice
Sarah A. Roberts1,
Amanda J. Davidson1,
Lynn McLean2,
Robert J. Beynon2, and
Jane L. Hurst1,*
1 Mammalian Behaviour and Evolution Group, Institute of Integrative Biology, University of Liverpool, Leahurst Campus, Neston CH64 7TE, UK.
2 Protein Function Group, Institute of Integrative Biology, University of Liverpool, Liverpool L69 7ZB, UK.
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Fig. 1. Female sexual attraction to male urine scent marks and conditioned place preference. After confirming no side bias (no urine), female mice were given test urine versus water in two dishes in 10-min daily learning sessions (L1 to L3). CPP was tested 24 hours later with no urine present (24-hour memory). Females were given (A, B, and C) unfamiliar male or (D) female urine for three [(A) n = 10 subjects], two [(B) n = 12 subjects] or one [(C) and (D) n = 12 subjects] learning sessions. Greater time spent in the urine (blue bars) versus control dish (yellow bars) was assessed using one-tailed paired t tests (data log transformed to meet parametric assumptions): *P < 0.05, **P < 0.01, ***P < 0.005, ****P < 0.001. Circles show matched-pair difference in time spent in test minus control dish. Data are means ± SEM.
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Fig. 2. Retention of female-remembered place preference conditioned by male urine. (A) CPP was tested 1 to 28 days after a single 10-min learning session with male urine (one test per female; full details in fig. S1). (B and C) CPP was tested 2 days after the learning session with no intervening exposure to the test arena [(B) n = 11 subjects], or both 1 day and 2 days after learning [(C) n = 20 subjects]. Key and statistical tests are as in Fig. 1.
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Fig. 3. Darcin stimulates female conditioned preference for male urine location. CPP was assessed 24 hours after a single 10-min learning session with test stimulus versus (A to C and F) control buffer or (D and E) water. (A) r-darcin (1 μg/μl buffer, n = 18 subjects); (B) r-MUP7 (1 μg/μl buffer, n = 37 subjects) (supplementary materials, materials and methods); (C) r-MUP11 (1 μg/μl buffer, n = 18 subjects); (D) C57BL/6 male urine containing 8 μg/μl protein, including 1 μg/μl natural darcin (n = 10 subjects); (E) BALB/c male urine containing <0.1 μg/μl darcin (n = 11 subjects); (F) BALB/c male urine plus r-darcin (1 μg/μl, n = 12 subjects). Greater time in test dish was assessed by Wilcoxon [(A) to (C)] or t tests (data log transformed). Key is as in Fig. 1.
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Fig. 4. Darcin stimulates conditioned preference for male urine location among competitor males. CPP was assessed 24 hours or 14 days after a single 10-min learning session with test stimulus versus (A to C, G, I, and J) control water or (D to F and H) buffer. Unfamiliar wild-stock male urine (A) 24 hours or (B) 14 days after contact, or (C) 24 hours after exposure to airborne urinary volatiles; (D) r-darcin (1 μg/μl buffer); (E) r-MUP7 (1 μg/μl buffer); (F) r-MUP11 (1 μg/μl buffer); (G) BALB/c male urine containing <0.1 μg/μl darcin; (H) BALB/c male urine plus r-darcin (1 μg/μl); n = 12 wild-stock males tested for each. Own urine (I) frozen prior to testing (n = 20 subjects) or (J) collected immediately before learning session (n = 11 subjects). Greater time in test dish was assessed by Wilcoxon [(D) to (H)] or t tests (data log transformed). Key is as in Fig. 1.
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