Fig. 4. Sequential comparison of the behavioral responses to an unexpected manual touch in socially inexperienced and experienced crayfish. (A) Average response frequencies of Orienting, Avoidance and No Response behavioral responses of Isolates, dominants and subordinates on Day 1 while separated from partners by an opaque barrier (condition as in Fig. 1D). The three behavioral responses of the Isolates and social pairs did not differ significantly (Kruskal–Wallis test, P=0.4570 for Orienting Responses, P=0.7133 for Avoidance Responses, P=0.6163 for No Responses). (B) Average response frequencies of the same animals on Day 2 when the dominant and subordinate crayfish were paired with their partners (condition as in Fig. 1G). The Orienting and the Avoidance responses of dominants and Isolates were significantly different from those of subordinates (for Orienting Responses, overall difference, P=0.0008; Isolates vs subordinates, P=0.0012; dominants vs subordinates, P=0.0002; for Avoidance Responses, overall difference, P=0.0003; Isolates vs subordinates, P=0.0003; dominants vs subordinates, P=0.0002) but were not different from each other (for Orienting Responses, P=0.6126; for Avoidance Responses, P=0.7789). The No Responses were not different in Isolates, dominants and subordinates (overall difference, P=0.2069). (C) Behavioral responses later on Day 2 when the dominants and subordinates were tested while separated by an open divider (condition as in Fig. 1I). The dominants' responses were not different from those of the subordinates (for Orienting Responses, P=0.2786; for Avoidance Responses, P=0.1605; for No Responses, P=0.7984). (D) Dominant and subordinate responses after re-isolation (condition as in Fig. 1K) are not significantly different (for Orienting Responses, P=0.2345; for Avoidance Responses, P=0.3282; for No Responses, P=0.9591). (E) The behavioral responses of the subordinates in the four consecutive social conditions (shown in A–D) significantly changed after pairing experience (Friedman test, P=0.0004 for both Orienting and Avoidance Responses). The subordinate animals when isolated on Day 1 displayed the Orienting Responses significantly more often than when paired on Day 2 (Wilcoxon test, P=0.0078), when semi-separated on Day 2 (P=0.0313) and when isolated on Day 2 (P=0.0313). The subordinate animals when paired on Day 2 displayed the Orienting Responses significantly less often than when semi-separated on Day 2 (P=0.0156) and when isolated on Day 2 (P=0.0156). The average frequency of the orienting response when semi-separated on Day 2 did not differ from those when isolated on Day 2 (P=0.5625). The subordinate animals when paired on Day 2 displayed the Avoidance Responses significantly more often than when isolated on Day 1 (P=0.0078), when separated by the open divider on Day 2 (P=0.0156), and when isolated on Day 2 (P=0.0078). The average frequency of No Responses did not change in the four social conditions examined (overall difference, P=0.0660). *P<0.05, **P<0.01, ***P<0.001. All values are means ± s.d.