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. 2020 Jul 29;40(31):6018–6034. doi: 10.1523/JNEUROSCI.2674-19.2020

Figure 2.

Figure 2.

npr-11 mutants are defective in diacetyl learning. A, Schematic of the diacetyl learning assay, modified from Vukojevic et al., (2012). Age-synchronized worms are washed from cultivation plates and immediately tested for naive CTX to diacetyl, while the remaining animals undergo 3 h of coupled (diacetyl + no food) or uncoupled (diacetyl-only or no food-only) conditioning. CTX to diacetyl is then quantified on CTX plates; and after 1 h, a CI is calculated. B, WT worms after coupled conditioning (cond.) show a significant drop in the CI compared with animals conditioned with diacetyl-only (left) or no food-only (right). Data were analyzed by Kruskal–Wallis (KW(2) = 24.15, p < 0.0001) and Dunn's multiple comparison tests (left, n ≥ 10) or one-way ANOVA (F(2,13) = 17.37, p = 0.0002) and Tukey's post hoc test (right, n ≥ 5). C, nmr-1 mutants show impaired diacetyl learning compared with WT. Two-way ANOVA revealed a significant effect of genotypes (F(1,30) = 13.83, p = 0.0008), of behavioral treatments (F(1,30) = 83.83, p < 0.0001), and of interaction between genotypes and behavioral treatments (F(1,30) = 13.14, p = 0.0011). Data were analyzed by Sidak's multiple comparison test (n ≥ 7). D, Shock treatment reverses the reduced attraction to diacetyl in animals conditioned for diacetyl learning (cond.), but not in a control group for desensitization exposed to undiluted diacetyl. Data were analyzed by one-way ANOVA (F(7,33) = 11.50, p < 0.0001) and Tukey's post hoc test (n ≥ 4). E, npr-11 mutants show normal attraction to diacetyl before conditioning but are defective in diacetyl learning. Two-way ANOVA revealed a significant effect of genotypes (F(1,46) = 7.666, p = 0.0081), of behavioral treatments (F(1,46) = 94.92, p < 0.0001), and of interaction between genotypes and behavioral treatments (F(1,46) = 14.61, p = 0.0004). Data were analyzed by Sidak's post hoc test (n ≥ 10). F, G, npr-11 mutants display normal locomotion speed on and off food. Activity is recorded for 10 min on E. coli OP50-seeded (F) or unseeded (G) NGM plates. The average speed of mutants is not significantly different from that of WT in both conditions. Data were analyzed by unpaired t test (F, w ≥ 23; G, w ≥ 39). B–G, Boxplots represent 25th (lower boundary) and 75th (upper boundary) percentiles. The 50th percentile (line) indicates the median. Whiskers represent the minimum and maximum values. Black dots represent individual CIs or worms. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. ns, not significant.