Fig. 4. Manganese treatment affects sucrose responsiveness. (A) Effects of MnCl₂, ZnCl₂ and 8-Br-cGMP treatments on sugar responsiveness using the proboscis extension assay (Ben-Shahar and Robinson, 2001). 8-Br-cGMP was used to explore the relationship between sucrose responsiveness and rate of behavioral maturation (Ben-Shahar et al., 2002b). Bees were exposed to a sequential series of increasing sucrose concentration (see Materials and methods); a higher response index indicates increased responsiveness to sucrose. (B) Effect of manganese treatment on head manganese levels. Manganese levels increased significantly in heads of treated bees relative to untreated controls (P<0.001, ANOVA), validating the treatment method. Bars represent means ± S.E.M. Numbers in bars represent sample sizes. Results of statistical analysis in text. Different letters above bars represent groups that were significantly different by ANOVA with Bonferroni post hoc analysis (P<0.05). Numbers in bars represent pooled data from six independent trials (A).