Patterns of PERIOD and pigment-dispersing hormone immunoreactivity in the brain of the European honeybee (Apis mellifera):: Age- and time-related plasticity

We explored the neural basis of age- and task-related plasticity in circadian patterns of activity in the honeybee. To identify putative circadian pacemakers in the bee brain, we used antibodies against Drosophila melanogaster and Antheraea pernyi PERIOD and an antiserum to crustacean pigment-dispersing hormone (PDH) known to cross-react with insect pigment-dispersing factors (PDFs). In contrast to previous results from Drosophila, PDH and PER immunoreactivity (-ir) were not colocalized in bee neurons. The most intense PER-ir was cytoplasmic, in two groups of large neurons in the protocerebrum. The number of protocerebral PER-ir neurons and PER-ir intensity within individual cells were highest in brains collected during subjective night and higher in old bees than in young bees. These results are consistent with previous analyses of brain per mRNA in honeybees. Nuclear PER-ir was found throughout the brain, including the optic and antennal lobes. A single group of PDH-ir neurons (approximately 20/optic lobe) was consistently and intensely labeled at the medial margin of the medulla, independent of age or time of day. The processes of these neurons extended to specific neuropils in the protocerebrum and the optic lobes but not to the deutocerebrum. The patterns displayed by PER- and PDH-ir do not completely match any patterns previously described. This suggests that, although clock proteins are conserved across insect groups, there is no universal pattern of coexpression that allows ready identification of pacemaker neurons within the insect brain. J. Comp. Neurol. (C) 2003 Wiley-Liss, Inc.