Immunohistochemical localization of clock proteins (DBT and PER), and [His<sup>7</sup>]- and [Arg<sup>7</sup>]-corazonins in the cerebral ganglia of Bombyx mori: Are corazonins downstream regulators of circadian clocks?

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The brain and subesophageal ganglion (BR-SG) of the commercial silk worm, Bombyx mori, were stained immunohistochemically at the larval stage for circadian clock neurons with antibodies against Doubletime (DBT) of B. mori and Period (PER) of Periplaneta americana. The BR-SGs were also stained with antisera against [Arg7]-corazonin, which has been known to be present in B. mori and co-localized with PER in Manduca sexta, and against [His7]-corazonin, a homolog identified in other species. From co-localization of [Arg7]-corazonin and PER-like reactivities in the pars lateralis, [Arg7]-corazonin is suspected to be a downstream regulator of the circadian clock in M. sexta. DBT- and corazonin-like immunohistochemical reactivities were found in both the neurosecretory cells of the pars intercerebralis (PIC) and pars lateralis (PL) in B. mori. Small numbers of neurons shared both reactivities against anti-DBT and anti-corazonin. The majority of the immunopositive cells were common to both corazonins, but some cells were unique in expressing either reactivity against [His7]-corazonin or [Arg7]-corazonin only. The results suggest that there is a diversity in the clock output pathway among lepidopterans and that [His7]-corazonin may be present in B. mori, as well as [Arg7]-corazonin, although the former has not been chemically identified in this species. Corazonin may be a downstream regulator of circadian clocks in B. mori because of the co-localization of [His7]-corazonin at PIC and [Arg7]-corazonin at PL with anti-DBT.