The spatial distribution of input and output neurons in the mushroom body (MB) calyx was investigated in the silkmoth Bombyx mori. In Lepidoptera, the brain has a specialized system for processing sex pheromones. How individual pheromone components are represented in the MB has not yet been elucidated. Toward this end, we first compared the distribution of the presynaptic boutons of antennal lobe projection neurons (PNs), which transfer odor information from the antennal lobe to the MB calyx. The axons of PNs that innervate pheromonal glomeruli were confined to a relatively small area within the calyx. In contrast, the axons of PNs that innervate nonpheromonal glomeruli were more widely distributed. PN axons for the minor pheromone component covered a larger area than those for the major pheromone component and partially overlapped with those innervating nonpheromonal glomeruli, suggesting the integration of the minor pheromone component with plant odors. Overall, we found that PN axons innervating pheromonal and nonpheromonal glomeruli were organized into concentric zones. We then analyzed the dendritic fields of Kenyon cells (KCs), which receive inputs from PNs. Despite the strong regional localization of axons of different PN classes, the dendrites of KCs were less well classified. Finally, we estimated the connectivity between PNs and KCs and suggest that the dendritic field may be organized to receive different amounts of pheromonal and nonpheromonal inputs. PNs for multiple pheromone components and plant odors enter the calyx in a concentric fashion, and they are read out by the elaborate dendritic field of KCs.

In insects, olfactory information in the glomeruli of the antennal lobe, the first olfactory center, is transmitted to the lateral protocerebrum and the calyx of the mushroom body via projection neurons. In male silkmoths (Bombyx mori), arborization patterns in the calyx differ markedly between projection neurons that respond to sex pheromones and those that respond to general odors. However, little is known about the organization of the mushroom body's intrinsic neurons, called Kenyon cells (KCs), which receive the inputs from the projection neurons. We investigated the silkmoth mushroom body and identified four parallel subdivisions in the lobes and pedunculus by immunolabeling with antibodies against the catalytic subunit of protein kinase A in Drosophila melanogaster (DC0) and the neuromodulatory peptide FMRFamide. To further understand the detailed organization of the mushroom body, we performed extensive labeling of individual KCs. We identified four morphological types whose axonal projections corresponded to the subdivisions in the lobes, and found that each type of KC had a characteristic dendritic morphology in the calyx. These results show a correlation between the axonal projections of KCs in the lobes and dendritic morphology in the calyx, and indicate different functional roles for the subdivisions.

The insect antennal lobe (AL) is the structural and functional analog of the olfactory bulb of mammals, in which odor information is spatially and/or temporally represented by functional glomerular units. Local interneurons (LNs) play critical roles through intra- and interglomerular communication to shape the output from the AL to higher brain centers; however, the function and even the components of LNs are unclear. We have used morphological and immunocytochemical approaches to examine LNs in the silkworm moth, Bombyx mori. First, we comprehensively analyzed the morphological variation of LNs. One hundred fifty-three AL LNs were intracellularly stained, analyzed in three dimensions with a confocal microscope, and subdivided into five morphological types based on differences in the arborization region in the AL and dendritic profiles within the glomeruli. Two global multiglomerular types arborized in the macroglomerular complex (MGC) and in most ordinary glomeruli, and the other three oligoglomerular types innervated some ordinary glomeruli with or without the MGC. Second, we performed double-labeling of Lucifer Yellow staining of a single LN combined with gamma-aminobutyric acid (GABA) immunocytochemistry. The two global multiglomerular types and two of the oligoglomerular types were GABA-immunoreactive, but the third oligoglomerular type, which innervates the MGC and some ordinary glomeruli, included some GABA-immunonegative neurons, suggesting the existence of a non-GABAergic subtype. These results suggest that the complex neural circuits of the AL are composed of several morphologically different types of LNs, most of which are inhibitory.