Azusa Kamikouchi

Comprehensive classification of the auditory sensory projections in the brain of the fruit fly Drosophila melanogaster

We established a comprehensive projection map of the auditory receptor cells (Johnstons organ neurons: JONs) from the antennae to the primary auditory center of the Drosophila brain. We found 477 ± 24 cell bodies of JONs, which are arranged like a “bottomless bowl” within the auditory organ. The target of the JONs in the brain comprises five spatially segregated zones, each of which is contributed by bundles of JON axons that gradually branch out from the antennal nerve. Four zones are confined in the antennal mechanosensory and motor center, whereas one zone further extends over parts of the ventrolateral protocerebrum and the subesophageal ganglion. Single‐cell labeling with the FLP‐out technique revealed that most JONs innervate only a single zone, indicating that JONs can be categorized into five groups according to their target zones. Within each zone, JONs innervate various combinations of subareas. We classified these five zones into 19 subareas according to the branching patterns and terminal distributions of single JON axons. The groups of JONs that innervate particular zones or subareas of the primary auditory center have their cell bodies in characteristic locations of the Johnstons organ in the antenna, e.g., in concentric rings or in paired clusters. Such structural organization suggests that each JON group, and hence each zone of the primary auditory center, might sense different aspects of sensory signals. J. Comp. Neurol. 499:317–356, 2006.

Distinct sensory representations of wind and near-field sound in the Drosophila brain

Behavioural responses to wind are thought to have a critical role in controlling the dispersal and population genetics of wild Drosophila species, as well as their navigation in flight, but their underlying neurobiological basis is unknown. We show that Drosophila melanogaster, like wild-caught Drosophila strains, exhibits robust wind-induced suppression of locomotion in response to air currents delivered at speeds normally encountered in nature. Here we identify wind-sensitive neurons in Johnston’s organ, an antennal mechanosensory structure previously implicated in near-field sound detection (reviewed in refs 5 and 6). Using enhancer trap lines targeted to different subsets of Johnston’s organ neurons, and a genetically encoded calcium indicator, we show that wind and near-field sound (courtship song) activate distinct populations of Johnston’s organ neurons, which project to different regions of the antennal and mechanosensory motor centre in the central brain. Selective genetic ablation of wind-sensitive Johnston’s organ neurons in the antenna abolishes wind-induced suppression of locomotion behaviour, without impairing hearing. Moreover, different neuronal subsets within the wind-sensitive population respond to different directions of arista deflection caused by air flow and project to different regions of the antennal and mechanosensory motor centre, providing a rudimentary map of wind direction in the brain. Importantly, sound- and wind-sensitive Johnston’s organ neurons exhibit different intrinsic response properties: the former are phasically activated by small, bi-directional, displacements of the aristae, whereas the latter are tonically activated by unidirectional, static deflections of larger magnitude. These different intrinsic properties are well suited to the detection of oscillatory pulses of near-field sound and laminar air flow, respectively. These data identify wind-sensitive neurons in Johnston’s organ, a structure that has been primarily associated with hearing, and reveal how the brain can distinguish different types of air particle movements using a common sensory organ.

Specification of auditory sensitivity by Drosophila TRP channels

Ears achieve their exquisite sensitivity by means of mechanical feedback: motile mechanosensory cells through their active motion boost the mechanical input from the ear. Examination of the auditory mechanics in Drosophila melanogaster mutants shows that the transient receptor potential (TRP) channel NompC is required to promote this feedback, whereas the TRP vanilloid (TRPV) channels Nan and Iav serve to control the feedback gain. The combined function of these channels specifies the sensitivity of the fly auditory organ.

Carbohydrate metabolism genes and pathways in insects: insights from the honey bee genome.

Carbohydrate‐metabolizing enzymes may have particularly interesting roles in the honey bee, Apis mellifera, because this social insect has an extremely carbohydrate‐rich diet, and nutrition plays important roles in caste determination and socially mediated behavioural plasticity. We annotated a total of 174 genes encoding carbohydrate‐metabolizing enzymes and 28 genes encoding lipid‐metabolizing enzymes, based on orthology to their counterparts in the fly, Drosophila melanogaster, and the mosquito, Anopheles gambiae. We found that the number of genes for carbohydrate metabolism appears to be more evolutionarily labile than for lipid metabolism. In particular, we identified striking changes in gene number or genomic organization for genes encoding glycolytic enzymes, cellulase, glucose oxidase and glucose dehydrogenases, glucose‐methanol‐choline (GMC) oxidoreductases, fucosyltransferases, and lysozymes.

Identification of a novel gene, Mblk-1, that encodes a putative transcription factor expressed preferentially in the large-type Kenyon cells of the honeybee brain

Mushroom bodies (MBs) are considered to be involved in higher‐order sensory processing in the insect brain. To identify the genes involved in the intrinsic function of the honeybee MBs, we searched for genes preferentially expressed therein, using the differential display method. Here we report a novel gene encoding a putative transcription factor (Mblk‐1) expressed preferentially in one of two types of intrinsic MB neurones, the large‐type Kenyon cells, which makes Mblk‐1 a candidate gene involved in the advanced behaviours of honeybees. A putative DNA binding motif of Mblk‐1 had significant sequence homology with those encoded by genes from various animal species, suggesting that the functions of these proteins in neural cells are conserved among the animal kingdom.

Concentrated expression of Ca2+/ calmodulin-dependent protein kinase II and protein kinase C in the mushroom bodies of the brain of the honeybee Apis mellifera L.

We have previously used the differential display method to identify a gene that is expressed preferentially in the mushroom bodies of worker honeybees and to show that it encodes a putative inositol 1,4,5‐trisphosphate receptor (IP3R) homologue (Kamikouchi et al. [ 1998 ] Biochem. Biophys. Res. Commun. 242:181–186). In the present study, we examined whether the expression of some of the genes for proteins involved in the intracellular Ca2+ signal transduction is also concentrated in the mushroom bodies of the honeybee by isolating cDNA fragments that encode the Ca2+/calmodulin‐dependent protein kinase II (CaMKII) and protein kinase C (PKC) homologues of the honeybee. In situ hybridization analysis revealed that the expression of these genes was also concentrated in the mushroom bodies of the honeybee brain: The CaMKII gene was expressed preferentially in the large‐type Kenyon cells of the mushroom bodies, whereas that for PKC was expressed in both the large and small types of Kenyon cells. The expression of the genes for IP3R and CaMKII was concentrated in the mushroom bodies of the queen and drone as well as in those of the worker bee. Furthermore, the enzymatic activities of CaMKII and PKC were found to be higher in the mushroom bodies/central bodies than in the optic and antennal lobes of the worker bee brain. These results suggest that the function of the intracellular Ca2+ signal transduction is enhanced in Kenyon cells in comparison to other neuronal cell types in the honeybee brain. J. Comp. Neurol. 417:501–510, 2000.

Identification of genes expressed preferentially in the honeybee mushroom bodies by combination of differential display and cDNA microarray1

To clarify the molecular basis underlying the neural function of the honeybee mushroom bodies (MBs), we identified three genes preferentially expressed in MB using cDNA microarrays containing 480 differential display‐positive candidate cDNAs expressed locally or differentially, dependent on caste/aggressive behavior in the honeybee brain. One of the cDNAs encodes a putative type I inositol 1,4,5‐trisphosphate (IP3) 5‐phosphatase and was expressed preferentially in one of two types of intrinsic MB neurons, the large‐type Kenyon cells, suggesting that IP3‐mediated Ca2+ signaling is enhanced in these neurons.

Identification and punctate nuclear localization of a novel noncoding RNA, Ks-1, from the honeybee brain

We identified a novel gene, Ks-1, which is expressed preferentially in the small-type Kenyon cells of the honeybee brain. This gene is also expressed in some of the large soma neurons in the brain and in the suboesophageal ganglion. Reverse transcription-polymerase chain reaction experiments indicated that Ks-1 transcripts are enriched in the honeybee brain. cDNA cloning revealed that the consensus Ks-1 cDNA is over 17 kbp and contains no significant open reading frames. Furthermore, fluorescent in situ hybridization revealed that Ks-1 transcripts are located in the nuclei of the neural cells, accumulating in some scattered spots. These findings demonstrate that Ks-1 encodes a novel class of noncoding nuclear RNA and is possibly involved in the regulation of neural functions.

The Nutrient-Responsive Hormone CCHamide-2 Controls Growth by Regulating Insulin-like Peptides in the Brain of Drosophila melanogaster

The coordination of growth with nutritional status is essential for proper development and physiology. Nutritional information is mostly perceived by peripheral organs before being relayed to the brain, which modulates physiological responses. Hormonal signaling ensures this organ-to-organ communication, and the failure of endocrine regulation in humans can cause diseases including obesity and diabetes. In Drosophila melanogaster, the fat body (adipose tissue) has been suggested to play an important role in coupling growth with nutritional status. Here, we show that the peripheral tissue-derived peptide hormone CCHamide-2 (CCHa2) acts as a nutrient-dependent regulator of Drosophila insulin-like peptides (Dilps). A BAC-based transgenic reporter revealed strong expression of CCHa2 receptor (CCHa2-R) in insulin-producing cells (IPCs) in the brain. Calcium imaging of brain explants and IPC-specific CCHa2-R knockdown demonstrated that peripheral-tissue derived CCHa2 directly activates IPCs. Interestingly, genetic disruption of either CCHa2 or CCHa2-R caused almost identical defects in larval growth and developmental timing. Consistent with these phenotypes, the expression of dilp5, and the release of both Dilp2 and Dilp5, were severely reduced. Furthermore, transcription of CCHa2 is altered in response to nutritional levels, particularly of glucose. These findings demonstrate that CCHa2 and CCHa2-R form a direct link between peripheral tissues and the brain, and that this pathway is essential for the coordination of systemic growth with nutritional availability. A mammalian homologue of CCHa2-R, Bombesin receptor subtype-3 (Brs3), is an orphan receptor that is expressed in the islet β-cells; however, the role of Brs3 in insulin regulation remains elusive. Our genetic approach in Drosophila melanogaster provides the first evidence, to our knowledge, that bombesin receptor signaling with its endogenous ligand promotes insulin production.

Identification of Honeybee Antennal Proteins/Genes Expressed in a Sex- and/or Caste Selective Manner

Abstract We identified three candidate proteins/genes involved in caste and/or sex-specific olfactory processing in the honeybee Apis mellifera L., that are differentially expressed between the antennae of the worker, queen, and drone honeybees using SDS-polyacrylamide gel electrophoresis or the differential display method. A protein was identified, termed D-AP1, that was expressed preferentially in drone antennae when compared to those of workers. cDNA cloning revealed that D-AP1 is homologous to carboxylesterases. Enzymatic carboxylesterase activity in the drone antennae was higher than in the workers, suggesting its dominant function in the drone antennae. In contrast, two proteins encoded by genes termed W-AP1 and Amwat were expressed preferentially in worker antennae when compared to those of queens. W-AP1 is homologous to insect chemosensory protein, and Amwat encodes a novel secretory protein. W-AP1 is expressed selectively in worker antennae, while Amwat is expressed both in the antennae and legs of the workers. These findings suggest that these proteins are involved in the antennal function characteristic to drone or worker honeybees.