Michiko Nishikawa

Projection neurons originating from thermo- and hygrosensory glomeruli in the antennal lobe of the cockroach.

Most insects are equipped with specialized thermo‐ and hygroreceptors to locate a permissible range of ambient temperature and distant water sources, respectively. In the cockroach, Periplaneta americana, cold, moist, and dry receptor cells in the antennae send axons to particular sets of two or three glomeruli in the dorsocentral part of the antennal lobe (primary olfactory center), designated DC1–3 glomeruli. However, it is not known how thermo‐ and hygrosensory signals from these glomeruli are represented in higher‐order centers, the protocerebrum, in any insect species. With the use of intracellular recording and staining techniques, we identified a new class of interneurons with dendrites almost exclusively in the DC1, DC2, or DC3 glomeruli and axons projecting to the protocerebrum in the cockroach. Remarkably, terminals of all these projection neurons (PNs) covered almost identical areas in the lateral protocerebrum (LP), although their termination areas outside the LP differed from neuron to neuron. The termination areas within the LP were distinct from, but close to, those of uniglomerular and macroglomerular PNs that transmitted signals concerning general odors and female sex pheromones, respectively. PNs originating from DC1, DC2, and DC3 glomeruli exhibited excitatory responses to cold, moist, and dry stimuli, respectively, probably due to excitatory synaptic input from cold, moist, and dry receptor cells, respectively, whereas their responses were often modulated by olfactory stimuli. These findings suggested that dorsocentral PNs participate in neural pathways that lead to behavioral responses to temperature or humidity changes. J. Comp. Neurol. 455:40–55, 2003.

Sex-specific antennal sensory system in the ant Camponotus japonicus: structure and distribution of sensilla on the flagellum

The antennae are a critically important component of the ant’s highly elaborated chemical communication systems. However, our understanding of the organization of the sensory systems on the antennae of ants, from peripheral receptors to central and output systems, is poorly understood. Consequently, we have used scanning electron and confocal laser microscopy to create virtually complete maps of the structure, numbers of sensory neurons, and distribution patterns of all types of external sensilla on the antennal flagellum of all types of colony members of the carpenter ant Camponotus japonicus. Based on the outer cuticular structures, the sensilla have been classified into seven types: coelocapitular, coeloconic, ampullaceal, basiconic, trichoid-I, trichoid-II, and chaetic sensilla. Retrograde staining of antennal nerves has enabled us to count the number of sensory neurons housed in the different types of sensilla: three in a coelocapitular sensillum, three in a coeloconic sensillum, one in an ampullaceal sensillum, over 130 in a basiconic sensillum, 50–60 in a trichoid-I sensillum, and 8–9 in a trichoid-II sensillum. The basiconic sensilla, which are cuticular hydrocarbon-receptive in the ant, are present in workers and unmated queens but absent in males. Coelocapitular sensilla (putatively hygro- and thermoreceptive) have been newly identified in this study. Coelocapitular, coeloconic, and ampullaceal sensilla form clusters and show biased distributions on flagellar segments of antennae in all colony members. The total numbers of sensilla per flagellum are about 9000 in unmated queens, 7500 in workers, and 6000 in males. This is the first report presenting comprehensive sensillar maps of antennae in ants.

Functional and topographic segregation of glomeruli revealed by local staining of antennal sensory neurons in the honeybee Apis mellifera

In the primary olfactory center of animals, glomeruli are the relay stations where sensory neurons expressing cognate odorant receptors converge onto interneurons. In cockroaches, moths, and honeybees, sensory afferents from sensilla on the anterodorsal surface and the posteroventral surface of the flagellum form two nerves of almost equal thicknesses. In this study, double labeling of the two nerves, or proximal/distal regions of the nerves, with fluorescent dyes was used to investigate topographic organization of sensory afferents in the honeybee. The sensory neurons of ampullaceal sensilla responsive to CO2, coelocapitular sensilla responsive to hygrosensory, and thermosensory stimuli and coeloconic sensilla of unknown function were characterized with large somata and supplied thick axons exclusively to the ventral nerve. Correspondingly, all glomeruli innervated by sensory tract (T) 4 received thick axonal processes exclusively from the ventral nerve. Almost all T1–3 glomeruli received a similar number of sensory afferents from the two nerves. In the macroglomerular complexes of the drone, termination fields of afferents from the two nerves almost completely overlapped; this differs from moths and cockroaches, which show heterogeneous terminations in the glomerular complex. In T1–3 glomeruli, sensory neurons originating from more distal flagellar segments tended to terminate within the inner regions of the cortical layer. These results suggest that some degree of somatotopic organization of sensory afferents exist in T1–3 glomeruli, and part of T4 glomeruli serve for processing of hygro‐ and thermosensory signals. J. Comp. Neurol. 515:161–180, 2009.

Dual, multilayered somatosensory maps formed by antennal tactile and contact chemosensory afferents in an insect brain.

The antennae of most insects move actively and detect the physical and chemical composition of objects encountered by using their associated tactile sensors. Positional information is required for these sensory modalities to interpret the physical environment. Although we have a good understanding of antennal olfactory pathways, little is known about the destinations of antennal mechanosensory and contact chemosensory (gustatory) receptor neurons in the central nervous system. The cockroach Periplaneta is equipped with a pair of long, thin antennae, which are covered in bristles. The distal portions of each antenna possess about 6,500 bimodal bristles that house one tactile sensory and one to four contact chemosensory neurons. In this study, we investigated the morphologies of bimodal bristle receptor afferents by staining individual or populations of bristles. Unlike olfactory afferents, which project exclusively into the glomeruli in the ventral region of the deutocerebrum, both the presumptive mechanosensory and the contact chemosensory afferents projected into the posterior dorsal region of the deutocerebrum and the anterior region of the subesophageal ganglion. Each afferent showed multilayered segmentation and spatial occupation reflecting its three‐dimensional position in the periphery. Presumptive contact chemosensory afferents, characterized by their thin axons and unique branching pattern, occupied more medioventral positions compared with the presumptive tactile afferents. Furthermore, projection fields of presumptive contact chemosensory afferents from single sensilla tended to be segregated from each other. These observations suggest that touch and taste positional information from the antenna is precisely represented in primary centers in a modality‐specific manner. J. Comp. Neurol. 493:291–308, 2005.

Topography of four classes of Kenyon cells in the mushroom bodies of the cockroach

Mushroom bodies (MBs), which are higher centers in the insect brain, are implicated in associative memory and in the control of some behaviors. Intrinsic neurons of the MB, called Kenyon cells, receive synaptic inputs from axon terminals of input neurons in the calyx. Axons of Kenyon cells project into the pedunculus and to the α and β lobes, where they make synaptic connections with dendrites of extrinsic (output) neurons. In this study, we examined the morphology of Kenyon cells in the cockroach by using Golgi stains and found that they can be classified into four classes (K1, K2, K3, and K4), according to the diameter, location, and morphology of the cell bodies, dendrites, and axons. The somata of Kenyon cells of different classes occupy different concentric zones; Kl cells occupy the most central zone, and K4 cells occupy the most peripheral zone. The main processes of Kenyon cells of different classes also occupy different concentric zones in the calyx. Dendrites of K2 and K3 cells are distributed throughout the calycal neuropil, whereas those of K1 and K4 cells cover the outer and inner halves of the depth of the neuropil, respectively. In the pedunculus and the α and β lobes, axons of Kenyon cells of different classes occupy different zones, although the separation is not complete. A class of extrinsic neurons in the α lobe has dendrite‐like arbors that cover the zones where either K1, K2, or K3 are located. These neurons probably transmit signals of each class of Kenyon cells. We conclude that, in the cockroach, four classes of Kenyon cells subdivide the cell body region, pedunculus, and lobes of the MBs, whereas subdivision is less prominent in the calycal neuropil. J. Comp. Neurol. 399:162–175, 1998.

Modular structures in the mushroom body of the cockroach

The mushroom body (MB) is a higher center of the insect brain and is critical to olfactory and other forms of associative memory. Here, we report that repetitive modular subunits, which we refer to as slabs, are present in the internal matrix of the alpha lobe, a major output neuropil of the MB in the cockroach. The methods employed were osmium-ethyl gallate, Bodian-reduced silver, and Golgi staining procedures. A total of 15 dark and 15 pale slabs, each consisting of specific subsets of intrinsic neurons (Kenyon cells), alternate throughout the length of the alpha lobe. One of the major classes of MB output neurons, which are postsynaptic to Kenyon cells, exhibited segmented dendritic arbors that interact with every other slabs, i.e. only either dark or pale slabs. As each output neuron interacts with each specific set of dark or pale slabs, the slab likely functions as a unit for transmitting MB output signals.

Three classes of GABA-like immunoreactive neurons in the mushroom body of the cockroach

The mushroom body (MB) is a higher center of the insect brain and is critical to some forms of associative memory. Each MB consists of calyces connected to alpha and beta lobes via pedunculus. In the calyces, input neurons make synaptic connections with intrinsic neurons. In the pedunculus and lobes, intrinsic neurons make synaptic connections with output neurons. Here, the distribution of gamma-aminobutyric acid (GABA)-like immunoreactivity in the MB of the cockroach Periplaneta americana was investigated, using an antiserum against a GABA-protein conjugate, to elucidate inhibitory pathways of the MB. We report that three classes of extrinsic neurons of the MB exhibit GABA-like immunoreactivity. The first is four large neurons which arborize in a diffuse neuropil surrounding the alpha lobe and project into whole areas of the calyces. Their cell bodies are 30-50 micron in diameter, among the largest in the brain. The second group is 7-9 neurons ascending from the circumesophageal connective and projecting into the calyces, which probably represent inhibitory input neurons. The third group is ca. 40 neurons with dendritic arborizations in the junction between the pedunculus and the lobes, which probably represent inhibitory output neurons.

Sexual Dimorphism in the Antennal Lobe of the Ant Camponotus japonicus

Abstract The carpenter ant, a social hymenopteran, has a highly elaborated antennal chemosensory system that is used for chemical communication in social life. The glomeruli in the antennal lobe are the first relay stations where sensory neurons synapse onto interneurons. The system is functionally and structurally similar to the olfactory bulbs of vertebrates. Using three-dimensional reconstruction of glomeruli and subsequent morphometric analyses, we found sexual dimorphism of the antennal lobe glomeruli in carpenter ants, Camponotus japonicus. Female workers and unmated queens had about 430 glomeruli, the highest number reported so far in ants. Males had a sexually dimorphic macroglomerulus and about 215 ordinary glomeruli. This appeared to result from a greatly reduced number of glomeruli in the postero-medial region of the antennal lobe compared with that in females. On the other hand, sexually isomorphic glomeruli were identifiable in the dorsal region of the antennal lobe. For example, large, uniquely shaped glomeruli located at the dorso-central margin of the antennal lobe were detected in all society members. The great sexual dimorphism seen in the ordinary glomeruli of the antennal lobe may reflect gender-specific tasks in chemical communications rather than different reproductive roles.

Response characteristics of two types of cold receptors on the antennae of the cockroach, Periplaneta americana L.

Summary1.There are two types of cold receptors on the antennae of Periplaneta americana. One type of cold receptor is present in a thermo- and hygroreceptive sensillum along with a moist receptor and a dry receptor, and the other is in a thermoreceptive and olfactory sensillum along with olfactory receptors. These two types of cold receptors were investigated using stimulations of constant temperature (T), stepwise decrease of temperature (ΔT), and constant rates of temperature decrease (dT/dt).2.For cold receptors in the thermo- and hygroreceptive sensilla (C-H cells), steady-state activities are higher at lower constant temperatures (T) within the range of 16–31 °C examined (n = 7). On the other hand, those of the cold receptors in the thermoreceptive and olfactory sensilla (C-O cells) have individually a broad activity maximum in the range of 18–27 °C (n = 7).3.The differential sensitivity to downward temperature steps (ΔT) of C-H cells was in the range of - 5.4 to - 12.4 (impulses/s)/°C (n = 11). It was higher at lower adapting temperature, from which changes were initiated, in the range of about 21 to 31 °C, that is, it was reflected by the steady-state activity at the initial and adapting temperature. On the other hand, the differential sensitivity of C-O cells was in the range of - 10.7 to- 15.2 (impulses/s)/°C (n = 9), and was little reflected by the steady-state activity.4.Responses to decreasing temperature in C-H cells (n = 14) are negatively related to the rate of temperature decrease (dT/dt) in the range of about 0 to - 0.16 °C/s at given temperatures, and are also affected by instantaneous temperature (T) in the range of 18 to 30 °C. This dependence is also observed in C-O cells (n = 10), but less pronounced than in C-H cells.

Complete mapping of glomeruli based on sensory nerve branching pattern in the primary olfactory center of the cockroach Periplaneta americana

Glomeruli are structural and functional units in the primary olfactory center in vertebrates and insects. In the cockroach Periplaneta americana, axons of different types of sensory neurons housed in sensilla on antennae form dorsal and ventral antennal nerves and then project to a number of glomeruli. In this study, we identified all antennal lobe (AL) glomeruli based on detailed innervation patterns of sensory tracts in addition to the shape, size, and locations in the cockroach. The number of glomeruli is ∼205, and no sex‐specific difference is observed. Anterograde dye injections into the antennal nerves revealed that axons supplying the AL are divided into 10 sensory tracts (T1–T10). Each of T1–T3 innervates small, oval glomeruli in the anteroventral region of the AL, with sensory afferents invading each glomerulus from multiple directions, whereas each of T4–T10 innervates large glomeruli with various shapes in the posterodorsal region, with a bundle of sensory afferents invading each glomerulus from one direction. The topographic branching patterns of all these tracts are conserved among individuals. Sensory afferents in a sub‐tract of T10 had axon terminals in the dorsal margin of the AL and the protocerebrum, where they form numerous small glomerular structures. Sensory nerve branching pattern should reflect developmental processes to determine spatial arrangement of glomeruli, and thus the complete map of glomeruli based on sensory nerve branching pattern should provide a basis for studying the functional significance of spatial arrangement of glomeruli and its developmental basis. J. Comp. Neurol. 518:3907–3930, 2010.