Tiling of the Drosophila epidermis by multidendritic sensory neurons

Insect dendritic arborization (da) neurons provide an opportunity to examine how diverse dendrite morphologies and dendritic territories are established during development. We have examined the morphologies of Drosophila da neurons by using the MARCM (mosaic analysis with a repressible cell marker) system. We show that each of the 15 neurons per abdominal hemisegment spread dendrites to characteristic regions of the epidermis. We place these neurons into four distinct morphological classes distinguished primarily by their dendrite branching complexities. Some class assignments correlate with known proneural gene requirements as well as with central axonal projections. Our data indicate that cells within two morphological classes partition the body wall into distinct, non-overlapping territorial domains and thus are organized as separate tiled sensory systems. The dendritic domains of cells in different classes, by contrast, can overlap extensively. We have examined the cell-autonomous roles of starry night (stan) (also known as flamingo (fmi)) and sequoia (seq) in tiling. Neurons with these genes mutated generally terminate their dendritic fields at normal locations at the lateral margin and segment border, where they meet or approach the like dendrites of adjacent neurons. However, stan mutant neurons occasionally send sparsely branched processes beyond these territories that could potentially mix with adjacent like dendrites. Together, our data suggest that widespread tiling of the larval body wall involves interactions between growing dendritic processes and as yet unidentified signals that allow avoidance by like dendrites.     

Dendrites of distinct classes of Drosophila sensory neurons show different capacities for homotypic repulsion

BACKGROUND: Understanding how dendrites establish their territory is central to elucidating how neuronal circuits are built. Signaling between dendrites is thought to be important for defining their territories; however, the strategies by which different types of dendrites communicate are poorly understood. We have shown previously that two classes of Drosophila peripheral da sensory neurons, the class III and class IV neurons, provide complete and independent tiling of the body wall. By contrast, dendrites of class I and class II neurons do not completely tile the body wall, but they nevertheless occupy nonoverlapping territories. RESULTS: By developing reagents to permit high-resolution studies of dendritic tiling in living animals, we demonstrate that isoneuronal and heteroneuronal class IV dendrites engage in persistent repulsive interactions. In contrast to the extensive dendritic exclusion shown by class IV neurons, duplicated class III neurons showed repulsion only at their dendritic terminals. Supernumerary class I and class II neurons innervated completely overlapping regions of the body wall, and this finding suggests a lack of like-repels-like behavior. CONCLUSIONS: These data suggest that repulsive interactions operate between morphologically alike dendritic arbors in Drosophila. Further, Drosophila da sensory neurons appear to exhibit at least three different types of class-specific dendrite-dendrite interactions: persistent repulsion by all branches, repulsion only by terminal dendrites, and no repulsion.     

Drosophila sensory neurons require Dscam for dendritic self-avoidance and proper dendritic field organization

A neuron's dendrites typically do not cross one another. This intrinsic self-avoidance mechanism ensures unambiguous processing of sensory or synaptic inputs. Moreover, some neurons respect the territory of others of the same type, a phenomenon known as tiling. Different types of neurons, however, often have overlapping dendritic fields. We found that Down's syndrome Cell Adhesion Molecule (Dscam) is required for dendritic self-avoidance of all four classes of Drosophila dendritic arborization (da) neurons. However, neighboring mutant class IV da neurons still exhibited tiling, suggesting that self-avoidance and tiling differ in their recognition and repulsion mechanisms. Introducing 1 of the 38,016 Dscam isoforms to da neurons in Dscam mutants was sufficient to significantly restore self-avoidance. Remarkably, expression of a common Dscam isoform in da neurons of different classes prevented their dendrites from sharing the same territory, suggesting that coexistence of dendritic fields of different neuronal classes requires divergent expression of Dscam isoforms.     

Morphology,Classification, and Distribution of the Projection Neurons in the Dorsal Lateral Geniculate Nucleus of the Rat

The morphology of confirmed projection neurons in the dorsal lateral geniculate nucleus (dLGN) of the rat was examined by filling these cells retrogradely with biotinylated dextran amine (BDA) injected into the visual cortex. BDA-labeled projection neurons varied widely in the shape and size of their cell somas, with mean cross-sectional areas ranging from 60–340 µm². Labeled projection neurons supported 7–55 dendrites that spanned up to 300 µm in length and formed dendritic arbors with cross-sectional areas of up to 7.0×10⁴ µm². Primary dendrites emerged from cell somas in three broad patterns. In some dLGN projection neurons, primary dendrites arise from the cell soma at two poles spaced approximately 180° apart. In other projection neurons, dendrites emerge principally from one side of the cell soma, while in a third group of projection neurons primary dendrites emerge from the entire perimeter of the cell soma. Based on these three distinct patterns in the distribution of primary dendrites from cell somas, we have grouped dLGN projection neurons into three classes: bipolar cells, basket cells and radial cells, respectively. The appendages seen on dendrites also can be grouped into three classes according to differences in their structure. Short “tufted” appendages arise mainly from the distal branches of dendrites; “spine-like” appendages, fine stalks with ovoid heads, typically are seen along the middle segments of dendrites; and “grape-like” appendages, short stalks that terminate in a cluster of ovoid bulbs, appear most often along the proximal segments of secondary dendrites of neurons with medium or large cell somas. While morphologically diverse dLGN projection neurons are intermingled uniformly throughout the nucleus, the caudal pole of the dLGN contains more small projection neurons of all classes than the rostral pole.     

Integrins regulate repulsion-mediated dendritic patterning of drosophila sensory neurons by restricting dendrites in a 2D space

Dendrites of the same neuron usually avoid each other. Some neurons also repel similar neurons through dendrite-dendrite interaction to tile the receptive field. Nonoverlapping coverage based on such contact-dependent repulsion requires dendrites to compete for limited space. Here we show that Drosophila class IV dendritic arborization (da) neurons, which tile the larval body wall, grow their dendrites mainly in a 2D space on the extracellular matrix (ECM) secreted by the epidermis. Removing neuronal integrins or blocking epidermal laminin production causes dendrites to grow into the epidermis, suggesting that integrin-laminin interaction attaches dendrites to the ECM. We further show that some of the previously identified tiling mutants fail to confine dendrites in a 2D plane. Expansion of these mutant dendrites in three dimensions results in overlap of dendritic fields. Moreover, overexpression of integrins in these mutant neurons effectively reduces dendritic crossing and restores tiling, revealing an additional mechanism for tiling.     

Development of morphological diversity of dendrites in Drosophila by the BTB-zinc finger protein abrupt

Morphological diversity of dendrites contributes to specialized functions of individual neurons. In the present study, we examined the molecular basis that generates distinct morphological classes of Drosophila dendritic arborization (da) neurons. da neurons are classified into classes I to IV in order of increasing territory size and/or branching complexity. We found that Abrupt (Ab), a BTB-zinc finger protein, is expressed selectively in class I cells. Misexpression of ab in neurons of other classes directed them to take the appearance of cells with smaller and/or less elaborated arbors. Loss of ab functions in class I neurons resulted in malformation of their typical comb-like arbor patterns and generation of supernumerary branch terminals. Together with the results of monitoring dendritic dynamics of ab-misexpressing cells or ab mutant ones, all of the data suggested that Ab endows characteristics of dendritic morphogenesis of the class I neurons.     

Presynaptic dendrite cells and two other classes of neurons in the superficial layers of the superior colliculus of the chimpanzee

In the superior colliculus of chimpanzee, three classes of neurons can be identified by ultrastructural criteria. They are 1) marginal cells located in the stratum zonale, 2) collicular relay cells in the stratum griseum superficiale and 3) presynaptic dendrite (PSD) cells, i.e., neurons with presynaptic specializations in soma and/or dendrites. PSD cells are the smallest neurons in the stratum griseum superficiale; they have a relatively large, deeply infolded nucleus and a small rim of cytoplasm rich in free ribosomes. PSD cells are sufficiently different from the two other classes of neurons to be reliably identified at the ultrastructural level. They closely resemble presynaptic neurons as described in the lateral geniculate nucleus of other mammalian species. Presynaptic dendrites in continuity with PSD cells are rich in organelles, especially ribosomal cluster, and establish en passage contact with other dendrites. Another type of presynaptic dendrite, poor in organelles, except for bundles of microtubules, could not be traced back to its parent neuron. Homo- or heterogeneity of PSD cells is discussed. No amxon was traced from a PSD cell.     

Shapes of nerve cells in the myenteric plexus of the guinea-pig small intestine revealed by the intracellular injection of dye

Summary The shapes of myenteric neurons in the guineapig small intestine were determined after injecting living neurons with the dye Lucifer yellow via a microelectrode. The cells were fixed and the distribution of Lucifer yellow rendered permanent by an immunohistochemical method. Each of 204 nerve cells was examined in whole-mount preparations of the myenteric plexus and drawn using a camera lucida at 1250 x magnification. Four cell shapes were distinguished: (1) neurons with several long processes corresponding to type II of Dogiel; (2) neurons with a single long process and lamellar dendrites corresponding to type I of Dogiel; (3) neurons with numerous filamentous dendrites; and (4) small neurons with few processes. About 15% of the neurons could not be placed into these classes or into any single class. The type II neurons (39% of the sample) had generally smooth somata and up to 7 (average 3.3) long processes, most of which ran circumferentially. Dogiel type I neurons (34% of sampled neurons) had characteristic lamellar dendrites, i.e., broad dendrites that were flattened in the plane of the plexus. The filamentous neurons (7% of the sample), had, on average, 14 fine processes up to about 50 m in length. Small neurons with smooth outlines and a few fine processes made up 5% of the neurons encountered. We conclude that myenteric neurons that have been injected with dye can be separated into morphologically distinct classes and that the different morphological classes probably correspond to different functional groupings of neurons.     

Microtubules have opposite orientation in axons and dendrites of Drosophila neurons

In vertebrate neurons, axons have a uniform arrangement of microtubules with plus ends distal to the cell body (plus-end-out), and dendrites have equal numbers of plus- and minus-end-out microtubules. To determine whether microtubule orientation is a conserved feature of axons and dendrites, we analyzed microtubule orientation in invertebrate neurons. Using microtubule plus end dynamics, we mapped microtubule orientation in Drosophila sensory neurons, interneurons, and motor neurons. As expected, all axonal microtubules have plus-end-out orientation. However, in proximal dendrites of all classes of neuron, approximately 90% of dendritic microtubules were oriented with minus ends distal to the cell body. This result suggests that minus-end-out, rather than mixed orientation, microtubules are the signature of the dendritic microtubule cytoskeleton. Surprisingly, our map of microtubule orientation predicts that there are no tracks for direct cargo transport between the cell body and dendrites in unipolar neurons. We confirm this prediction, and validate the completeness of our map, by imaging endosome movements in motor neurons. As predicted by our map, endosomes travel smoothly between the cell body and axon, but they cannot move directly between the cell body and dendrites.     

Golgi analysis of tangential neurons in the lobula plate of<Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The lobula plate (LP), which is the third order optic neuropil of flies, houses wide-field neurons which are exquisitely sensitive to motion. Among Diptera, motion-sensitive neurons of larger flies have been studied at the anatomical and physiological levels. However, the neurons ofDrosophila lobula plate are relatively less explored. AsDrosophila permits a genetic analysis of neural functions, we have analysed the organization of lobula plate ofDrosophila melanogaster. Neurons belonging to eight anatomical classes have been observed in the present study. Three neurons of the horizontal system (HS) have been visualized. The HS north (HSN) neuron, occupying the dorsal lobula plate is stunted in its geometry compared to that of larger flies. Associated with the HS neurons, thinner horizontal elements known as h-cells have also been visualized in the present study. Five of the six known neurons of the vertical system (VS) have been visualized. Three additional neurons in the proximal LP comparable in anatomy to VS system have been stained. We have termed them as additional VS AVS)-like neurons. Three thinner tangential cells that are comparable to VS neurons, which are elements of twin vertical system (tvs); and two cells with wide dendritic fields comparable to CH neurons of Diptera have been also observed. Neurons comparable to VS cells but with ‘tufted’ dendrites have been stained. The HSN and VS1-VS2 neurons are dorsally stunted. This is possibly due to the shape of the compound eye ofDrosophila which is reduced in the fronto-dorsal region as compared to larger flies     

Early events in insect neurogenesis. II. The role of cell interactions and cell lineage in the determination of neuronal precursor cells

The insect central nervous system (CNS) is composed of a brain and a chain of segmental ganglia; each hemiganglion contains about 1000 individually identifiable neurons. How is the enormous neuronal diversity and specificity generated? Neurons of a hemiganglion largely arise during embryogenesis from a stereotyped pattern of individually identified neuronal precursor cells, called neuroblasts (NBs). The transition from ectoderm to individual neurons thus involves two major steps: first, an undifferentiated ectodermal cell sheet produces the stereotyped pattern of 30 NBs per hemisegment; second, each of these NBs contributes a specific family of neuronal progeny to the developing CNS. We have used a laser microbeam to ablate individual cells in the grasshopper embryo in order to study the initial events of neuronal determination. In particular, how does a layer of apparently equivalent ectodermal cells produce a highly stereotyped pattern of unique NBs? Our results suggest the following mechanism for NB determination. (1) Cell interactions between the approximately 150 equivalent ectodermal cells of a hemisegment allow 30 cells to enlarge into NBs. (2) As these young NBs enlarge they inhibit adjacent ectodermal cells from becoming NBs; the adjacent cells then either differentiate into nonneuronal support cells or die. (3) Each NB is assigned a unique identity due to its position of enlargement within the neuroepithelium. (4) The NB then generates its characteristic family of neurons by an invariant cell lineage. Development of the insect CNS depends on cell interactions and positional cues to create a pattern of NBs, and then on cell lineage to restrict the fate of the NB progeny.     

Early differences between alternate n blast cells in leech embryo

Segmentally iterated tissues of the mature leech comprise five distinct sets of definitive progeny that arise from chains of blast cells (m, n, o, p, and q bandlets) produced by five bilateral pairs of stem cells (M, N, O/P, O/P, and Q teloblasts). In each n and q bandlet, two blast cells are needed to generate one set of hemisegmental progeny, and two alternating classes of blast cells (nf and ns, qf and qs) can be distinguished after their first divisions. Furthermore, two distinct subsets of definitive N and Q progeny exist within each hemisegment. Here we first show that there is fixed correspondence between the class of blast cell and the subset of final progeny: ns cells contribute mainly anterior ganglionic neurons and epidermal cells; nf cells contribute mainly posterior ganglionic neurons, peripheral neurons and neuropil glia; qs cells contribute both ventral and dorsal progeny; and qf cells contribute only dorsal progeny. Second, ablation studies indicate that the two classes of n blast cells do not behave as an equivalence group in the germinal band. Finally, we show that the cycles giving rise to nf and ns blast cells differ. These data suggest that cellular interactions within the germinal band may not be critical in establishing the distinct nf and ns cell fates and that, conversely, differences between the two classes of n blast cells may be established at birth.     

Neuronal types in the lateral geniculate nucleus of man

Summary Nerve cell types of the lateral geniculate body of man were investigated with the use of a transparent Golgi technique that allows study of not only the cell processes but also the pigment deposits. Three types of neurons have been distinguished:Type-I neurons are medium-to large-sized multipolar nerve cells with radiating dendrites. Dendritic excrescences can often be encountered close to the main branching points. Type-I neurons comprise a variety of forms and have a wide range of dendritic features. Since all intermediate forms can be encountered as well, it appears inadequate to subdivide this neuronal type. One pole of the cell body contains numerous large vacuolated lipofuscin granules, which stain weakly with aldehyde fuchsin.Type-II and type-III neurons are small cells with few, sparsely branching and extended dendrites devoid of spines. In Golgi preparations they cannot be distinguished from each other. Pigment preparations reveal that the majority of these cells contains small and intensely stained lipofuscin granules within their cell bodies (type II), whereas a small number of them remains devoid of any pigment (type III). Intermediate forms do not occur.     

The hair-peg organs of the shore crab,Carcinus maenas (Crustacea,Decapoda): Ultrastructure and functional properties of sensilla sensitive to changes in seawater concentration

Summary The hair-peg organs of the shore crab, Carcinus maenas, are modified hair-sensilla. A small hair shaft (peg) is surrounded by a tuft of solid cuticular bristles (hairs). Each hair-peg organ is innervated by 6 sensory neurons, 2 of which have scolopidial (type-I) dendrites. The outer segments of all dendrites pass through a cuticular canal extending to the articulated hair base in which the 2 type-I dendrites terminate. The other 4 (type-II) dendrites reach the clavate tip of the hair shaft and have access to a terminal pore and a large sickle-shaped aperture. Three inner and 8–12 outer enveloping cells belong to a hair-peg organ. The innermost enveloping cell contains a scolopale, which has desmosomal connections to the ciliary rootlets of the type-I dendrites. An inner and an outer sensillum lymph space are present. The ultrastructural features of the dendrites and the cuticular apparatus indicate that the hair-peg organs are bimodal sensilla, comprising 2 mechano- and 4 chemosensitive sensory neurons. Extracellular recordings from the leg nerve indicate that the chemosensitive neurons of the hair-peg organs respond to changes in seawater concentration in the physiological range of Carcinus maenas.Supported by the Deutsche Forschungsgemeinschaft (SFB 45/A1; W. Gnatzy)     

Tracing cells throughout development: Insights into single glial cell differentiation

In the article “Predetermined embryonic glial cells form the distinct glial sheaths of the Drosophila peripheral nervous system” we combined our expertise to identify glial cells of the embryonic peripheral nervous system on a single cell resolution with the possibility to genetically label cells using Flybow. We show that all 12 embryonic peripheral glial cells (ePG) per abdominal hemisegment persist into larval (and even adult) stages and differentially contribute to the three distinct glial layers surrounding peripheral nerves. Repetitive labelings of the same cell further revealed that layer affiliation, morphological expansion, and control of proliferation are predetermined and subject to an intrinsic differentiation program. Interestingly, wrapping and subperineurial glia undergo enormous hypertrophy in response to larval growth and elongation of peripheral nerves, while perineurial glia respond to the same environmental changes with hyperplasia. Increase in cell number from embryo (12 cells per hemisegment) to third instar (up to 50 cells per hemisegment) is the result of proliferation of a single ePG that serves as transient progenitor and only contributes to the outermost perineurial glial layer.     

Morphology of neurons in the torus semicircularis of the northern leopard frog,Rana pipiens pipiens

The neuronal morphology of the torus semicircularis of the northern leopard frog, Rana pipiens pipiens, was examined in Golgi-impregnated material. Neurons in each of the five subdivisions of the torus semicircularis (Potter, '65a) have distinct morphologies which are characteristic of the subdivision. Laminar nucleus neurons are mostly multipolar with spherical or ovoidal somata and smooth dendrites oriented primarily parallel and perpendicular to the cell laminae. Principal nucleus neurons have variable soma shapes with short dendrites ( < 100 μm) radiating in all directions. In the magnocellular nucleus, there are three major cell types: neurons characterized by small, spherical-shaped somata, with short, thin, radiating dendrites and many varicosities; bi- or tripolar neurons with ovoidal somata, and long (100–200 μm) and smooth dendrites orienting primarily dorsoventrally and mediolaterally; and multipolar neurons with triangular-shaped somata and very long (200–350 μm) dendrites, which are either smooth or highly spiny. Neurons in the commissural nucleus are mostly multipolar cells with ovoidal somata and beaded dendrites projecting mostly dorsally and ventrally. The subependymal midline nucleus contains mostly uni- or bipolar neurons with small ovoidal somata and straight, spiny dendrites. In addition to revealing the morphological features of neurons in the torus, the counterstained material shows further cytoarchitectural organization of the principal nucleus, i.e., the presence of a circular lamellar organization. The functional significance of these anatomical features is discussed.     

The target of rapamycin complex 2 controls dendritic tiling of Drosophila sensory neurons through the Tricornered kinase signalling pathway

To cover the receptive field completely and non‐redundantly, neurons of certain functional groups arrange tiling of their dendrites. In Drosophila class IV dendrite arborization (da) neurons, the NDR family kinase Tricornered (Trc) is required for homotypic repulsion of dendrites that facilitates dendritic tiling. We here report that Sin1, Rictor, and target of rapamycin (TOR), components of the TOR complex 2 (TORC2), are required for dendritic tiling of class IV da neurons. Similar to trc mutants, dendrites of sin1 and rictor mutants show inappropriate overlap of the dendritic fields. TORC2 components physically and genetically interact with Trc, consistent with a shared role in regulating dendritic tiling. Moreover, TORC2 is essential for Trc phosphorylation on a residue that is critical for Trc activity in vivo and in vitro. Remarkably, neuronal expression of a dominant active form of Trc rescues the tiling defects in sin1 and rictor mutants. These findings suggest that TORC2 likely acts together with the Trc signalling pathway to regulate the dendritic tiling of class IV da neurons, and thus uncover the first neuronal function of TORC2 in vivo.     

Apterous is a Drosophila LIM domain gene required for the development of a subset of embryonic muscles.

The recently discovered LIM motif is found in a set of homeodomain-containing proteins thought to mediate the generation of particular cell types. Of the four LIM domain family members described to date, mec-3 and lin-11 determine cell lineages in C. elegans. Isl-1 and Xlim-1 may play similar roles in vertebrates. We have identified a Drosophila member of this class, the product of the apterous (ap) gene. During embryogenesis, ap is expressed in a small subset of fusing mesodermal precursors that give rise to 6 muscles in each abdominal hemisegment and in 5 neurons within each corresponding CNS hemisegment. Lack of ap function results in loss of ap-expressing muscles, while misexpression of ap using a heterologous promoter produces ectopic muscles.     

Are the most numerous sensilla of terrestrial isopods hygroreceptors? Ultrastructure of the dorsal tricorn sensilla of Porcellio scaber

The ultrastructure of the tricorn sensilla of the woodlouse Porcellio scaber was investigated in cryofixed and freeze-substituted, or chemically fixed specimens. The tricorn sensilla have a foramenized triangular-shaped outer hair and bear a poreless rod-like inner hair. The conical base of the inner hair is connected to the base of the outer hair by a complex cuticular structure. Each sensillum contains three sensory cells. The tip of one of the three dendrites contains a tubular body and is clamped between two bulges of the dendritic sheath. The two other dendrites protrude to the tip of the inner hair, flush against the cuticular wall. The microtubules in the ciliary segments are arranged in nine double tubuli that have neither osmiophilic cores nor arms. The ciliary rootlets are small. The inner segment of the largest dendrite wraps around the two smaller dendrites and one of seven enveloping cells in a mesaxon-like manner. Although this ultrastructure deviates considerably from most crustacean mechanosensitive sensilla, it nevertheless suggests a mechanosensitive function, at least for one of the sensory cells. In many aspects, the tricorn sensilla resemble the thermohygrosensilla of insects. However, our results suggest that the structural criteria for thermo-hygro-sensitivity used in insects cannot simply be applied to crustaceans.