New microsatellite loci for the European bushcricket,Ephippiger ephippiger (Orthoptera: Tettigoniidae)

Ephippiger ephippiger is a model organism for studies of sexual selection and phylogeography but little is known about fine‐scale population structure. Available microsatellite loci have null allele problems so we used an enrichment technique to isolate 21 new microsatellite loci for E. ephippiger. We present primer pairs for 10 polymorphic loci (3–11 alleles per locus). Observed heterozygosities at polymorphic loci ranged from 0.118 to 0.787, but several were significantly lower than expected.     

Photoreceptor development in the pineal organ and the eye of Plecoglossus altivelis and Paralichthys olivaceus (Teleostei)

Summary The ontogenetic apperance of pineal photo-receptors was compared with that of retinal photoreceptors in the ayu Plecoglossus altivelis and the lefteye flounder Paralichthys olivaceus, which hatched 10 days and 3 days after fertilization, respectively. Despite the disparity in incubation time, the outer segments (containing membranous lamellae) of the pineal photoreceptors first appeared from 3 to 4 days after fertilization in both species. In contrast, the outer segments of the retinal photoreceptors first became visible 5 to 6 days after fertilization, although a characteristic retinal stratification and the optic tract leaving the ganglion cell layer were already found 4 days after fertilization in both species. The functional significance of these temporal disparities and/or similarities in photoreceptor development are discussed with special reference to the timing of daily rhythmic activities during the early developmental period of the teleosts.The results were presented at the Annual Meeting of the Japanese Society of Scientific Fisheries on April 2, 1990 (Tokyo)     

Postembryonic development of leucokinin I-immunoreactive neurons innervating a neurohemal organ in the turnip moth Agrotis segetum

Summary In the abdominal ganglia of the turnip moth Agrotis segetum, an antibody against the cockroach neuropeptide leucokinin I recognizes neurons with varicose fibers and terminals innervating the perisympathetic neurohemal organs. In the larva, the abdominal perisympathetic organs consist of a segmental series of discrete neurohemal swellings on the dorsal unpaired nerve and the transverse nerves originating at its bifurcation. These neurohemal structures are innervated by varicose terminals of leucokinin I-immunoreactive (LKIR) fibers originating from neuronal cell bodies located in the preceding segment. In the adult, the abdominal segmental neurohemal units are more or less fused into a plexus that extends over almost the whole abdominal nerve cord. The adult plexus consists of peripheral nerve branches and superficial nerve fibers beneath the basal lamina of the neural sheath of the nerve cord. During metamorphosis, the LKIR fibers closely follow the restructuration of the perisympathetic organs. In both larvae and adults the LKIR fibers in the neurohemal structures originate from the same cell bodies, which are distributed as ventrolateral bilateral pairs in all abdominal ganglia. The transformation of the series of separated and relatively simple larval neurohemal organs into the larger, continuous and more complex adult neurohemal areas occurs during the first of the two weeks of pupal life. The efferent abdominal LKIR neurons of the moth Agrotis segetum thus belong to the class of larval neurons which persist into adult life with substantial peripheral reorganization occurring during metamorphosis.     

Ultrastructure of the chemosensitive basiconic single-walled wall-pore sensilla on the antennae in adults and embryonic stages of Locusta migratoria L. (Insecta,Orthoptera)

Summary The structure and embryonic development of the two types (A, B) of basiconic sensilla on the antennae of Locusta migratoria were studied in material that had been cryofixed and freeze-substituted, or chemically fixed and dehydrated. Both types are single-walled wall-pore sensilla. Type-A sensilla comprise 20–30 sensory and 7 enveloping cells. One enveloping cell (thecogen cell secretes the dendrite sheath); four are trichogen cells, projections of which form the trichogen process during the 2nd embryonic molt. The trichogen cells form two concentric pairs proximally. Two tormogen cells secrete the cuticular socket of the sensillum. The dendritic outer segments of the sensory cells are branched. Bifurcate type-A sensilla have also been observed. Type-B sensilla comprise three sensory and four enveloping cells (one thecogen, two trichogen and one tormogen). The trichogen process is formed by the two trichogen cells, each of which gives rise to two projections. The trichogen cells are concentrically arranged. The dendritic outer segments of the sensory cells are unbranched. In the fully developed sensillum, all trichogen and tormogen cells border on the outer receptor lymph cavity. It is suggested that the multicellular organization of the type-A sensilla can be regarded as being advanced rather than primitive.Supported by the Dcutschc Forschungsgemeinschaft (SFB 4/G1)     

Sensory cell degeneration in the ontogeny of the chemosensitive sensilla in the labial palp-pit organ of the butterfly,Pieris rapae L. (Insecta,Lepidoptera)

Summary The ontogeny of the chemoreceptive sensilla in the labial palp-pit organ was studied in Pieris rapae by examining twelve successive stages between pupation and emergence of the imago, which takes a period of 160 h under the experimental conditions. Mitoses occur until 20 h after pupation. They lead to anlagen of sensilla, 91% of which are comprised of three sensory cells. However, two sensory cells degenerate in each sensillum during a period of 28 h. The same process occurs in anlagen with four sensory cells resulting in bicellular sensilla. Axons grow out only after the number of sensory cells has been reduced. Further consecutive steps in sensory cell differentiation are: (a) outgrowth of dendritic outer segment and dendrite sheath; (b) outgrowth of trichogen process and change in structure of elongating dendrite sheath; (c) deposition of cuticle and pore tubules in the pegs; (d) retraction of trichogen process; (e) increase in diameter of dendritic outer segment accompanied by increase of microtubule number and appearance of regularly spaced electron-dense bodies at tubular doublets; (f) branching of dendritic outer segment; and (g) transformation of the dendritic branches into curled lamellae and partial destruction of the dendrite sheath. The unique process of sensory cell degeneration is interpreted as an event that revokes a step towards a possible functional improvement of the labial palp-pit organ during further evolutionSupported by the Deutsche Forschungsgemeinschaft (SFB 4/G1)     

The embryonic development of the Drosophila visual system

We have used electron-microscopic studies, bromodeoxyuridine (BrdU) incorporation and antibody labeling to characterize the development of the Drosophila larval photoreceptor (or Bolwig's) organ and the optic lobe, and have investigated the role of Notch in the development of both. The optic lobe and Bolwig's organ develop by invagination from the posterior procephalic region. After cells in this region undergo four postblastoderm divisions, a total of approximately 85 cells invaginate. The optic lobe invagination loses contact with the outer surface of the embryo and forms an epithelial vesicle attached to the brain. Bolwig's organ arises from the ventralmost portion of the optic lobe invagination, but does not become incorporated in the optic lobe; instead, its 12 cells remain in the head epidermis until late in embryogenesis when they move in conjunction with head involution to reach their final position alongside the pharynx. Early, before head involution, the cells of Bolwig's organ form a superficial group of 7 cells arranged in a rosette pattern and a deep group of 5 cells. Later, all neurons move out of the surface epithelium. Unlike adult photoreceptors, they do not form rhabdomeres; instead, they produce multiple, branched processes, which presumably carry the photopigment. Notch is essential for two aspects of the early development of the visual system. First, it delimits the number of cells incorporated into Bolwig's organ. Second, it is required for the maintenance of the epithelial character of the optic lobe cells during and after its invagination.     

Postembryonic development of serotonin-immunoreactive neurons in the central nervous system of the blowfly

Summary Neurons immunoreactive with antibodies to serotonin (5-HT) were mapped in the thoracico-abdominal ganglia of the blowfly, Calliphora erythrocephala, during postembryonic development. Reconstructions from serial sections of tissue processed with a preincubation PAP-method permitted a detailed analysis of the morphological changes occurring in 5-HT-immunoreactive (5-HTi) neurons.All the 5-HTi cell bodies in the thoracico-abdominal ganglia of the 3rd instar larva, except two in the metathoracic ganglion, retain their immunochemical phenotype throughout pupal development. Hence, all the adult 5-HTi neurons in these ganglia differentiate during embryonic development. The finer processes of the larval 5-HTi neurons undergo a substantial regression during the first 24 h of pupal development, and thereafter new branches form on the primary processes of the same cell bodies. The slight change in relative position of 5-HTi cell bodies and the reorganization of the neuropil into an adult pattern occur during the first half of pupal development. The neuropil mass and extent of 5-HTi processes continue to increase during the following days and appear to be fully developed two days (80% of pupal development) before hatching.On the basis of the presented data, some of the basic processes are discussed that lead to the transformation of the larval nervous system into its adult form.     

Sensory basis for sound intensity discrimination in the bushcricket Requena verticalis (Tettigoniidae, Orthoptera)

The ability of the female bushcricket, Requena verticalis, to discriminate between two conspecific sound signals that differed in sound pressure level (SPL) was tested in a two-choice paradigm. Significant discrimination was achieved with a 2-dB difference. The property of each pair of receptors to establish binaural discharge differences was investigated in electrophysiological experiments. The threshold to the conspecific signal varies for each fibre from about 40 to 90 dB SPL, allowing for a range fractionation of the hearing organ. Each pair of receptors establishes significant binaural discharge differences only within a restricted intensity range about 10 dB above threshold. Based on a model of the intensity response function of a receptor the total discharge of the 22 receptors in both ears was calculated with monaural and binaural stimulation. The profile of receptors exhibiting significant discharge differences changes with increasing SPL, from the most sensitive fibres with a characteristic frequency between 12 kHz and 35 kHz at low SPLs to the least sensitive fibres at very low and high characteristic frequencies at medium to high SPLs. The discharge difference with an intensity difference of 2 dB is rather small (4% of the total receptor activity) and limited only to a few pairs of receptors. Accepted: 8 November 1997     

Inferring the patterns and causes of geographic variation in Ephippiger ephippiger (Orthoptera, Tettigoniidae) using geographical information systems (GIS)

Ephippiger ephippiger is a tcltigoniid cricket that is fairly widespread in Europe. In southern France, between the Alps and the Pyrenees, it is geographically highly variable lor a number of different traits including morphology, allozymcs, behaviour and genetic markers. This variation has resulted in considerable confusion over the taxonomic and evolutionary status of forms of the organism. Through the use of geographical information systems (GIS) it has been possible to integrate both trait and environmental data sets from a variety of sources. The GIS is used to interpolate trait and environmental surfaces that are then examined for coincidence. Four general spatial patterns are identified. These can be explained through a combination of secondary contact, environmental adaptation and drift. Covariance between trait and environmental surfaces point to patterns which are primary in origin, whereas other dines probably reflect secondary contact following divergence in rcfugiae. Multivariate statistics support the validity of a major subdivision identified by GIS. This study is an example of the considerable potential for GIS as an investigative tool in evolutionary studies.     

The development of GABA-like immunoreactivity in the thoracic ganglia of the locust Schistocerca gregaria

Summary The development of GABA-like immunoreactivity was investigated in embryonic and juvenile locusts using an antibody raised against GABA-protein conjugates. GABA-like immunoreactivity was first detectable in the neuropile of embryonic ganglia at 55% development, and in neuronal somata at 62% development. The total number of immunoreactive somata increased between 62% and 85% embryonic development, and followed an anterio-posterior pattern of expression. At 85% development, the number of immunoreactive somata reached adult levels and no change in number was then seen. In embryonic stages and first and second juvenile instars two dorsal and four ventral groups of somata were labeled in all three thoracic ganglia, whilst in later juvenile instars one of the dorsal groups was visible as a separate entity only in the metathoracic ganglion. These early patterns were modified by alterations in the positions of some of the groups during late embryogenesis and during juvenile development to produce the adult pattern. The results show that the development of GABA expression is similar to that of other neurotransmitters. The characteristics of the development of immunoreactivity indicate that some of these immunoreactive clusters may be derived from clonally related neurones. Finally, we demonstrate the presence of immunoreactive somata and processes in embryos, which correspond to those of identified local and intersegmental interneurones studied in the adult.Abbreviations Ab1–3 first-third abdominal ganglion - CON connective - CI _1–3 common inhibitors 1–3 - CTC tract - DC I–VII dorsal commissures I–VII - DIT dorsal intermediate tract - DMT dorsal median tract - LDT lateral dorsal tract - LF lateral fibres - o, iLVT outer and inner lateral ventral tract - MVT median ventral tract - N1–5 nerves 1–5 - aPT anterior perpendicular tract - PT perpendicular tract - aRT anterior ring tract - R1–5 nerve roots 1–5 - PVC posterior ventral commissure - SMC supra-median commissure - T3 metathoracic neuromere - TT T tract - aVAC anterior ventral association centre - VC I ventral commissure I - d,vVCII dorsal and ventral parts of ventral commissure II - VF ventral fibres - VIT ventral intermediate tract - VLT ventral lateral tract - VMT ventral median tract - (d,v)LAG (dorsal and ventral) lateral anterior group - LDG lateral dorsal group - LVG lateral ventral group - MDG medial dorsal group - MPG medial posterior group - MVG medial ventral group     

Postsmolt change in numbers of acetylcholinesterase-positive cells in the pineal organ of the Pacific coho salmon

Summary We have examined the occurrence of acetylcholinesterase (AChE)-positive cells in the pineal organ of different developmental stages of the Pacific coho salmon. Large numbers of AChE cells were present in fresh-water living alevins, in all stages of presmolts (n=307–544), and in adult spawners (n=696–1774) whereas seawater-living postmolts displayed a total lack of labeled cells. The AChE-reactive cells were evently distributed within the pineal end-vesicle and stalk of the presmolts and adults. However, the AChE-positive cells that occurred in the pineal stalk were of a smaller type and more uniform in shape than the cells of the pineal endvesicle. The dense populations of AChE-stained cells in the alevins, were all situated in the caudal part of the pineal end-vesicle. We conclude that changes in pineal metabolism occur in postsmolt salmon that liver in saltwater. It is not clear whether the observed change in pineal AChE expression is an unspecific change caused by life in the sea, reflecting alterations that are related to aspects of osmoregulation, and/or is involved in the visual function of the pineal organ resulting from changes in the environmental lighting conditions, e.g., photoperiod, light-intensity, or spectral composition. This study adds to our previous findings of changes that occur in the central nervous system of the salmon during the time of the parr-smolt transformation and migration between limnic and marine environments and indicates a possible central role of the pineal organ in the control of these events.     

Pathfinding of peripheral neurons in the central nervous system of an embryonic grasshopper (Chorthippus biguttulus)

The peripheral leg nerves of grasshoppers are initially formed by a set of pioneer neurons and guidepost cells. These cells are used as guiding structures for later-arising axons of sensory neurons. The development of the central projections of the pioneer cells, the guidepost cells and some sensory cells is shown with Lucifer Yellow injection or with DiI application. The axons of the pioneer cells Ti1 enter the central nervous system at 38% of embryonic development. They turn anteriorly close to the midline and ascend with no major branching to the brain. The axons of the guidepost cells Fe1 and Tr1 follow the same path but do not ascend to the brain. Sensory axons of the subgenual organ and the femoral organ probably do not follow the central path pioneered by the former neurons. They end ipsilaterally in the respective thoracic neuromere, as is found in the adult.     

Evidence for a frontal-organ homologue in the pineal complex of the salamander,Hynobius dunni

The pineal complex of larval and adult salamanders, Hynobius dunni, was examined by light and scanning electron microscopy. This pineal complex displays an anterior and a posterior portion, both of which possess a lumen. The anterior lumen is small and closed, whereas the posterior lumen is in open communication with the third ventricle. Cell processes of the photoreceptor cells and microvilli of the supportive cells are visible in both lumina. The anterior part of the complex is formed by an independent, second evagination from the common pineal anlage; this process takes place immediately after hatching. The anterior body of the pineal complex of H. dunni appears to be homologous to the frontal organ of anurans.     

Comparison of auditory sense organs in parasitoid Tachinidae (Diptera) hosted by Tettigoniidae (Orthoptera) and homologous structures in a non-hearing Phoridae (Diptera)

The dipteran parasitoids Therobia leonidei and Homotrixa alleni (Tachinidae) use acoustic cues to locate their calling tettigoniid (Ensifera, Orthoptera) hosts. The sexually dimorphic tympanal organs of both fly species are located at the prosternum. For comparison a homologous chordotonal organ in the non-hearing fly Phormia regina, Meigen (Phoridae) is also described. The scolopidial sense organs of the ears have approximately 180 sensory cells in Th. leonidei and 250 cells in H. alleni. Interspecific analysis indicates that the cell number and arrangement might be genus specific in Tachinidae. The mononematic scolopidia, each with one sensory cell, are of different sizes and insert at the tympanal membrane. Large scolopidial units (diameter of sensory cells up to 50 μm) extend longitudinally from the centre of the sensory organ towards the ligament, whereas small units (sensory cell diameter up to 10 μm) are arranged sequentially within the sensory organ. This arrangement is discussed to be a possible basis for frequency discrimination. The ultrastructure of the scolopidia is similar in the hearing and non-hearing flies. In both groups, the majority of scolopales has a diameter from 2 to 2.9 μm, although hearing species have additionally wider scolopales. The homologous chordotonal organ of Ph. regina consists of approximately 55 sensory cells of uniform direction. The data are discussed in comparison to the ears of other Diptera.     

Postembryonic development of Arg-Phe-amide-like and cholecystokinin-like immunoreactive neurons in the blowfly optic lobe

Summary The adult optic lobes of the blowfly Calliphora erythrocephala were found to be innervated by more than 2000 neurons immunoreactive to antisera raised against the neuropeptides FMRFamide, its fragment RFamide, and gastrin/cholecystokinin (CCK). All of the CCK-like immunoreactive (CCK-IR) neurons also reacted with antisera to RFamide, FMRFamide and pancreatic polypeptide. A few RFamide/FMRFamide-like immunoreactive (RF-IR) neurons did not react with CCK antisera; they reacted instead with antisera to Leu-enkephalin and Met-enkephalin-Arg⁶-Phe⁷. The RF-IR neurons are, thus, heterogeneous with respect to their contents of immunoreactive peptides. Two of the RF-IR neuron types innervating the adult optic lobes could be traced in their entirety only after following their postembryonic development, because of the complexity of the trajectories of the immunoreactive neuronal process in the adult insect. The majority of the cell bodies of the RF-IR and CCK-IR neurons lie within the optic lobes and are derived from imaginal neuroblasts of the inner and outer optic anlagen. Six of the peptidergic neurons are, however, metamorphosing larval neurons with their cell bodies in the central part of the protocerebrum. The full extent of immunoreactivitiy is not attained in some of the neurons until the late pupal or early adult stage. The larval optic center was also found to be innervated by neurons immuno-reactive with both RFamide and CCK antisera. The cell bodies of these RF-IR/CCK-IR neurons are located near the developing lamina (one on each side). In the 24 h pupa, the cell bodies of these neurons are still immunoreactive, but thereafter they cannot be immunolabeled apparently due to cell death or a change in transmitter phenotype.     

Ultrastructure of the taste disc in the red-bellied toad Bombina orientalis (Discoglossidae,Salientia)

The taste disc of the red-bellied toad Bombina orientalis (Discoglossidae) has been investigated by light and electron microscopy and compared with that of Rana pipiens (Ranidae). Unlike the frog, B. orientalis possesses a disc-shaped tongue that cannot be ejected for capture of prey. The taste discs are located on the top of fungiform papillae. They are smaller than those in Ranidae, and are not surrounded by a ring of ciliated cells. Ultrastructurally, five types of cells can be identified (mucus cells, wing cells, sensory cells, and both Merkel cell-like basal cells and undifferentiated basal cells). Mucus cells are the main secretory cells of the taste disc and occupy most of the surface area. Their basal processes do not synapse on nerve fibers. Wing cells have sheet-like apical processes and envelop the mucus cells. They contain lysosomes and multivesicular bodies. Two types of sensory cells reach the surface of the taste disc; apically, they are distinguished by either a brush-like arrangement of microvilli or a rod-like protrusion. They are invaginated into lateral folds of mucus cells and wing cells. In contrast to the situation in R. pipiens, sensory cells of B. orientalis do not contain dark secretory granules in the perinuclear region. Synaptic connections occur between sensory cells (presynaptic sites) and nerve fibers. Merkel cell-like basal cells do not synapse onto sensory cells, but synapse-like connections exist between Merkel cell-like basal cells (presynaptic site) and nerve fibers.     

Proctolin-immunoreactive neurons persist during metamorphosis of an insect: A developmental study of the ventral nerve cord of Tenebrio molitor(Coleoptera)

Summary Proctolin-immunoreactive neurons in all neuromers of the ventral nerve cord of Tenebrio molitor L. have been quantitatively demonstrated and mapped throughout metamorphosis. Each neuromer contains an anterior and a posterior group of neurons with light and dark staining properties as revealed by peroxidase-antiperoxidase labeling. Serial homologous subsets of dark staining neurons with central and peripheral projections have been identified and found to persist during morphogenetic changes from the larva to the adult. Most neurons maintain their topological and structural characteristics throughout metamorphosis. The identified proctolin-immunoreactive neurons exhibit structures similar to those described in other insect species; some may correspond known motoneurons.     

Axonal pathfinding in developing labial palps of the butterflies,Pieris rapae and Pieris brassicae

Summary Electron microscopy was used to study the developmental changes in the pupal labial palp of Pieris rapae L. and P. brassicae L. prior to axogenesis of the peripheral receptor organs. Two cells emigrate from the anlage of the labial palp-pit organ (LPPO). They develop growth cones and filopodia, which are directed distally. The filopodia insert into stem cells of the LPPO. The perikarya of the cells proceed into the hemolymph space of the palp such that, eventually, these cells bridge the gap between the LPPO and the apical scolopidial organ (ASO) of the labial palp. This bridge is established at about 11% of total pupal development. Only 6 h later, at 15% of pupal development, the cells are no longer visible. We suggest that these cells are luminal neurons and name them pit-organ luminal (POL) cells. These POL cells may act as primary guiding structures for the axons of the sensory cells of the LPPO, which are assumed to orientate along this structure once they reach the ASO.Supported by the Deutsche Forschungsgemeinschaft (H.A.: SFB4/G1).     

Structure,development and death of sensory cells and neurons in the pupal labial palp of the butterflies Pieris rapae L. and Pieris brassicae L. (Insecta,Lepidoptera)

Summary The development of neurons possibly related to the outgrowth of axons from the labial palp-pit organ was studied in Pieris rapae. Serial sections of six successive stages between pupation and emergence of the imago were examined with the electron microscope. At pupation the palp contains an apical scolopidial organ (ASO) and cellular strands connected to it. The ASO consists of three type-1 scolopidia, which are characterized by the presence of a ciliary 9 × 2 + 0 pattern throughout the dendritic outer segment and a ciliary dilation beneath the cap. The scolopidia show two special features: (i) the dendritic outer segments reach beyond the cap, and (ii) an intricate junctional complex develops between the dendritic inner segments and the scolopale cells. The cellular strands comprise two types of cells: (1) bipolar cells regarded as neurons due to their cytological features, and (2) enveloping cells, which are wrapped around the bipolar cells. The strands degenerate about 10 h after pupation. The sensory cells of the ASO degenerate consecutively between 28 h and 130 h after pupation. However, their enveloping cells survive and endure in the imago, which emerges about 160 h after pupation. An ASO similarly lacking sensory cells was observed in imagines of Pieris brassicae. It is hypothesized that the ASO and the bipolar neurons of the strands play a role in pathfinding of the axons of the labial palp-pit organ.Supported by the Deutsche Forschungsgemeinschaft (SFB 4/G1)