Muscles and nerves of the posterior abdomen and genitalia of male Periplaneta americana (L.) (Dictyoptera : Blattidae)

Segmental and intersegmental muscles of abdominal segments 7–10 are described for adult, male Periplaneta americana (L.) (Dictyoptera : Blattidae). Locations of extrinsic and intrinsic genitalic muscles are documented, and the actions of those associated with the right phallomere are hypothesized. Muscles of the 5 abdominal segments are innervated by branches from 5 pairs of segmental nerves and 3 pairs of transverse nerves. These stem from a terminal synganglion, formed during embryogenesis by fusion of neuromeres of abdominal segments 7–11. One pair of segmental nerves issues from each of the 5 neuromeres, and one pair of transverse nerves arises from neuromeres of abdominal segments 7–9. The nerves are traced to the muscles, integument, and reproductive glands, and their peripheral unions are characterized. Serial homologies of the nerves and muscles are proposed, and comparisons are made with neuromusculature of the female.     

Regulation of egg laying and in vitro oviductal contractions in Gryllus bimaculatus

Oviductal contractions and the control of oviposition were investigated in vivo and in vitro in Gryllus bimaculatus females. In vivo experiments showed that oviposition is controlled nervously by both the brain and the last abdominal ganglion, and that one or more neurohormones cause ovipositor movements and abdominal contractions. In vitro, the assay of nerve ganglia and corpora cardiaca extracts on the isolated oviduct showed that they markedly increase the frequency of oviductal contractions. However, the action of thoracic ganglia extracts varies according to a circadian cycle. This observation, combined with the finding that the effects of the corpora cardiaca differ from those of the brain, suggests that each of these organs contains distinct neurohormones. None of the neurotransmitters tested was as potent as brain or ganglia extracts, although octopamine, l-glutamate and proctolin do stimulate oviductal contractions at low concentrations.     

N-β-alanyldopamine and N-acetyldopamine occurrence and synthesis in the central nervous system of Manduca sexta (L.)

N-β-Alanyldopamine (NBAD), N-acetyldopamine (NADA), dopamine (DA), and 3,4-dihydroxyphenylalanine (DOPA) were detected in the brain and ganglia of the central nervous system of larval and adult tobacco hornworms, Manduca sexta, by reversed phase HPLC with electrochemical detection. NBAD predominated in larval neural tissue during development of the fifth instar and increased to peak concentrations of 936, 650 and 263 nmol g⁻¹ in the abdominal ganglia, subesophageal plus thoracic ganglia and brain, respectively, at the wandering stage of development. Concentrations of all catecholamines decreased in the pharate pupa and were generally lowest in the adult nerve cord. DA was the second most abundant catecholamine in larval ganglia, but the primary catecholamine in adult ganglia. Relatively high levels of DOPA also occurred in the ganglia of wandering larvae but not at other times. NADA was detected only in the abdominal ganglia of day 3 larvae. N-Acyltransferases that catalyze synthesis of NBAD and NADA from DA also were present in abdominal ganglia, as demonstrated by analysis of in vitro cultures in which exogenous DA stimulated synthesis of both N-acylated catecholamines. Maximal NBAD synthesis occurred in ganglia removed from wandering stage larvae (9.3 nmol g⁻¹ day⁻¹), whereas NADA synthesis was highest in ganglia from pharate pupae (7.3 nmol g⁻¹ day⁻¹). Thus, N-β-alanylation and N-acetylation are competing metabolic reactions for DA in the hornworm's nervous system. The role played by the N-acylated catecholamines in M. sexta neurophysiology is unknown, but these compounds may be storage or inactive forms of the putative neurotransmitter DA.     

A Matter of Contrast: Yellow Flower Colour Constrains Style Length in Crocus species

Most flowers display distinct colour patterns comprising two different areas. The peripheral large-area component of floral colour patterns attracts flower visitors from some distance and the central small-area component guides flower visitors towards landing sites. Whereas the peripheral colour is largely variable among species, the central colour, produced mostly by anthers and pollen or pollen mimicking floral guides, is predominantly yellow and UV-absorbing. This holds also for yellow flowers that regularly display a UV bull’s eye pattern. Here we show that yellow-flowering Crocus species are a noticeable exception, since yellow-flowering Crocus species–being entirely UV-absorbing–exhibit low colour contrast between yellow reproductive organs and yellow tepals. The elongated yellow or orange-yellow style of Crocus flowers is a stamen-mimicking structure promoting cross-pollination by facilitating flower visitors’ contact with the apical stigma before the flower visitors are touching the anthers. Since Crocus species possess either yellow, violet or white tepals, the colour contrast between the stamen-mimicking style and the tepals varies among species. In this study comprising 106 Crocus species, it was tested whether the style length of Crocus flowers is dependent on the corolla colour. The results show that members of the genus Crocus with yellow tepals have evolved independently up to twelve times in the genus Crocus and that yellow-flowering Crocus species possess shorter styles as compared to violet- and white-flowering ones. The manipulation of flower visitors by anther-mimicking elongated styles in Crocus flowers is discussed.     

Turn the temperature to turquoise: cues for colour change in the male chameleon grasshopper (Kosciuscola tristis) (Orthoptera: Acrididae)

Rapid, reversible colour change is unusual in animals, but is a feature of male chameleon grasshoppers (Kosciuscola tristis). Understanding what triggers this colour change is paramount to developing hypotheses explaining its evolutionary significance. In a series of manipulative experiments the author quantified the effects of temperature, and time of day, as well as internal body temperature, on the colour of male K. tristis. The results suggest that male chameleon grasshoppers change colour primarily in response to temperature and that the rate of colour change varies considerably, with the change from black to turquoise occurring up to 10 times faster than the reverse. Body temperature changed quickly (within 10 min) in response to changes in ambient temperature, but colour change did not match this speed and thus colour is decoupled from internal temperature. This indicates that male colour change is driven primarily by ambient temperature but that their colour does not necessarily reflect current internal temperature. I propose several functional hypotheses for male colour change in K. tristis.     

Distribution of allatostatin in the adult cockroach,Diploptera punctata and effects on corpora allata in vitro

Direct radiochemical measurements of juvenile hormone synthesis showed that corpora allata from adult female Diploptera punctata can be inhibited in vitro by neuropeptides extracted from several ganglia of the central nervous system of females at many stages of the reproductive cycle. Extracts of protocerebra, corpora cardiaca, suboesophageal, thoracic and ventral ganglia all elicited dose-depedent reductions in juvenile hormone synthesis. On a ‘per organ’ basis, the protocerebrum contains the most extractable material. Inhibitory activity of extracts of suboesophageal, thoracic and 6th abdominal ganglia, like that of protocerebra (Rankin et al., 1986) was trypsin sensitive.Glands of high activity were less sensitive to protocerebral extract than those of low activity. The inhibitory effect on glands of low activity was maximal within 1 h, persisted in the presence of protocerebral extract for at least 46 h, and was abolished within 1 h after corpora allata were placed in normal medium. The inhibitory effect of protocerebral extract was not altered by the addition of magnesium to the medium. The extract had a specific effect on synthetic step(s) prior to methylation and epoxidation as demonstrated by enhanced juvenile hormone synthesis in the presence of inhibitory factor and the juvenile hormone precursor, farnesoic acid.     

Pigment‐dispersing factor in the locust abdominal ganglia may have roles as circulating neurohormone and central neuromodulator

Pigment‐dispersing factor (PDF) is a neuropeptide that has been indicated as a likely output signal from the circadian clock neurons in the brain of Drosophila. In addition to these brain neurons, there are PDF‐immunoreactive (PDFI) neurons in the abdominal ganglia of Drosophila and other insects; the function of these neurons is not known. We have analyzed PDFI neurons in the abdominal ganglia of the locust Locusta migratoria. These PDFI neurons can first be detected at about 45% embryonic development and have an adult appearance at about 80%. In each of the abdominal ganglia (A3–A7) there is one pair of lateral PDFI neurons and in each of the A5–A7 ganglia there is additionally a pair of median neurons. The lateral neurons supply varicose branches to neurohemal areas of the lateral heart nerves and perisympathetic organs, whereas the median cells form processes in the terminal abdominal ganglion and supply terminals on the hindgut. Because PDF does not influence hindgut contractility, it is possible that also these median neurons release PDF into the circulation. Release from one or both the PDFI neuron types was confirmed by measurements of PDF‐immunoreactivity in hemolymph by enzyme immunoassay. PDF applied to the terminal abdominal ganglion triggers firing of action potentials in motoneurons with axons in the genital nerves of males and the 8th ventral nerve of females. Because this action is blocked in calcium‐free saline, it is likely that PDF acts via interneurons. Thus, PDF seems to have a modulatory role in central neuronal circuits of the terminal abdominal ganglion that control muscles of genital organs. © 2001 John Wiley & Sons, Inc. J Neurobiol 48: 19–41, 2001     

Surgical generation of supernumerary appendages for studying neuronal specificity in Drosophila melanogaster

The ability of sensory neurons to establish specific synaptic contacts in the central nervous system (CNS) can be studied by changing the spatial relationship between the periphery and the CNS. In contrast to the genetic displacement of appendages by homoeotic mutations, the surgical approach used in this study allows one to place homologous as well as heterologous appendages to the same site on the body surface. Using an improved technique of “surface transplantation,” we generated supernumerary appendages of any desired type in a particular abdominal position. The sensory axons originating from these grafts enter the CNS through the main abdominal nerve and arborize in the fused abdominal ganglia; many fibers extend also into thoracic centers. In the abdominal ganglia, terminals from dorsal transplants (wings and halteres) stay on the ipsilateral side, whereas terminals from ventral transplants (legs and antennae) distribute ipsi- and contralaterally. The same preference holds true for dorsal and ventral abdominal bristles, respectively, whose projection patterns served as a reference. In thoracic ganglia, axons from dorsal and ventral grafts yield completely different terminal patterns. Dorsal grafts project into the ipsilateral wing center, even in the mutant wingless, in which normal wing afferents are suppressed. In contrast, fibers from ventral grafts often extend along the thoracic midline. These data indicate that sensory axons of homologous appendages on the one hand, and their central targets on the other, share serially repeated surface markers. This may enable sensory fibers to recognize centers of homologous appendages.     

The nature of the seasonal colour dimorphism in the scorpion fly,Panorpa japonica thunberg

The scorpion fly, Panorpa japonica, displays a seasonal colour dimorphism by changing from black to yellow in the adult state. The yellow pigment in the integument and haemolymph of the adult fly was identified as sepiapterin, while the black integument pigment was found to be melanin. After analysis of sepiapterin content by high performance liquid chromatography and determination of total haemolymph volume by [carboxyl-¹⁴C]inulin, it was shown that sepiapterin levels in the haemolymph and integument varied greatly both in the two colour types of insects and in the two sexes. Photometric analysis of melanin content showed that melanin levels correlated negatively with sepiapterin levels. These quantitative differences in sepiapterin and melanin fully explain the colour dimorphism in the insect.     

Structure of cephalic ganglia and their changes in the post-embryonic development of <Emphasis Type="Italic">Calliphora vomitoria</Emphasis> (L.) (Diptera,Calliphoridae)

The central nervous system of Calliphora vomitoria larvae is situated in the metathoracic and the first abdominal segments and is characterized by a high degree of oligomerization. It consists of only two ganglia: the supraoesophageal ganglion, or brain, and one large synganglion, a product of fusion of the suboesophageal ganglion, three thoracic, and all the abdominal ganglia. Weak development of the neuropil of the larval optic and olfactory lobes in the supraoesophageal ganglion is the result of a significant reduction of the head capsule and sensory organs in the larvae. The formation of the imaginal optic lobes begins at the III larval instar. The commissure of the future central body is present in the I instar larva, but formation of the imaginal structure of the central complex proceeds in the 3-day pupae and ends at the late pupal stage. The mushroom bodies are represented in the I instar larvae only by the pedunculi; the calyces can be distinguished in the II instar larvae but the final formation of their structure and the lobes of the imaginal type occurs at the pupal stage. The glomeruli in the deutocerebrum are also formed at a late stage of pupal development. Based on the degree of development of ganglia of the central nervous system, we can conclude that individual development of higher Diptera is characterized by deep de-embryonization.     

Peptidomic survey of the locust neuroendocrine system

Neuropeptides are important controlling agents in animal physiology. In order to understand their role and the ways in which neuropeptides behave and interact with one another, information on their time and sites of expression is required. We here used a combination of MALDI-TOF and ESI-Q-TOF mass spectrometry to make an inventory of the peptidome of different parts (ganglia and nerves) of the central nervous system from the desert locust Schistocerca gregaria and the African migratory locust Locusta migratoria. This way, we analysed the brain, suboesophageal ganglion, retrocerebral complex, stomatogastric nervous system, thoracic ganglia, abdominal ganglia and abdominal neurohemal organs. The result is an overview of the distribution of sixteen neuropeptide families, i.e. pyrokinins, pyrokinin-like peptides, periviscerokinins, tachykinins, allatotropin, accessory gland myotropin, FLRFamide, (short) neuropeptide F, allatostatins, insulin-related peptide co-peptide, ion-transport peptide co-peptide, corazonin, sulfakinin, orcokinin, hypertrehalosaemic hormone and adipokinetic hormones (joining peptides) throughout the locust neuroendocrine system.     

Location of the reproductive timer in the male cricket Gryllus bimaculatus DeGeer as revealed by local cooling of the central nervous system

The location of the reproductive timer for the post-copulatory, time-fixed, sexually refractory stage was investigated in the male cricket Gryllus bimaculatus. This stage was defined as the interval between spermatophore protrusion and recommencement of copulation or a calling song. To inactivate the central nervous system locally and reversibly, different body regions were cooled to 10°C for 20-30 min after spermatophore protrusion. A behavioural test then measured the duration of the refractory stage after males recovered from cooling. Males with the head, thorax and anterior abdomen cooled did not show a lengthening of that stage. In contrast, males with the entire abdomen or even the posterior abdominal segments containing only the 6th and terminal (7th-11th) abdominal ganglia showed a lengthening of the refractory stage up to, but not exceeding, the cooling duration. When 20-min cooling was interposed twice after spermatophore protrusion, the refractory stage was lengthened by about 40 min, indicating that interposed cooling did not reset the timer. These results are in agreement with our previous hypothesis that the reproductive timer for the refractory stage in the male cricket is located in the posterior abdominal ganglia, possibly within the terminal abdominal ganglion.     

Convergent evolution of neuroendocrine control of phenotypic plasticity in pupal colour in butterflies

Phenotypic plasticity in pupal colour occurs in three families of butterflies (the Nymphalidae, Papilionidae and Pieridae), typically in species whose pupation sites vary unpredictably in colour. In all species studied to date, larvae ready for pupation respond to environmental cues associated with the colour of their pupation sites and moult into cryptic light (yellow–green) or dark (brown–black) pupae. In nymphalids and pierids, pupal colour is controlled by a neuroendocrine factor, pupal melanization-reducing factor (PMRF), the release of which inhibits the melanization of the pupal cuticle resulting in light pupae. In contrast, the neuroendocrine factor controlling pupal colour in papilionid butterflies results in the production of brown pupae. PMRF was extracted from the ventral nerve chains of the peacock butterfly Inachis io (Nymphalidae) and black swallowtail butterfly Papilio polyxenes (Papilionidae). When injected into pre-pupae, the extracts resulted in yellow pupae in I. io but brown pupae in P. polyxenes. These results suggest that the same neuroendocrine factor controls the plasticity in pupal colour, but that plasticity in pupal colour in these species has evolved independently (convergently).     

Tyrosine and phenylalanine concentrations in haemolymph and tissues of the American cockroach, Periplaneta americana, during metamorphosis

Phenylalanine and tyrosine concentrations were measured in the haemolymph, fat body, and abdominal integument of the American cockroach, Periplaneta americana, during the pre- and post-ecdysial periods of cuticle formation and sclerotization.Gas-liquid chromatography of trimethylsilyl derivatives of phenylalanine, tyrosine, and their metabolites provided a very sensitive and rapid method for determining those amino acids in small haemolymph and tissue samples.Haemolymph tyrosine increased in two stages: initially near apolysis and 16 to 25 hr pre-ecdysis, reaching its highest concentration at ecdysis (3·5 μg tyrosine/mg haemolymph). During that time, total haemolymph tyrosine increased by approximately 700 μg/insect. Fat body and abdominal integument began to accumulate tyrosine near apolysis. Fat body tyrosine peaked between ecdysis and 3·3 hr post-ecdysis whereas abdominal integument tyrosine peaked at ecdysis. Maximum concentrations were 6·0 μg and 4·1 μg tyrosine/mg wet wt. of tissue, respectively. Between ecdysis and 24 hr post-ecdysis, the period of maximum sclerotization, total tyrosine in haemolymph and fat body decreased by approximately 600 μg and 420 μg/insect, respectively. Phenylalanine concentrations did not change significantly in the haemolymph, fat body, or abdominal integument during the pre- and post-ecdysial periods.The cockroach apparently does not store free phenylalanine or tyrosine in the fat body during larval development as compared to tyrosine storage in some Diptera. The rapid increase of haemolymph, fat body, and integument tyrosine just prior to ecdysis suggests another form of storage for this important amino acid.     

Incorporation of arylphorins (LSP-1) and LSP-2 like protein into the integument of Ceratitis capitata during pupariation

The arylphorins and LSP-2 like polypeptide were detected by immunoblotting analysis during development in the integument of C. capitata. In vitro translation of total RNA from fat body and integument during pupariation, clearly revealed that the polypeptides under consideration were exclusively synthesized in the fat body. Furthermore, in vitro experiments demonstrated that radiolabeled arylphorins and LSP-2 like polypeptide were taken up by the integument, in an undegraded state. Immunofluorescence experiments in cross sections of wandering stage larvae and white pupae revealed that the LSP-2 like polypeptide was mainly localized in the epidermal cells, and a very weak signal was also given by the cuticle. Furthermore, the presented results indicated that a small portion of the extracted proteins exist in high molecular weight aggregate(s).     

Head and tail development of the Drosophila embryo involves spalt, a novel homeotic gene

Mutations in spalt (sal), a novel homeotic gene on the second chromosome of Drosophila, cause opposite transformations in two subterminal regions of the embryo: posterior head segments are transformed into anterior thoracic structures and anterior tail segments are transformed into posterior abdominal structures. The embryonic phenotypes of double mutants for sal and various Antennapedia (ANT-C) or bithorax (BX-C) genes indicate that sal acts independently of the hierarchical order of the latter gene complexes. Trans-regulatory gene mutations causing ectopic expression of ANT-C and BX-C genes do not change the realms of sal action. It is proposed that the region-specific action of the sal gene primarily promotes head as opposed to trunk development, while the BX-C gene AbdB distinguishes tail from head.     

The synthesis of hemolymph proteins by the larval epidermis of an insect Calpodes ethlius (Lepidoptera: Hesperiidae)

Sheets of the dorsal abdominal integument from fifth instar larvae of Calpodes ethlius (Lepidoptera: Hesperiidae) were incubated in artificial hemolymph in the presence of [³⁵S]methionine to investigate protein synthesis and vectorial secretion. The epidermis synthesizes and secretes at least 13 polypeptides basally and 15 apically. Two dimensional analysis of proteins labeled in vitro and in vivo showed that (a) most of the polypeptides secreted on apical and basal surfaces are different, (b) in vitro apical secretions are the same as in vivo cuticular proteins, (c) at least four of the basal secretions can be demonstrated in hemolymph labeled in vivo.Antibodies made against whole hemolymph recognized five basally secreted polypeptides and one apically secreted polypeptide both on fluorograms of immunoprecipitates and immunoblots. Arylphorin is secreted from both surfaces. Arylphorin synthesized in vitro has been identified through its precipitation by antibodies to hemolymph arylphorin in epidermis, cuticle and medium. We conclude that insect epidermis has bi-directional secretion. Cuticular proteins are carried to the apical face. A different set of proteins are carried basally to the hemolymph.     

Time studies of ecdysone-action on in vitro apolysis of Chilo suppressalis integument

The relationship between induction of in vitro apolysis and the duration of hormone treatment, and the effects of metabolic inhibitors on the ecdysone-induced apolysis were investigated in the cultured integument taken from the rice stem borer larva, Chilo suppressalis. When fragments of integument were subjected to 0.3 μg/ml β-ecdysone for more than 5 hr and then transferred to hormone free medium, they were induced to apolyse one day after treatment. If the fragments of integument were treated with hormone for 1 to 4 hr at first and then treated with hormone for 2 to 5 hr again after a 5 day interval in hormone free medium, almost all the fragments were induced to apolyse one day after treatment. This result suggests that the action of β-ecdysone on the cultured integument is accumulative. If the fragments of integument were cultured in the medium containing actinomycin-D and then transferred to medium containing β-ecdysone, a strong inhibitory effect on the apolysis of the integument was observed. Similarly, an inhibitory effect appeared when fragments of integument were treated first with hormone and then with puromycin. These results show that the m-RNA synthesis necessary for apolysis was completed within 6 hr after hormone treatment. However, the protein synthesis required for apolysis was not. The relationship of the results obtained from these in vitro experiments to the mode of action of ecdysone is discussed.     

Myogenesis and neuromuscular junction formation in cultures of Periplaneta americana myoblasts and neurones

Myoblasts isolated from thoracic and abdominal muscle somites of 11-day-old Periplaneta americana grew and differentiated in vitro in a manner that resembled that of in vitro vertebrate myogenesis. Rounded myoblasts became bipolar and then fused to form multinucleated myotubes. The syncitia became cross-striated and responded actively to depolarising injected current. When these muscle cells were grown in the presence of neurones from cerebral or thoracic ganglia they formed neuromuscular junctions that structurally resembled those on adult insect muscle. Excitatory post-synaptic potentials (E.P.S.Ps.) were frequently encountered in these co-cultures suggesting that the junctions were physiologically functional.