The interaction of external and internal receptors on the feeding behaviour of the blowfly, Phormia regina

The interaction of sensory activity from internal gut stretch receptors and from external labellar chemosensory hairs has been studied both behaviourally and electrophysiologically in the control of proboscis extension of the blowfly, Phormia regina. Labellar thresholds for proboscis extension, tested behaviourally, do not change significantly up to an hour after feeding in contrast to tarsal thresholds which rise quickly after feeding. Motor activity of the extensor muscle of the haustellum was recorded simultaneously with sensory activity from labellar sensilla. The mean number of muscle spikes per response and the sensory input necessary to trigger a response do not vary with starvation, feeding, or sectioning of the recurrent nerve. Activity of internal stretch receptors seem to interact with tarsal sensory input but apparently do not modulate motor responses triggered by labellar sensory input.     

The Effects of Amino Acids on the Labellar Hair Chemosensory Cells of the Fly

The effects of amino acids on the labellar hair chemosensory cells were examined with two kinds of flies (the fleshfly, Boettcherisca peregrina, and the blowfly, Phormia regina). As a result of this examination, the effects of amino acids were divided into four main classes. Amino acids in class 1 did not stimulate any chemoreceptor cell. Amino acids in class 2 inhibited nonspecifically the discharges from three kinds of chemosensory cells. Amino acids in class 3 stimulated the salt receptor cell. Amino acids in class 4 stimulated the sugar receptor cell. A possibility that a fourth neuron in the labellar hair chemosensory cell might be a protein or an amino acid receptor cell was eliminated.     

Properties of membrane-bound α-glucosidases: Possible sugar receptor protein of the blowfly, Phormia regina

Previously reported PII-type α-glucosidase located in the precipitate of the labellar homogenate of the blowfly Phormia regina was solubilized by sodium deoxycholate (DOC) and further separated into three isozymes with different molecular weight: PII-M (mol. wt 9 × 10⁴). PII-D (mol. wt 2 × 10⁵) and PII-T (mol. wt 8 × 10⁵) by molecular sieve chromatography on Biogel P-300 or Ultragel AcA-34. These three isozymes had almost the same K_m's and relative values of V_m's for several substrates, suggesting that they had the same common active site.PII-D and PII-T are more strongly embedded in the membrane than PII-M, because the proportion of PII-D and PII-T was much increased when the remaining glucosidase in the precipitate after the first solubilization was reextracted by DOC. A large peak of α-glucosidase isozyme P-IV which preferentially hydrolyze sucrose eluted just after P-II (soluble P-II) when the supernatant fraction of the labellar homogenate was chromatographed on DEAE-Sephadex A-50. P-IV was scarcely present in the precipitate fraction.Soluble P-II had the same mol. wt as PII-M and had similar properties to PII-M except for the ratio of V_m's.A large proportion of PII-D was contained in the well washed labellar integuments, a preparation rich in labellar chemosensilla. It suggests that most of the insoluble α-glucosidase contained in the dendrite in labellar chemosensilla is PII-D. PII-D (and PII-T) are possible sites of the pyranose receptor molecule because their properties and localization agree well with those of the receptor.     

Comparative labellar micromorphology of Zygopetalinae (Orchidaceae)

Background and Aims

Molecular evidence indicates that the Neotropical sub-tribe Zygopetalinae is sister to Maxillariinae. Most members of the latter sub-tribe have deceit pollination strategies, but some species produce rewards such as nectar, pseudopollen, resin and wax, and are pollinated by a range of pollinators that include stingless bees (Meliponini), wasps and hummingbirds. By contrast, relatively little is known about the pollination of Zygopetalinae species. However, some are pollinated by fragrance-gathering, male euglossine bees or employ nectar deceit strategies. The aim of this study is to describe the labellar micromorphology of Zygopetalinae and to compare it with that of Maxillariinae sensu lato (s.l.) as part of an ongoing project to record the range of labellar characters found within the tribe Maxillarieae, and to assess whether these characters represent synapomorphies or homoplasies resulting from similar pollination pressures.

Methods

The labella of 31 species of Zygopetalinae, including Cryptarrhena R. Br. and representatives of the Zygopetalum, Huntleya and Warrea clades, were examined using light microscopy and scanning electron microscopy, and the range of labellar characters was recorded. These characters were subsequently compared with those of Maxillariinae s.l. which formed the subject of our previous investigations.

Key Results and Conclusions

The labellar micromorphology of Zygopetalinae is less diverse than that of Maxillariinae and does not reflect the currently accepted phylogeny of the former sub-tribe based on molecular studies. Instead, the relative uniformity in labellar micromorphology of Zygopetalinae is probably due to homoplasies resulting from similar pollinator pressures. Labellar trichomes are relatively uncommon in Zygopetalinae, but occur in certain members of both the Zygopetalum and Huntleya clades. Trichomes are unbranched, uniseriate and multicellular with rounded apices, or unbranched and unicellular, with tapering, pointed and flexuose apices. Hitherto, unicellular trichomes of this kind have been observed only for euglossophilous orchid taxa, and the adoption of a relatively limited range of pollination strategies by Zygopetalinae may have resulted in reduced investment in micromorphological labellar characters.     

Behavioral responses of Phormia regina (Meigen) to labellar stimulation with amino acids

Behavioral responses to labellar stimulation with 19 L-amino acids were predicted on the basis of electrophysiological responses of largest labellar hairs. With the exceptions alanine, aspartic and glutamic acids, and valine tests of these predictions confirmed that Phormia can discriminate amino acids, and that these acids may be grouped according to their effects. Electrophysiological investigation of the four exceptions was repeated and results were consistent with the behavioral data. In particular, these acids elicited previously unreported responses from the salt receptor. The discrepancies between this and earlier studies may be explainable, in part, on methodological grounds. There was evidence for response differences among hairs of different sizes and among the largest labellar hairs themselves. The significance of amino acid discrimination for the problem of protein recognition can only be speculated upon until more complete electrophysiological and nutritional information is available.     

Ingestion of sugar diets correlates with spike activity in labellar chemosensilla of the flesh-fly, Neobellieria (= Sarcophaga ) bullata

Abstract .Female 2-day-old Neobellieria (= Sarcophaga ) bullata (Parker) (Diptera: Sarcophagidae) were exposed to different concentrations of sucrose, glucose and fructose in a single-choice potometer, and the volume ingested in the first hour was measured. Nerve spike activity in response to the same sugars was recorded from medium labellar taste hairs of similar flies by tip-recording. Two classes of chemosensory cells responded to sucrose, glucose and fructose. Cell 1 showed an increasing spike activity with sugar concentration, whereas cell 2 did not; cell 1 was identified as the 'sugar cell'.
For both spike activity in cell 1 and feeding, sucrose was the most stimulatory sugar. The dose–response curves for glucose and fructose crossed over at about 200 m m . At higher concentrations, glucose was more stimulatory for both cell 1 and for feeding, and at lower concentrations, fructose. The pattern of spike activity supports a separate location on the sensory cells of receptors for pyranose and fructose forms of sugar. The strong correlation between volume ingested and spike activity indicates that sugar feeding is controlled by sensory input from the 'sugar' cells of labellar chemosensilla. Moreover, the results suggest that the flies do not distinguish between these sugars except by apparent 'sweetness'.     

Labellar anatomy and secretion in Bulbophyllum Thouars (Orchidaceae: Bulbophyllinae) sect. Racemosae Benth. & Hook. f.

Background and Aims

Floral secretions are common in Bulbophyllum Thouars, and the labella of a number of Asian species are said to produce secretions rich in lipids that act as food rewards for insect pollinators. Although some of these reports are based on simple histochemical tests, a much greater number are anecdotal and, hitherto, neither the ultrastructure of the labellum nor the secretory process has been investigated in detail. Furthermore, sophisticated histochemical approaches have generally not been applied. Here, both the labellar structure and the secretory process are investigated for four species of Asian Bulbophyllum sect. Racemosae Benth. & Hook. f., namely Bulbophyllum careyanum (Hook.) Spreng., B. morphologorum Kraenzl., B. orientale Seidenf. and B. wangkaense Seidenf., and compared with those of unequivocal lipid-secreting orchids.

Methods

Labellar, secretory tissue was investigated using light microscopy, scanning electron microscopy, transmission electron microscopy and histochemistry.

Key Results

The adaxial median longitudinal groove of the labellum contained secretory tissue comprising palisade-like epidermal cells, similar to those of certain lipid-secreting Oncidiinae Benth. However, these cells and their secretions gave positive results mainly for protein and mucilage, and their organelle complement was consistent with that of cells involved in protein and mucilage synthesis. Sub-cuticular accumulation of secretion resulted in cuticular distension and blistering. The sub-epidermal layer of isodiametric parenchyma contained starch and, like the epidermal cells, ultrastructure consistent with mucilage synthesis. Lipids were mainly confined to the cuticle, and hardly any intracellular lipid droplets were observed.

Conclusions

It is proposed that mucilage is produced by dictyosomes present in the palisade-like epidermal cells. Mucilage precursors may also be produced by these same organelles in sub-epidermal cells and are thought to pass along the symplast via plasmodesmata into the adjoining palisade-like secretory cells, which contain abundant arrays of rough endoplasmic reticulum. Here, they become chemically modified and form a protein-rich, mucilaginous secretion that, following vesicle-mediated transport across the cytoplasm, traverses the cell wall and accumulates in blisters formed from the distended cuticle. Rupture of these blisters releases the secretion onto the labellar surface. However, in certain species, there is some evidence that the secretion may traverse the cuticle via cuticular pores, and micro-channels may permit the passage of fragrance. Hydrolysis of sub-epidermal starch probably generates the carbohydrate and, together with mitochondria, much of the energy required for the secretory process. This anatomical organization resembles that found in certain lipid-secreting, Neotropical species of Bulbophyllum and Oncidiinae, but since the chemical composition of their secretions is different, and these taxa occur on a separate continent and have different insect pollinators, parallelism of floral anatomy is likely.     

Elaiophore diversity in three contrasting members of Oncidiinae (Orchidaceae)

The elaiophores of Trichocentrum cavendishianum (Bateman) M.W. Chase & N.H. Williams, Oncidium loefgrenii Cogn., and Gomesa recurva R. Br. display considerable morphological and anatomical diversity. Oil secretion by flowers of T. cavendishianum and O. loefgrenii can be related to the presence of saddle-like, labellar elaiophores and the labellar callus, respectively, whereas, in G. recurva , although oil is present, no obvious structure appears to be involved in its secretion. In the first two species, the secretory tissue consists of palisade-like cells, whereas, in G. recurva , these cells are oval. Many Oncidiinae are thought to mimic members of the Malpighiaceae, and the elaiophores of that family also contain palisade-like cells that may indicate evolutionary convergence. As oils accumulate below the elaiophore cuticle, that of T. cavendishianum becomes distended, whereas that of the other two species does not. Full discharge of oil from the elaiophores of T. cavendishianum probably occurs only after the cuticle is ruptured by a visiting insect, and this may contribute towards pollinator selection.  © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 155 , 135–148.     

α-Glucosidase at the tip of the contact chemosensory seta of the blowfly, Phormia regina

α-Glucosidase activity was detected at the tip of the labellar contact chemosensory hair of the blowfly, Phormia regina. The enzyme split about 1 pmole of sucrose per hr per hair on average and the Michaelis constant for sucrose was about 50 mM. The activity of the enzyme was not solubilized into the incubation solution, but stuck stably to the tip of the sensory hair. From the cut end of the sensory hair a high activity of α-glucosidase eluted out. But its Michaelis constant was smaller by far than the one at the tip, suggesting that different types of α-glucosidase isozymes exist in the hair. The possibility that the enzyme at the tip of the sensory hair could be the sugar receptor is discussed.     

Electrical activity in the chemoreceptors of the blowfly. II. Responses to electrical stimulation

An analysis of the various parts of the electrical responses to the chemical and electrical stimulation of a single labellar chemosensory hair of the blowfly, Phormia regina, indicates that the recording conditions for the spike potentials approximate the intracellular recordings made in other types of sense cells. The large positive resting potential probably arises from the basement membrane of the hypodermal cells and neurilemma rather than from the neurons at the base of the chemosensory hair. The responses to polarizing currents passed through single chemosensory hairs support this analysis. The behavioral responses to similar polarizing currents are shown to result from the action of the current on the neurons at the bases of the adjacent chemosensory hairs. The reported neural interaction of the two chemosensory neurons associated with the chemosensory hair is probably due to the physical-chemical attributes of the stimulating solution rather than to any real neural interaction. Observations on the latency of the initial nerve impulse in response to chemical stimulation indicate that the chemosensory neurons are normally free from spontaneous spike activity.     

Comparative histology of floral elaiophores in the orchids Rudolfiella picta (Schltr.) Hoehne (Maxillariinae sensu lato) and Oncidium ornithorhynchum H.B.K. (Oncidiinae sensu lato)

Background and Aims

Floral elaiophores, although widespread amongst orchids, have not previously been described for Maxillariinae sensu lato. Here, two claims that epithelial, floral elaiophores occur in the genus Rudolfiella Hoehne (Bifrenaria clade) are investigated. Presumed elaiophores were compared with those of Oncidiinae Benth. and the floral, resin-secreting tissues of Rhetinantha M.A. Blanco and Heterotaxis Lindl., both genera formerly assigned to Maxillaria Ruiz & Pav. (Maxillariinae sensu stricto).

Methods

Putative, floral elaiophore tissue of Rudolfiella picta (Schltr.) Hoehne and floral elaiophores of Oncidium ornithorhynchum H.B.K. were examined by means of light microscopy, histochemistry, scanning electron microscopy and transmission electron microscopy.

Key Results and Conclusions

Floral, epithelial elaiophores are present in Rudolfiella picta, indicating, for the first time, that oil secretion occurs amongst members of the Bifrenaria clade (Maxillariinae sensu lato). However, whereas the elaiophore of R. picta is borne upon the labellar callus, the elaiophores of O. ornithorhynchum occur on the lateral lobes of the labellum. In both species, the elaiophore comprises a single layer of palisade secretory cells and parenchymatous, subsecretory tissue. Cell wall cavities are absent from both and there is no evidence of cuticular distension in response to oil accumulation between the outer tangential wall and the overlying cuticle in R. picta. Distension of the cuticle, however, occurs in O. ornithorhynchum. Secretory cells of R. picta contain characteristic, spherical or oval plastids with abundant plastoglobuli and these more closely resemble plastids found in labellar, secretory cells of representatives of Rhetinantha (formerly Maxillaria acuminata Lindl. alliance) than elaiophore plastids of Oncidiinae. In Rhetinantha, such plastids are involved in the synthesis of resin-like material or wax. Despite these differences, the elaiophore anatomy of both R. picta (Bifrenaria clade) and O. ornithorhynchum (Oncidiinae) fundamentally resembles that of several representatives of Oncidiinae. These, in their possession of palisade secretory cells, in turn, resemble the floral elaiophores of certain members of Malpighiaceae, indicating that convergence has occurred here in response to similar pollination pressures.Key words: Bifrenaria clade, elaiophore, floral oil, Heterotaxis, Maxillariinae, Oncidiinae, Oncidium ornithorhynchum, Rhetinantha, Rudolfiella picta, secretion     

Characterization of alpha-glucosidase at the tips of the chemosensory setae of the fly, Phormia regina

The properties of α-glucosidase activity located at the tip of the labellar chemosensory seta of the blowfly were examined using the ultramicro method for determination of hexose. The enzyme activity was independent over a wide range of pH (3·0–8·0) and inhibited by Tris in a competitive manner and by Ca²⁺ ion in low concentration. By comparison of the present results with those for the α-glucosidase isozymes in the extract from the proboscis, it became clear that the enzyme in the intact state had properties to distinguish it from the enzyme in the soluble form. Furthermore, apparent similarities were observed between the properties of the enzyme in the intact state and those of the labellar sugar receptor examined electrophysiologically.     

Impulse frequency and action potential amplitude in labellar chemosensory neurones of Drosophila melanogaster

The electrophysiological responses of water, salt and sugar receptors in the labellar chemosenory hair of Drosophila were investigated. In contrast to the responses of large flies such as blowfly and fleshfly, spike height changed in parallel with the spike frequencies in all the three kind of receptors, and at the same time, the spike also changed in shape: when the receptor potential was small, the spike was small and biphasic, but when the receptor potential was large, the spike was large and monophasic. These phenomena are consistently explained by assuming that antidromic conduction of spikes in the distal process of the receptor cells is blocked due to inactivation of Na channels by the depolarizing receptor potential.     

The sensitivity of the labellar sugar receptors of Phormia regina in relation to feeding

The hypothesis that behavioural feeding threshold of P. regina modulates and is modulated by the sensitivity of the labellar contact chemoreceptors is not supported by the examination of individual flies.Sensitivity (number of impulses fired in the first second of stimulation) of the sugar receptors in selected labellar hairs of flies of known age was recorded periodically on a fixed schedule for up to 72 hr. Experimental flies were hand fed 2.0 M fructose once every 24 hr. No correlations between feeding or age and receptor sensitivity were found. Statistically significant changes in firing frequency did occur but were unrelated to feeding and age.The proboscis extension response, on which behavioural threshold measures are based, is known to be triggered by the first few sugar receptor spikes of sufficient frequency. Neither age nor feeding significantly affected the number or frequency of impulses during the first 50 msec of stimulation.     

The chemosensitivity of labellar sugar receptor in female Phormia regina is paralleled with ovary maturation: Effects of serotonin

Oogenesis in most adult insects is a nutrient-dependent process involving ingestion of both proteins and carbohydrates that ultimately depends on peripheral input from chemoreceptors.The main goal of this study was to characterize, in the female blowfly Phormia regina, the responsive changes of the labellar chemoreceptors to carbohydrates and proteins in relation to four different stages along the ovarian cycle: (1) immature ovaries, (2) mid-mature ovaries, (3) mature ovaries and ready for egg-laying and (4) post egg-laying ovaries. Then, the possible effects exerted by exogenous serotonin on the chemoreceptor sensitivity profiles were investigated.Our results show that ovary length, width and contraction rate progressively increase from stage 1 to 3, when all these parameters reach their maximum values, before declining in the next stage 4.The sensitivity of the labellar “sugar” chemoreceptors to both sucrose and proteins varies during the ovarian maturation stages, reaching a minimum for sucrose in stage 3, while that to proteins begins. Exogenous 5-HT supply specifically increases the chemoreceptor sensitivity to sugar at the stages 3 and 4, while it does not affect that to proteins.In conclusion, our results provide evidence that in female blowflies the cyclic variations in the sensitivity of the labellar chemosensilla to sugars and proteins are time-related to ovarian development and that during the stages 3 and 4 the responsiveness of the sugar cell to sucrose is under serotonergic control.     

Protein-sensitive elements of the labellar sensillas of Musca domestica flies

Using electrophysiological technique of registration of impulse activity in chemoreceptive cells of the labellar sensillae of the housefly, it has been demonstrated that taste hairs are not uniform in their properties. They differ from each other by the set of receptive elements which exhibit different sensitivity, range of selectivity and pattern of impulse activity. It was shown that albumen solution (10(-5) M) evokes the activity in 1--2 cells of a sensilla which are classified as water and sugar receptors. Among these receptors, protein-sensitive and protein-insensitive cells may be distinguished. Considering the inhomogeneity of sugar receptor sites, it was suggested that chemo receptive membranes in most sensitive to protein cells contain more numerous fructose receptive sites, that glucose ones.     

The innervation of some proboscis structures involved in feeding behavior of the blowfly (Calliphora vicina)

The microtopology of the motoneurons involved in protraction and retraction of the proboscis of the blowfly (Calliphora vicina) has been studied. In addition, taste input from the labellar hairs was investigated. As a result of this study it appears that protraction movements are controlled by two while retraction movements are guided by three motoneurons on each side. The neurons in each group apear to be in ipsicontralateral communication with each other. The musculi protractores fulcri (MPF) probably contain a proprioceptive cell group which projects to the MPF motoneurons. It is proposed that the proboscis motor system can be modulated by proprioception as well as by chemosensory labellar input. Neurosecretory cells may be involved in adjusting muscle power.     

Sugar response differences related to sensillum type and location on the labella of Protophormia terraenovae: a contribution to spatial representation of the stimulus

The chemoreceptor spike activity in response to sucrose in the concentration range 1-500mM was recorded from each of the 11 Intermediate and 13 Large labellar sensilla in the blowfly Protophormia terraenovae. The results showed that: (1) three of the four cells present in each sensillum are activated by sucrose stimulation; (2) differences between the Large and Intermediate types exist in the dose-response profiles of one of these cells (the 'water' cell), possibly reflecting different sugar receptor site populations on the dendritic membranes of homologous cells in the two types; (3) sensilla of both types are differentially responsive to sucrose solutions according to their location on the labellum. These differences may provide elements for a spatial representation of the stimulus source within the sensory coding process.