Biomedical subjects

R A Steinbrecht

Publications and source records attributed to R A Steinbrecht.

At least 19 recordsLinked to original sources

PubMed2001

Expression of SNMP-1 in olfactory neurons and sensilla of male and female antennae of the silkmoth Antheraea polyphemus.

SNMP-1 (sensory neuron membrane protein 1) is an olfactory-specific membrane-bound protein which is homologous with the CD36 receptor family. Previous light level immunocytochemical studies suggested that SNMP-1 was localized in the dendrites and distal cell body of sex-pheromone-specific olfactory receptor neurons (ORN); these studies further suggested SNMP-1 was expressed in only one of two to three neurons in male-specific pheromone-sensitive trichoid sensilla. To better understand the expression and localization of SNMP-1, an immunocytochemical study was performed using electron microscopy to visualize the distribution of SNMP-1 among the neurons of several classes of olfactory sensilla of both male and female antennae of the silkmoth Antheraea polyphemus. SNMP-1 antigenicity was primarily restricted to the receptive dendritic membranes of ORNs of all sensilla types examined and was observed in cytosolic granules, but not plasma membranes, of the cell soma. Mean labeling densities ranged from 1 to 16 gold particles per micrometer of dendrite circumference; dendrites of trichoid and intermediate sensilla showed significantly higher labeling densities than those of basiconic sensilla. Larger dendrites of trichoid sensilla showed significantly higher mean labeling densities (13-16/micron) than smaller diameter dendrites (3-7/micron). Immunofluorescence studies using baculovirus expressed SNMP-1 and multiphoton photon laser scanning microscopy (MPLSM) indicated that rSNMP-1, which was post-translationally processed to the in vivo molecular weight, was inserted into the plasma membrane in a topography presenting extracellular epitopes. These studies suggest SNMP-1 is a common feature of the ORNs, is asymmetrically expressed among functionally distinct neurons, and possesses a topography which permits interaction with components of the extracellular sensillum lymph.

Animals↗

PubMed2001

Gustatory organs of Drosophila melanogaster: fine structure and expression of the putative odorant-binding protein PBPRP2.

In Drosophila, as in most insects, gustation is mediated by sensory hairs located on the external and internal parts of the proboscis and on the legs and wings. We describe in detail the organization and ultrastructure of the gustatory sensilla on the labellum and legs and the distribution of PBPRP2, a putative odorant-binding protein, in the gustatory organs of Drosophila. The labellum carries two kinds of sensilla: taste bristles and taste pegs. The former have the typical morphology of gustatory sensilla and can be further subdivided into three morphological subtypes, each with a stereotyped distribution and innervation. Taste pegs have a unique morphology and are innervated by two receptor cells: one mechanoreceptor and the other a putative chemoreceptor cell. PBPRP2 is abundantly expressed in all adult gustatory organs on labellum, legs, and wings and in the internal taste organs on the proboscis. In contrast to olfactory organs, where PBPRP2 is expressed in the epidermis, this protein is absent from the epidermis of labial palps and legs. In the taste bristles of the labellum and legs, PBPRP2 is localized in the crescent-shaped lumen of the sensilla, and not in the lumen where the dendrites of the gustatory neurons are found, making a function in stimulus transport unlikely in these sensilla. In contrast, PBPRP2 in peg sensilla is expressed in the inner sensillum-lymph cavity and is in contact with the dendrites. Thus, PBPRP2 could be involved as a carrier for hydrophobic ligands, e.g., bitter tastants, in these sensilla.

Animals↗

PubMed2001

Immunolocalization of odorant-binding proteins in noctuid moths (Insecta, Lepidoptera).

Odorant-binding proteins were studied in the noctuid moths Agrotis segetum, Autographa gamma, Helicoverpa armigera, Heliothis virescens and Spodoptera littoralis using antisera raised against the pheromone-binding protein (PBP) and general odorant-binding protein 2 (GOBP2) of Antheraea polyphemus (Saturniidae). Proteins immunoreacting with these antisera were only found on the antennae and PBP and GOBP2 could be identified on western blots of males and females of all five species. PBPs were predominantly localized in sensilla trichodea and GOBP2 in sensilla basiconica, in good correlation with the stimulus specificity of the receptor cells in these sensilla. In H. armigera and H. virescens the majority of the s. trichodea immunoreacted with the antiserum against PBP of A. polyphemus; in A. segetum, A. gamma and S. littoralis, on the other hand, a high percentage of s. trichodea remained unlabelled. Probably, the PBP expressed in these sensilla is so different that it does not immunoreact with the antiserum used. Such a protein was found by native PAGE of antennal extracts of A. segetum and S. littoralis. These data correlate with the fact that the two heliothine species use pheromones with the same alkyl chain length as A. polyphemus, while the other three species use pheromones with shorter chains. In H. armigera, H. virescens, A. gamma and S. littoralis female antennae were also immunolabelled and a large number of PBP-expressing s. trichodea was consistently found. In S.littoralis this fits with the electrophysiologically recorded high pheromone sensitivity of female s. trichodea, whereas in females of H. armigera and H. virescens no or only weak responses to pheromone stimulation have been reported. Therefore, PBP expression in a sensillum does not necessarily imply pheromone sensitivity of its receptor cells.

Animals↗

PubMed2000

Expression patterns of two putative odorant-binding proteins in the olfactory organs of Drosophila melanogaster have different implications for their functions.

The aqueous medium bathing the dendrites of olfactory neurons contains high concentrations of odorant-binding proteins (OBPs) whose role is still unclear. OBPs may facilitate interactions between odorants and their membrane-bound receptors, perhaps by increasing the water solubility of hydrophobic molecules. Alternatively, OBPs may be involved in the inactivation of odorants and other volatile molecules, preventing desensitization and/or protecting olfactory neurons from toxic chemicals. We report here novel features of the localization of two putative OBPs, PBPRP2 and PBPRP5, that have important and different implications for their role in olfaction. Unlike several other putative OBPs of Drosophila melanogaster that are only found in adult olfactory organs, PBPRP5 is also expressed in the larval olfactory organs, suggesting that it plays a common role in olfaction at both stages. In the adult, PBPRP5 expression is restricted to the sensillum lymph that bathes the olfactory dendrites of a subset of olfactory hairs, the basiconic sensilla. Since individual basiconic sensilla differ in olfactory specificity, PBPRP5 may be able to bind to and mediate olfactory responses to a wide range of odorants. In contrast, PBPRP2 is present in the space immediately below the antennal cuticle and in the outer cavity of approximately 30% of the double-walled coeloconic sensilla on the antennal surface. In neither case is PBPRP2 in contact with the dendritic membranes of olfactory neurons, making a carrier function unlikely for this protein. Instead, PBPRP2 may act as a sink, binding to odorants and other volatile chemicals and limiting their interactions with olfactory neurons.

Animal Structures↗

PubMed2000

Chemosensory proteins from the proboscis of mamestra brassicae.

Soluble, low molecular weight proteins were immunodetected in proboscis extracts of Mamestra brassicae males by Western blot, using antibodies raised against the general odorant-binding protein of the moth Antheraea polyphemus. The same antibodies weakly labelled the sensillum lymph and subcuticular space of sensilla styloconica on ultrathin sections of the proboscis. The morphology of sensilla styloconica is described. The immunodetected proteins yielded several N-terminal sequences, three of which showed strong affinity for tritiated analogues of pheromonal compounds of M. brassicae in binding assays. The cDNAs coding for these sequences were cloned and it was shown that the new proteins are related to the OS-D protein of Drosophila. They are named chemosensory proteins (CSP-MBRA:A1-CSP-MBRA:A5 and CSP-MBRA:B1 and CSP-MBRA:B2) and may have an odorant-binding protein-like function. A common localization in both olfaction and taste organs suggests a physiological role depending on the cellular environment.

Amino Acid Sequence↗

PubMed1998

Olfactory coding in a compound nose. Coexpression of odorant-binding proteins in Drosophila.

Odorant-binding proteins (OBPs) are small, soluble proteins present in the aqueous medium surrounding olfactory receptor neurons. Their function in olfaction is unknown: they have been proposed to facilitate the transit of hydrophobic molecules to olfactory receptors, to deactivate the odorant stimulus, and/or to play a role in chemosensory coding. We have examined the genomic organization and expression patterns of two olfactory-specific genes (OS-E and OS-F) of Drosophila melanogaster, the products of which are members of a protein family in Drosophila sharing sequence similarity with moth OBPs. We found that the OS-E and OS-F transcription units are located < 1 kb apart. They are oriented in the same direction and display a similar intron-exon organization. Expression of both OS-E and OS-F proteins is spatially restricted to the ventrolateral region of the Drosophila antenna. Within this region, both OS-E and OS-F proteins are expressed within two different types of sensory hairs: in most, if not all, sensilla trichodea and in approximately 40% of the interspersed small sensilla basiconica. We consistently observe that OS-E and OS-F are coexpressed, indicating that an individual sensillum can contain more than one odorant-binding protein. This finding has potential implications for the roles of odorant-binding proteins in olfactory coding.

Animals↗

PubMed1998

Odorant-binding proteins: expression and function.

Odorant-binding proteins (OBPs) are a major constituent of the aqueous perireceptor compartment in vertebrates and in insects. Although different in primary structure, they are supposed to serve similar functions in both animal groups: (i) OBPs may act as solubilizers and carriers of the lipophilic odorants in the aqueous mucus or sensillum lymph; (ii) OBPs may act in addition as peripheral filters in odor discrimination by selectively binding certain classes of odorants; (iii) OBPs may present the stimulus molecule in a particular way to the receptor proteins to facilitate signal transduction; (iv) OBPs may clean the perireceptor space from unwanted and toxic compounds; (v) OBPs may rapidly deactivate odorants after stimulation of the receptors. Experimental evidence in favor of this multiple role of OBPs is reviewed.

Animals↗

PubMed1997

Coexpression of two odorant-binding protein homologs in Drosophila: implications for olfactory coding.

Odorant-binding proteins (OBPs) are small soluble proteins present in the aqueous medium surrounding olfactory receptor neurons. Their function in olfaction is still unknown: they have been proposed to facilitate the transit of hydrophobic molecules to olfactory receptors, to deactivate the odorant stimulus, and/or to play a role in chemosensory coding. In this study we examine the genomic organization and expression patterns of two olfactory-specific genes (OS-E and OS-F) of Drosophila melanogaster, the products of which are members of a protein family in Drosophila sharing sequence similarity with moth OBPs. We show that the OS-E and OS-F transcription units are located <1 kb apart. They are oriented in the same direction and display a similar intron-exon organization. Expression of both OS-E and OS-F proteins is restricted spatially to the ventrolateral region of the Drosophila antenna. Within this region both OS-E and OS-F proteins are expressed within two different types of sensory hairs: in most, if not all, sensilla trichodea and in approximately 40% of the interspersed small sensilla basiconica. We consistently observe that OS-E and OS-F are coexpressed, indicating that an individual sensillum can contain more than one odorant-binding protein. The functional significance of the observed expression pattern and its implications for olfactory coding are discussed.

Animals↗

PubMed1996

Structure and function of insect olfactory sensilla.

Olfactory sensilla show a large diversification of sensillum types even in the same species. Thus, double-walled and single-walled sensilla with highly different wall pores are usually found on the same antenna, and these may appear in the form of long slender hairs, pore plates or pit pegs. The selective constraints leading to this diversification are evident only in a few cases, e.g. the demand for extreme sensitivity in moth pheromone communication supported the evolution of long sensilla trichodea with high efficiency of capturing odour molecules. The structural diversity continues with the odorant-binding proteins (OBPs) in the sensillum lymph surrounding the sensory dendrites. These proteins may be subdivided into pheromone-binding proteins and two classes of general odorant-binding proteins according to their primary sequence. Different sensilla of the same morphological type may contain different OBPs of the same or of different subclasses. However, OBPs of different subclasses are not co-localized in the same individual sensory hair. The presence of a given OBP is related more to the functional specificity of the receptor cells than to the morphological type of the sensillum, suggesting a role of OBPs in stimulus recognition.

Animals↗

PubMed1996

Are odorant-binding proteins involved in odorant discrimination?

Pheromone-sensitive sensilla trichodea of nine moth species belonging to six families and three superfamilies of Lepidoptera were immunolabelled with an antiserum against the pheromone-binding protein of Antheraea polyphemus. Strong immunolabelling of the sensillum lymph was observed in all long sensilla trichodea of A. polyphemus, A. pernyi (Saturniidae), Bombyx mori (Bombycidae) and Manduca sexta (Sphingidae). Very weak labelling was found with all sensilla trichodea of Dendrolimus kikuchii (Lasiocampidae) and Lymantria dispar (Lymantriidae). In three noctuid species, some long sensilla trichodea were labelled strongly, some only weakly and some were not labelled at all. The fraction of long sensilla trichodea that were strongly labelled was large in Helicoverpa armigera, but small in Spodoptera littoralis and Autographa gamma. The observed cross-reactivity was not correlated with taxonomic relatedness of the species but rather with chemical relatedness of the pheromones used by these species, as a high labelling density was consistently observed in sensilla tuned to pheromones with an alcyl chain of 16 carbon atoms. The highly divergent specificity of pheromone-receptor cells in Noctuidae appears to be mirrored by a similar diversity of the pheromone-binding proteins in the sensilla trichodea. These data support the notion that pheromone-binding proteins participate in odorant discrimination.

Animals↗

PubMed1994

The tuft organs of the human body louse, Pediculus humanus corporis--cryofixation study of a thermo-/hygrosensitive sensillum.

The tuft organs of the human body louse, Pediculus humanus corporis, display a complex cuticular apparatus without wall pores, consisting of the cuticular cone, cuticular pocket, and the sensillar peg with 5-6 tuft processes. The cuticular wall of the sensillar peg exhibits several layers of differing electron density tightly surrounding the dendrites. The sensillar peg is innervated by the unbranched dendrites of two receptor cells which do not reach into the tuft processes. A third receptor cell forms a lamellar outer dendritic segment which wraps around the two other dendrites and terminates before reaching the base of the sensory peg. This type of receptor cell is named stratoterminal while for those which end within the cuticular apparatus the term conoterminal is proposed. Five auxiliary cells are associated with each tuft organ. The pore organs display a porous cuticular plate innervated by the branching dendrites of four receptor cells. A fifth receptor cell terminates more proximal and resembles the stratoterminal cell of the tuft organ. From the fine structure it is evident that the tuft organs represent thermo/hygrosensitive sensilla whereas the pore organs may have an olfactory function.

Animals↗

PubMed1993

Freeze-substitution for morphological and immunocytochemical studies in insects.

Methods of plunge freezing and freeze-substitution (FS) for insect antennae and similar body appendages are described. In these more or less cylindrical specimens, usually a layer below the cuticular surface of 10-15 microns thickness is well preserved without freezing damage, further inwards ice-crystal ghosts of increasing size are encountered, but in the very centre of antennal branches (diameter approximately 80 microns) of the silkmoth, Bombyx mori, freezing damage is usually reduced again. The frost-hardy species, Poecilocampa populi and Boreus hiemalis, exhibit regions free from freezing damage up to 40 microns below the cuticular surface. Secondary freezing damage in silkmoth sensory hairs is observed only after deliberately warming the specimens to -43 degrees C for >> 10 min before FS. Secondary artefacts due to the substitution process are investigated by comparison with freeze-etching and by comparing different FS media and protocols. Methanol is not recommended as a substitution medium for insect specimens. Structures particularly liable to substitution damage are the stimulus-conducting pore tubules of olfactory sensilla and the receptor cell membrane. Extraction of soluble components is more likely with pure organic solvents without added chemical fixing agents and with prolonged substitution at elevated temperatures. Such extraction may also be a possible artefact with soluble antigens in immunocytochemical studies. A review is given of the major achievements attained with these techniques is insect functional morphology and immunocytochemistry.

Animals↗

PubMed1992

Experimental morphology of insect olfaction: tracer studies, X-ray microanalysis, autoradiography, and immunocytochemistry with silkmoth antennae.

The general morphology and methodological peculiarities of insect sensilla are briefly reviewed. The stimulus conducting pore-tubule systems of pheromone-sensitive sensilla of the silkmoths Bombyx mori and Antheraea polyphemus are described. Lipophilic tracers readily enter the hair lumen, while hydrophilic tracers do so only after prolonged extraction with lipid solvents and/or pronase. X-ray microanalysis demonstrates a high potassium content of the sensillum lymph; calcium was only found in the haemolymph above detection limit. Auxiliary cells rapidly take up radioactive leucine administered via the haemolymph. Antibodies against pheromone-binding protein of Antheraea polyphemus label the sensillum lymph of sensilla trichodea, but not of sensilla basiconica in A. polyphemus as well as in B. mori. The cytoplasm of auxiliary cells of the sensilla trichodea is also labelled. The results are discussed in context with present hypotheses on the role of sensillum lymph in stimulus transport and inactivation.

Animals↗

PubMed1989

Fine structure of a sensory organ in the arista of Drosophila melanogaster and some other dipterans.

The arista, a characteristic appendage of dipteran antennae, consists of 2 short segments at the base and a long distal shaft. A small sensory ganglion, from which arises the aristal nerve, is located proximally in the shaft. The fine structure of the aristal sensory organ was studied in detail in the fruitfly (Drosophila) and for comparison in the housefly (Musca) and the blowfly (Calliphora). In Drosophila, the aristal sense organ consists of 3 identical sensilla that terminate in the hemolymph space of the aristal shaft, and not in an external cuticular apparatus. Each sensillum comprises 2 bipolar neurons and 2 sheath cells; a third sheath cell envelops the somata of all six neurons of the ganglion. The neurons have long slender dendrites with the usual subdivision into an inner and an outer segment. One of the outer segments is highly lamellated and bears small particles (BOSS-structures) on the outside of its cell membrane; the other outer segment is unbranched and has a small diameter. The fine structure of the first dendrite is strongly reminiscent of thermoreceptors known from the antennae of other insects. These thermoreceptors are often coupled with hygroreceptors; however, we can only speculate whether the second dendrite of the aristal organ also has this function. Our present results argue against mechanoreceptive functions, as formerly postulated. The aristal sense organs in Musca and Calliphora are similar to those in Drosophila, but contain more sensilla (12 in Musca, 18 in Calliphora.

Animals↗

PubMed1989

Ions and mucoid substances in sensory organs--microanalytical data from insect sensilla.

In insect sensory organs, the receptor endings are surrounded by an extracellular subcuticular matrix, the sensillum lymph. This matrix provides the correct aqueous and ionic milieu around the sensory processes, plays a role in the generation of the receptor current, and is also important for stimulus transport and/or stimulus inactivation in chemoreceptors. Histochemical evidence indicates the presence of proteoglycans (probably hyaluronate and/or chondroitin sulfate) in the sensillum lymph. X-ray micro-analytical studies demonstrate a very low sodium content, whereas potassium is present in high concentrations, comparable to those in intracellular compartments. The total positive charges of cationic elements are not balanced by the negative charges of chlorine, but by polyanions of the mucoid substances present in sensillum lymph. These fixed negative charges probably play a key-role in the establishment of the non-neural transepithelial voltage of insect sensilla, which in turn is a major source of the receptor current. Moreover, the mucoid substances of the sensillum lymph are probably involved in water conservation, since sensilla are prone to water loss, because the overlying cuticle must be permeable to the chemical stimuli. Functional analogies are pointed out between insect sensillum lymph and vertebrate olfactory mucus and, in particular, the endolymph of the vertebrate inner ear.

Animals↗

PubMed1984

Pheromone receptors in Bombyx mori and Antheraea pernyi. I. Reconstruction of the cellular organization of the sensilla trichodea.

The cellular organization of freeze-substituted antennal sensilla trichodea, which contain the sex pheromone receptors, was studied in male silkmoths of two species (Bombyx mori, Bombycidae; Antheraea pernyi, Saturniidae). The cellular architecture of these sensilla is complex, but very similar in both species. A three-dimensional reconstruction of a sensillum trichodeum of B. mori is presented. Two receptor cells (in A. pernyi 1-3) and three auxiliary cells are present. Of the latter, only the thecogen cell forms a true sheath around the receptor cells. A unique thecogen-receptor cell junction extends over the entire area of contact. Septate junctions occur between all sensillar cells apically, and in the region of the axonal origin basally. Gap junctions are also found between all cells except the receptor cells. The trichogen and tormogen cells show many structural indications of secretory activity and are thought to secrete the receptor lymph. Their apical membrane bordering the receptor-lymph space is enlarged by microvilli and microlamellae, but only those of the trichogen cell show regularly arranged membrane particles (portasomes), indicating secretory specialization among the auxiliary cells. Epidermal cells are found as slender pillars between sensilla, but extend apically along the non-sensillar cuticle and basally along the basal lamina.

Animals↗

PubMed1984

Pheromone receptors in Bombyx mori and Antheraea pernyi. II. Morphometric analysis.

Sensilla trichodea of the silk moths, Antheraea pernyi and Bombyx mori, were reconstructed from serial sections after freeze substitution. The volume and surface area of the different sensillar cells were calculated from the area and circumference of consecutive section profiles. A. pernyi and B. mori differ largely in the size of the sensory hair and the larger outer dendritic segments as well as in the volume of the receptor lymph within the hair, while there are only small differences regarding inner dendritic segments, receptor-cell somata, trichogen and tormogen cells and the volume of the receptor lymph below the hair base. In each sensillum the two (or three) receptor-cell somata, dendrites, and initial axonal segments differ significantly in volume and surface. The apical cell membranes of the trichogen and tormogen cells, which border the receptor-lymph cavity and which are the presumed site of electrogenic cation pumps, are deeply invaginated and enlarged by microlamellae and microvilli, so that their area is twice that of the remaining basolateral cell membrane. In contrast to mechanoreceptors, the trichogen cell is the largest auxiliary cell and has the largest apical membrane surface. The morphometric data are discussed with regard to recent electrophysiological observations.

Animals↗

PubMed1984

A cryoembedding method for cutting ultrathin cryosections from small frozen specimens.

A technique is described which allows one to embed small frozen specimens for easy sectioning in a cryoultramicrotome. n-Heptane is recommended as a cryoembedding medium due to its suitable melting point (182.4K), high vapour pressure, chemical inertia, and good sectioning properties when solid. Freeze dried cryosections of cultured muscle cells, slime moulds, insect antennae and unicellular algaee are presented as examples. There is no evidence that the preparation disturbs the distribution of soluble elements in the specimen, since X-ray spectra of muscle cells exposed to heptane do not differ from those of samples not exposed.

Animals↗