Processing of antennular input in the brain of the spiny lobster, Panulirus argus

Neurons in the olfactory deutocerebrum of the spiny lobster, Panulirus argus, were recorded intracellularly and filled with biocytin. Recorded neurons arborized in the olfactory lobe (OL), a glomerular neuropil innervated by olfactory and some presumptive mechanosensory antennular afferents. The neurons responded to chemosensory input from the lateral antennular flagellum bearing the olfactory sensilla but not the medial flagellum bearing many non-olfactory chemosensory sensilla. Many neurons received additional mechanosensory input. Thus the OL integrates specifically olfactory with mechanosensory input. OL neurons had multiglomerular arborizations restricted to one or two of the three horizontal layers of the columnar glomeruli. OL local interneurons comprised core neurons with tree-like neurites and terminals in the base of the glomeruli and rim neurons with neurites surrounding the OL and terminals in the cap/subcap. The somata of OL local interneurons lay in the medial soma cluster (100000 somata). OL projection neurons arborized in the base of the glomeruli and ascended via the olfactory glomerular tract to the lateral protocerebrum. A parallel projection pathway is constituted by projection neurons of the accessory lobe, a glomerular neuropil without afferent innervation but intimate links to the OL. The projection neuron somata constituted the lateral soma cluster (200000 somata).Abbreviations AC anterior cluster (cluster 6,7) - AL accessory lobe - aMC anterior subcluster of medial cluster (cluster 9) - A _lNv main antenna I (antennular) nerve - A _lNM antenna I (antennular) motor nerve - A _llNv main antenna II (antennal) nerve - CB central body - CL central layer of accessory lobe - DC deutocerebral commissure - DCN deutocerebral commissure neuropil - dDUMC dorsal subcluster of dorsal unpaired median cluster (cluster 17) - dMC dorsal subcluster of medial cluster (cluster 11) - dVPALC dorsal subcluster of ventral paired anterolateral cluster (cluster 8) G glomerulus - IDUMC lateral subcluster of dorsal unpaired median cluster (cluster 16) - LC lateral cluster (cluster 10) - LF lateral flagellum of antenna I (antennule) - LL lateral layer of accessory lobe - MF medial flagellum of antenna I (antennule) - ML medial layer of accessory lobe - MPN anterior and posterior median protocerebral neuropils - OGT olfactory globular tract - OGTN olfactory globular tract neuropil - OL olfactory lobe - OLALT olfactory lobe-accessory lobe tract - PB protocerebral bridge - pMC posterior subcluster of medial cluster (cluster 9) - PT protocerebral tract - TNv tegumentary nerve - VPMC ventral paired medial cluster (cluster 12) - VUMC ventral unpaired medial cluster (cluster 13) - vVPALC ventral subcluster of ventral paired anterolateral cluster (cluster 8) - ASW artificial sea water - M3 mixture 3 - PRO L-proline - TM TetraMarin extract     

Tuning posture to body load: decreases in load produce discrete sensory signals in the legs of freely standing cockroaches

Decreases in load are important cues in the control of posture and walking. We recorded activities of the tibial campaniform sensilla, receptors that monitor forces as strains in the exoskeleton, in the middle legs of freely moving cockroaches. Small magnets were attached to the thorax and body load was changed by applying currents to a coil below the substrate. Body position was monitored by video recording. The tibial sensilla are organized into proximal and distal subgroups that have different response properties and reflex effects: proximal sensilla excite extensor motoneurons while distal receptors inhibit extensor firing. Sudden load decreases elicited bursts from distal sensilla, while increased load excited proximal receptors. The onset of sensory discharges closely approximated the time of peak velocity of body movement in both load decreases and increases. Firing of distal sensilla rapidly adapted to sustained unloading, while proximal sensilla discharged tonically to load increases. Load decreases of small amplitude or at low rates produced only inhibition of proximal activity while decrements of larger size or rate elicited distal firing. These response properties may provide discrete signals that either modulate excitatory extensor drive during small load variations or inhibit support prior to compensatory stepping or initiation of swing.     

Primary sensory neurons containing command neuron peptide constitute a morphologically distinct class of sensory neurons in the terrestrial snail

In the central nervous system of the terrestrial snail Helix, the gene HCS2, which encodes several neuropeptides of the CNP (command neuron peptide) family, is mostly expressed in cells related to withdrawal behavior. In the present work, we demonstrate that a small percentage (0.1%) of the sensory cells, located in the sensory pad and in the surrounding epithelial region ("collar") of the anterior and posterior tentacles, is immunoreactive to antisera raised against the neuropeptides CNP2 and CNP4, encoded by the HCS2 gene. No CNP-like-immunoreactive neurons have been detected among the tentacular ganglionic interneurons. The CNP-like-immunoreactive fiber bundles enter the cerebral ganglia within the nerves of the tentacles (tentacular nerve and medial lip nerve) and innervate the metacerebral lobe, viz., the integrative brain region well-known as the target area for many cerebral ganglia nerves. The procerebral lobe, which is involved in the processing of olfactory information, is not CNP-immunoreactive. Our data suggest that the sensory cells, which contain the CNP neuropeptides, belong to a class of sensory neurons with a specific function, presumably involved in the withdrawal behavior of the snail.     

Effect of dietary nickel deprivation on vision,olfaction, and taste in rats

Early studies on dietary nickel deprivation found decreased reproduction rate in pigs and decreased insemination and conception rates in goats. Studies from our laboratory demonstrated that nickel deprivation impaired male reproductive function of rats. A physiological amount of nickel modulates the function of cyclic nucleotide-gated cation channels (CNG channels) in vitro. Thus, because CNG channels have important roles in spermatozoa function, it was speculated that the impairment of reproduction by nickel deprivation was through an effect on CNG channels. Because CNG channels are found in retinal photoreceptor, olfactory receptor, and taste receptor cells, we hypothesized that nickel deprivation would also alter light/dark preference, odor preference to female rat urine, and taste preference/aversion in rats. In the light/dark Y-maze, nickel deprivation significantly decreased time spent in the dark arm by rats. The number of sniffs to estrous female urine was significantly increased only in nickel-supplemented rats. The number of licks at the saccharin bottle was significantly decreased by dietary nickel deprivation. These findings suggest that nickel has a biological role in the special senses: vision, olfaction and taste.     

Microtubule fascicles in the stem processes of cultured sensory ganglion cells

Summary Microtubule fascicles, resembling those characterizing the initial segment of multipolar neurons, have been observed by electron microscopy within and close to the origin of the stem process of some unipolar ganglion cells in explant cultures of embryonic chick dorsal root ganglia. Each fascicle comprised 2–6 closely spaced parallel microtubules linked by electron dense cross-bridges. Since similar observations have been made on stem processes in vivo, the possibility that linked microtubules occur commonly in this site is considered. The observations are discussed in relation to a possible correlation between the presence of microtubule fascicles and the initiation of action potentials.We thank Messrs. S. Waterman, P. Felton and D. Fraser for technical assistance and Prof. D.W. James for laboratory facilities     

Induced preference for host plant chemicals in the tobacco hornworm: contribution of olfaction and taste

Many herbivorous insects induce preferences for host plants. Recent work in Manduca sexta indicates that induced preferences are mediated by a “tuning” of the peripheral taste system to chemicals within host plant foliage. We tested this hypothesis by rearing caterpillars on artificial diet or potato foliage, and then examining olfactory- and taste-mediated responses to potato foliage extract. First, we confirmed earlier reports that consumption of potato foliage tunes the peripheral taste system by reducing responsiveness to glucose and increasing responsiveness to foliage extract. Second, we offered caterpillars a choice between disks treated with foliage extract (experimental) or solvent alone (control). The foliage-reared caterpillars approached and consumed the experimental disks disproportionately, whereas the diet-reared caterpillars approached and consumed both disks indiscriminately. This indicated that induced preferences involve olfaction and taste. Third, we ran choice tests with foliage-reared caterpillars deprived of either olfactory or gustatory input. Caterpillars lacking olfactory input approached both disks indiscriminately, but fed selectively on experimental disks. In contrast, caterpillars lacking gustatory input approached experimental disks selectively, but fed indiscriminately on both types of disk. We conclude that even though olfaction helps caterpillars locate potato foliage, it is the “tuned” gustatory response that ultimately mediates the induced preference.     

Rational design and optimization of downstream processes of virus particles for biopharmaceutical applications: current advances

The advent of advanced therapies in the pharmaceutical industry has moved the spotlight into virus-like particles and viral vectors produced in cell culture holding great promise in a myriad of clinical targets, including cancer prophylaxis and treatment. Even though a couple of cases have reached the clinic, these products have yet to overcome a number of biological and technological challenges before broad utilization. Concerning the manufacturing processes, there is significant research focusing on the optimization of current cell culture systems and, more recently, on developing scalable downstream processes to generate material for pre-clinical and clinical trials. We review the current options for downstream processing of these complex biopharmaceuticals and underline current advances on knowledge-based toolboxes proposed for rational optimization of their processing. Rational tools developed to increase the yet scarce knowledge on the purification processes of complex biologicals are discussed as alternative to empirical, “black-boxed” based strategies classically used for process development. Innovative methodologies based on surface plasmon resonance, dynamic light scattering, scale-down high-throughput screening and mathematical modeling for supporting ion-exchange chromatography show great potential for a more efficient and cost-effective process design, optimization and equipment prototyping.     

Merkel-like basal cells in the nasal septal island of dromedaries: Ultrastructure and possible functions

Unlike other Merkel cell types, the morphology and functions of the Merkel-like basal cells remain unclear. The aim of the present study was to investigate the ultrastructural features of Merkel-like basal cells in the nasal septal island (NSI) of dromedaries (Camelus dromedarius) using transmission electron microscopy and to speculate their potential functions. Ten pairs of nasal septal islands obtained from ten heads of dromedary camels were used for the current study. Interestingly, these cells have been identified in the basal layer of the neuroepithelium of the dromedary nasal septal island near the sensory nerve endings. These cells were ovoid to elliptical in shape and rested on the basal lamina. Their surface had spine like cytoplasmic processes which interwined with the adjacent basal cells. Their nuclei were large lobulated with 2–3 deep notches. Moreover, numerous dense-core granules surrounded by electron-lucent halo were aggregated in the basal portion of the cells close to the nerve ending as well as melanin pigments in the apical portion. The ultrastructural characteristics of the Merkel-like basal cells of NSI were typical to those of Merkel cells, but with some morphological differences, including their location, cellular attachments, and connections to other structures. The potential functions were discussed in the light of the cellular context and architecture. The Merkel-like basal cells of the NSI neuroepithelium might play a role in nociception and magnetoreception in dromedaries.     

The carrot,not the stick: appetitive rather than aversive gustatory stimuli support associative olfactory learning in individually assayed <Emphasis Type="Italic">Drosophila</Emphasis> larvae

The ability to learn is universal among animals; we investigate associative learning between odors and tastants in larval Drosophila melanogaster. As biologically important gustatory stimuli, like sugars, salts, or bitter substances have many behavioral functions, we investigate not only their reinforcing function, but also their response-modulating and response-releasing function. Concerning the response-releasing function, larvae are attracted by fructose and repelled by sodium chloride and quinine; also, fructose increases, but salt and quinine suppress feeding. However, none of these stimuli has a nonassociative, modulatory effect on olfactory choice behavior. Finally, only fructose but neither salt nor quinine has a reinforcing effect in associative olfactory learning. This implies that the response-releasing, response-modulating and reinforcing functions of these tastants are dissociated on the behavioral level. These results open the door to analyze how this dissociation is brought about on the cellular and molecular level; this should be facilitated by the cellular simplicity and genetic accessibility of the Drosophila larva.     

Ultrastructural evidence for the occurrence of three types of mechanosensitive cells in the tentacles of the cubozoan polypCarybdea marsupialis

Summary The ectodermal cell layer in the tentacles of the cubozoan polypCarybdea marsupialis contains four types of cells (types 1–4) bearing specialized cilia. Epitheliomuscular cells (type 1) are characterized by motile cilia with dynein-decorated axonemes. 200 nm long extramembranous filaments of unknown function are restricted to a belt-like region distal to the transition zone. Up to 40 rn long rigid cilia formed by a slender epithelial cell type (type 2) are surrounded by rings of short microvilli. The axonemes of these cilia are composed of incomplete microtubules and lack dynein. Microvilli and cilia are linked by intermembrane connectors. Microtubuledoublets and ciliary membrane are interconnected by microtubule-associated cross-bridges only within this contact region. At the tip of each tentacle a single nematocyte (type 3) is surrounded by 7–10 accessory cells (type 4). These both cell types are equipped with similar cilium-stereovilli-complexes consisting of a cone-like arrangement of stereovilli and a modified cilium. The axonemal modifications of the cilium, its interconnections with the surrounding stereovilli and the linkages between ciliary axoneme and ciliary membrane are similar to those known from the cnidocil-complexes of hydrozoons and other epithelial mechanosensitive cells of the collar-receptor type. Our data indicate that besides the nematocyte two other types of mechanosensory cells (types 2 and 4) are integrated in the ectodermal cell layer ofCarybdea which possibly affect the triggering mechanism of nematocyst discharge.     

Evolution of gustatory sensitivity in Yponomeuta caterpillars: sensitivity to the stereo-isomers dulcitol and sorbitol is localised in a single sensory cell

The cellular localisation of the sensitivity to two host-plant specific larval phagostimulants was determined in Yponomeuta evonymellus and in inter-specific hybrids of Y. cagnagellus and Y. padellus. The combined results of cross-adaptation experiments, analysis of mixture responses and inter-spike interval distributions indicated that the sensitivity for sorbitol and dulcitol is localised in the same cell in the investigated species. This result suggests that during the evolution of the genus the main alteration in the mechanism of sugar-alcohol detection occurred at the level of the receptor proteins. Relatively simple modification of this kind can, however, have large effects, since larvae with modified receptor proteins will directly respond to the new stimulus while all central processing and resulting behaviour can remain unmodified. Accepted: 7 October 1998     

Monitoring biological wastewater treatment processes: recent advances in spectroscopy applications

Biological processes based on aerobic and anaerobic technologies have been continuously developed to wastewater treatment and are currently routinely employed to reduce the contaminants discharge levels in the environment. However, most methodologies commonly applied for monitoring key parameters are labor intensive, time-consuming and just provide a snapshot of the process. Thus, spectroscopy applications in biological processes are, nowadays, considered a rapid and effective alternative technology for real-time monitoring though still lacking implementation in full-scale plants. In this review, the application of spectroscopic techniques to aerobic and anaerobic systems is addressed focusing on UV–Vis, infrared, and fluorescence spectroscopy. Furthermore, chemometric techniques, valuable tools to extract the relevant data, are also referred. To that effect, a detailed analysis is performed for aerobic and anaerobic systems to summarize the findings that have been obtained since 2000. Future prospects for the application of spectroscopic techniques in biological wastewater treatment processes are further discussed.     

Encoding of forces by cockroach tibial campaniform sensilla: implications in dynamic control of posture and locomotion

Forces exerted by a leg in support and propulsion can vary considerably when animals stand upon or traverse irregular terrains. We characterized the responses of the cockroach tibial campaniform sensilla, mechanoreceptors which encode force via strains produced in the exoskeleton, by applying forces to the leg at controlled magnitudes and rates. We also examined how sensory responses are altered in the presence of different levels of static load. All receptors exhibit phasico-tonic discharges that reflect the level and rate of force application. Our studies show that: (1) tonic discharges of sensilla can signal the level of force, but accurate encoding of static loads may be affected by substantial receptor adaptation and hysteresis; (2) the absolute tonic sensitivities of receptors decrease when incremental forces are applied at different initial load levels; (3) phasic discharges of sensilla accurately encode the rate of force application; and (4) sensitivities to changing rates of force are strictly preserved in the presence of static loads. These findings imply that discharges of the sensilla are particularly tuned to the rate of change of force at all levels of leg loading. This information could be utilized to adapt posture and walking to varying terrains and unexpected perturbations. Accepted: 8 January 2000     

Cytokeratin 14 is expressed in immature cells in rat taste buds

We analyzed the differentiation of taste bud cells, by precisely describing expression profiles of cytokeratins (CKs) 8 and 14 in relation to those of marker molecules including label of 5-bromo-2′-deoxy uridine (BrdU) injected. In rat circumvallate papillae, cell division was observed at the basal layer of the epithelium expressing CK14 and located outside taste buds. The progenitor cells began to migrate toward the apical surface and maintained CK14 expression at 1 day after BrdU injection (day 1). On the other hand, a minor population of newly divided cells was infrequently incorporated into taste buds and also maintained CK14 expression at day 1. In taste buds, the conversion of CK subtypes occurred from CK14 to cytokeratin 8 (CK8) at day 2–3, showing the differentiation from immature cells expressing CK14 into mature or maturing cells expressing CK8. Functionally matured cells such as taste receptor cells expressing inositol triphospate receptor type 3 (IP₃R3) never expressed CK14, suggesting that CK14 would be expressed only in immature cells. On the other hand, a small but distinct population of BrdU-positive cells still showed CK14 immunoreactivity in taste buds even at day 12, which might correspond to the cells that remain undifferentiated for a long period within taste buds.     

Primary processes in heme-based sensor proteins

A wide and still rapidly increasing range of heme-based sensor proteins has been discovered over the last two decades. At the molecular level, these proteins function as bistable switches in which the catalytic activity of an enzymatic domain is altered mostly by binding or dissociation of small gaseous ligands (O₂, NO or CO) to the heme in a sensor domain. The initial “signal” at the heme level is subsequently transmitted within the protein to the catalytic site, ultimately leading to adapted expression levels of specific proteins. Making use of the photolability of the heme-ligand bond that mimics thermal dissociation, early processes in this intra-protein signaling pathway can be followed using ultrafast optical spectroscopic techniques; they also occur on timescales accessible to molecular dynamics simulations. Experimental studies performed over the last decade on proteins including the sensors FixL (O₂), CooA (CO) and soluble guanylate cyclase (NO) are reviewed with an emphasis on emerging general mechanisms. After heme-ligand bond breaking, the ligand can escape from the heme pocket and eventually from the protein, or rebind directly to the heme. Remarkably, in all sensor proteins the rebinding, specifically of the sensed ligand, is highly efficient. This ”ligand trap” property possibly provides means to smoothen the effects of fast environmental fluctuations on the switching frequency. For 6-coordinate proteins, where exchange between an internal heme-bound residue and external gaseous ligands occurs, the study of early processes starting from the unliganded form indicates that mobility of the internal ligand may facilitate signal transfer. This article is part of a Special Issue entitled: Oxygen Binding and Sensing Proteins.