Functions of Opsins in Drosophila Taste

1. Download : Download high-res image (230KB)
2. Download : Download full-size image

     

Analysis of Slow Hyperpolarizing Potentials in Frog Taste Cells Induced by Glossopharyngeal Nerve Stimulation

Chemical Senses, 29,     

Constrained Ordination Analysis with Enrichment of Bell-Shaped Response Functions

Constrained ordination methods aims at finding an environmental gradient along which the species abundances are maximally separated. The species response functions, which describe the expected abundance as a function of the environmental score, are according to the ecological fundamental niche theory only meaningful if they are bell-shaped. Many classical model-based ordination methods, however, use quadratic regression models without imposing the bell-shape and thus allowing for meaningless U-shaped response functions. The analysis output (e.g. a biplot) may therefore be potentially misleading and the conclusions are prone to errors. In this paper we present a log-likelihood ratio criterion with a penalisation term to enforce more bell-shaped response shapes. We report the results of a simulation study and apply our method to metagenomics data from microbial ecology.     

The Distribution of Afferent Neurons in the Trigeminal Mesencephalic Nucleus and the Central Projection of Afferent Fibers of the Mylohyoid Nerve in the Rat

Horseradish peroxidase conjugated to wheatgerm agglutinin (HRP:WGA) was injected into the proximal cut ends of three branches of the mylohyoid nerve in rats: the branch to the mylohyoid muscle (BrMh), the branch to the anterior belly of the digastricus muscle (BrDg), and the cutaneous branch (BrCu). HRP-labeled cells were detected in the ipsilateral caudal portion of the trigeminal mesencephalic nucleus (Vmes) and the ipsilateral ventromedial division of the trigeminal motor nucleus, except when HRP:WGA was applied to the BrCu. Morphologically, all labeled Vmes cells were of the pseudounipolar type.

Projections of the primary afferents of the BrMh were observed in the ipsilateral trigeminal nucleus caudalis, the upper cervical dorsal horns of laminae I -III, and the dorsolateral recticular formation (Rf), whereas the primary afferents of the BrDg terminated in the ipsilateral trigeminal nucleus principalis and Rf. These observations suggest that the role of the afferent inputs of the mylohyoid muscle differs from that of those of the anterior belly of the digastricus muscle in terms of several functions associated with jaw-closing and infrahyoid muscles.     

Postnatal Changes in Enzyme Activities in Synaptosomes Isolated from Hamster Optic Nerve Endings

Subsynaptosomal fractions isolated from optic terminal nuclei of adult and neonatal hamsters exhibited developmental changes in specific density, mitochondrial activity, and K+-stimulated, ouabain-inhibited p-nitrophenylphosphatase (K-pNPPase) activity around the time of eye opening. The specific activity of K-pNPPase was six- to sevenfold higher after eye opening (14-16 days postnatal). A significant proportion of high-specific- activity K-pNPPase was recovered from the lightest subsynaptosomal fraction at all ages. This fraction contained very little external membrane by galactose oxidase - NaB3H4 labeling, suggesting that it may represent an internal pool, possibly the axonally transported form of the enzyme. Synaptic mitochondrial cytochrome c. oxidase activity also approximately doubled in the period between 12 and 16 days. The specific density of the external membrane increased very slowly, banding at 1.0 M sucrose at 12 and 16 days, and at 1.2 M in adults. These maturational events may reflect increased energetic needs for optic nerve endings following eye opening.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

C57BL/6 and AKR mice were treated with hamster erythrocytes (HRBC) in complete Freund's adjuvant (CFA) or incomplete Freund's adjuvant (IFA) and the development of delayed hypersensitivity and antibody production were examined. 1) Delayed hypersensitivity against HRBC antigen, as determined by the peritoneal macrophage disappearance test, was detected in mice sensitized with HRBC in CFA but not in those sensitized with HRBC in IFA. 2) Antibody production against HRBC or hapten TNP after a booster injection of HRBC or trinitrophenylated HRBC (TNP-HRBC) in saline was enhanced by pretreatment with HRBC in CFA or IFA. 3) Delayed hypersensitivity was not detectable after a booster sensitization with HRBC in CFA in mice which had been pretreated with HRBC in IFA 2 weeks earlier. In the mice treated with both HRBC in IFA (day ?21) and in CFA (day ?7), however, an enhanced antibody production against HRBC or TNP was detected after an intravenous injection with HRBC or TNP-HRBC in saline (day 0). These results suggest that sensitized effector lymphocytes in delayed hypersensitivity and helper cells in antibody production may be derived from the same pool of unprimed T cells. The pool of unprimed T cells with a capacity to differentiate into either type of primed T cells may be exhausted after pretreatment with the antigen in IFA, and the primed helper T cells may not be able to differentiate into sensitized lymphocytes even after sensitization with the antigen in CFA, which favors development of delayed hypersensitivity in normal controls.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

Antibody response against hamster red blood cells (H-RBC) was examined in inbred strains of C57BL/6, AKR, C3H/He, DDD and SL mice, and outbred CF1 mice. 1) There were strain differences in antibody response after a primary intravenous injection of H-RBC. DDD, SL and CF1 mice belonged to high-responder strains, while C57BL/6, AKR and C3H/He to low-responder strains. In the spleens of immunized CF1 and SL, 40 to 70 times as many plaque-forming cells (PFC) as those in C57BL/6 mice were detected. The magnitudes of the response were: CF1 ≒ SL>DDD>>C3H/He ? AKR>C57BL/6. 2) 2-mercaptoethanol resistant (MER) antibody was detected in neither low- nor high-responders after a primary intravenous antigen-injection. 3) After a secondary intravenous antigen-injection, MER antibody was detected in all the SL mice, but only in 30 to 50% of AKR and C57BL/6 mice. 4) A subcutaneous injection of H-RBC in Freund's complete adjuvant (FCA) did not elicit antibody production within 10 days. When mice pre-sensitized 7 days in advance w^Tith H-RBC in FCA were intravenously injected with H-RBC, enhanced antibody production of the primary type was observed in all the mouse strains. 5) In pre-sensitized mice, the extent of the enhancement of antibody production was the highest in low-responder C57BL/6 mice and the lowest in high-responder SL and CF1 strains. Thus, there was no strain difference in antibody titers or the numbers of PFC after the booster.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

In a previous paper we reported that an inbred strain of SL mice and an outbred strain of CF1 mice belonged to the high-responder strains in antibody production after primary immunization with hamster erythrocytes (H-RBC), while inbred strains of C57BL/6, AKR and C3H/He mice belonged to low-responder strains. In the present study we obtained the following results. 1) Pre-sensitization with hamster lymphoma enhanced antibody production after an intravenous injection of H-RBC. There was no strain difference in the pattern of antibody production against H-RBC among pre-sensitized mice. 2) The pattern of enhanced antibody production after an intravenous injection of H-RBC into pre-sensitized mice assumed the primary type in terms of time of appearance of hemolysin plaque-forming cells (PFC) in the spleens and the conversion from 2-mercaptoethanol sensitive to 2-mercaptoethanol resistant antibody production, when the intervals between both treatments were within 7 days. 3) Pre-sensitization with lymphoma induced not only an increase in numbers of PFC after an intravenous injection of H-RBC, but also an increase in the size of the hemolysin plaques. These results suggested that sensitization with hamster lymphoma stimulated some kinds of immuno-competent cells, which could contribute to antibody production against H-RBC after a booster injection of H-RBC.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

Patterns of proliferation of antibody-forming cells after an intravenous immunization with hamster erythrocytes (HRBC) were compared in groups of mice possessing different activities of thymus-derived lymphocytes (T cells). 1) Marked differences in the numbers of hemolysin plaque-forming cells (PFC) after HRBC injection were found among the low- and high-responder normal mice and those pretreated with HRBC in complete Freund's adjuvant (CFA) or incomplete adjuvant (IFA), and they appeared to depend primarily upon the different rates of proliferation of antibody-forming cells rather than on the numbers of antigen-specific lymphocytes initiating the antibody response. 2) The numbers of hemolytic foci were slightly larger in mice with large numbers of PFC (normal SL mice, the pretreated SL and C57BL/6 mice) than in those with small numbers of PFC (normal C57BL/6 mice). The numbers of hemolytic foci increased at almost the same rate from day 2 to day 3 in both groups, while the numbers of PFC increased more efficiently in mice with large numbers of PFC than in those with small numbers of PFC from day 2 to day 3. Individual hemolytic foci appeared to contain larger numbers of PFC in mice with large total numbers of PFC than in those with small total numbers of PFC. 3) The numbers of rosette-forming cells (RFC) were increased by pretreatment with HRBC in CFA and by pretreatment with HRBC in IFA to almost the same extent. Rates of increases in PFC were, however, larger by pretreatment with HRBC in CFA than with HRBC in IFA. These results suggested that the activity of the T cell determined not only the rates of proliferation of antibody-forming cells but also the antibody-producing capacity of each cell.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

Enhancing and suppressing effects of microbial adjuvants were studied in female mice of the C3H/He, AKR and SL strains. Propionibacterium acnes, Bordetella pertussis, BCG and yeast cell wall (YCW) were chosen as adjuvants. As antigens, we chose hamster erythrocytes (HRBC) which proved to be a weak antigen for mice. Adjuvants were given on day —7, day 0 or day 3, and HRBC were injected on day 0. The results were as follows. 1) P. acnes facilitated IgM and IgG antibody production in AKR mice and suppressed IgM antibody production in SL mice, when given on day —7. When P. acnes was given on day 0, they suppressed IgM antibody production in all of the strains used. 2) When B. pertussis was given on day 0, it exhibited enhancing effects on IgG antibody production in all of the strains and a suppressing effect on IgM antibody production in SL mice. 3) BCG suppressed IgM antibody production in all strains when given on day 0. 4) YCW showed no influence on antibody production in any combination used in this work. 5) SL mice were very sensitive to suppressing effects by adjuvants. Strain differences in the expression of enhancing and suppressing effects by adjuvants appear to be under some control independent of antigen-specific immune response genes.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

Delayed hypersensitivity against hamster erythrocyte antigen was examined after sensitization with hamster erythrocytes (HRBC) in Freund's complete adjuvant (FCA). Extent of the delayed hypersensitivity was determined by the migration inhibition test, the peritoneal macrophage disappearance test and the skin test in the ear using solubilized HRBC as the test antigen. 1) Delayed hypersensitivity against HRBC developed earlier in high-responder SL mice than in low-responder C57BL/6 mice after sensitization. The period required for development of the delayed hypersensitivity in AKR mice was intermediate between periods in high-responder SL mice and low-responder C57BL/6 mice. 2) After sensitization with HRBC in FCA, a delayed hypersensitive state without detectable antibody production persisted until day 12 in high-responder SL mice and until day 16 or later in low-responder C57BL/6 mice. 3) Delayed hypersensitivity against HRBC antigen persisted even after the appearance of circulating antibody which occurred late after sensitization with HRBC in FCA or after intravenous injection of HRBC into sensitized mice.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

It was confirmed by passive transfer experiments that the function of thymus-derived cells specific for hamster erythrocytes (HRBC) was deficient in the low-responder mouse strains. 1) Antibody production against HRBC was enhanced by passive transfer of thymus cells from normal SL mice (high-responder) to normal C57BL/6 mice (low-responder). 2) The enhancing effect of passive transfer of thymus cells from SL mice was abrogated by pre-sensitization of the recipients (C57BL/6) with thymus cells from SL mice. 3) In C57BL/6 mice, antibody production against HRBC was enhanced by the transfer of lymph node or spleen cells from C57BL/6 mice which had been sensitized with HRBC in Freund's complete adjuvant or hamster lymphoma cells.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

Golden hamsters were used as hosts in this work, and mice of various strains as donors of antigens. 1) There were no strain differences in immunogenicity of erythrocytes from C57BL/6, AKR, SL and CF1 mice. 2) Primary intravenous immunization with mouse erythrocytes (MRBC) induced the production of hemolysin plaque-forming cells (PFC) in a large number, but elicited only in a negligible titer production of 2-mercaptoethanol-resistant antibody. 3) 2-Mercaptoethanol-resistant antibody was produced more efficiently in hamsters pre-sensitized with mouse lymph node (MLN) cells rather than those pre-immunized with MRBC after a booster with MRBC. 4) Numbers of PFC in pre-sensitized hamsters were three-times that of the non-sensitized hamsters after a booster with MRBC, when pre-sensitization was performed intradermally with a small number of MLN cells. 5) Average diameter of the hemolysin plaques in pre-sensitized hamsters was one and a half times larger than that in non-sensitized hamsters. Conclusions agree well with the results in our previous papers that the reversed combination of hosts and antigen donors employed support the concept that certain processes required for delayed hypersensitivity contributed to antibody production under a condition suitable for antibody response.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

The production of anti-hapten antibody after immunization with trinitrophenylated (TNP) hamster erythrocytes (HRBC) or sheep erythrocytes (SRBC) was determined in high- and low-responder mouse strains against HRBC antigen. 1) Anti-TNP antibody was detected in sera of high-responder DDD and CF1 mice after primary immunization with TNP-HRBC, but not in those of low-responder C57BL/6 mice. 2) Anti-TNP antibody was detectable in sera of all the strains after primary immunization with TNP-SRBC. 3) Production of anti-TNP antibody was elicited after a booster injection of TNP-HRBC in low-responder C57BL/6 mice pre-sensitized with HRBC in Freund's complete adjuvant. These results suggest that functions of thymus-derived cells specific for HRBC antigen are deficient in low-responder mice.     

Immune Response against Hamster Erythrocytes in the Low-Responder Mouse Strains

1) A subcutaneous injection of hamster erythrocytes (HRBC) in Freund's complete adjuvant (FCA) or an intravenous injection of hamster lymph node (HLN) cells suppressed antibody production against HRBC in the low-responder C57BL/6 and AKR mice, when HRBC in saline were given on the same day; 2) The suppressing effect of such treatments was neither detectable in the high-responder SL mice, nor in the C57BL/6 mice, which had been pre-sensitized with HRBC in FCA or hamster lymphoma cells; 3) Positive reactions of the peritoneal macrophage disappearance test and the enhanced antibody production were detected seven days after treatment with HRBC in FCA and HRBC in saline, or HLN cells and HRBC in saline; 4) The suppressing effect of such simultaneous treatments on anti-HRBC antibody production was eliminated by a transfer of normal syngeneic thymus cells to AKR mice or a transfer of thymus cells from SL to C57BL/6 mice. Suppression of the antibody production in the low-responder mice by the described simultaneous treatments may be due to a competitive involvement of HRBC-specific thymus-derived cells (T cells) in the developmental stages of delayed hypersensitivity and antibody production. High-responder SL mice appear to have enough T cells for development of the delayed hypersensitivity and as helper cells in antibody production. These results appear to support the concept that T cells for delayed hypersensitivity and antibody production to HRBC antigen are derived from the same original pool.     

Characterization of the Host Response to Pichinde Virus Infection in the Syrian Golden Hamster by Species-Specific Kinome Analysis

The Syrian golden hamster has been increasingly used to study viral hemorrhagic fever (VHF) pathogenesis and countermeasure efficacy. As VHFs are a global health concern, well-characterized animal models are essential for both the development of therapeutics and vaccines as well as for increasing our understanding of the molecular events that underlie viral pathogenesis. However, the paucity of reagents or platforms that are available for studying hamsters at a molecular level limits the ability to extract biological information from this important animal model. As such, there is a need to develop platforms/technologies for characterizing host responses of hamsters at a molecular level. To this end, we developed hamster-specific kinome peptide arrays to characterize the molecular host response of the Syrian golden hamster. After validating the functionality of the arrays using immune agonists of defined signaling mechanisms (lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α), we characterized the host response in a hamster model of VHF based on Pichinde virus (PICV¹) infection by performing temporal kinome analysis of lung tissue. Our analysis revealed key roles for vascular endothelial growth factor (VEGF), interleukin (IL) responses, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling, and Toll-like receptor (TLR) signaling in the response to PICV infection. These findings were validated through phosphorylation-specific Western blot analysis. Overall, we have demonstrated that hamster-specific kinome arrays are a robust tool for characterizing the species-specific molecular host response in a VHF model. Further, our results provide key insights into the hamster host response to PICV infection and will inform future studies with high-consequence VHF pathogens.Viruses of the Arenaviridae family cause persistent and asymptomatic infections of rodents throughout the world (1). However, infection with select members of the Arenaviridae can result in viral hemorrhagic fever (VHF) in humans and non-human primates. Lassa virus (LASV), a Risk Group 4 Select Agent, is responsible for severe febrile illness in 15–20% of those infected (2, 3). While pathophysiological studies for LASV are limited to high-containment laboratories, Pichinde virus (PICV), a South American arenavirus, has been investigated as a potential surrogate for LASV and can be handled under biosafety level 2 conditions (4). Further, the histopathological effects of PICV infection of guinea pigs and hamsters are similar to those induced by LASV in humans (4–6). Thus, PICV is attractive as a surrogate virus for modeling the hemorrhagic disease process of LASV.Recently, there has been interest in the use of the Syrian golden hamster as a model for high-consequence pathogens (7–9). In contrast to mice and guinea pigs, hamsters more closely recapitulate the pathological effects of VHF infections in non-human primates, particularly with regard to coagulopathy. It has been posited that VHF infection studies in hamsters will supplant those of mouse and guinea pigs (7–9). However, such investigations are limited by the lack of available hamster-specific molecular reagents (9). Thus, there is a great need for technologies that can characterize the molecular host response of hamsters.Despite the potential threat to global health posed by high-consequence pathogens, there is limited information regarding the role of host cell signaling network dysregulation in the molecular pathogenesis of these pathogens. As the molecular events associated with disease are essential to understanding pathogenesis and identifying drug targets, it is important to examine the dynamic cellular changes that occur during infection. From the perspective of host cellular responses, phosphorylation of host proteins is the most well-characterized post-translational modification (PTM) with regard to cell signaling regulation, as well as the most ubiquitous PTM found within host cells (10). While gene expression data are informative, defining host responses at the level of host cell kinases (the kinome) provides a unique perspective regarding host-pathogen interactions. In particular, kinase-mediated phosphorylation events provide the host with a mechanism to rapidly respond to environmental changes (stress, infection, etc.) through the modulation of cell signaling networks as compared with more long-term changes in gene or protein expression. Further, there is increasing interest in the use of kinome analyses for the identification of novel therapeutic targets, including malaria and other parasitic diseases (11). Importantly, kinases are a primary target for the development of therapeutics with 25 kinase inhibitors currently licensed for the treatment of various malignancies by the U.S. Food and Drug Administration (12). Further, the investigation of kinase inhibitors as therapeutics for infectious disease fulfills a National Institutes of Health mandate focused on the repurposing of drugs (12, 13).Recently, there has been an increased appreciation for the role of kinase-mediated signaling events in the host response to pathogens (14–17). Members of the Arenaviridae are known to modulate normal host immune signaling function (18, 19). As many immune responses are modulated through kinase-mediated signaling cascades, it is essential to examine the host response at this level. Previous investigations have attempted to characterize these responses; however, these investigations have been limited by the lack of availability of species-specific reagents and the relative scarcity of information regarding the host-specific molecular responses (18). Thus, there has been little investigation into the molecular response of hamster to high-consequence pathogens (5, 6).Here, we present a novel analysis for characterizing hamster-specific kinome responses that overcome these challenges. Through our analysis, we have demonstrated the utility of hamster-specific kinome arrays for assessing species-specific host responses to innate immune agonists of well-defined signaling pathways, bacterial lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α. Moreover, we have characterized temporal host responses in the lungs of PICV-infected hamsters and demonstrated for the first time that vascular endothelial growth factor (VEGF), interleukin (IL) responses, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling, and Toll-like receptor (TLR) signaling form central components of the host response to PICV infection in the lungs of the Syrian golden hamster. Further, we also demonstrated that the modulation of VEGF- and angiogenic-related cell signaling processes correlated with modulation of downstream cell adhesion molecule expression. Importantly, these events were unique to late-stage disease in PICV-infected animals as compared with sham-infected animals. Taken together, our investigation provides a methodology for the global analysis of host responses of the Syrian golden hamster to pathogen insult and illustrates the potential to use data from this method to characterize the activity state of functional signaling networks useful for identifying therapeutic targets.     

双斑蟋触角传入神经纤维在脑内投射分布研究

目的:研究昆虫触角感受器传入神经末梢在脑内投射的空间布局,揭示触角感觉信息传入的神经结构.方法:使用氯化镍神经元示踪标记技术,对双斑蟋触角感受嚣传入神经纤维进行可视化标记,观察研究触角传入神经末梢在脑内的走行形态及分布规律.结果:双斑蟋触角感受器传入神经纤维进入中脑后大量的神经末梢终止在同侧的触角叶和触角机械感觉运动中枢,部分神经纤维向前走行,其神经末梢终止在前脑,还有部分神经纤维向后下行,经同侧神经索,其神经末梢终止在食道下神经节.结论:双斑蟋触角感受器传入神经纤维进入脑后主要投射到触角叶和触角机械感觉运动中枢,少部分投射到前脑和食道下神经节.这种多重投射模式可能在双斑蟋嗅觉信息传递整合、触角运动调节、味觉和摄食活动等方面发挥重要作用.     

An Electron Microscope Study of Polyoma Virus in Hamster Kidney

Electron microscope studies were made of hamster kidneys taken at daily intervals after injection of a variant of polyoma virus into newborn animals. Particular attention was paid to the period 5 to 6 days after injection at which time the necrotizing response was at its peak and virus particles were seen in greatest numbers. The most numerous particles were about 28 mµ in diameter. They were observed mainly within nuclei of stromal cells and are similar to the particles seen in large numbers in polyoma-infected mouse cells growing in vitro. They were not observed in cells of fully developed tumors. Filamentous or tubular structures closely associated with the 28 mµ particles and probably concerned in their formation are described. Considerable quantities of viral material were contained within cytoplasmic inclusions. In some of the inclusions larger particles of diameter 60 mµ were observed. The origin of these particles and their relation to the 28 mµ particles is discussed.     

Evoked Activity of Afferent and Efferent Fibers of the Sciatic Nerve in Rats under Conditions of Experimental Hyperthyroidism

In Wistar albino rats with experimental hyperthyroidism (HTh) and control animals, we measured parameters of the responses evoked in peripheral segments of the ventral and dorsal roots (VR and DR, respectively) by stimulation of the sciatic nerve. We found that the chronaxia of the afferent fibers of the sciatic nerve in HTh animals is shorter, while the duration of the mass action potential (AP) in the DR is somewhat longer than in the control. Under conditions of HTh, the excitation threshold of the efferent fibers became higher, the chronaxia decreased, and the second high-amplitude component could appear in the AP recorded from the VR. Possible mechanisms of changes in the excitability of afferent and efferent fibers of the sciatic nerve and specific features of the AP recorded from the VR under HTh conditions are discussed. In particular, we consider the possibility of ephaptic spreading of excitation in VR fibers under HTh conditions.