Confirmation of sable QTL that modifies the effects of the Ay allele on yellow coat color on mouse chromosome 1

F₁-A^y mice between RR (aabbCC) and C57BL/6J-A^y (A^yaBBCC) have a much darker yellow coat color than do C57BL/6J-A^y. Quantitative trait locus (QTL) analysis was carried out to identify genes responsible for the darker modification of the yellow coat color (this has been traditionally termed “sable”). A significant sable QTL was identified on chromosome 1 (Dmyaq4, LOD score 15.5 for lightness, and 13.4 for color difference), in a chromosomal position similar to that of Dmyaq2, a sable QTL previously identified in C3H/HeJ. Another significant sable QTL was identified on chromosome 4 (Dmyaq5, LOD score 5.6 for lightness, and 4.3 for color difference) near the tyrosinase-related protein 1 (Tyrp1) locus. The effect of Dmyaq5 was significant only in the presence of the RR allele at Dmyaq4, suggesting that the Dmyaq4 (as well as Dmyaq2) is a novel coat color gene that may act up-stream of Tyrp1 signaling to increase eumelanin production.     

STDP-driven networks and the C. elegans neuronal network

We study the dynamics of the structure of a formal neural network wherein the strengths of the synapses are governed by spike-timing-dependent plasticity (STDP). For properly chosen input signals, there exists a steady state with a residual network. We compare the motif profile of such a network with that of a real neural network of C. elegans and identify robust qualitative similarities. In particular, our extensive numerical simulations show that this STDP-driven resulting network is robust under variations of the model parameters.     

Ultrastructural immunogold localization of major sperm protein (MSP) in spermatogenic cells of the nematode Acrobeles complexus (Nematoda,Rhabditida)

The nematode spermatozoa represent a highly modified (aberrant) type of male gametes that lack a flagellum but for which the process of spermatogenesis culminates in the production of a crawling spermatozoon on the basis of the cytoskeletal component known as “major sperm protein”, or MSP. MSP is also known as an important hormone triggering oocyte maturation and ovulation in the model nematode Caenorhabditis elegans, where this protein was first identified. However, direct evidence of MSP localization and of its fate in nematode spermatogenic cells is rare. In this study, the spermatogenesis and sperm structure in the rhabditid nematode Acrobeles complexus (Rhabditida: Tylenchina: Cephalobomorpha: Cephaloboidea: Cephalobidae) has been examined with electron microscopy. Morphological observations were followed by high-pressure freezing and freeze-substitution fixation which allows post-embedding immunogold localization of MSP in all stages of sperm development using antibodies raised for MSP of C. elegans. In spermatocytes, synthetic activity results in the development of specific cellular components, fibrous bodies (FB) and membranous organelles (MO), which appear as FB-MO complexes where the filamentous matter of FB has been MSP-labeled. The spermatids subdivide into a residual body with superfluous cytoplasm, and a main cell body which contains nucleus, mitochondria and FB-MO complexes. These complexes dissociate into individual components, MO and FB, with the MSP being localized in FB. Immature spermatozoa from testes are opaque cells where a centrally located nucleus is surrounded by mitochondria, MO and FB clustered together, the MSP still being localized only in FB. Cytoplasm of mature spermatozoa from spermatheca is segregated into external pseudopods lacking organelles and a central cluster of mitochondria with intact MO surrounding the central nucleus. The FB ultimately disappear, and the MSP labeling becomes concentrated in the filamentous content of pseudopods and cytoplasm of the main cell body. Although the spermatogenesis and sperm structure of A. complexus is similar to that of many other rhabditid nematodes, their intact MO makes the morphology of the mature spermatozoa distinct from the “rhabditid pattern” and may be considered as a synapomorphy. The MSP localization in spermatogenic cells of A. complexus also follows the “rhabditid pattern” described in C. elegans and Ascaris spp. Our results and techniques of MSP labeling of A. complexus spermatogeneous cells reveal new possibilities to elucidate different research questions on MSP localization in nematodes related to C. elegans. Furthermore, the laboratory-cultured A. complexus strains can be used as a new and fascinating model to study MO and MSP functions in nematode reproduction.     

The nestin-expressing and non-expressing neurons in rat basal forebrain display different electrophysiological properties and project to hippocampus

Background

In vertebrates and invertebrates, sensory neurons adapt to variable ambient conditions, such as the duration or repetition of a stimulus, a physiological mechanism considered as a simple form of non-associative learning and neuronal plasticity. Although various signaling pathways, as cAMP, cGMP, and the inositol 1,4,5-triphosphate receptor (InsP₃R) play a role in adaptation, their precise mechanisms of action at the cellular level remain incompletely understood. Recently, in Drosophila, we reported that odor-induced Ca²⁺-response in axon terminals of olfactory receptor neurons (ORNs) is related to odor duration. In particular, a relatively long odor stimulus (such as 5 s) triggers the induction of a second component involving intracellular Ca²⁺-stores.

Results

We used a recently developed in-vivo bioluminescence imaging approach to quantify the odor-induced Ca²⁺-activity in the axon terminals of ORNs. Using either a genetic approach to target specific RNAs, or a pharmacological approach, we show that the second component, relying on the intracellular Ca²⁺-stores, is responsible for the adaptation to repetitive stimuli. In the antennal lobes (a region analogous to the vertebrate olfactory bulb) ORNs make synaptic contacts with second-order neurons, the projection neurons (PNs). These synapses are modulated by GABA, through either GABAergic local interneurons (LNs) and/or some GABAergic PNs. Application of GABAergic receptor antagonists, both GABA_A or GABA_B, abolishes the adaptation, while RNAi targeting the GABAB_R (a metabotropic receptor) within the ORNs, blocks the Ca²⁺-store dependent component, and consequently disrupts the adaptation. These results indicate that GABA exerts a feedback control. Finally, at the behavioral level, using an olfactory test, genetically impairing the GABA_BR or its signaling pathway specifically in the ORNs disrupts olfactory adapted behavior.

Conclusion

Taken together, our results indicate that a relatively long lasting form of adaptation occurs within the axon terminals of the ORNs in the antennal lobes, which depends on intracellular Ca²⁺-stores, attributable to a positive feedback through the GABAergic synapses.     

Elucidating a normal function of huntingtin by functional and microarray analysis of huntingtin-null mouse embryonic fibroblasts

Background

Multi-sensory integration is necessary for organisms to discriminate different environmental stimuli and thus determine behavior. Caenorhabditis elegans has 12 pairs of amphid sensory neurons, which are involved in generating behaviors such as thermotaxis toward cultivation temperature, and chemotaxis toward chemical stimuli. This arrangement of known sensory neurons and measurable behavioral output makes C. elegans suitable for addressing questions of multi-sensory integration in the nervous system. Previous studies have suggested that C. elegans can process different chemoattractants simultaneously. However, little is known about how these organisms can integrate information from stimuli of different modality, such as thermal and chemical stimuli.

Results

We studied the behavior of a population of C. elegans during simultaneous presentation of thermal and chemical stimuli. First, we examined thermotaxis within the radial temperature gradient produced by a feedback-controlled thermoregulator. Separately, we examined chemotaxis toward sodium chloride or isoamyl alcohol. Then, assays for simultaneous presentations of 15°C (colder temperature than 20°C room temperature) and chemoattractant were performed with 15°C-cultivated wild-type worms. Unlike the sum of behavioral indices for each separate behavior, simultaneous presentation resulted in a biased migration to cold regions in the first 10 min of the assay, and sodium chloride-regions in the last 40 min. However, when sodium chloride was replaced with isoamyl alcohol in the simultaneous presentation, the behavioral index was very similar to the sum of separate single presentation indices. We then recorded tracks of single worms and analyzed their behavior. For behavior toward sodium chloride, frequencies of forward and backward movements in simultaneous presentation were significantly different from those in single presentation. Also, migration toward 15°C in simultaneous presentation was faster than that in 15°C-single presentation.

Conclusion

We conclude that worms preferred temperature to chemoattractant at first, but preferred the chemoattractant sodium chloride thereafter. This preference was not seen for isoamyl alcohol presentation. We attribute this phase-dependent preference to the result of integration of thermosensory and chemosensory signals received by distinct sensory neurons.     

A Novel Technique of Synthesis of Highly Fluorescent Carbon Nanoparticles from Broth Constituent and In-vivo Bioimaging of <Emphasis Type="Italic">C. elegans</Emphasis>

Here we have demonstrated a novel single step technique of synthesis of highly fluorescent carbon nanoparticles (CNPs) from broth constituent and in vivo bioimaging of Caenorhabditis elegans (C. elegans) with the synthesized CNPs has been presented. The synthesized CNPs has been characterized by the UV-visible (UV-Vis) absorption spectroscopy, transmission electron microscopy (TEM) and Raman studies. The sp ² cluster size of the synthesized samples has been determined from the measured Raman spectra by fitting it with the theoretical skew Lorentzian (Breit-Wigner- Fano (BWF)) line shape. The synthesised materials are showing excitation wavelength dependent tunable photoluminescence (PL) emission characteristics with a high quantum yield (QY) of 3 % at a very low concentration of CNPs. A remarkable increase in the intensity of PL emission from 16 % to 39 % in C. elegans has also been observed when the feeding concentration of CNPs to C. elegans is increased from 0.025 % to 0.1 % (w/v). The non-toxicity and water solubility of the synthesized material makes it ideal candidate for bioimaging.     

Imaging Caenorhabditis elegans embryogenesis by third-harmonic generation microscopy

In this study, third-harmonic generation (THG) imaging measurements were performed to characterize different developmental stages of the nematode Caenorhabditis elegans (C. elegans) embryos. Femtosecond laser pulses (1028 nm) were utilized for excitation. THG image contrast modality proved as a powerful diagnostic tool, providing valuable information and offering new insights into the complex developmental process of C. elegans embryogenesis.     

Dynamical complexity in the C.elegans neural network

We model the neuronal circuit of the C.elegans soil worm in terms of a Hindmarsh-Rose system of ordinary differential equations, dividing its circuit into six communities which are determined via the Walktrap and Louvain methods. Using the numerical solution of these equations, we analyze important measures of dynamical complexity, namely synchronicity, the largest Lyapunov exponent, and the Φ_AR auto-regressive integrated information theory measure. We show that Φ_AR provides a useful measure of the information contained in the C.elegans brain dynamic network. Our analysis reveals that the C.elegans brain dynamic network generates more information than the sum of its constituent parts, and that attains higher levels of integrated information for couplings for which either all its communities are highly synchronized, or there is a mixed state of highly synchronized and desynchronized communities.     

A new paradigm of bacteria-gut interplay brought through the study of Shigella

Bacteria-gut epithelial interplay and the mucosal immune response are the most critical issues in determining the fate of bacterial infection and the severity of diseases. Shigella species (abbreviated here as Shigella), the causative agent of bacillary dysentery (shigellosis), are highly adapted human pathogens that are capable of invading and colonizing the intestinal epithelium, which results in severe inflammatory colitis. Shigella secrete a large and diverse number (more then 50) of effectors via the type III secretion system (TTSS) during infection, some of which are delivered into the surrounding bacterial space and some others into the host cell cytoplasm and nucleus. The delivered effectors mimic and usurp the host cellular functions, and modulate host cell signaling and immune response, thus playing pivotal roles in promoting bacterial infection and circumventing host defense systems. This article overviews the pathogenic characteristics of Shigella, and highlights current topics related to the bacterial infectious stratagem executed by the TTSS-secreted effectors. Though bacterial stratagems and the molecular mechanisms of infection vary greatly among pathogens, the current studies of Shigella provide a paradigm shift in bacterial pathogenesis.     

Neuron participation in a synchrony-encoding assembly

Background

Caenorhabditis elegans actively crawls down thermal gradients until it reaches the temperature of its prior cultivation, exhibiting what is called cryophilic movement. Implicit in the worm's performance of cryophilic movement is the ability to detect thermal gradients, and implicit in regulating the performance of cryophilic movement is the ability to compare the current temperature of its surroundings with a stored memory of its cultivation temperature. Several lines of evidence link the AFD sensory neuron to thermotactic behavior, but its precise role is unclear. A current model contends that AFD is part of a thermophilic mechanism for biasing the worm's movement up gradients that counterbalances the cryophilic mechanism for biasing its movement down gradients.

Results

We used tightly-focused femtosecond laser pulses to dissect the AFD neuronal cell bodies and the AFD sensory dendrites in C. elegans to investigate their contribution to cryophilic movement. We establish that femtosecond laser ablation can exhibit submicrometer precision, severing individual sensory dendrites without causing collateral damage. We show that severing the dendrites of sensory neurons in young adult worms permanently abolishes their sensory contribution without functional regeneration. We show that the AFD neuron regulates a mechanism for generating cryophilic bias, but we find no evidence that AFD laser surgery reduces a putative ability to generate thermophilic bias. In addition, although disruption of the AIY interneuron causes worms to exhibit cryophilic bias at all temperatures, we find no evidence that laser killing the AIZ interneuron causes thermophilic bias at any temperature.

Conclusion

We conclude that laser surgical analysis of the neural circuit for thermotaxis does not support a model in which AFD opposes cryophilic bias by generating thermophilic bias. Our data supports a model in which the AFD neuron gates a mechanism for generating cryophilic bias.     

Biochemical principle of Limulus test for detecting bacterial endotoxins

A hemocyte lysate from horseshoe crab (Limulus) produced a gel, when exposed to Gram-negative bacterial endotoxins, lipopolysaccharides (LPS). This gelation reaction of the lysate, so-called Limulus test, has been widely employed as a simple and very sensitive assay method for endotoxins. Recent biochemical studies on the principle of Limulus test indicate that the hemocytes contain several serine protease zymogens, which constitute a coagulation cascade triggered by endotoxins, and that there is a (1,3)-β-D-glucan-mediated coagulation pathway which also results in the formation of gel. Up to now, six protein components, designated coagulogen, proclotting enzyme, factor B, factor C, and factor G, all of which are closely associated with the endotoxin-mediated coagulation pathway, have been purified and biochemically characterized. The molecular structures of these proteins have also been elucidated. Moreover, the reconstitution experiments using the isolated clotting factors, factor C, factor B, proclotting enzyme and coagulogen in the presence of endotoxin, leads to the formation of coagulin gel. Here, I will focus on the biochemical principle of Limulus test for detecting bacterial endotoxins, and its activation and regulation mechanism on the LPS-mediated coagulation cascade.     

Calculation of on-state I–V characteristics of LDMOSFETs based on an accurate LDD resistance modeling

In this paper, we present an I–V model for LDMOSFETs. It is based on modeling the Lightly-Doped Drain (LDD) region of the device as voltage-controlled resistors where velocity saturation effect is also taken into account. Using the LDD region model along with a model for the channel region of the device, the on-state I–V characteristic of the transistor is accurately calculated. The models for the LDD region resistors can be incorporated into a circuit simulator such as HSPICE which has an accurate model for the channel region of the transistor. The accuracy of the models is verified by comparing its results with those of a device simulator. The results show a maximum error of 1% for a wide range of voltages and overlapped LDD region lengths.     

Removal of lead(II) by adsorption onto Viscumalbum L.: Effect of temperature and equilibrium isotherm analyses

The removal efficiency of Viscumalbum L. from lead containing aqueous solutions was investigated. The effect of adsorbent mass, pH of solution, initial Pb(II) concentration and temperature was investigated using a batch adsorption technique. The optimum pH for Pb(II) adsorption was found as 3.0 for Viscumalbum L. Results were analyzed by the Langmuir, Freundlich, Temkin and Harkins-Jura, equation using linearized correlation coefficient at different temperature. The characteristic parameters for each isotherm have been determined. The Langmuir model agrees very well with experimental data than the other models. According to Langmuir isoterm, the monolayer saturation capacity (Q_o) is 769.23 mg/g at 25 °C. Models and the isotherm constant were evaluated depending on temperature. Thermodynamic parameters such as ΔH^o, ΔS^o and ΔG^o were calculated. The adsorption process was found to be endothermic and spontaneous. The experimental data were analyzed using the first- and the second-order kinetic models. The rate constants of adsorption for both kinetics models have been calculated. The second-order model provides the best correlation of the data.     

In vivo imaging of cell morphology and cellular processes in Caenorhabditis elegans, using non-linear phenomena

We present the detailed imaging of structures and processes of the nematode Caenorhabditis elegans (C. elegans) using non-linear microscopy. Complementary information about the anatomy of the nematode was collected by implementing a combination of two photon excitation fluorescence (TPEF), second and third harmonic generation (SHG and THG) image contrast modes on the same microscope. Three-dimensional (3D) reconstructions of TPEF, SHG and THG images were also performed. Moreover, THG imaging technique has been tested as a potential, novel, non-destructive diagnostic tool for monitoring cellular processes in vivo, such as neuronal degeneration.     

Google matrix analysis of C.elegans neural network

We study the structural properties of the neural network of the C.elegans (worm) from a directed graph point of view. The Google matrix analysis is used to characterize the neuron connectivity structure and node classifications are discussed and compared with physiological properties of the cells. Our results are obtained by a proper definition of neural directed network and subsequent eigenvector analysis which recovers some results of previous studies. Our analysis highlights particular sets of important neurons constituting the core of the neural system. The applications of PageRank, CheiRank and ImpactRank to characterization of interdependency of neurons are discussed.     

Physiological characteristics of tropical rain forest tree species: a basis for the development of silvicultural technology

The physiological characteristics of the dominant tree species in the tropical rain forest mainly belonging to dipterocarps as well as the environmental conditions especially for the light in the forest were studied to establish the silvicultural system for the forest regeneration in the tropical South Asia. The flowering patterns of the dipterocarp trees are usually irregular and unpredictable, which make difficult to collect sufficient seeds for raising the seedlings. The field survey revealed the diverged features of the so-called gregarious or simultaneous flowering of various species of this group. Appropriate conditions and methods for the storage of the seeds were established according to the detailed analyses of the morphological and physiological characteristics of the seeds such as the low temperature tolerance and the moisture contents. The intensity and spectra of the light in the forest primarily determine the growth and the morphological development of the seedlings under the canopy. Based on the measurements of the diffused light at the sites in the tropical forest in the varying sunlight, the parameters such as "the steady state of the diffuse light" and "the turning point" were defined, which were useful to evaluate the light conditions in the forest. To improve the survival of the transplanted seedlings, a planting method of "the bare-root seedlings", the seedlings easy to be handled by removal of all leaves, soil and pots, was developed. Its marked efficiency was proved with various dipterocarps and other tropical trees by the field trial in the practical scale. Tolerance of the various species to the extreme environmental conditions such as fires, acid soils and drought were examined by the experiments and the field survey, which revealed marked adaptability of Shorea roxburghii as a potential species for regeneration of the tropical forests.     

8-Hydroxyguanine: From its discovery in 1983 to the present status

8-Hydroxyguanine (8-OH-G) was discovered in 1983 in our laboratory at the National Cancer Center Research Institute, Tokyo. Since it could be formed in DNA not only in vitro but also in vivo by oxygen radical forming agents, we immediately hypothesized the importance of this discovery in connection with its biological consequence. Further intensive efforts by us from 1983 to 1990 confirmed that 8-OH-G is a highly significant oxidated DNA lesion involved in mutation and/or carcinogenesis in mammals, including humans. With the subsequent entry of many investigators to this research field the number of publications on 8-OH-G increased exponentially, reaching more than several thousands by the end of 2005. In this article, a summary is given of the important works carried out in the early days, and further notable contributions by many investigators are reviewed, focusing on 8-OH-G in the mammalian system. A special emphasis is given to research on knockout mice that are deficient in genes involved in the repair systems of the 8-OH-G lesion. Lastly, our own recent work is summarized involving a one-year carcinogenesis study of Ogg1 (the gene for 8-OH-G specific glycosylase/AP lyase^*1) knockout mice that have been exposed to oxidative stress.     

High spin chemistry underlying organic molecular magnetism: Topological symmetry rule as the first principle of spin alignment in organic open-shell systems of π-conjugation and their ions

This review paper deals with an overview of molecule-based magnetism as a rapidly developing interdisciplinary field, topological symmetry rule as the first principle of spin alignment in organic open-shell systems in the ground state, the proposal of organic through-bond 1D and 2D ferro- and superparamagnets and the detection of the first organic high-spin molecule, m-phenylenebis(phenylmethylene) in the quintet ground state (S = 2), followed by extended organic high-spin systems with π-conjugation such as aromatic hydrocarbons having S = 3, 4, 5. The paper also describes a theoretical approach to the understanding of electronic spin structures of organic high-spin molecules by invoking both Heisenberg and Hubbard model Hamiltonians, weakly interacting intramolecular high-spin systems from both experimental and theoretical sides, the spin density distribution of the first organic high-spin molecule in terms of electron- nuclear multiple resonance spectroscopy and the detection and characterization of ionic high-spin hydrocarbons, emphasizing the establishment of high spin chemistry underlying organic molecular magnetism.     

Influence of fixed electric charges on potential profile across the squid axon membrane

The potential profile for a model of squid axon membrane has been determined for two physiological states: resting and action states. The non-linear Poisson-Boltzmann equation has been solved by considering the volumetric charge densities due to charges dissolved in an electrolytic solution and fixed on both glycocalyx and cytoplasmatic proteins. Results showing the features of the potential profile along the outer electrolytic region are similar for both resting and action states. However, the potential fall along glycocalyx at action state is lower than at resting. A small variation in the Na⁺ concentration drastically affects the surface membrane potentials and vice versa. We conclude that effects on the potential profile due to surface lipidic bilayer charge and contiguous electric double layers are more relevant than those provoked by fixed charges distributed along the cell cytoplasm.