植食性昆虫的嗅觉选食过程及其机制研究进展

植食性昆虫的嗅觉在其选择食物的过程中发挥了重要的作用,它能通过对植物挥发物的感受来定向和定位食物源并产生趋近行为,进而根据特殊的化合物或者多种化合物的特异浓度组合来区分寄主和非寄主植物.在这个过程中,昆虫嗅觉器官上相关的嗅觉感受蛋白被植物挥发物激活,形成特异的嗅觉感受通路,在行为上调控昆虫嗅觉选食的能力.本文主要从植食性昆虫嗅觉选食过程中植物挥发物的散布特征、昆虫识别植物信息的嗅觉感受机制及其相关的分子基础等方面进行叙述,同时讨论了近年的研究成果并展望了下一步的研究方向.     

Clonal analysis of adult human olfactory neurosphere forming cells

Olfactory neuroepithelium (ONe) is unique because it contains progenitor cells capable of mitotic division that replace damaged or lost neurons throughout life. We isolated populations of ONe progenitors from adult cadavers and patients undergoing nasal sinus surgery that were heterogeneous and consisted of neuronal and glial progenitors. Progenitor lines have been obtained from these cultures that continue to divide and form nestin positive neurospheres. In the present study, we used clonal and population analyses to probe the self-renewal and multipotency of the neurosphere forming cells (NSFCs). NSFCs plated at the single cell level produced additional neurospheres; dissociation of these spheres resulted in mitotically active cells that continued to divide and produce spheres as long as they were subcultured. The mitotic activity of clonal NSFCs was assessed using bromodeoxyuridine (BrdU) incorporation. Lineage restriction of the clonal cultures was determined using a variety of antibodies that were characteristic of different levels of neuronal commitment: β-tubulin isotype III, neural cell adhesion molecule (NCAM) and microtubule associated protein (MAP2), or glial restriction: astrocytes, glial fibrillary acidic protein (GFAP); and oligodendrocytes, galactocerebroside (GalC). Furthermore, nestin expression, a marker indicative of progenitor nature, decreased in defined medium compared to serum-containing medium. Therefore, adult human ONe-derived neural progenitors retain their capacity for self-renewal, can be clonally expanded, and offer multipotent lineage restriction. Therefore, they are a unique source of progenitors for future cell replacement strategies in the treatment of neurotrauma and neurodegenerative diseases.     

The impact of a brominated flame retardant on smoltification and olfactory function in Atlantic salmon (Salmo salar L.) smolts

Despite a reduction in exploitation of salmon stocks throughout the NE Atlantic, there continues to be a decline in many populations. The factors regulating these populations remain poorly understood, although there is evidence that environmental conditions experienced in freshwater can effect survival in the marine environment. The aim of this study was to investigate the impact of a brominated flame retardant (hexabromocyclododecane, HBCD) on the parr-smolt transformation in juvenile salmon. It is during this pre-adaptive period to marine life that olfactory imprinting to the natal river is considered to occur. Fish were exposed to low levels of HBCD for 30 days over the peak smoltification period in freshwater, and then transferred to clean seawater for 20 days. Fish were sampled weekly to assess changes in some of the physiological parameters associated with smoltification, and olfactory response to conspecific smolt urine was measured using an electro-olfactogram (EOG). Exposure to HBCD did not affect seawater adaptability, although there was some disruption of plasma thyroid hormone levels, as well as a reduction in olfactory function to conspecific smolt urine. The results are discussed in relation to the marine survival and successful homing of adult salmon.     

The sox gene Dichaete is expressed in local interneurons and functions in development of the Drosophila adult olfactory circuit

In insects, the primary sites of integration for olfactory sensory input are the glomeruli in the antennal lobes. Here, axons of olfactory receptor neurons synapse with dendrites of the projection neurons that relay olfactory input to higher brain centers, such as the mushroom bodies and lateral horn. Interactions between olfactory receptor neurons and projection neurons are modulated by excitatory and inhibitory input from a group of local interneurons. While significant insight has been gleaned into the differentiation of olfactory receptor and projection neurons, much less is known about the development and function of the local interneurons. We have found that Dichaete, a conserved Sox HMG box gene, is strongly expressed in a cluster of LAAL cells located adjacent to each antennal lobe in the adult brain. Within these clusters, Dichaete protein expression is detected in both cholinergic and GABAergic local interneurons. In contrast, Dichaete expression is not detected in mature or developing projection neurons, or developing olfactory receptor neurons. Analysis of novel viable Dichaete mutant alleles revealed misrouting of specific projection neuron dendrites and axons, and alterations in glomeruli organization. These results suggest noncell autonomous functions of Dichaete in projection neuron differentiation as well as a potential role for Dichaete‐expressing local interneurons in development of the adult olfactory circuitry. © 2012 Wiley Periodicals, Inc. Develop Neurobiol, 2013     

Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits

Detection and interpretation of olfactory cues are critical for the survival of many organisms. Remarkably, species across phyla have strikingly similar olfactory systems suggesting that the biological approach to chemical sensing has been optimized over evolutionary time¹. In the insect olfactory system, odorants are transduced by olfactory receptor neurons (ORN) in the antenna, which convert chemical stimuli into trains of action potentials. Sensory input from the ORNs is then relayed to the antennal lobe (AL; a structure analogous to the vertebrate olfactory bulb). In the AL, neural representations for odors take the form of spatiotemporal firing patterns distributed across ensembles of principal neurons (PNs; also referred to as projection neurons)^2,3. The AL output is subsequently processed by Kenyon cells (KCs) in the downstream mushroom body (MB), a structure associated with olfactory memory and learning^4,5. Here, we present electrophysiological recording techniques to monitor odor-evoked neural responses in these olfactory circuits.First, we present a single sensillum recording method to study odor-evoked responses at the level of populations of ORNs^6,7. We discuss the use of saline filled sharpened glass pipettes as electrodes to extracellularly monitor ORN responses. Next, we present a method to extracellularly monitor PN responses using a commercial 16-channel electrode³. A similar approach using a custom-made 8-channel twisted wire tetrode is demonstrated for Kenyon cell recordings⁸. We provide details of our experimental setup and present representative recording traces for each of these techniques.     

二点委夜蛾非典型嗅觉受体AlepOrco的基因克隆、原核表达及多克隆抗体制备

【目的】非典型嗅觉受体(olfactory receptor co-receptor, Orco)与典型嗅觉受体共同形成离子通道,在昆虫嗅觉识别中具有至关重要的作用。本研究旨在克隆和表达二点委夜蛾Athetis lepigone Orco基因,明确其分子特性,为进一步研究该基因在二点委夜蛾中的功能奠定基础。【方法】将二点委夜蛾雌雄成虫触角转录组数据建立本地数据库,通过生物信息学分析获得二点委夜蛾Orco同源基因AlepOrco;利用RT-PCR方法克隆二点委夜蛾AlepOrco基因全长,并在pGEX-6P-1/BL21(DE3)系统中进行了该基因开放阅读框(ORF)的原核表达,制备多克隆抗体,用Western blot检测抗体特异性;利用qPCR技术检测该基因在二点委夜蛾雌雄成虫不同组织(喙、触角、去除触角和喙的头、胸、腹、足和翅)中的表达谱。【结果】获得了二点委夜蛾AlepOrco的cDNA(GenBank登录号:MN583125)全长序列,开放阅读框长1 422 bp,编码473个氨基酸,序列中有7个跨膜结构区,预测等电点为8.59,分子量为53.40 kD。SDS-PAGE和Western blot分析结果表明AlepOrco能够在大肠杆菌Escherichia coli中高效表达。利用制备的多克隆抗体对二点委夜蛾AlepOrco进行Western blot检测,其能够特异识别成虫触角中AlepOrco蛋白。qPCR结果表明,AlepOrco在二点委夜蛾雌雄成虫不同组织间具有相似的表达模式,都是在触角中的相对表达量最大,在翅中的表达量最小。【结论】克隆并原核表达了二点委夜蛾非典型嗅觉受体基因AlepOrco,制备的多克隆抗体能够特异识别二点委夜蛾成虫触角中的AlepOrco。结果为深入了解二点委夜蛾AlepOrco基因的结构和功能奠定了基础。     

Time-restricted foraging under natural light/dark condition shifts the molecular clock in the honey bee,Apis mellifera

ABSTRACT

Honey bees have a remarkable sense of time and individual honey bee foragers are capable of adjusting their foraging activity with respect to the time of food availability. Although, there is compelling experimental evidence that foraging behavior is guided by the circadian clock, nothing is known about the underlying molecular mechanisms. Here we present for the first time a study that explores whether time-restricted foraging under natural light-dark (LD) condition affects the molecular clock in honey bees. Food was presented in an enclosed flight chamber (12 m × 4 m × 4 m) either for 2 hours in the morning or 2 hours in the afternoon for several consecutive days and daily cycling of the two major clock genes, cryptochrome2 (cry2) and period (per), were analyzed for three different parts of the nervous system involved in feeding-related behaviors: brain, subesophageal ganglion (SEG), and the antennae with olfactory sensory neurons. We found that morning and afternoon trained foragers showed significant phase differences in the cycling of both clock genes in all three tissues. In addition, the phase differences were more pronounced when the feeder was scented with the common plant odor, linalool. Together our findings suggest that foraging time may function as a Zeitgeber that might have the capability to modulate the light entrained molecular clock.     

In vivo NMR studies of regional cerebral energetics in MPTP model of Parkinson's disease: recovery of cerebral metabolism with acute levodopa treatment

In this study, we have evaluated cerebral atrophy, neurometabolite homeostasis, and neural energetics in 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridin (MPTP) model of Parkinson's disease. In addition, the efficacy of acute l ‐DOPA treatment for the reversal of altered metabolic functions was also evaluated. Cerebral atrophy and neurochemical profile were monitored in vivo using MRI and ¹H MR Spectroscopy. Cerebral energetics was studied by ¹H‐[¹³C]‐NMR spectroscopy in conjunction with infusion of ¹³C labeled [1,6^‐13C₂]glucose or [2‐¹³C]acetate. MPTP treatment led to reduction in paw grip strength and increased level of GABA and myo‐inositol in striatum and olfactory bulb. ¹³C Labeling of glutamate‐C4 (1.93 ± 0.24 vs. 1.48 ± 0.06 μmol/g), GABA‐C2 (0.24 ± 0.04 vs. 0.18 ± 0.02 μmol/g) and glutamaine‐C4 (0.26 ± 0.04 vs. 0.20 ± 0.04 μmol/g) from [1,6‐¹³C₂]glucose was found to be decreased with MPTP exposure in striatum as well as in other brain regions. However, glutamine‐C4 labeling from [2‐¹³C]acetate was found to be increased in the striatum of the MPTP‐treated mice. Acute l ‐DOPA treatment failed to normalize the increased ventricular size and level of metabolites but recovered the paw grip strength and ¹³C labeling of amino acids from [1,6‐¹³C₂]glucose and [2‐¹³C]acetate in MPTP‐treated mice. These data indicate that brain energy metabolism is impaired in Parkinson's disease and acute l ‐DOPA therapy could temporarily recover the cerebral metabolism.

     

Phenylacetaldehyde attracts male and female apple clearwing moths,Synanthedon myopaeformis,to inflorescences of showy milkweed,Asclepias speciosa

Synanthedon myopaeformis Borkhausen (Lepidoptera: Sesiidae) is a diurnal clearwing moth native to Eurasia that was recently introduced into British Columbia (BC) and Ontario, Canada, where it has become a serious pest in apple orchards. In BC, these moths commonly feed on nectar of inflorescences, particularly that of showy milkweed, Asclepias speciosa Torrey (Apocynaceae). We investigated the relative importance of visual and olfactory cues, and the key floral semiochemical(s) mediating attraction of S. myopaeformis to A. speciosa. In field experiments, inflorescences left exposed or enclosed in cheesecloth bags dyed green induced similar visitation rates by moths, indicating that olfactory cues are attractive. Among the >10 floral odourants that elicited responses from moth antennae in coupled gas chromatographic‐electroantennographic detection analyses, phenylacetaldehyde induced the most frequent proboscis extension reflexes of male and female moths. Among eight floral odourants that were field‐tested singly, phenylacetaldehyde attracted 35 times more male and female moths than any other candidate semiochemical. Attractiveness of phenylacetaldehyde could not be enhanced by admixture with other floral odourants at the ratios or concentrations tested indicating that it alone may mediate attraction of S. myopaeformis to the inflorescences of A. speciosa. The potential use of phenylacetaldehyde as bait to monitor or mass‐trap populations of male and female S. myopaeformis should be investigated.