The number of morphological synapses between neurons does not predict the strength of their physiological synaptic interactions: a study of dendrites in the nematode Ascaris suum

Previous electrophysiological and anatomical studies of Ascaris suum motor neurons demonstrated a strong correlation between functional interactions and the presence of anatomically defined synapses. However, one example of a physiologically robust synaptic connection was encountered for which no anatomical evidence of direct chemical synapses was found. This involved synaptic transmission from an identified excitatory motor neuron to its inhibitory partner. In this study, pressure injection of horseradish peroxidase or nickel lysine into inhibitory motor neurons revealed numerous spines projecting from the main neuronal process toward the neuromuscular surface that then branched and extended fine, longitudinal processes up to 130 microm in length. Subsequent examination of nickel lysine-injected spines by electron microscopy revealed numerous chemical synapses, including for the first time direct inputs from the excitatory neuron. However, the numbers of synapses from this motor neuron were very small relative to inputs from other identified cells. Thus, direct synapses are unlikely to explain the robust nature of this physiological interaction.     

Localization and differential expression of FMRFamide-like immunoreactivity in the nematode Ascaris suum

By immunocytochemical and immunohistochemical methods, FMRFamide-like immunoreactivity (FLI) was localized to many neurons and processes in the Ascaris nervous system, including the head, tail, and lateral lines. Some of these cells were identified; they included sensory neurons, interneurons, and motor neurons. FLI was also present in the pharyngeal neurons and in their varicosities near the surface of the pharynx. By HPLC analysis of extract, only a subset of the FMRFamide-like peptides (FLPs) expressed in Ascaris heads, and heads from which the pharynx had been removed, were expressed in the pharynx. Furthermore, FLPs appeared to be differentially expressed in female heads and tails and male heads and tails. Acetone and acid methanol differentially extracted subforms of FLI from Ascaris heads and from C. elegans. © 1993 Wiley-Liss, Inc.     

模式动物线虫的学习与学习抉择行为

线虫因为其简单的体系和对于诸如接触、味道、气味和温度等外界刺激的敏感反映能力,日益成为受人瞩目的研究行为可塑性的模式动物。目前认为,线虫中的学习包括非联想与联想学习两大类,且存在至少6种形式化学趋向性介导的联想学习。同时,三种研究体系已经被开发出来用于学习抉择的研究。本文讨论了线虫中学习与学习抉择的形式、研究模型及其遗传与分子调控机制。     

Sensory neuroanatomy of Parastrongyloides trichosuri, a nematode parasite of mammals: Amphidial neurons of the first-stage larva

Owing to its ability to switch between free-living and parasitic modes of development, Parastrongyloides trichosuri represents a valuable model with which to study the evolution of parasitism among the nematodes, especially aspects pertaining to morphogenesis of infective third-stage larvae. In the free-living nematode Caenorhabditis elegans, developmental fates of third-stage larvae are determined in part by environmental cues received by chemosensory neurons in the amphidial sensillae. As a basis for comparative study, we have described the neuroanatomy of the amphidial sensillae of P. trichosuri. By using computational methods, we incorporated serial electron micrographs into a three-dimensional reconstruction of the amphidial neurons of this parasite. Each amphid is innervated by 13 neurons, and the dendritic processes of 10 of these extend nearly to the amphidial pore. Dendritic processes of two specialized neurons leave the amphidial channel and terminate within invaginations of the sheath cell. One of these is similar to the finger cell of C. elegans, terminating in digitiform projections. The other projects a single cilium into the sheath cell. The dendritic process of a third specialized neuron terminates within the tight junction of the amphid. Each amphidial neuron was traced from the tip of its dendrite(s) to its cell body in the lateral ganglion. Positions of these cell bodies approximate those of morphologically similar amphidial neurons in Caenorhabditis elegans, so the standard nomenclature for amphidial neurons in C. elegans was adopted. A map of cell bodies within the lateral ganglion of P. trichosuri was prepared to facilitate functional study of these neurons.     

模式动物线虫的学习与学习抉择行为(英文)

线虫因为其简单的体系和对于诸如接触、味道、气味和温度等外界刺激的敏感反映能力,日益成为受人瞩目的研究行为可塑性的模式动物。目前认为,线虫中的学习包括非联想与联想学习两大类,且存在至少6种形式化学趋向性介导的联想学习。同时,三种研究体系已经被开发出来用于学习抉择的研究。本文讨论了线虫中学习与学习抉择的形式、研究模型及其遗传与分子调控机制。     

Neuronal localization of serotonin in Ascaris suum

We have used immunocytochemical techniques to investigate the distribution of serotonin-like immunoreactivity in the nematode Ascaris suum. Antisera raised against serotonin (5-hydroxytryptamine, 5-HT) conjugated to bovine serum albumin (BSA) labelled a pair of neurons in the pharynx of both sexes and five cells in the ventral cord of the male tail. The labelling was blocked by 5-HT or by 5-HT conjugated to BSA. The 5-HT-immunoreactive cells in the pharynx resemble neurosecretory cells and are probably homologous to the neurosecretory motor neurons (NSM) in Caenorhabditis elegans; the cells in the male tail appear to be motor neurons that are homologous to CP neurons in C. elegans. Other cells that stain with 5-HT antisera have been observed in C. elegans but are not seen in Ascaris. © 1996 Wiley-Liss, Inc.     

秀丽线虫记忆的分子调控机制

模式无脊椎动物秀丽线虫已经成为揭示记忆复杂行为的理想研究模型之一.线虫具有三种简单的记忆形式对温度感知的记忆、对化学物质感知的记忆以及对于机械刺激感知的记忆.在对机械刺激感知的记忆研究中,短时程、中时程与长时程记忆均得到了系统的分析.其中短时程与中时程记忆可能定位于感觉神经元的前突触,而长时程记忆可能定位于中间神经元的后突触.本文针对线虫中记忆的遗传与分子调控机制近些年的研究进展进行了总结与讨论.     

Evaluation of the long-term memory for thermosensation regulated by neuronal calcium sensor-1 in Caenorhabditis elegans

Objective To evaluate whether the thermotaxis tracking model is suitable for assessing long-term memory （LTM） in the nematode Caenorhabditis elegans. Methods Animals were trained at 20℃ overnight in presence of food. The percentage of animals performing isothermal tracking （IT） behavior was measured at different time intervals after the training. Results The percentage of animals performing IT behavior, the numbers of body bends inside and outside the training temperature, and the expression patterns of AFD and AIY neurons were similar to those in control animals at 36 and 48 h after training; whereas when extending to 60, 72, and 84 h, locomotory behavior defects were observed in the assayed animals, suggesting that this thermal tracking model is feasible for analyzing LTM at 36 and 48 h after training. Moreover, the percent-age of animals performing IT behavior was reduced at 18, 36, and 48 h after training in neuronal calcium sensor-1 gene （nsc-1） mutant animals compared with that in wild-type N2 animals. In addition, exposure to plumbum （Pb） significantly repressed the LTM at 18, 36, and 48 h after training in both wild-type N2 and ncs-1 mutant animals. Conclusion The thermotaxis tracking model is suitable for evaluating the LTM regulated by NCS-1, and can be employed for elucidating regulatory functions of specific genes or effects of stimuli on memory in C. elegans.     

对秀丽线虫模型NCS-1调控的温度感知长时程记忆的评价研究

目的 评价线虫温度趋向性模型是否适用于长时程记忆的研究.方法 在20℃含有新鲜食物过夜培养的训练条件下,考察在不同时间点执行温度趋向性(isothermal tracking,IT)行为的虫体百分比.结果 训练后36与48h,线虫的IT百分数、温度趋向性模型内外的头部摆动频率、以及AFD和AIY神经元的表达模式与对照组相似,而训练后60、72和84 h的线虫则表现出运动行为上的缺陷,表明温度趋向性模型适合于在训练36和48 h后的线虫中进行长时程记忆的考察.此外,与野生型N2相比,神经元钙传感蛋白(neuronal calcium sensor-1,NCS-1)的突变能够降低训练后18、36和48 h的虫体的IT百分数.而铅暴露可以显著抑制训练后18、36和48 h野生型和ncs-1突变体动物在不同时间点下的IT百分数.结论 温度趋向性模型适用于评价秀丽线虫中NCS-1参与调控的温度感知长时程记忆,且该模型可用于针对线虫中参与调节或影响记忆行为的某一特定基因或刺激信号功能的揭示.     

Sensory neuroanatomy of a skin-penetrating nematode parasite: Strongyloides stercoralis. I. Amphidial neurons

The Strongyloides stercoralis infective larva resumes feeding and development on receipt of signals, presumably chemical, from a host. Only two of the anterior sense organs of this larva are open to the external environment. These large, paired goblet-shaped sensilla, known as amphids, are presumably, therefore, the only chemoreceptors. Using three-dimensional reconstructions made from serial electron micrographs, amphidial structure was investigated. In each amphid, cilialike dendritic processes of 11 neurons extend nearly to the amphidial pore; a twelfth terminates at the base of the amphidial channel, behind an array of lateral projections on the other processes. A specialized dendritic process leaves the amphidial channel and forms a complex of lamellae that interdigitate with lamellae of the amphidial sheath cell. This “lamellar cell” is similar to one of the “wing cells” or possibly the “finger cell” of Caenorhabditis elegans. Each of the 13 amphidial neurons was traced to its cell body. Ten neurons, including the lamellar cell, connect to cell bodies in the lateral ganglion, posterior to the nerve ring. The positions of these cell bodies were similar to those of the amphidial cell bodies in C. elegans. Therefore, they were named by using C. elegans nomenclature. Three other amphidial processes connect to cell bodies anterior to the nerve ring; these have no homologs in C. elegans. A map allowing identification of the aimphidial cell bodies in the living worm was prepared. Consequently, laser ablation studies can be conducted to determine which neurons are involved in the infective process. © 1995 Wiley-Liss, Inc.     

Generation of monoclonal antibodies against a nematode peptide extract: another approach for identifying unknown neuropeptides

Monoclonal antibodies that cross-react with Ascaris neural antigens were generated in mice immunized with a conjugate made with keyhole limpet hemocyanin (KLH) linked to a crude peptide extract from Caenorhabditis elegans. The response to KLH was suppressed by injection of cyclophosphamide 3 days after immunization with a gamma-aminobutyric acid (GABA)-KLH conjugate. Screening of hybridomas was carried out by enzyme-linked immunosorbent assay and whole mount immunocytochemistry. Two similar clones produced antibodies that recognized a small subset of Ascaris neurons. This result suggests that the monoclonal antibody technique might be useful for identifying new neuropeptides since the antibodies can be used for localization of the neuropeptidelike substances and, potentially, for immunoaffinity chromatography. As a by-product of this experiment, monoclonal antibodies that recognize GABA-like immunoreactivity in whole mounts and plastic sections were also obtained.     

Sensory neuroanatomy of a skin-penetrating nematode parasite Strongyloides stercoralis. II. labial and cephalic neurons

Host recognition, contact, and skin-penetration by Strongyloides stercoralis infective larvae are crucially important behavioral functions mediating transition from free-living to parasitic life. The sensilla of the worm's anterior tip presumably play an important role in these processes. Besides the main chemosensilla, the amphids, which are of central importance, the larva has 16 putative mechanosensilla. There are six inner labial sensilla: two dorsal, two ventral, and two lateral. The two dorsal and ventral pairs are each innervated by two neurons, whereas each lateral sensillum is singly innervated. The six outer labial and four cephalic sensilla are all singly innervated. All of these have the characteristics of mechanoreceptors: they are closed to the external environment, and closely associated with the overlying cuticle. Distally, their dendritic processes contain granular material and associated microtubules. With two exceptions, the relevant neuronal cell bodies lie in lateral ganglia adjacent to the nerve ring, their positions remarkably similar to those of their homologues in the free-living nematode, Caenorhabditis elegans. Cell bodies of two neuronal pairs, one of two dorsal inner labial neurons and one of two ventral inner labial neurons per side, are however, found far anterior to the remaining cell bodies. All labial and cephalic sensilla are apparently mechanoreceptors, complementing the well-developed chemosensilla. Presumably infective larvae require touch and stretch receptors, not only to initiate skin penetration by finding irregularities as points of access, but also to bore through tissue to reach their ultimate enteral destination. J. Comp. Neurol. 389:212–223, 1997. © 1997 Wiley-Liss, Inc.     

The connectome of the Caenorhabditis elegans pharynx

Detailed anatomical maps of individual organs and entire animals have served as invaluable entry points for ensuing dissection of their evolution, development, and function. The pharynx of the nematode Caenorhabditis elegans is a simple neuromuscular organ with a self-contained, autonomously acting nervous system, composed of 20 neurons that fall into 14 anatomically distinct types. Using serial electron micrograph (EM) reconstruction, we re-evaluate here the connectome of the pharyngeal nervous system, providing a novel and more detailed view of its structure and predicted function. Contrasting the previous classification of pharyngeal neurons into distinct inter- and motor neuron classes, we provide evidence that most pharyngeal neurons are also likely sensory neurons and most, if not all, pharyngeal neurons also classify as motor neurons. Together with the extensive cross-connectivity among pharyngeal neurons, which is more widespread than previously realized, the sensory-motor characteristics of most neurons define a shallow network architecture of the pharyngeal connectome. Network analysis reveals that the patterns of neuronal connections are organized into putative computational modules that reflect the known functional domains of the pharynx. Compared with the somatic nervous system, pharyngeal neurons both physically associate with a larger fraction of their neighbors and create synapses with a greater proportion of their neighbors. We speculate that the overall architecture of the pharyngeal nervous system may be reminiscent of the architecture of ancestral, primitive nervous systems.     

Heterogeneity of cholecystokinin/gastrin-like immunoreactivity in the nervous system of the nematode Ascaris suum

A wholemount immunocytochemical method was used for the localization of cholecystokinin (CCK8)-like and gastrin-like immunoreactivity in Ascaris. The patterns of specific neuronal staining given by two antisera and four monoclonal antibodies made against CCK8, and one antiserum made against gastrin were investigated. Preabsorption of these antibodies with CCK8 or gastrin 17 resulted in complete loss of immunoreactivity in almost all of the neurons (two antisera also contained nonspecific antibodies), suggesting that all of the antibodies recognize epitopes, in Ascaris neurons, that include some or all of the C-terminal five amino acids that are identical in CCK8 and gastrin 17. However, the seven different antibodies showed immunoreactivity in different subpopulations of neurons, implying that there are at least seven different species of CCK-like molecules in Ascaris. Fractionation of Ascaris peptide extracts by high performance liquid chromatography (HPLC), monitoring fractions with a CCK8 radioimmunoassay (RIA), also shows heterogeneity of molecules immunologically related to CCK8. © 1996 Wiley-Liss, Inc.     

Dystrobrevin- and dystrophin-like mutants display similar phenotypes in the nematode Caenorhabditis elegans

Dystrophin, the protein disrupted in Duchenne muscular dystrophy, forms a transmembrane complex with dystrophin-associated proteins. Dystrobrevins, proteins showing homology to the C-terminal end of dystrophin, and whose function is unknown, are part of the dystrophin complex. We report here that, in the nematode Caenorhabditis elegans, animals carrying mutations in either the dystrophin-like gene dys-1 or the dystrobrevin-like gene dyb-1 display similar behavioral and pharmacological phenotypes consistent with an alteration of cholinergic signalling. These findings suggest that: (1) dystrobrevin and dystrophin are functionally related and (2) their disruption impairs cholinergic signalling. Received: November 25, 1998 / Accepted: January 7, 1999 / Published online: March 18, 1999     

Transplantation of autologous dorsal root ganglia into the peroneal nerve of adult rats: uni- and bidirectional axonal regrowth from the grafted DRG neurons

Previous studies have demonstrated that transplanted dorsal root ganglion neurons (DRGNs) can survive and differentiate in a variety of orthotopic and heterotopic locations. In order to develop strategies aimed at restoring the sensory function following traumatic injury to the spinal cord and to its peripheral sensory connections, we have transplanted adult autologous dorsal root ganglia (DRGs) into the peroneal nerve of adult rats. Twelve female Sprague-Dawley rats were used. A segment of the peroneal nerve was isolated by double transection and ligature to prevent undesirable reinnervation. The left fifth cervical (C5) DRG was removed from its normal location and inserted into the midportion of the isolated nerve segment. One month after the grafting procedure, a morphological study included axonal retrograde labeling with True Blue (TB) and/or Diamidino Yellow (DY) applied on each cut end of the nerve segment, cell counting, and cell measurement after staining with cresyl violet. Compared to the C5 ganglion maintained in situ, the mean number of surviving DRGNs in the transplant was 1381, corresponding to a survival rate of 20%. Both singly (TB or DY) and doubly (TB + DY) stained DRGNs were encountered. The proportion of surviving neurons that appeared to be doubly labeled was 23%. These neurons were considered as having grown two opposite axonal projections, one into the "central" part of the nerve segment and a second one into its "peripheral" part. The present results give new insights and interesting prospects concerning the possibilities of reconstructing the sensory circuitry after central and/or peripheral injuries.     

Subpopulations of rat dorsal root ganglion neurons express active vesicular acetylcholine transporter

The vesicular acetylcholine transporter (VAChT) is a transmembrane protein required, in cholinergic neurons, for selective storage of acetylcholine into synaptic vesicles. Although dorsal root ganglion (DRG) neurons utilize neuropeptides and amino acids for neurotransmission, we have previously demonstrated the presence of a cholinergic system. To investigate whether, in sensory neurons, the vesicular accumulation of acetylcholine relies on the same mechanisms active in classical cholinergic neurons, we investigated VAChT presence, subcellular distribution, and activity. RT-PCR and Western blot analysis demonstrated the presence of VAChT mRNA and protein product in DRG neurons and in the striatum and cortex, used as positive controls. Moreover, in situ hybridization and immunocytochemistry showed VAChT staining located mainly in the medium/large-sized subpopulation of the sensory neurons. A few small neurons were also faintly labeled by immunocytochemistry. In the electron microscope, immunolabeling was associated with vesicle-like elements distributed in the neuronal cytoplasm and in both myelinated and unmyelinated intraganglionic nerve fibers. Finally, [(3)H]acetylcholine active transport, evaluated either in the presence or in the absence of ATP, also demonstrated that, as previously reported, the uptake of acetylcholine by VAChT is ATP dependent. This study suggests that DRG neurons not only are able to synthesize and degrade ACh and to convey cholinergic stimuli but also are capable of accumulating and, possibly, releasing acetylcholine by the same mechanism used by the better known cholinergic neurons.     

质膜相关的SNAREs syntaxin/UNC-64与SNAP-25/RIC-4蛋白调控秀丽线虫的脂肪积累(英文)

目的研究神经突触组装与功能调控相关基因是否参与秀丽线虫的脂肪积累调节。方法分析神经突触组装与功能调控相关基因突变体的脂肪积累变化,进而观察SNAREs syntaxin/unc-64和SNAP-25/ric-4基因与daf-2、daf-7、nhr-49、sbp-1及mdt-15所介导的信号通路在调控脂肪积累上的遗传关系。对unc-64与ric-4基因进行组织特异性活性分析,以确定它们在神经系统与肠道内对脂肪积累的影响。结果突触前为神经突触组装所必需的基因的突变并未明显影响脂肪积累。对调控神经突触功能的基因进行分析的结果显示,SNAREssyntaxin/unc-64与SNAP-25/ric-4基因均参与了脂肪积累的调节。利用苏丹黑染色与尼罗红标记法观察到unc-64与ric-4突变体肠道中脂肪积累显著增加,而unc-64与ric-4突变体中积累的脂肪颗粒并未出现尺寸的显著变化。在神经系统与肠道中,unc-64基因的表达均能显著降低unc-64突变体动物的脂肪积累,而基因ric-4在神经系统的表达则可以完全恢复ric-4突变体动物的脂肪积累。遗传分析表明,unc-64与ric-4对脂肪积累的调控独立于daf-2...     

GABA-immunoreactive neurons in the nematode Ascaris

gamma-Aminobutyric acid (GABA) immunoreactive neurons in the cephalic, somatic, and caudal regions of the Ascaris nervous system were visualized with serial section and whole-mount GABA immunocytochemistry. In the ventral and dorsal nerve cords, GABA-like immunoreactivity (GLIR) is localized to the neurites and cell bodies of identified inhibitory motor neurons and to two fibers, one in each cord, that arise from neurons in the nerve ring. GLIR is absent from identified excitatory motor neurons and from ventral cord interneurons. In neurons containing GLIR, immunoreactivity was present throughout the cell, which argues against an exclusive localization of GABA at conventional synapses. In whole mounts, ten GABA-immunoreactive neurons were present in the cephalic region. These include four nerve ring-associated cells (the RME-like cells), two bilaterally symmetrical pairs of lateral ganglia neurons (the amphid-GABA and deirid-GABA cells) and one bilaterally symmetrical pair of ventral ganglion cells (the VG-GABA cells). In sections, the RME-like cells and the VG-GABA cells were consistently stained through the cephalic region. However, anti-GABA staining of the lateral ganglia cells in sections was light, thus suggesting that they contain less GLIR than the other more intensely stained GABA-immunoreactive neurons. In the caudal region, a single GABA-immunoreactive neuron was present in the dorsal rectal ganglion. Our data suggest that these ten cephalic neurons, and a single dorsal rectal ganglion neuron, use GABA as a neurotransmitter.     

A quantitative electron microscope study of the perikaryal projections of sensory ganglion neurons. II. Gecko and lizard

The perikaryal projections of the neurons in the thoracic spinal ganglia of gecko and lizard usually appear as finger-shaped evaginations running roughly parallel to the surface of the nerve cell body; they show a nearly circular cross section with a rather uniform transverse diameter having an average value of about 0.2 micron. In both gecko and lizard a very high correlation was found between the surface area of perikaryal projections and both the volume and smoothed surface area of the corresponding nerve cell body. The results of the present research agree with those obtained in a previous study on two mammal species (cat and rabbit) and lend further support to the hypothesis advanced in that study; i.e., that perikaryal projections in sensory ganglion neurons are normal formations which maintain the surface-to-volume ratio above the critical level for metabolic exchanges. Perikaryal projections increase the surface area of the nerve cell body by 32.5% in gecko and 30% in lizard, while they increase it by 43% in cat and 39.5% in rabbit. This difference may be related to the lower metabolic rate of the neurons in poikilotherms than in mammals.