Comprehensive morphological identification and GABA immunocytochemistry of antennal lobe local interneurons in Bombyx mori

The insect antennal lobe (AL) is the structural and functional analog of the olfactory bulb of mammals, in which odor information is spatially and/or temporally represented by functional glomerular units. Local interneurons (LNs) play critical roles through intra- and interglomerular communication to shape the output from the AL to higher brain centers; however, the function and even the components of LNs are unclear. We have used morphological and immunocytochemical approaches to examine LNs in the silkworm moth, Bombyx mori. First, we comprehensively analyzed the morphological variation of LNs. One hundred fifty-three AL LNs were intracellularly stained, analyzed in three dimensions with a confocal microscope, and subdivided into five morphological types based on differences in the arborization region in the AL and dendritic profiles within the glomeruli. Two global multiglomerular types arborized in the macroglomerular complex (MGC) and in most ordinary glomeruli, and the other three oligoglomerular types innervated some ordinary glomeruli with or without the MGC. Second, we performed double-labeling of Lucifer Yellow staining of a single LN combined with gamma-aminobutyric acid (GABA) immunocytochemistry. The two global multiglomerular types and two of the oligoglomerular types were GABA-immunoreactive, but the third oligoglomerular type, which innervates the MGC and some ordinary glomeruli, included some GABA-immunonegative neurons, suggesting the existence of a non-GABAergic subtype. These results suggest that the complex neural circuits of the AL are composed of several morphologically different types of LNs, most of which are inhibitory.     

Localized changes in olfactory bulb morphology associated with early olfactory learning

Young rats exposed to an odor while receiving reinforcing stimulation come to approach that odor upon subsequent presentation. In addition, such pups have increased 14C-2-deoxyglucose (2DG) uptake within focal areas of the glomerular layer in response to that odor, compared to control animals experiencing the odor for the first time. In this study, the morphology of the glomerular areas underlying these 2DG foci was examined to determine whether early olfactory learning imposed local structural changes that could produce the enhanced 2DG uptake. Alternate sections either were processed with a silver and a Nissl stain to label both cell bodies and their processes or were histochemically treated for the mitochondrial enzymes cytochrome oxidase (CO) or succinic dehydrogenase (SDH) to define the glomerular core of the bulb; 2DG autoradiographs were aligned with adjacent stained sections, and regions underlying the high 2DG uptake foci were examined. In odor-familiar animals, large glomerular clusters that protruded into the external plexiform layer or the olfactory nerve layer were associated with the focal areas of increased 2DG uptake. Morphometric analysis of these regions revealed that the glomerular layer underlying the foci of high 2DG uptake was 30% wider in odor-familiar animals than comparable areas in odor-unfamiliar animals; the cross-sectional areas of individual glomeruli were 21% larger in odor-familiar animals. The foci of enhanced 2DG uptake therefore appear to be associated with groups of enlarged glomeruli. These data demonstrate that early olfactory learning influences the morphology of the olfactory bulb.     

用磷钨酸染色对实验性克汀病大鼠小脑小球突触数的电镜分析

用Wistar大鼠制成的实验性克汀病模型,出生后5—6个月处死,进行电镜观察;发现实验组小脑皮质颗粒层细胞密度比对照组稍高,细胞间隙即小脑小球占的比例小,E—PTA染色阳性突触数少。这些变化表明,克汀病时小脑小球发育迟缓,突触形成不全。     

细胞因子对体外培养大鼠肾小球及肾小球系膜细胞分泌TNF的影响

采用ＥＬＩＳＡ法，观察了正常大鼠肾小球及ＧＭＣ在体外培养期间分泌ＴＮＦ的变化规律及不同细胞因子对其分泌ＴＮＦ的影响。结果表明：体外培养的肾小球与ＧＭＣ具有分泌ＴＮＦ的功能，而且不同的时相ＴＮＦ分泌的量不等。ＩＬ－１对其分泌ＴＮＦ无影响。ＩＬ－６抑制肾小球分泌ＴＮＦ，但促进ＧＭＣ分泌ＴＮＦ。ＥＴ－１能明显促进ＴＮＦ的分泌，而且促进的幅度与其浓度成正比。     

Sorting and convergence of primary olfactory axons are independent of the olfactory bulb

Primary olfactory axons expressing the same odorant receptor gene sort out and converge to fixed sites in the olfactory bulb. We examined the guidance of axons expressing the P2 odorant receptor when they were challenged with different cellular environments in vivo. In the mutant extratoes mouse, the olfactory bulb is lacking and is replaced by a fibrocellular mass. In these animals, primary olfactory axons form glomerular-like loci despite the absence of normal postsynaptic targets. P2 axons are able to sort out from other axons in this fibrocellular mass and converge to form loci of like axons. The sites of these loci along mediolateral and ventrodorsal axes were highly variable. Similar convergence was observed for larger subpopulations of axons expressing the same cell surface carbohydrates. The sorting out and convergence of like axons also occurred during regeneration following bulbectomy. Olfactory axon behaviour in these models demonstrates that sorting and convergence of axons are independent of the target, which instead provides distinct topographic cues for guidance.     

Stereological estimation of volume-weighted mean glomerular volume from arbitrary sections of the equine kidney

Mean glomerular volume has previously been estimated, using stereological techniques, specifically the point-sampled intercept (PSI), either from isotropic or from vertical sections. As glomeruli are approximately spherical structures, the same stereological technique was carried out on vertical and arbitrary sections to determine whether section orientation had any effect on mean glomerular volume estimation. Equine kidneys from 10 individuals were analysed using the PSI method of estimating volume-weighted mean glomerular volume (MGV); for each kidney, arbitrary and vertical sections were analysed. MGVs were not significantly different between arbitrary and vertical sections ( P = 0.691) when analysing the data with the paired t test; when plotting MGV estimates from arbitrary sections against those from vertical sections the intercept was found not to be significantly different from zero ( P > 0.8) and the slope of the regression line not to be significantly different from 1.0 ( P > 0.4). For the estimation of MGV in equine kidneys using PSI, arbitrary sections may be used if it is not possible to use isotropic or vertical sections, but some caution must be exercised in the interpretation of results so gained.     

Digital in vivo 3D atlas of the antennal lobe of Drosophila melanogaster

As a model for primary olfactory perception, the antennal lobe (AL) of Drosophila melanogaster is among the most thoroughly investigated and well‐understood neuronal structures. Most studies investigating the functional properties and neuronal wiring of the AL are conducted in vivo, although so far the AL morphology has been mainly analyzed in vitro. Identifying the morphological subunits of the AL—the olfactory glomeruli—is usually done using in vitro AL atlases. However, the dissection and fixation procedure causes not only strong volumetric but also geometrical modifications; the result is unpredictable dislocation and a distortion of the AL glomeruli between the in vitro and in vivo brains. Hence, to characterize these artifacts, which are caused by in vitro processing, and to reliably identify glomeruli for in vivo applications, we generated a transgenic fly that expresses the red fluorescent protein DsRed directly fused to the presynaptic protein n‐synaptobrevin, under the control of the pan‐neuronal promotor elav to label the neuropil in the live animal. Using this fly line, we generated a digital 3D atlas of the live Drosophila AL; this atlas, the first of its kind, provides an excellent geometric match for in vivo studies. We verified the identity of 63% of AL glomeruli by mapping the projections of 34 GAL4‐lines of individual chemosensory receptor genes. Moreover, we characterized the innervation patterns of the two most frequently used GAL4‐lines in olfactory research: Orco‐ and GH146‐GAL4. The new in vivo AL atlas will be accessible online to the neuroscience community. J. Comp. Neurol. 523:530–544, 2015. © 2014 Wiley Periodicals, Inc.