Three-dimensional structure of the gustatory cell in the mouse fungiform taste buds: a computer-assisted reconstruction from serial ultrathin sections

Taste buds in fungiform papillae of the mouse were examined with transmission electron microscopy and computer-assisted, three-dimensional reconstructions from serial ultrathin sections. In accord with observation by Murray (1971), four distinct cell types, type I, II, III and basal cells, were identified. Of these, only the type III cell made synaptic contacts with nerve terminals and contained both small, clear vesicles and dense-cored granules. The former vesicles were synaptic-type and accumulated in the cytoplasm just below the synaptic in membrane thickenings. This finding clearly indicates a sensory function for the type III cell. One to three type III cells were identified within a taste bud. The type III cell had at most eight synapses with nerve terminals. One nerve fiber making two synapses with the type III cell was occasionally observed in its terminal region.     

Coincidence detection and changes of synaptic efficacy in spiny stellate neurons in rat barrel cortex

Paired whole-cell voltage recordings were made from synaptically connected spiny stellate neurons in layer 4 of the barrel field in young (P14) rat somatosensory cortex. When postsynaptic action potentials (APs) followed each of 5 presynaptic APs in a 10- or 20-Hz train by less than 25 ms, subsequent unitary EPSP amplitudes were persistently reduced. Induction of long-term depression (LTD) depended on activation of group II metabotropic glutamate receptors, but not on NMDA or AMPA receptors. Reducing postsynaptic increases in intracellular calcium ([Ca2+]i) by intracellular loading with a fast- (BAPTA) or a slow- (EGTA) acting Ca2+ buffer blocked synaptic depression. Analysis of EPSP failures suggested mediation of LTD by a reduction in release probability. We propose a mechanism by which coincident activity results in long-lasting reduction of synaptic efficacy between synaptically connected neurons.     

数字化可视人体连续断层标本的参数测量

目的：探讨连续薄层断层标本的计算机测量方法。方法：利用冰冻连续铣切方法铣切一已知形态参数的工件,根据数学计算方法在图像处理软件中测量已知形态参数的模件,计算其误差值,并讨论图像处理软件中的计算方法。结果：断层图像上能清晰显示被测结构的断面图像,用计算机计算的测量值与实际值相差不显著。结论：利用连续断层可快速、精确测量人体各结构的空间形态参数,连续断层标本的空间参数测量精度和误差取决于被测结构在图像上的有效分辨率和连续断层的间距。     

大鼠肾标本连续切片的三维重建及形态学测量

目的 综合运用目前较成熟的技术,建立一个易施行的、可对大组织标本连续切片整体图像进行三维重建及形态学测量的方法。 方法 对正常大鼠肾的石蜡包埋标本进行全序列连续切片,HE染色后对切片图像进行数字化采集,经过拼接、配准步骤,建立切片整体图像的数字化数据集,对其内部的血管系统及肾盂进行分割及三维重建,并对重建结果进行形态学测量。 结果 切片的整体图像清晰,数字化图像数据集配准准确,三维重建结果再现了各种结构的立体特征及其在原组织块内的位置,在三维空间内,可进行无阻挡的自由观察,并且测量出了脉管系统的多项三维形态学指标,包括组织体积构成百分比、动脉长度及分支角度等。 结论 采用这种方法取得了较为满意的重建结果,可为其他组织连续切片的三维重建研究提供借鉴。     

Ultrastructure of Golgi-impregnated and gold-toned spiny and aspiny neurons in the monkey neostriatum

Summary Golgi-impregnated, gold-toned spiny and aspiny neurons in the monkey neostriatum were deimpregnated and examined at the electron microscope level.Spiny type I neurons have relatively large nuclei with few indentations and aggregates of chromatin under the nuclear membrane which in some regions give the appearance of a dark rim. The small quantity of surrounding cytoplasm is poor in organelles.Aspiny type I neurons have eccentric, highly indented nuclei. The relatively large proportion of cytoplasm is rich in organelles especially Golgi apparatus and rough endoplasmic reticulum which often appears in stacks.Synapses with symmetric membrane densities are common on the somata of spiny type I neurons. Those on the proximal and distal dendritic shafts are few in number and asymmetric, and those on spines more frequent and primarily asymmetric. Aspiny type I neurons have few synapses on their cell bodies. Proximal and distal dendrites, however, are contacted by numerous profiles which contain small round vesicles and make both symmetric and asymmetric synapses. The same axon terminals also synapse with dendritic spines of spiny neurons, indicating that an input, most likely of afferent origin, is shared by both cell types. Other less frequently occurring profiles forming symmetric membrane densities also contact the dendrites of aspiny and spiny neurons. The axon hillocks and initial segments of both neuronal types receive a synaptic input, which is more common on spiny cells.Results offer unequivocal evidence for the differences in the ultrastructure of these two most common categories of medium-size neostriatal neurons, which may help in their proper identification in standard material, as well as information on the types and distributions of synaptic inputs onto these neurons. Moreover, the findings clarify some controversies in the literature probably originating from observations on a mixed population of cells of medium size.     

Three-dimensional reconstruction from serial sections: II. A microcomputer-based facility for rapid data collection

A microcomputer-based facility is described that permits the data required for three-dimensional reconstructions to be collected quickly and inexpensively from serial sections. The facility consists of a microcomputer, a digitizer tablet, a graphics terminal, a printer, a plotter, and telephone coupler. Images of serial sections are superimposed on the digitizer tablet. Contours of interest on each section are digitized and the coordinates are stored on "floppy" disks. The problems of putting successive sections in correct register and of taking into account magnification factors are discussed briefly. Use of the facility for high-resolution applications is also considered.     

Spatial accuracy of 3D reconstructed radioluminographs of serial tissue sections and resultant absorbed dose estimates

Many agents using tumour-associated characteristics are deposited heterogeneously within tumour tissue. Consequently, tumour heterogeneity should be addressed when obtaining information on tumour biology or relating absorbed radiation dose to biological effect. We present a technique that enables radioluminographs of serial tumour sections to be reconstructed using automated computerized techniques, resulting in a three-dimensional map of the dose-rate distribution of a radiolabelled antibody. The purpose of this study is to assess the reconstruction accuracy. Furthermore, we estimate the potential error resulting from registration misalignment, for a range of beta-emitting radionuclides. We compare the actual dose-rate distribution with that obtained from the same activity distribution but with manually defined translational and rotational shifts. As expected, the error produced with the short-range 14C is much larger than that for the longer range 90Y; similarly values for the medium range 131I are between the two. Thus, the impact of registration inaccuracies is greater for short-range sources.     

Distribution of thalamic input to different dendrites of a spiny stellate cell in mouse sensorimotor cortex

A Golgi impregnated, gold-toned [2] spiney stellate cell from layer IV of mouse SmI cortex was reconstructed in three dimensions from serial thin sections to assess the apatial relationships of the synapses onto its dendrites. The distribution of thalamocortical (TC) synapses with the reconstructed dendrites is presented in this report. Thalamocortical axon terminals were labeled by lesion induced degeneration which, in mouse SmI cortex, may reliably indicate the numbers of thalamocortical axon terminals. Results indicate that thalamocortical synapses, which are distributed over most regions of the dendritic tree, are arranged in a regular, periodic fashion on parts of two of the reconstructed dendrites. In these regions, the necks of spines receiving thalamocortical input attach to the dendrite shaft at intervals of about 5 micrometers. In many other regions of the dendritic tree, two spines receiving thalamocortical synapses are separated by a similar interval. Further studies are expected to determine the extent to which dendrites of spiny stellate cells and of other kinds of cortical neurons are contacted in a periodic fashion by thalamocortical axon terminals.     

Synaptic relationships of a type of GABA-immunoreactive neuron (clutch cell), spiny stellate cells and lateral geniculate nucleus afferents in layer IVC of the monkey striate cortex

The precise stimulus specificity of striate cortical neurons is strongly influenced by processes involving gamma-aminobutyric acid (GABA). In the visual cortex of the monkey most afferents from the lateral geniculate nucleus terminate in layer IVC. We identified a type of smooth dendritic neuron (clutch cell) that was immunoreactive for GABA, and whose Golgi-impregnated dendrites and axon were largely restricted to layer IVC beta. The slightly ovoid somata were 8-12 micron by 12-15 micron and the dendritic field was often elongated, extending 80-200 micron in one dimension. The axonal field was 100-150 micron in diameter and densely packed with large bulbous boutons. Although mainly located in IVC beta both the dendritic and axonal processes entered IVC alpha. Fine structural features of GABA-immunoreactive and-impregnated clutch cells and impregnated spiny stellate cells were compared. Clutch cells had more cytoplasm, more densely packed mitochondria and endoplasmic reticulum, and made type II as opposed to type I synapses. A random sample of 159 elements postsynaptic to three clutch cells showed that they mainly terminated on dendritic shafts (43.8-58.5%) and spines (20.8-46.3%), rather than somata (10-17%). The majority of the postsynaptic targets were characteristic of spiny stellate cells. This was directly demonstrated by studying synaptic contacts between an identified GABA positive clutch cell and the dendrites and soma of an identified spiny stellate cell. The termination of clutch cells mainly on dendrites and spines of spiny stellate cells suggests that they interact with other inputs to the same cells. Following an electrolytic lesion in the ipsilateral lateral geniculate nucleus we examined the distribution of degenerating terminals on three identified spiny stellate neurons in layer IVC beta. Out of eight synapses from the lateral geniculate nucleus one was on a dendritic shaft, the rest on spines. Only a small fraction of all synapses on the cells were from degenerating boutons. A clutch cell within the area of dense terminal degeneration was not contacted by terminals from the lateral geniculate nucleus. The results show that layer IVC in the monkey has a specialized GABAergic neuron that terminates on spiny stellate cells monosynaptically innervated from the lateral geniculate nucleus. Possible functions of clutch cells may include inhibitory gating of geniculate input to cortex; maintenance of the antagonistic subregions in the receptive fields; and the creation from single opponent of double colour opponent receptive fields.     

Comparative electrophysiology of pyramidal and sparsely spiny stellate neurons of the neocortex

Slices of sensorimotor and anterior cingulate cortex from guinea pigs were maintained in vitro and bathed in a normal physiological medium. Electrophysiological properties of neurons were assessed with intracellular recording techniques. Some neurons were identified morphologically by intracellular injection of the fluorescent dye Lucifer yellow CH. Three distinct neuronal classes of electrophysiological behavior were observed; these were termed regular spiking, bursting, and fast spiking. The physiological properties of neurons from sensorimotor and anterior cingulate areas did not differ significantly. Regular-spiking cells were characterized by action potentials with a mean duration of 0.80 ms at one-half amplitude, a ratio of maximum rate of spike rise to maximum rate of fall of 4.12, and a prominent afterhyperpolarization following a train of spikes. The primary slope of initial spike frequency versus injected current intensity was 241 Hz/nA. During prolonged suprathreshold current pulses the frequency of firing adapted strongly. When local synaptic pathways were activated, all cells were transiently excited and then strongly inhibited. Bursting cells were distinguished by their ability to generate endogenous, all-or-none bursts of three to five action potentials. Their properties were otherwise very similar to regular-spiking cells. The ability to generate a burst was eliminated when the membrane was depolarized to near the firing threshold with tonic current. By contrast, hyperpolarization of regular-spiking (i.e., nonbursting) cells did not uncover latent bursting tendencies. The action potentials of fast-spiking cells were much briefer (mean of 0.32 ms) than those of the other cell types.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dendritic and synaptic properties of collicular neurons: a quantitative light and electron microscopical study of Golgi-impregnated cells.

The present study deals with a light- and electron microscopic morphometric analysis of Golgi-impregnated neurons in the superior colliculus of rats with the purpose to unravel inter- and intralaminar differences in their dendritic and synaptic organization. In particular, layer IV was studied and compared with its boundary layers III and V. The results show that collicular cells in layer IV basically form a homogeneous population with respect to the number of primary dendrites, the total length of impregnated dendrites, and the diameter, ellipticity, and orientation of dendritic fields and somata of Golgi-impregnated neurons. Somata of reconstructed small cells in layer III and IV as well as V have all a similar density of about 40 synaptic contacts per 100 microns2 surface. However, the cell bodies of large multipolar cells in layer V have a slightly but significantly larger synaptic density (about 50 per 100 microns2). Dendrites of large and small collicular cells had no significantly different synaptic densities (43 and 48 per 100 microns2, respectively). In conclusion, the present results show only minor dendritic and synaptic differences between individual cells in the same layer, as well as in neighboring layers, which implies a low degree of cellular and synaptic intra- and interlaminar differentiation. It is discussed that this organization differs markedly from that in other visual centers, including the collicular homologue, the tectum of lower vertebrates, and the mammalian visual cortex, where pronounced inter- and intralaminar differentiations exist. Such an organization may provide a framework of laminar specificity by which distinct cell types may select a restricted set of input out of all information available. The present quantitative investigation suggests that a similar framework is not present in the superior colliculus.     

Imaging and modelling from serial microscopic sections for the study of anatomy

A system is considered for segmenting noisy intensity images and consequent three-dimensional object reconstruction from a set of planar contours. A new semiautomatic method for the extraction of contours from a sequence of cross-sectional images based on an active contour model (ACM) is proposed. The dynamic ACM proceeds along the sequence of cross-sections following a non-rigid motion, in accordance with the organ boundary. Image texture information is also employed in the model. Problems associated with topological reconstruction from planar contours are addressed, and several criteria promoting semi-automatic topological reconstruction are introduced. The proposed system is successfully applied to the processing of real data related to animal embryonic organs, proving that the system allows detailed modelling of irregular objects. The reconstructed models can be observed in wire-frame, solid, transparent or stereoscopic semi-transparent format. The human-computer interaction implemented in the procedure assists with problems of feature identification and object manipulation about an arbitrary axis.     

Transport pathways for macromolecules in the aortic endothelium. II. The distribution analysis of plasmalemmal vesicles reconstructed by serial sections

Three-dimensional organization of vesicles was examined to elucidate the transendothelial transport properties for macromolecules in rat aortic endothelium using ultrathin serial sections and horseradish peroxidase (HRP) as a tracer molecule. A total number of reconstructing vesicles was 1,298 in nine series electron micrographs, if each vesicular entity was counted as one regardless of composite number of vesicles. The vesicles could be classified into the following six types: HRP-positive luminal, abluminal and intercellular invaginations; HRP-positive channels; HRP-positive and -negative vesicles. The vesicular invaginations and the channels occupied 97.8% and 0.9% of the total vesicles, whereas HRP-positive and -negative free vesicles were found in 0.8% and 0.5%, respectively. The average numerical density of the luminal invagination was 41.1/μm² and approximately equal to that of the abluminal invagination (42.0/μm²), whereas the frequency of the latter was 1.4 times higher than that of the former since the abluminal surface of the endothelium was more irregular to increase a surface area. Each endothelial region varied in the vesicular density and the peripheral region generally showed the higher density, although the transendothelial channels composed of vesicles were not always found in every peripheral region. These results suggest that the shuttle hypothesis is unsuitable to explain “vesicular transport” in the arterial endothelium as well as in the capillary endothelium and that the channels in the peripheral region mainly control transendothelial transport for macromolecules via vesicles. © 1993 Wiley-Liss, Inc.