The morphogenesis of Wulst neurons in the pied flycatcher under conditions of limited afferent inflow]

Golgi preparations of the pied flycatcher Wulst region (the structure analogous to the mammalian visual cortex) were analyzed using the method of computerized morphometry, to study the influence of visual deprivation on the development of different types of neurons selected previously. Deprivation was accomplished by covering the young's eye with nontransparent caps. The experiments were carried out in 10-day-old nestlings (the onset of patterned vision) and 13-day-old nestlings (functioning patterned vision). In 10-day-old nestlings, the deprivation produced constructive changes in dendritic apparatus of projective stellate cells (among them, the most pronounced was more than three-fold increase in the number of foci of maximal branching) practically not affecting the small stellate-like cells. In 13-day-old nestlings, cells belonging to all selected cell types underwent destructive changes: their quantitative characteristics were decreased as compared to those in control nestlings. A large number of tree-like neurons were revealed in the Wulst in the deprived 10-day-old nestlings while in the control age-matched nestlings they were virtually never found. This phenomenon may be explained by the increased affinity to impregnation evoked by deprivation-induced biochemical changes in the tree-like neurons or to increase in their number. In the latter case, the phenomenon may be considered as compensatory, directed at the establishing of contacts with nonvisual afferents.     

Wulst Neurons in Nestlings of the Pied Flycatcher Ficedula hypoleuca at the Pre-Visual Stage of Development

Neurons of the Wulst region, an analog of the mammalian visual cortex, were studied in Golgi-impregnated preparations of brain of non-precocial 1-day old nestlings of the pied flycatcher Ficedula hypoleuca. At this age, vision does not function in nestlings, their behavior is provided by an acoustic analyzer. Two populations of nerve cells, large and small juvenile neurons, were differentiated at visual examination. The comparative morphometry has shown these groups to differ significantly from each other by most studied parameters: the area of the profile field of their soma, the total length of dendrites, branching of the cell, the number of the maximal branching foci, the character of distribution of dendrite free endings in three concentric zones of the cell dendritic field. The distribution of dendrites in the dendritic neuron field was similar in the both groups of neurons. An increased density of dendrites was observed from the side of the afferent input. At the same time, study of orientation of the longest dendrite has shown it to be located parallel to the plane of the afferent input practically in all cells of the both groups. It is suggested that such orientation of the longest dendrite broadens the area of cell contacts, which is necessary for search for the maximal number of afferents. The predominant orientation of dendrites in the direction to the afferent input forms foundation for establishing a more extended zone of contacts with growing visual afferents.     

Quantitative morphological characteristics of developing neurons of the sensory trigeminal nuclei in kittens

Five types of neurons were distinguished in the sensory nuclei of the trigeminal nerve, stained by Golgi's method, in kittens aged 1–5 days and 30 days: reticular and short-dendritic cells (with few branches), and multipolar giant cells, arborescent, and bushy neurons (densely branching). Yet another special type of cell, with a few short dendrites and one long dendrite, was distinguished in preparations from the brain of newborn kittens. Analysis of the dimensions of the bodies, the number, length, and ramification of the dendrites, and the total ramification of the cell yielded quantitative morphological characteristics of these neurons at different times of development. These types of neurons differed in their qualitative and quantitative parameters and in the features of their maturation.Bushy neurons underwent regressive changes during development. Foci of maximal ramification of dendrites of densely branched neurons changed their location during the first months of life relative to the cell body, moving into the more distal regions of the dendrites. Differences in orientation of dendrites with foci of maximal ramification were found relative to neighboring brain formations, which depended on the types of cells and the animal's age. The high level of maturity of trigeminal neurons at birth was demonstrated.Institute of Higher Nervous Activity and Neurophysiology, Academy of Sciences of the USSR, Moscow. Brain Institute, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 14, No. 6, pp. 592–600, November–December, 1982.     

3-D Golgi and image analysis of the olfactory tubercle in schizophrenia

OBJECTIVE: To examine morphologic changes in the olfactory tubercle (OT) spiny neurons and astrocytes in schizophrenia (Sch) by means of quantitative 3-D Golgi and immunocytochemical studies. STUDY DESIGN: Free-floating vibrotome sections of postmortem brain tissue from 10 controls and 12 Sch cases were used for Golgi study and glial fibrillary acidic protein (GFAP) immunocytochemistry. A gray level image analysis was applied for quantitative estimation of GFAP-positive astrocytes on uniform, randomly sampled sections. This method is effective for low-contrast objects on an uneven background. Golgi-impregnated OT spiny neurons were analyzed both qualitatively and quantitatively in three dimensions with a semiautomated microscope-computer system. From digitized image of the neurons, various metric parameters were estimated to characterize the dendritic tree. RESULTS: In cases of Sch, degenerative changes in the dendrites of OT spiny neurons were revealed. A decrease in the maximal radius of the dendritic tree and total length of dendrites were accompanied by an increase in the length density of dendrites. Hypertrophy and a more-intensive GFAP reaction of astrocytes were found in OT of Sch. CONCLUSION: Based on these results, one can hypothesize that OT spiny neurons in Sch are involved in the process of dendritic reorganization, including degenerative changes in dendrites.     

Genetic manipulation of single neurons in vivo reveals specific roles of flamingo in neuronal morphogenesis

To study the roles of intracellular factors in neuronal morphogenesis, we used the mosaic analysis with a repressible cell marker (MARCM) technique to visualize identifiable single multiple dendritic (MD) neurons in living Drosophila larvae. We found that individual neurons in the peripheral nervous system (PNS) developed clear morphological polarity and diverse dendritic branching patterns in larval stages. Each MD neuron in the same dorsal cluster developed a unique dendritic field, suggesting that they have specific physiological functions. Single-neuron analysis revealed that Flamingo did not affect the general dendritic branching patterns in postmitotic neurons. Instead, Flamingo limited the extension of one or more dorsal dendrites without grossly affecting lateral branches. The dendritic overextension phenotype was partially conferred by the precocious initiation of dorsal dendrites in flamingo mutant embryos. In addition, Flamingo is required cell autonomously to promote axonal growth and to prevent premature axonal branching of PNS neurons. Our molecular analysis also indicated that the amino acid sequence near the first EGF motif is important for the proper localization and function of Flamingo. These results demonstrate that Flamingo plays a role in early neuronal differentiation and exerts specific effects on dendrites and axons.     

Statistical evaluation of dendritic growth models

A mathematical model (Kliemann, W. 1987.Bull. math. Biol. 49, 135–152.) that predicts the quantitative branching pattern of dendritic tree was evaluated using the apical and basal dendrites of rat hippocampal neurons. The Wald statistics for χ²-test was developed for the branching pattern of dendritic trees and for the distribution of the maximal order of the tree. Using this statistic, we obtained a reasonable, but not excellent, fit of the mathematical model for the dendritic data. The model's predictability of branching patterns was greatly enhanced by replacing one of the assumptions used for the original model “splitting of branches for all dendritic orders is stochastically independent”, with a new assumption “branches are more likely to split in areas where there is already a high density of branches”. The modified model delivered an excellent fit for basal dendrites and for the apical dendrites of hippocampal neurons from young rats (30–34 days postpartum). This indicates that for these cells the development of dendritic patterns is the result of a purely random and a systematic component, where the latter one depends on the density of dendritic branches in the brain area considered. For apical dendrites there is a trend towards decreasing pattern predictability with increasing age. This appears to reflect the late arrival of afferents and subsequent synaptogenesis proximal on the apical dendritic tree of hippocampal neurons.     

Golgi-type I and Golgi-type II Neurons in the Ventral Anterior Thalamic Nucleus of the Adult Human: Morphological Features and Quantitative Analysis

The morphological and quantitative features of neurons in the adult human ventral anterior thalamic nucleus were studied in Golgi preparations. Two neuronal types were found and their quantitative features were studied. Golgi-type I neurons were medium to large cells with dense dendritic trees and dendritic protrusions and short hair-like appendages. They have somatic mean diameter of 30.8 μm (±9.4, n = 85). They have an average 100.3 dendritic branches, 48.97 dendritic branching points, and 58.85 dendritic tips. The mean diameters of their primary, secondary, and tertiary dendrites were 3.1 μm (±1, n = 80), 1.85 μm (±0.8, n = 145), and 1.5 μm (±0.4, n = 160), respectively. Golgi-type II neurons were small to medium cells with few sparsely branching dendrites and dendritic stalked appendages with or without terminal swellings. They have somatic mean diameters of 22.2 μm (±5.8, n = 120). They have an average 33.76 dendritic branches, 16.49 dendritic branching points, and 21.97 dendritic tips. The mean diameters of their primary, secondary, and tertiary dendrites were 1.6 μm (±0.86, n = 70), 1.15 μm (±0.55, n = 118), and 1 μm (±0.70, n = 95), respectively. These quantitative data may form the basis for further quantitative studies involving aging or some degenerative diseases that may affect cell bodies and/or dendritic trees of the Golgi-type I and/or Golgi-type II thalamic neurons.     

Control of dendritic field formation in Drosophila: the roles of flamingo and competition between homologous neurons

Neurons elaborate dendrites with stereotypic branching patterns, thereby defining their receptive fields. These branching patterns may arise from properties intrinsic to the neurons or competition between neighboring neurons. Genetic and laser ablation studies reported here reveal that different multiple dendritic neurons in the same dorsal cluster in the Drosophila embryonic PNS do not compete with one another for dendritic fields. In contrast, when dendrites from homologous neurons in the two hemisegments meet at the dorsal midline in larval stages, they appear to repel each other. The formation of normal dendritic fields and the competition between dendrites of homologous neurons require the proper expression level of Flamingo, a G protein-coupled receptor-like protein, in embryonic neurons. Whereas Flamingo functions downstream of Frizzled in specifying planar polarity, Flamingo-dependent dendritic outgrowth is independent of Frizzled.     

Effects of lingual nerve section on morphometric characteristics of reticular nucleus neurons in the kitten brain stem

Computerized morphometric techniques were used to investigate each of 23 parameters in three types of brain stem reticular nucleus neurons in Golgi-stained frontal slices from the brain of 30-day-old kittens after uni- and bilateral lingual nerve section 5–7 days after birth. Particular statistically significant differences in some parameters were discovered in all types of cell. Certain group-specific differences in parameters could be most frequently distinguished in each category: distribution of loose dendritic endings through the dendritic area in reticular neurons, length of dendritic segments in branching cells, and distribution of foci of dendritic arborization in giant multipolar neurons. Unilateral lingual nerve section results in quantitatively more marked deviation from the normal state. It was only under these circumstances, moreover, that differences in overall length of dendrites could be seen, which could indicate a difference in the surface area of the cell.Institute of Higher Nervous Activity and Neurophysiology, Academy of Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 23, No. 4, pp. 409–418, July–August, 1991.     

Degeneration of Axotomized Projection Neurons in the Rat dLGN: Temporal Progression of Events and Their Mitigation by a Single Administration of FGF2

Removal of visual cortex in the rat axotomizes projection neurons in the dorsal lateral geniculate nucleus (dLGN), leading to cytological and structural changes and apoptosis. Biotinylated dextran amine was injected into the visual cortex to label dLGN projection neurons retrogradely prior to removing the cortex in order to quantify the changes in the dendritic morphology of these neurons that precede cell death. At 12 hours after axotomy we observed a loss of appendages and the formation of varicosities in the dendrites of projection neurons. During the next 7 days, the total number of dendrites and the cross-sectional areas of the dendritic arbors of projection neurons declined to about 40% and 20% of normal, respectively. The response of dLGN projection neurons to axotomy was asynchronous, but the sequence of structural changes in individual neurons was similar; namely, disruption of dendrites began within hours followed by cell soma atrophy and nuclear condensation that commenced after the loss of secondary dendrites had occurred. However, a single administration of fibroblast growth factor-2 (FGF2), which mitigates injury-induced neuronal cell death in the dLGN when given at the time of axotomy, markedly reduced the dendritic degeneration of projection neurons. At 3 and 7 days after axotomy the number of surviving dendrites of dLGN projection neurons in FGF-2 treated rats was approximately 50% greater than in untreated rats, and the cross-sectional areas of dendritic arbors were approximately 60% and 50% larger. Caspase-3 activity in axotomized dLGN projection neurons was determined by immunostaining for fractin (fractin-IR), an actin cleavage product produced exclusively by activated caspase-3. Fractin-IR was seen in some dLGN projection neurons at 36 hours survival, and it increased slightly by 3 days. A marked increase in reactivity was seen by 7 days, with the entire dLGN filled with dense fractin-IR in neuronal cell somas and dendrites.     

The neurons in layer 1 of cat visual cortex.

We have examined the morphology of neurons in layer 1 by injecting them intracellularly with lucifer yellow in lightly fixed brain slices (250 microns thick) taken from the medial bank of area 17 in adult cats. Of 22 neurons with well-filled dendrites, 16 had smooth dendrites, two had sparsely spiny dendrites (less than 200 spines) and, unexpectedly, four had spiny dendrites typical of pyramidal cells. The axon was generally not well filled. Computer reconstructions showed that parts of the dendritic tree had been lost in the sectioning. Nevertheless, measurements of the length of intact dendrites suggested an average diameter of the dendritic tree of 220 microns. The density of the neurons was such that the dendritic trees of about six neurons cover each point in layer 1. Thus, despite the very low density of neurons that characterizes layer 1, there are more than sufficient neurons to sample from the entire representation of the visual field in area 17.     

The Morphological Identity of Insect Dendrites

Dendrite morphology, a neuron's anatomical fingerprint, is a neuroscientist's asset in unveiling organizational principles in the brain. However, the genetic program encoding the morphological identity of a single dendrite remains a mystery. In order to obtain a formal understanding of dendritic branching, we studied distributions of morphological parameters in a group of four individually identifiable neurons of the fly visual system. We found that parameters relating to the branching topology were similar throughout all cells. Only parameters relating to the area covered by the dendrite were cell type specific. With these areas, artificial dendrites were grown based on optimization principles minimizing the amount of wiring and maximizing synaptic democracy. Although the same branching rule was used for all cells, this yielded dendritic structures virtually indistinguishable from their real counterparts. From these principles we derived a fully-automated model-based neuron reconstruction procedure validating the artificial branching rule. In conclusion, we suggest that the genetic program implementing neuronal branching could be constant in all cells whereas the one responsible for the dendrite spanning field should be cell specific.     

The shape and arrangement of the cholinergic neurons in the rabbit retina

The acetylcholine-synthesizing neurons of the rabbit retina were selectively stained by intraocular injection of the fluorescent dye 4,6-diamidino-2-phenylindole (DAPI). Retinas were then isolated from the eye, fixed for 10-30 min with 4% paraformaldehyde, and mounted flat on the stage of a fluorescence microscope. The acetylcholine-synthesizing cells were penetrated under visual control by microelectrodes filled with lucifer yellow CH. When the dye was electrophoretically injected into the cells, complete filling of their dendrites often occurred. Cells were successfully injected as long as one month after fixation of the tissue. Complete or nearly complete filling of 281 cells was accomplished, at retinal locations systematically covering the retinal surface. The cells stained with DAPI were found to form a single morphological population. They have two to seven primary dendrites, which branch repeatedly within a narrow plane and form a round or slightly oval dendritic tree. The branching becomes very fine for the distal one third of the dendritic tree, and the dendrites there are studded with small swellings. The distal dendritic tree lies mainly within one of the two thin strata of the inner plexiform layer where acetylcholine is present. The shape and size of the dendritic tree are continuously graded across the retina, the dendritic tree is narrower and the branching denser in the central retina, wider and sparser in the periphery. From knowledge of the population density and the shape of the neurons, one can reconstruct the array of dendrites that exists within the inner plexiform layer. The overlap of the dendritic fields is an order of magnitude greater than of any other retinal neuron previously described. Because the cells not only overlap widely but branch quite profusely, a very dense plexus of cholinergic dendrites is created.     

Quantitative morphological charateristics of developing neurons of the brain-stem reticular formation

Two types of neurons — reticular (with few branches) and multipolar giant (densely ramified) were distinguished in the brain-stem reticular nuclei of the brain in Golgi preparations from cat fetuses aged 45–55 days and kittens aged 1–5 and 30 days. The quantitative morphological characteristics of these neurons at different stages of development were determined from the dimensions of their bodies, the number, length, and ramification of their dendrites, and the overall ramification of the cell. The types of neurons described above differed in both qualitative and quantitative indices and in the character of their maturation. Maximal ramification of dendrites of giant multipolar neurons was observed in the embryonic period. Foci of maximal ramification in reticular neurons were close to the cell bodies. In gaint multipolar neurons in fetuses and 30-day-old kittens foci of maximal ramification were located on the proximal and distal portions of the dendrites, but in the newborn kittens on the proximal segments only. These facts are examined in connection with differences in the spike activity of the growing neuron.Institute of Higher Nervous Activity and Neurophysiology, Academy of Sciences of the USSR, Moscow. Brain Institute, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 12, No. 1, pp. 53–61, January–February, 1980.     

Control of dendritic branching and tiling by the Tricornered-kinase/Furry signaling pathway in Drosophila sensory neurons

To cover the receptive field completely but without redundancy, neurons of certain functional groups exhibit tiling of their dendrites via dendritic repulsion. Here we show that two evolutionarily conserved proteins, the Tricornered (Trc) kinase and Furry (Fry), are essential for tiling and branching control of Drosophila sensory neuron dendrites. Dendrites of fry and trc mutants display excessive terminal branching and fail to avoid homologous dendritic branches, resulting in significant overlap of the dendritic fields. Trc control of dendritic branching involves regulation of RacGTPase, a pathway distinct from the action of Trc in tiling. Timelapse analysis further reveals a specific loss of the ability of growing dendrites to turn away from nearby dendritic branches in fry mutants, suggestive of a defect in like-repels-like avoidance. Thus, the Trc/Fry signaling pathway plays a key role in patterning dendritic fields by promoting avoidance between homologous dendrites as well as by limiting dendritic branching.     

Morphological characteristics of different types of neurons in the ventrooralis anterior,ventrooralis internus,and ventrooralis posterior nuclei in the human thalamus

1. Golgi-Kopsch preparations of the oral ventral nuclei of human thalamus were analyzed in an attempt to classify the neuronal types. 2. Three types of neurons are described for the first time in humans. Type I neurons are large or medium in size and bear dendrites with protrusions, spines, and short hair-like appendages. Some have a radiate dendritic arbor and others have dendrites grouped in tufts. The dendritic trees of these neurons are dense. 3. Type II neurons are medium or small in size with less dense dendritic trees. These cells have somatic as well as dendritic appendages of different forms. 4. Relatively rare is a type of very small neurons, type III, with few and sparsely branching dendrites.     

Neuronal types in the lateral geniculate nucleus of man

Summary Nerve cell types of the lateral geniculate body of man were investigated with the use of a transparent Golgi technique that allows study of not only the cell processes but also the pigment deposits. Three types of neurons have been distinguished:Type-I neurons are medium-to large-sized multipolar nerve cells with radiating dendrites. Dendritic excrescences can often be encountered close to the main branching points. Type-I neurons comprise a variety of forms and have a wide range of dendritic features. Since all intermediate forms can be encountered as well, it appears inadequate to subdivide this neuronal type. One pole of the cell body contains numerous large vacuolated lipofuscin granules, which stain weakly with aldehyde fuchsin.Type-II and type-III neurons are small cells with few, sparsely branching and extended dendrites devoid of spines. In Golgi preparations they cannot be distinguished from each other. Pigment preparations reveal that the majority of these cells contains small and intensely stained lipofuscin granules within their cell bodies (type II), whereas a small number of them remains devoid of any pigment (type III). Intermediate forms do not occur.     

Glycinergic transmission regulates dendrite size in organotypic culture

We previously demonstrated that inhibitory synaptic transmission influences dendrite development in vivo. We now report an analogous finding in an organotypic culture of a glycinergic projection nucleus, the medial nucleus of the trapezoid body (MNTB), and its postsynaptic target, the lateral superior olive (LSO) of gerbils. Cultures were generated at 6–7 days postnatal and grown in serum containing medium with or without the glycine receptor antagonist, strychnine (SN), at 2 μM. LSO neurons were then labeled with biocytin, and the dendritic arbors were analyzed morphometrically. Compared to neurons from age-matched in vivo tissue, the neurons cultured in control media were somewhat atrophic, including decreases in dendritic branching and length. Incubation in strychnine led to a dramatic increase in dendritic branching and total dendritic length. Control neurons averaged 6.3 branches, compared to 18 branches/neuron in SN-treated cultures. There was a similar increase in primary dendrites and total dendritic length. The physical elimination of MNTB cells did not mimic SN treatment, presumably because glycinergic LSO neurons generated intrinsic connections. In fact, the LSO soma area was significantly greater following MNTB removal, suggesting that these afferents provide a second signal to postsynaptic neurons. These results suggest that spontaneous glycinergic transmission regulates the growth of postsynaptic processes. © 1996 John Wiley & Sons, Inc.     

A Golgi study of the isthmic nuclei in the pigeon (Columba Iivia)

Summary The isthmic nuclei of the pigeon were studied with the use of three different Golgi techniques. The nucleus isthmo-opticus (IO) consists of a single cell type in which all dendrites of one neuron take the same direction and ramify at identical distances from the perikaryon to form dense dendritic arborizations. The cell bodies of the IO neurons form two parallel layers. The dendrites of these neurons always extend to the area between the two layers so that the dendritic arborizations of opposite neurons overlap. A model of the cellular organization of the IO was constructed based upon these morphological characteristics. The neurons of the n. isthmi/pars parvocellularis (Ipc) have oval perikarya and long, smooth, infrequently branching dendrites. All neurons except those at the borders of the nucleus show the same dorsoventral orientation in their dendritic arborizations and together with their afferents seem to have a columnar organization. The dendrites of the neurons located at the margin of the nucleus ramify within the Ipc along its border. The n. semilunaris (Slu) consists of neurons with round somata that have on an average three dendrites with small spines. The axons leave the nucleus from the medial side and join the lemniscus lateralis. The neurons of the n. isthmi/pars magnocellularis (Imc) comprise a generalized isodendritic type resembling the cells of the reticular formation. Axons from the tectum penetrate the nucleus, making numerous en-passant contacts with several neurons.     

Nature of the distribution of synapses in the neurons of the reticular formation and some afferent nuclei]

Under study was the morphology of synaptic terminations of the brain reticular formation in cats and dogs as well as that of the afferent nuclei of the cat's posterior columns. The neurons of the Goll's and Burdach's nuclei have a richly ramified dendritic network. In this connection the main mass of synapses is disposed on the dendrites and their different branchings. The dendrites of the multipolar cells of the reticular formation have the main type of branching, but the cells are distributed from the nerve cell body at a considerable distance (up to 50 mu and more). The synapses are observed at the total length of the dendrite, so the axo-dendritic contacts quantitatively prevail over axo-somatic ones. The morphological data are compared with physiological axo-somatic and axo-dendritic concepts of the role of synapses in conducting the nerve impulse.