Chromation at low temperatures improves impregnation of neurons in Golgi-aldehyde methods.

A modification of the Golgi-aldehyde methods is proposed in which the tissue is maintained at 4 degrees C during chromation. This prevents precipitation of the solution, and significantly improves the quality of the Golgi stain, probably owing to better fixation of the tissue.

A combination of Golgi impregnation and fluorescent retrograde labeling.

Central nervous system structures containing neurons labeled by the fluorescent tracers Fast blue (FB), Diamidino yellow dihydrochloride (DY), Rhodamine B isothiocyanate (RITC) and Rhodamine-labeled latex microspheres (RLM) were processed with the Golgi method. The goal was to improve the visualization of the fluorescent labeled neurons and to allow their ultrastructural examination. While the fluorescence of FB and RITC is greatly attenuated by the Golgi method, RLM and DY are still visible in Golgi-impregnated neurons. However, it is usually necessary to remove the silver precipitate by gold-toning.

A method for the study of the spatial distribution of the neuronal dendritic tree using a universal stage.

The use of a microscope equipped with a universal stage with 4 rotation axes, drawing tube and photographic system for the 3-dimensional study of neuronal morphology is considered. Two-dimensional projections of the neuronal tree are obtained by rotating the stage, and the application of coordinate transformations results in 3-dimensional mapping of neuronal topography. Algorithms used for these transformations are presented and the method applied to Golgi-impregnated neurons of the cerebral cortex of adult lizards. The advantages, limitations and sources of error of this method are discussed.

Effect of hypothyroidism on the size of spines of pyramidal neurons of the cerebral cortex.

We have previously shown that hypothyroidism produces a decrease in the number of spines counted along the apical shafts of pyramidal neurons of the cortex. Nevertheless, other authors have found that when an animal is subjected to some adverse living conditions the size of the spines decreases, making them invisible to the light microscope. The question arises then of whether the decrease in the number of spines reported by us in hypothyroid animals is real or is due to a shrinking effect. In order to elucidate this question the cross-surface area of dendritic spines of apical shafts belonging to 20- and 60-day-old rats, thyroidectomized at 10 days of age, as well as those of their corresponding controls were measured in different layers of their cortex, studied using conventional electron microscopic techniques. The application of the three-way analysis of variance model to these data has shown us that while the age of the animal produces a definite increase in the size of the spines, hypothyroidism does not produce any change in their size, leading us to the conclusion that the decrease in the number of spines previously reported is due to an actual loss of these elements.

Handling Golgi-impregnated tissue for light microscopy.

The use of cyanocrylic glue to fix pieces of Golgi-stained nervous tissue on a paraffin blank is proposed for obtaining thick sections of unembedded tissue with a sliding microtome. This procedure makes correct orientation of the tissue easy during sectioning and makes it possible to obtain tissue sections quickly. The sections are flat-mounted using epoxy resin, resulting in permanent preparations with excellent optical properties and enabling further thin-sectioning for light and electron microscopic studies.

Apical dendritic spines and axonic terminals in the bipyramidal neurons of the dorsomedial cortex of lizards (Lacerta).

Gold-toned bipyramidal neurons of the dorsomedial cortex of Lacerta have been studied using light and electron microscopy. The spines have been classified as stubby, mushroom-shaped or thin. Thin and mushroom-shaped spines are only found on proximal and intermediate dendritic segments, whereas stubby spines are found on distal dendritic segments. A Timm's method modification for electron microscopy (sulphide-osmium procedure) has been used. Timm-positive axonal endings usually synapse on thin and mushroom-shaped spines, whereas Timm-negative axonal endings usually synapse on stubby spines. Timm-positive afferents and their post-synaptic spines on bipyramidal neurons of Lacerta's dorsomedial cortex are compared with the corresponding elements on pyramidal neurons of the CA3 region of the hippocampus of mammals, on the basis of several histochemical and morphological studies. The possibility that these two neuronal types may be homologous is discussed.

Intrinsic organization of the medial cerebral cortex of the lizard Lacerta pityusensis: A golgi study.

The morphology of cells and the organization of axons were studied in Golgi-Colonnier and toluidine blue stained preparations from the medial cerebral cortex of the lizard Lacerta pityusensis. In the medial cortex, six strata were distinguished between the superficial glial membrane and the ependyma. Strata I and II formed the outer plexiform layer, stratum III formed the cellular layer, and strata IV go VI the inner plexiform layer. The outer plexiform layer contained smooth bipolar neurons; their dendrites were oriented anteroposteriorly and their axons were directed towards the posterior zone of the brain. Five neuronal types were observed in the cellular layer. The spinous pyramidal neurons had well-developed apical dendrites and poorly developed basal ones. Their axons entered the inner plexiform layer and gave off collaterals oriented anteroposteriorly. The small, sparsely spinous pyramidal neurons had poorly developed dendrites and their axons entered the inner plexiform layer. The spinous bitufted neurons had well-developed apical and basal dendritic tufts. Their axons gave off collaterals that reached the outer and inner plexiform layers of both the dorsomedial and dorsal cortices. The sparsely spinous horizontal neurons had dendrites restricted to the outer plexiform layer. Their axons entered the inner plexiform layer. The sparsely spinous, multipolar neurons had their soma close to stratum IV and their axons entered the outer plexiform layer. In stratum V of the inner plexiform layer were large, spiny polymorphic neurons; they had dendrites with long spines, and their axons reached the cellular layer. On the basis of these results, we have subdivided the medial cortex into two subregions: the superficial region, which contains the neurons of the cellular layer and their dendritic domains, and the deep region, strata V and VI, which contains the large, spiny polymorphic neurons. The neurons in the medial cortex of these lizards resembles those in the area dentata of mammals. On this basis, the superficial region may be compared to the dentate gyrus and the deep region to the hilar region of the hippocampus of mammals.