William Bondareff

Decrease in the number of synapses in the senescent brain: A quantitative electron microscopic analysis of the dentate gyrus molecular layer in the rat

Axo-dendritic synapses were counted in electron micrographs taken from the middle third of the dentate gyrus molecular layer of young adult and senescent Fischer 344 rats. A significant decrease in the number of synapses was found in senescent animals relative to young ones. This loss of synapses, which involved all the morphological varieties of axo-dendritic synaptic contacts in the dentate gyrus molecular layer, appeared to be unrelated to changes in dimensions of synapses, tissue volume or number of postsynaptic granule cells. It is proposed that the age-related loss of synaptic contacts might be attributed to a reduced capacity of senescent brains for synaptic regeneration and remodelling.

SUBMICROSCOPIC MORPHOLOGY OF GRANULAR VESICLES IN SYMPATHETIC NERVES OF RAT PINEAL BODY

SummaryIn the rat pineal body sympathetic nerve endings are characterized by numerous agranular and granular vesicles. The latter mainly include vesicles of about 450 Å diameter, each of which is bounded by a trilaminar membrane of about 75 Å total thickness. The cores of these granular vesicles appear structurally similar to the bounding membranes and are formed by a less dense, globular or vacuolar component around which a more dense component is distributed. The relative distributions of these two structural elements result in a variety of submicroscopic configurations which suggest a morphological continuum that may be related to the cyclical uptake, storage and release of biogenic amines. A tripartite classification of granular vesicles, such as proposed previously, is not supported by these data.

Intrastriatal spreading of biogenic amines

Abstract Biogenic amines were injected into the caudate nuclei of awake, freely moving rats through cannulae implanted 1 week previously. After injections the brains were freeze-dried and treated with formaldehyde vapors according to the method of Falck and Hillarp. Intrastriatal spreading was determined by fluorescence microscopy. Intrastriatal spread of catecholamines related directly to both concentration and osmolality of injected amine. Dopamine and norepinephrine, 1 μg in 1 μl saline, for example, were found to spread rostral-caudally mean total distances of 2.11 and 1.88 mm, respectively. Intrastriatal spreading was affected by dexamethasone and mannitol, which reduced rostral-caudal spreading of dopamine by more than 50% when administered prior to intrastriatal injection. Spreading of serotonin through brain tissue could not be determined. These data suggest that the spreading of exogenous catecholamines through cerebral tissues may occur through extracellular channels. The observed rate of spreading (about 0.5 mm/min) and electron microscopic analysis were consonant with this interpretation.

Synaptic atrophy in the senescent hippocampus

Quantitative analyses of electron micrographs have shown a decrease in the number of synapses in the dentate gyrus of the senescent Fischer-344 rat. The loss of synapses, involving both dendritic spines and shafts and axon terminals of more than one population of presynaptic neurons, did not depend upon the antecedent loss of postsynaptic neurons or their dendrites. These findings suggest that the age-related loss of synapses in the dentate gyrus may depend upon an inability of presynaptic elements to maintain the structural integrity of synapses in senescence. It is proposed that a change in the glycoprotein component of presynaptic plasma membranes resulting from a deficiency in axonal transport mechanisms in the septo-hippocampal pathway may underly this presynaptic malfunction. The resulting partial deafferentation of neurons in the dentate gyrus in senescence appears to be associated with a secondary atrophy of dendrites, which results in a loss of postsynaptic membranes before a loss of postsynaptic neurons can be documented.

Cytochemistry of synaptosomes

Abstract The effects of neuraminidase and testicular hyaluronidase on the ultrastructure of synaptosomes, prepared from hippocampi of female Sprague-Dawley rats, were evaluated in sections of glutaraldehyde-fixed specimens by electron microscopy. Whereas incubation in testicular hyaluronidase (80 μl/ml) was without apparent effect, neuraminidase (100 U/ml) caused a decrease in the visualized densities of presynaptic and postsynaptic processes and, especially, the intervening synaptic cleft. The decrement in density of the synaptic cleft, also demonstrated microdensitometrically, suggested that at least some of the sialic acid freed from synaptosomes by incubation in neuraminidase is located extracellularly, within the synaptic cleft.

Diminished axonal transport of glycoproteins in the senescent rat brain.

At various time intervals (10, 15, 20, 25, 30 min) after injection of 3H-fucose into the medial septal nucleus of young adult (3 months old) and senescent (25 months old) Fischer-344 rats, the specific activities of trichloroacetic acid-phosphotungstic acid (TCA-PTA) soluble and insoluble fractions were determined in the medial area of the septum and in three successive rostro-caudal sections of the hippocampal formation containing mainly the dentate gyrus, but also its hilus with fields CA4 and CA3c of the hippocampus. The rate of 3H-fucose incorporation into glycoproteins of the septum did not differ in young adult and senescent rats. Part of the TCA-PTA soluble and insoluble radioactive material was transported through the septo-hippocampal pathway to the dentate gyrus. This transport was inhibited by the injection of colchicine into the septum prior to 3H-fucose injection and was completely blocked by electrolytic lesion of the medial septal nucleus. The arrival time and the amount of the TCA-PTA soluble radioactive material transported to the dentate gyrus did not differ in young adult and senescent rats. However, the TCA-PTA insoluble labelled glycoprotein was transported to the dentate gyrus in a significantly smaller amount and during a longer period of time in the senescent animals. This age-related change may reflect a reduction in amount and/or in rate of axonal transport of glycoproteins in the septo-hippocampal pathway of senescent rats.

Demonstration of an intercellular substance in mouse cerebral cortex

SummarySpecimens of mouse cerebral cortex were preserved by freezing and drying and treated in vacuo with osmium tetroxide vapors. In the neuropil of the molecular layer dense, osmiophilic laminae, presumably plasma membranes of immediately adjacent cell processes, were found to be separated by an intercellular space, which appeared relatively electron lucent in unstained sections. In sections stained with uranyl acetate an intercellular substance was demonstrated, the visualized density of which was reduced when staining was accomplished at low pH. These reactions suggest the presence of a non-lipid, anionic intercellular substance which may contain acid mucopolysaccharide.

Brain extracellular space in monkey fetuses subjected to prolonged partial asphyxia

Abstract Rhesus monkey fetuses, closely approaching term, were subjected to episodes of prolonged partial asphyxia while still in utero . Maternal blood presure was gradually decreased by increasing the concentration of inspired fluothane until a moderate degree of fetal compromise, indicated by a pH of 6.9–7.1 and a Po 2 of 15–18 mm Hg was sustained for 3–4 hr. Immediately after delivery by cesarian section, specimens of fetal cerebral cortex were frozen in liquid Freon-22, pre-chilled to —78C for 10 days. Electron micrographs of these specimens were prepared from which the relative volume of extracellular space was estimated stereometrically. A mean extracellular space of 4.1% in fetal animals subjected to prolonged partial asphyxia differed significantly from an extracellular space of 9.1% found in cerebral cortex from normal fetuses, similarly delivered. Additionally, mitochondria from asphyxiated fetuses were swollen and characterized by a dispersed, dilute-appearing matrix. These data suggested that the brain swelling concomitant with prolonged partial asphyxia is associated with a translocation of extracellular fluid, which appears to depend upon an altered mitochondrial metabolism and which results in a reduced volume of extracellular space.

Distribution of ferritin in the cerebral cortex of the mouse revealed by electron microscopy

Abstract In the cerebral cortex of the mouse, ferritin, administered intravenously or intraperitoneally, readily passes from blood to brain. It is observed to enter the neuropil in an apparently nonspecific manner and is found free in the cytoplasmic ground substance of astrocytes and other cells adjacent to the capillary basement membrane. Its passage through the neuropil appears to occur with water from one cell to another and is not restricted to astrocytes, other cell species, or extracellular space. The data presented suggest that ferritin may specifically activate cell membranes and induce an uptake of molecular species similar to, but more rapid than, that accomplished by pinocytosis.

Submicroscopic structure of single neurons isolated from rabbit lateral vestibular nucleus

Giant neurons from the lateral vestibular nuclei of rabbits have been isolated in 0.25 M sucrose by freehand dissection. They were fixed by immersion in glutaraldehyde followed by osmium tetroxide and dehydrated with Durcupan A prior to embedding in Araldite 502. By means of this simple method it was possible to achieve a preservation of isolated cells which was comparable to that obtained when small pieces of tissue are preserved, sectioned, and examined in the electron microscope. Mitochondria, however, were characterized by a condensed, granular matrix and more closely resembled those found in fractions of cells homogenized in 0.25 M sucrose than in tissue sections. Plasma membranes were essentially intact, and in fortuitous sections a trilaminar structure was demonstrated. These data support the assumption that disruption of plasma membranes and other morphological changes previously reported occur primarily after isolation of nerve cells and during their preparation for examination in the electron microscope. Although these changes could be minimized, it was difficult to estimate accurately their extent.