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PubMed2004

Active caspase-6 and caspase-6-cleaved tau in neuropil threads, neuritic plaques, and neurofibrillary tangles of Alzheimer's disease.

Previously, we have shown that caspase-6 but not caspase-3 is activated by serum deprivation and induces a protracted cell death in primary cultures of human neurons (LeBlanc AC, Liu H, Goodyer C, Bergeron C, Hammond J: Caspase-6 role in apoptosis of human neurons, amyloidogenesis and Alzheimer's disease. J Biol Chem 1999, 274:23426-23436 and Zhang Y, Goodyer C, LeBlanc A: Selective and protracted apoptosis in human primary neurons microinjected with active caspase-3, -6, -7, and -8. J Neurosci 2000, 20:8384-8389). Here, we show with neoepitope antibodies that the p20 subunit of active caspase-6 increases twofold to threefold in the affected temporal and frontal cortex but not in the unaffected cerebellum of Alzheimer's disease brains and is present in neurofibrillary tangles, neuropil threads, and the neuritic plaques. Furthermore, a neoepitope antibody to caspase-6-cleaved Tau strongly detects intracellular tangles, extracellular tangles, pretangles, neuropil threads, and neuritic plaques. Immunoreactivity with both antibodies in pretangles indicates that the caspase-6 is active early in the pathogenesis of Alzheimer's disease. In contrast to the nuclear and cytosolic localization of active caspase-6 in apoptotic neurons of fetal and adult ischemic brains, the active caspase-6 in Alzheimer's disease brains is sequestered into the tangles or neurites. The localization of active caspase-6 may strongly jeopardize the structural integrity of the neuronal cytoskeletal system leading to inescapable neuronal dysfunction and eventual cell death in Alzheimer's disease neurons. Our results suggest that active caspase-6 is strongly implicated in human neuronal degeneration and apoptosis.

Aged↗
PubMed1998

Tracing of a neuronal network in the locust by pressure injection of markers into a synaptic neuropil.

Central neuronal circuits of vertebrates have often been investigated using injection of markers into synaptic neuropils, whereas similar techniques have rarely been applied in invertebrates. In this study we tested several neuroanatomical tracers for their ability to mark central neuronal circuits in insects, using the well described auditory network of the locust, Locusta migratoria. After physiological localization of an auditory neuropil various tracers were pressure injected. Horseradish peroxidase, dextrans (3 and 10 kDa) and especially biocytin and neurobiotin were effectively incorporated by auditory interneurons, which resulted in their extensive labeling. Postsynaptic regions turned out to be the major, if not exclusive sites of uptake of injected markers, which is deduced from two lines of evidence: (i) for labeling of identified auditory neurons it was necessary to apply the tracer to postsynaptic sites of the neuron; (ii) only a few non-auditory neurons were labeled (probably by lesioning axons during electrode impalement). No evidence could be found for an activity dependent uptake. We conclude that pressure injection of certain tracers into synaptic areas can be used to identify central nervous circuits in insects.

Acoustic Stimulation↗
PubMed1997

Expansion of the neuropil of the mushroom bodies in male honey bees is coincident with initiation of flight.

The mushroom bodies (MB), the insect brain structures most often associated with learning, have previously been shown to exhibit structural plasticity during the adult behavioral development of female worker and queen honey bees. We now show that comparable morphological changes occur in the brains of male honey bees (drones). The volume of the MB in the brains of drones was estimated from tissue sections using the Cavalieri method. Brains were obtained from six groups of drones that differed in age and flight experience. Circulating levels of juvenile hormone (JH) in these drones were determined by radioimmunoassay (RIA). There was an expansion of the neuropil of the MB that was temporally associated with drone behavioral development, as in female queens and workers. The observed changes in drones were maintained in the presence of low levels of JH, also as in females. These results suggest that expansion of the neuropil of the MB in honey bees is associated with learning the location of the nest, because this learning is the most prominent aspect of behavioral development common to all members (workers, drones, queen) of the honey bee colony.

Age Factors↗
PubMed2004

NADPH-diaphorase histochemical labeling patterns in the hippocampal neuropil and visual cortical neurons in weaned rats reared during lactation on different litter sizes.

Tissue distribution of nitric oxide-synthases was investigated in the rat hippocampus and visual cortex under nutritional changes induced by modification of the litter size. Young (30-45-days-old) rats, suckled in litters formed by 3,6 or 12 pups (called small, medium and large litters, respectively), were studied by using nicotine-adenine-dinucleotide phosphate-diaphorase histochemistry (shortly, diaphorase), a simple and robust procedure to characterize tissue distribution of nitric oxide-synthases. We assessed morphometric features of the diaphorase-positive cells in visual cortex, and the neuropil histochemical activity in hippocampal CA1 and dentate gyrus using densitometry analysis. In the large-litter group, the labeled-cell density in white matter of area 17 was higher, as compared to the small-litter group. There was a clear trend, in the large-litter group, to lower values of soma area, dendritic field and branches per neuron, but the differences were not significant. Densitometry analysis of hippocampus revealed a significant increase in the relative neuropil histochemical activity of the dentate gyrus molecular layer in the larger litters, which may be associated to increased compensatory blood flow in the hippocampus. The pathophysiological mechanisms of the observed changes remain to be investigated.

Animals↗
PubMed2004

Neuropil and neuronal changes in hippocampal NADPH-diaphorase histochemistry in the ME7 model of murine prion disease.

Nitric oxide (NO) has been implicated in neurotoxicity and cerebral blood flow changes in chronic neurodegeneration, but its activity in the mammalian prion diseases has not been studied in detail. Nicotine adenine dinucleotide phosphate (NADPH)-diaphorase (NADPH-d) histochemistry is a simple and robust histochemical procedure that allows localization of the tissue distribution of NO synthases. The aim of the present study is to assess whether NADPH-d histochemical activity is altered in the hippocampus in the ME7 model of prion disease in C57BL/6J mice. At early and late stages after the initiation of the disease we assessed features of the NADPH-d positive cells and the neuropil histochemical activity in CA1 and dentate gyrus using densitometric analysis. In C57BL/6J mice 13 weeks postinjection of the prion agent ME7, when behavioural changes first become apparent, neuropil NADPH-d histochemical staining increases, whereas at late stages it decreases dramatically. Both type I and type II NADPH-d positive cells were found to survive throughout the hippocampal formation into the late stages of the disease, but diaphorase activity was reduced in dendritic branches and abnormal varicosities were present in both dendritic and axonal processes of NADPH-d positive type I cells. The pathophysiological implications of the results remain to be investigated but both blood flow alteration and NO neurotoxicity may be features of the disease.

Algorithms↗
PubMed1996

Differential glutamatergic innervation in cytochrome oxidase-rich and -poor regions of the macaque striate cortex: quantitative EM analysis of neurons and neuropil.

One of the hallmarks of the primate striate cortex is the presence of cytochrome oxidase (CO)-rich puffs and CO-poor interpuffs in its supragranular layers. However, the neurochemical basis for their differences in metabolic activity and physiological properties is not well understood. The goals of the present study were to determine whether CO levels in postsynaptic neuronal compartments were correlated with the proportion of excitatory glutamate-immunoreactive (Glu-IR) synapses they received and if Glu-IR terminals and synapses in puffs differed from those in interpuffs. By combining CO histochemistry and postembedding Glu immunocytochemistry on the same ultrathin sections, the simultaneous distribution of the two markers in individual neuronal profiles was quantitatively analyzed. As a comparison, adjacent sections were identically processed for the double labeling of CO and GABA, an inhibitory neurotransmitter. In both puffs and interpuffs, most axon terminals forming asymmetric synapses (84%)--but not symmetric ones, which were GABA-IR--were intensely immunoreactive for Glu. GABA-IR neurons received mainly Glu-IR synapses on their cell bodies, and they had three times as many mitochondria darkly reactive for CO than Glu-rich neurons, which received only GABA-IR axosomatic synapses. In puffs, GABA-IR neurons received a significantly higher ratio of Glu-IR to GABA-IR axosomatic synapses and contained about twice as many darkly CO-reactive mitochondria than those in interpuffs. There were significantly more Glu-IR synapses and a higher ratio of Glu- to GABA-IR synapses in the neuropil of puffs than of interpuffs. Moreover, Glu-IR axon terminals in puffs contained approximately three times more darkly CO-reactive mitochondria than those in interpuffs, suggesting that the former may be synaptically more active. Thus, the present results are consistent with our hypothesis that the levels of oxidative metabolism in postsynaptic neurons and neuropil are positively correlated with the proportion of excitatory synapses they receive. Our findings also suggest that excitatory synaptic activity may be more prominent in puffs than in interpuffs, and that the neurochemical and synaptic differences may constitute one of the bases for physiological and functional diversities between the two regions.

Animals↗
PubMed1996

Ultrastructure of the stomatogastric ganglion neuropil of the crab, Cancer borealis.

The stomatogastric ganglion (STG) of the crab, Cancer borealis, contains the neural networks responsible for rhythmic pattern generation of the foregut. Neuron counts indicate that the STG of C. borealis has 25-26 neurons, 4-5 fewer than that found in lobsters. We describe the ultrastructural features of the ganglion by focusing on those that may be involved in storage, release, or range of action of peptide modulators, including a lacunar system and multiple types of intercellular junctions. In the neuropil, we identify five synaptic profile classes that contain the invertebrate presynaptic apparatus (dense bars, small clear vesicles), two of which also contain dense core (modulator-containing) vesicles. These latter two are comprised of multiple immunocytochemical classes that are not easily distinguished by structural criteria. In addition, we find neurohemal-like profiles that contain primarily dense core vesicles. Our finding that multiple profile types in the STG possess modulator-containing vesicles coincides with immunocytochemical results better than do previous ultrastructural studies that report only one such profile type. We show that a single modulatory input, stomatogastric nerve axon 1, makes only classical synapses and not neurohemal-like profiles, although some modulators are found in both these profile types. These data provide the groundwork for understanding the architecture of modulatory input-target interactions and suggest ways that the specificity of modulatory effects within a complex neuropil may be attained.

Animals↗
PubMed1997

Genetic influences on cellular reactions to brain injury: activation of microglia in denervated neuropil in mice carrying a mutation (Wld(S)) that causes delayed Wallerian degeneration.

This study examines the relationship between the appearance of degenerative changes in synaptic terminals and axons and the activation of microglia in denervated neuropil regions of normal mice of the C57BL/6 strain and mutant mice (Wld(S)), in which Wallerian degeneration is substantially delayed. The time course of degenerative changes in synaptic terminals and axons was assessed using selective silver staining. Microglial cells were identified by immunostaining for Mac-1, a monoclonal antibody to the CR3 complement receptor, and by histochemical staining for nucleoside diphosphatase (NDPase). Increased argyrophilia, indicative of degenerative changes, was evident as early as 1 day postlesion in normal mice, but was not seen until 6-8 days in mice with the Wld(S) mutation. Microglial activation in normal C57BL/6 mice was evident by 24 hours postlesion, as evidenced by increased immunostaining for Mac-1, increased histochemical staining for NDPase, and morphological changes indicative of an activated phenotype (short, thick processes). Quantitative evaluation of immunostaining for Mac-1 revealed that peak activation occurred between 2 and 6 days postlesion with a return to a quiescent phenotype by 12 days. In contrast, the microglial response was significantly delayed and prolonged in mice bearing the Wld(S) mutation. Activated microglia were not seen within the deafferented area until 6 to 8 days postlesion and peak activation occurred between 12 and 20 days postlesion. These data suggest that the response of microglia in denervated neuropil zones is triggered by the same types of degenerative changes that cause increased argyrophilia as detected by selective silver staining methods.

Acid Anhydride Hydrolases↗
PubMed1999

Daily rhythmic changes of cell size and shape in the first optic neuropil in Drosophila melanogaster.

Daily rhythms of changes in axon size and shape are seen in two types of monopolar cell-L1 and L2-that are unique cells within each of the modules or cartridges of the first optic neuropil or lamina in the fly's optic lobe. In the fruit fly Drosophila, L1 and L2's axons swell at the beginning of both day and night, with larger size increases occurring at the beginning of night. Later, they shrink during the day and night, respectively. Simultaneously, they change shape from an inverted conical form during the day to a cylindrical one at night. This is because the axonal cross section of L1 increases during the night, especially at proximal depths of the lamina, closest to the brain, whereas the axon of L2 increases in size at distal lamina depths. The cross-sectional areas of the L1 cell and of an individual cartridge both change under constant darkness (DD), indicating the circadian origin of changes observed under day/night (LD) conditions. We sought to see whether such changes impart a net change to the entire lamina's volume or shape that is visible by light microscopy, but oscillations in the volume or the curvature of the whole lamina neuropil are found neither in LD nor in DD. These size changes are discussed in relation to previous findings in the housefly Musca, with respect to differences in L1 and L2 between the two species, and to differences in the time course of their circadian changes.

Animals↗
PubMed2007

Neuronal morphology and neuropil structure in the stomatogastric ganglion of the lobster, Homarus americanus.

The stomatogastric nervous system (STNS) has long been used as a model system for the study of central pattern generation, neuromodulation, and network dynamics. Anatomical studies of the crustacean stomatogastric ganglion (STG) in different species have mostly been restricted to subsets of neurons and/or general structural features. For the first time, we describe the morphology of all STG neurons belonging to the two circuits that produce the well-described pyloric and gastric rhythms in the lobster, Homarus americanus. Somata sit on the dorsal and lateral surface of the STG and send a single primary neurite into the core of the neuropil, which is mostly made up of larger lower order branches. The perimeter of the neuropil consists mostly of finer higher order branches. Immunohistochemical labeling for synaptic proteins is associated with the small diameter branches. Somata positions are not constant but show preferred locations across individuals. The number of copies is constant for all neuron types except the PY and GM neurons (PY neuron number ranges from 3 to 7, and GM neuron number ranges from 6 to 9). Branch structure is largely nondichotomous, and branches can deviate substantially from cylindrical shape. Diameter changes at branch points can be as large as 20-fold. Clearly, the morphology of a specific neuron type can be quite variable from animal to animal.

Animals↗
PubMed1975

Origin of specific synaptic types in the motoneuron neuropil of the monkey.

An attempt has been made to establish, so far as possible, the specific types of synaptic bulbs on motoneurons which arise from extrinsic (descending and dorsal root) and intrinsic sources (interneurons). This has involved the laborious analysis of thousands of electron micrographs of material from normal motoneuron neuropil, and motoneuron neuropil of animals with lesions of spinal tracts, spinal roots, and motor cortex. Our studies have established that the large synaptic bulbs on dendrites, which are the only type in the spinal cord to possess pre-synaptic synapses (serial synapses), are derived from monosynaptic dorsal root fibers (R bulbs). The presynaptic component of the serial synapse appeared to degenerate at levels below spinal cord transections (P bulbs). By means of transections of spinal cord we have found that descending fibers in the spinal cord terminate as one of at least two distinct classes of synaptic bulbs, one with spheroid synaptic vesicles (S) and one with flattened vesicles (F). There is evidence from several sources that the first type may often be excitatory in function and the second type inhibitory. Only synaptic bulbs with spheroid vesicles show definite signs of degeneration after lesions of the opposite motor cortex. The large (L) synaptic bulbs on motoneuron somata, associated with subsynaptic cisterns, are clearly not derived from the descending systems, or from posterior root fibers.

Animals↗
PubMed1987

Development of synaptic connections between muscle sensory and motor neurons: anatomical evidence that postsynaptic dendrites grow into a preformed sensory neuropil.

The anatomical development of muscle sensory arbors and dendrites of brachial motoneurons in the spinal cord of the bullfrog was studied by labeling both types of cells with horseradish peroxidase. Sensory and motoneurons were labeled in tadpoles (stages XV-XVIII) by backfilling the triceps nerve in vivo with HRP throughout the stages in development when functional monosynaptic connections between these cells are first being formed. Individual triceps motoneurons were injected with HRP in other tadpoles at the same developmental stages. By stage XV, triceps sensory afferents already projected to and arborized in the ventral sensory neuropil region of the spinal cord where sensory-motor connections are made. In contrast, the dendrites of triceps motoneurons rarely were present in this region until stage XVI. By stage XVII, triceps dendrites in this region were common and they intermingled with the collaterals of muscle sensory axons. Thus, sensory axons supplying limb muscles grow into the future neuropil region well in advance of the arrival of motoneuronal dendrites. Electrophysiological studies have shown that the connections between triceps sensory and motor neurons are already specific at stage XVII, as soon as monosynaptic potentials between these cells can be detected (Frank and Westerfield: J. Physiol. (Lond.) 343:593-610, '83). The present anatomical results demonstrate that the processes of sensory and motor cells are not in close anatomical proximity before this time; thus the selection of appropriate synaptic partners must occur from the outset.

Animals↗
PubMed1990

Electrophysiological measurements of volume changes in leech neuropile glial cells.

Double-barrelled microelectrodes, sensitive to quaternary ammonium ions, were used for simultaneous measurements of the intracellular free concentrations of choline ([Ch]i) or tetramethylammonium ([TMA]i) as well as membrane potential (Em) in neuropile glial cells of the leech, Hirudo medicinalis. Bath application of Ch or TMA (5 mM, 1 min) resulted in a transient membrane depolarization accompanied by a long-lasting (0.5-1 h) intracellular accumulation of these compounds to levels of between 5 and 15 mM. Changes in [Ch]i or [TMA]i were used for the calculation of changes in relative cell volume. Elevation of the extracellular K+ concentration [( K+]e) from 4 to 9, 15, 21, 27.5, or 40 mM elicited a membrane depolarization and a reversible cell swelling by about 7.5, 14, 18.5, 27 and 50%, whereas reduction of [K+]e to 1.5 mM as well as bath application of serotonin (5-HT) produced a membrane hyperpolarization and a concomitant shrinkage by about 6 and 14.3%, respectively. The measured alterations in cell volume were compared with calculated data based on the assumption of an osmotic equilibrium disturbed by potential-dependent changes of the intracellular Cl- concentration. The results indicate, that K(+)- and serotonin-induced changes in the cell volume of the neuropile glial cells are due to passive KCl and water fluxes.

Animals↗
PubMed1993

Optic synapses are found in diencephalic neuropils before development of the tectum in Xenopus.

The position of the earliest optic synapses in Xenopus and the stage at which they developed were studied with the electron microscope after labelling of optic axons with horseradish peroxidase. In addition, tritiated thymidine autoradiography and bromodeoxyuridine immunohistology were used to identify the birth dates of cells in the regions where the synapses had been found. The earliest mature optic synapses were found in the mid-diencephalic region, where the major diencephalic optic neuropils were beginning to develop. These synapses were seen at stage 35/36, before cells in the tectal precursor region had become postmitotic. In other animals labelling with tritiated thymidine or bromodeoxyuridine showed that cells in the diencephalon, close to where the synapses had been seen, were becoming postmitotic at the time the earliest optic axons arrived. The first optic synapses to form in the developing Xenopus visual system thus appeared to do so in the neuropil of Bellonci and the rostral visual nucleus.

Animals↗
PubMed1986

Differential immunocytochemical staining for glial fibrillary acidic (GFA) protein, S-100 protein and glutamine synthetase in the rat subcommissural organ, nonspecialized ventricular ependyma and adjacent neuropil.

Antibodies raised against glial fibrillary acidic protein (GFA), S-100 protein (S100) and glutamine synthetase (GS) are currently used as glial markers. The distribution of GFA, S100 and GS in the ependyma of the rat subcommissural organ (SCO), as well as in the adjacent nonspecialized ventricular ependyma and neuropil of the periaqueductal grey matter, was studied by use of the immunocytochemical peroxidase-antiperoxidase technique. In the neuropil, GFA, S100 and GS were found in glial elements, i.e., in fibrous (GFA, S100) and protoplasmic astrocytes (S100, GS). The presence of S100 in the majority of the ventricular ependymal cells and tanycytes, and the presence of GFA in a limited number of ventricular ependymal cells and tanycytes confirm the glial nature of these cells. The absence of S100, GFA and GS from the ependymocytes of the SCO, which are considered to be modified ependymal cells, suggests either a non-astrocytic lineage of these cells or an extreme specialization of the SCO-cells as glycoprotein-synthesizing and secreting elements, a process that may have led to the disappearance of the glial markers.

Animals↗
PubMed1976

High voltage electron microscopy of the optic neuropile of the housefly, Musca domestica.

Synaptic cartridges of the first optic neuropile (lamina ganglionaris) of the housefly were examined by high voltage electron microscopy (HVEM). Stereo pairs (from thick, i.e., 0.25 mum, sections viewed at 1,000 kV) provided a three dimensional representation of cartridge neurons and clearly revealed the lateral spread, bifurcation and some functional associations of Type I (L1, L2) monopolar interneurons. Slightly proximal to cartridge neck level, pairs of retinular (R) axons made contact with each other and it appeared that R processes projected through the cleft between the Type I interneurons. No junctional modifications were seen between contiguous R axon terminals. The speculation was made that functional contact might exist between neighboring R axons prior to their extensive synapses with principal first order interneurons. Such alleged coupling between R axons would account for several electrophysiological findings from other laboratories. Modifications in EM technique applicable for HVEM were detailed. The value of obtaining thick serial sections and the use of the HVEM in expediting three dimensional reconstructions of neuropile were demonstrated.

Animals↗
PubMed1982

Ultrastructural pathology of the compound eye and optic neuropiles of the retinal degeneration mutant (w rdg BKS222) Drosophila melanogaster.

The compound eye and the two most distal optic neuropils (lamina ganglionaris and medulla externa) of the Drosophila mutant w rdg BKS222 were examined with transmission electron microscopes at conventional (60 kV) and high (0.8-1 MV) voltages. Eye tissue was sampled in the newly emerged and at 3, 7, and 21 days following eclosion. This mutant is known to show a light-induced degeneration of the peripheral retinular cells (R1-6); the spectral sensitivity is altered and the threshold is increased reflecting the function of the central cells (R7, 8) which do not degenerate. A totally normal appearing visual system (peripheral retina and optic neuropiles) was found in newly emerged adults. After 3 days the somata of some of the peripheral retinal cells are affected and all of their axons show degeneration. At one week the R1-6 pathology is well advanced in both somal and axonal regions. In affected cells the cytoplasm is more or less uniformly electron dense and contains liposomes, lysosome-like bodies, myeloid figures and vacuoles suggesting autophagy. Such cytoplasm (noted at 3 and 7 days post-eclosion) exhibits an electron dense reticulum and degenerate mitochondria. Microvilli become more electron dense. Retinular axon terminals are electron opaque and lack synaptic vesicles with few if any presynaptic structures. Mitochondrial remains are barely recognizable. Transsynaptic degeneration was not found. After 3 weeks, the structure of R1-6 in the peripheral retina (somata and rhabdomeres) is greatly reduced or lost while R7 and R8 and higher order neurons are not affected. The debris from cell bodies and axon terminals or R1-6 seems diminished, so that some phagocytosis probably takes place along with gliosis in the lamina.

Animals↗
PubMed1984

Immunocytochemistry and ultrastructure of the neuropil located ventral to the rat supraoptic nucleus.

The neuropil located ventral to the SON was investigated by the use of immunoperoxidase staining for neurophysins, oxytocin and vasopressin, and electron microscopy. The study was performed in six groups of rats: 1) control; 2) infusion of isotonic saline into the CSF; 3) infusion of hypertonic saline into the CSF; 4) drinking hypertonic saline for 4 days; 5) same as group 4 but injection of colchicine into the CSF on second day of dehydration; 6) salt loading for 3 months. In the control rats the ventral neuropil contained a few immunoreactive processes, the general morphology of which was completely different from that of the neurosecretory axons emerging from the SON at its dorsal aspect. In rats of groups 3 to 6 the ventral processes (VP) became loaded with neurosecretory granules, whereas the perikarya and axons were depleted. Based on their general morphology and reactivity pattern it is suggested that the VP are dendrites. Most of these "dendrites" were embedded in a glial cushion formed by the processes of a particular type of marginal glia. Some of these "dendrites" enveloped an arteriole penetrating the optic tract. All VP were rich in synaptic contacts. The possibility that the VP of neurosecretory cells may be functionally related to the subarachnoid CSF and the arteriolar blood flow is discussed.

Animals↗