Biomedical subjects

F Eckenstein

Publications and source records attributed to F Eckenstein.

At least 19 recordsLinked to original sources

PubMed2001

Beta-amyloid plaques induce neuritic dystrophy of nitric oxide-producing neurons in a transgenic mouse model of Alzheimer's disease.

A causative role for nitric oxide has been postulated in a number of neurodegenerative diseases. Using histochemical and immunohistochemical methods, we examined the effect of beta-amyloid plaques on nitric oxide-producing cells in transgenic mice which overexpress a mutant human amyloid precursor protein (APP). In 14-month-old animals, nitric oxide synthase (NOS)-positive dystrophic neurites were observed frequently in the cerebral cortex and hippocampus of all of 16 plaque-bearing transgenic animals and in none of 16 wild-type animals. Double labeling of NOS and beta-amyloid revealed that 90% of beta-amyloid plaques were associated with NOS-containing dystrophic neurites. In 7-month-old animals, beta-amyloid plaques were very rare, but those present were frequently associated with NOS-positive neuritic dystrophy. We conclude that beta-amyloid plaques induce neuritic dystrophy in cortical neurons containing NOS in this model of AD, and hypothesize that this finding may be relevant to the mechanism of beta-amyloid neurotoxicity in human AD.

Alzheimer Disease↗

PubMed2000

Lack of interleukin-6 expression is not protective against focal central nervous system ischemia.

BACKGROUND AND PURPOSE: Interleukin-6 (IL-6) appears to be involved in the inflammatory response associated with central nervous system (CNS) ischemia. Although IL-6 levels increase after stroke, it is not known whether IL-6 directly influences CNS ischemic injury. In this study, we used a focal reversible stroke model to investigate whether mice lacking IL-6 were protected against acute ischemic injury. METHODS: We bred IL-6-deficient C57 black mice (I-129 IL-6 KO back-crossed with C57), including homozygous knockouts (IL-6 -/-), heterozygous littermates (IL-6 +/-), and normal littermates (IL-6 +/+). The status of all animals was confirmed by DNA sampling and polymerase chain reaction analysis. Reversible middle cerebral artery occlusion was produced by advancing a silicone-coated 8-0 filament into the internal carotid artery for 2 hours (experiment 1) or 45 minutes (experiment 2). At 24 hours, animals were evaluated on a 28-point clinical scale, blood and cerebrospinal fluid were obtained, and the brains were evaluated for infarct volume and IL-6 mRNA levels. RESULTS: In experiment 1 (severe ischemia), no differences were seen in lesion size or neurological function between the groups: lesion volume was IL-6 -/- (n=15), 57+/-13 mm(3); IL-6 +/- (n=15), 58+/-23 mm(3); and IL-6 +/+ (n=15), 58+/-18 mm(3) (P=NS). ELISA testing confirmed very low to absent levels of IL-6 in the serum and cerebrospinal fluid of knockout animals. Brain mRNA levels of the other proinflammatory cytokines, including tumor necrosis factor-alpha, IL-1beta, and IL-1 receptor antagonist, were 50% lower in IL-6-deficient ischemic animals than in normal animals. In experiment 2 (mild ischemia), no differences were seen in lesion size or neurological function between the groups: lesion volume was IL-6 -/- (n=10), 16+/-8 mm(3); IL-6 +/- (n=10), 14+/-4 mm(3); and IL-6 +/+ (n=10), 19+/-12 mm(3) (P=NS). CONCLUSIONS: In this study, infarct size and neurological function at 24 hours were not different in animals deficient in IL-6 after transient CNS ischemia. This suggests that IL-6 does not have a direct influence on acute ischemic injury. Further study investigating the role of IL-6 on long-term recovery after stroke is in progress.

Acute Disease↗

PubMed1999

Time course of IL-6 expression in experimental CNS ischemia.

Interleukin-6 (IL-6) appears to be an important modulator of the inflammatory response associated with CNS ischemia. Clinically, IL-6 values obtained in the first week post-stroke have been shown to correlate with infarct size and outcome. In this study we used a focal reversible stroke model to investigate the time course and relationship to outcome of IL-6 production in plasma, brain and CSF. Reversible middle cerebral artery occlusion or sham surgery was produced in 50 adult Swiss Webster mice by advancing an 8-0 filament into the internal carotid artery for 2 h (sham 1 min). At 3, 6, 12, 24, and 72 h (8 each ischemia; 2 each sham) groups of animals were evaluated on a 28 point clinical scale, blood and CSF obtained, and the brains were evaluated for infarct volume and IL-6 mRNA levels. Serum levels of IL-6 (ELISA mean +/- SD; undetectable in controls) overall sham group, 102 +/- 87; 3 h, 908 +/- 494* pg ml-1; 6 h, 1079 +/- 468* pg ml-1; 12 h, 980 +/- 221* pg ml-1; pg ml-1; 24 h, 320 +/- 314* pg ml-1; 72 h, 20 +/- 30* pg ml-1 (*p < or = 0.05 to sham). CSF levels (ELISA) overall sham group, 10 +/- 18; 3 h, 379 +/- 210* pg ml-1; 6 h, 157 +/- 61* pg ml-1; 12 h, 136 +/- 88* pg ml-1; 24 h, 127 +/- 99 pg ml-1; 72 h, 72 +/- 9* pg ml-1 (*p < or = 0.05 to sham). Brain IL-6 mRNA levels overall sham group, 20; 3 h, 480; 6 h, 599; 12 h, 7960; 24 h, 20267; 72 h, 0. There was an overall R2 of 0.20 between plasma and CSF IL-6. There was an overall R2 of 0.13 and 0.20 between infarct size and serum and CSF IL-6 level respectively, and an overall R2 of 0.10 and 0.17 between neurologic function and serum and CSF IL-6 level respectively. These findings confirm that IL-6 values increase following CNS ischemia with peak serum and CSF levels occurring before brain values. CSF IL-6 levels had a stronger correlation with neurologic function and infarct size than serum.

Age Factors↗

PubMed1997

Monofilament intraluminal middle cerebral artery occlusion in the mouse.

The rat middle cerebral artery (MCA) occlusion model with an intraluminal filament is well characterized with a two hour period of occlusion in widespread use. The recent availability of transgenic animals has led to an interest in adapting the MCA model in the mouse. To date the model has not been well characterized in the mouse. We performed the present study to compare different durations of MCA occlusion and to validate new functional assessments in this model. The MCA occlusion model (5-0 filament) was used. Swiss-Webster mice, 24-44 g, were randomly assigned to four groups: one hour of occlusion; two hours of occlusion; three hours of occlusion; or permanent occlusion. At 48 hours post-ischemia, the animals were rated on three neurologic function scales, and then the brains were removed for lesion size determination. Overall, there was a significant difference in lesion volume (p < 0.001) between the groups. In the permanent group of mice, the average lesion volume was 78.41 +/- 17.47 mm (n = 12); two and three hours of ischemia produced 51.29 +/- 29.82 mm3 (n = 11) and 54.85 mm3 (n = 13), respectively, significantly different than the one hour group 14.84 +/- 31.34 mm3 (n = 11). All three functional scoring systems found significant overall differences between the four groups with our detailed General and Focal scores producing more robust between group treatment differences and showing correlation coefficients of r = 0.766 and r = 0.788, respectively to infarct volume. The MCA filament occlusion model can be successfully adapted in the mouse with either two or three hour occlusions producing reliable infarcts. New functional scoring systems unique to the mouse appear to add additional information.

Analysis of Variance↗

PubMed1991

Differential localization and possible functions of aFGF and bFGF in the central and peripheral nervous systems.

We investigated the relative distribution of acidic and basic FGF (aFGF and bFGF) in the nervous system of the rat, using a combination of biological, biochemical, immunochemical, and immunohistochemical methods that can differentiate unambiguously between aFGF and bFGF. We found that different regions of the nervous system contained varying levels of aFGF and bFGF. In the central nervous system, bFGF was present nearly exclusively in astrocytes. Most neurons did not contain detectable amounts of bFGF immunoreactivity, with the notable exception of pyramidal cells in hippocampal area CA2. Interestingly, bFGF immunoreactivity was localized to the nucleus of both CA2 neurons and astrocytes. Astrocytes in vitro were also found to express bFGF, whereas cortical neurons in culture did not contain detectable amounts of bFGF. Transection of the optic nerve led to an approximately twofold increase of bFGF in the distal stump, which is consistent with the observation that bFGF is expressed by astrocytes. Transection of rat and chicken sciatic nerve resulted in a rapid and complete disappearance of aFGF from the distal nerve stump, suggesting that aFGF is present in axons projecting through the sciatic nerve. We observed, in agreement with this notion, that cultured sensory neurons contain reasonably high levels of FGF-like bioactivity. Similar levels of activity were found in developing sciatic nerve, suggesting that neuronal aFGF might be involved in regulating the development of the peripheral nervous system.

Animals↗

PubMed1988

Transient expression of NGF-receptor-like immunoreactivity in postnatal rat brain and spinal cord.

The pattern of expression of nerve growth factor (NGF)-receptor-like immunoreactivity during postnatal development in rat central nervous system (CNS) was analyzed using immunohistochemical localization of the receptor. Interestingly, in addition to the expected staining in basal forebrain, several structures were strongly labelled only during specific postnatal developmental stages. These structures included fibers in the thalamus, the external granule cell layer of the cerebellum and motor neurons, indicating that specific neurotrophic mechanisms might play an important role for the labelled cells during a precisely defined period.

Aging↗

PubMed1987

Choline acetyltransferase immunocytochemistry of Edinger-Westphal and ciliary ganglion afferent neurons in the cat.

The distribution of cholinergic neurons in the region of the cat Edinger-Westphal nucleus (EW) was determined by immunocytochemical localization of the acetylcholine-synthesizing enzyme choline acetyltransferase (ChAT). Neurons containing ChAT-like immunoreactivity (ChAT-LI) were densely distributed within EW, the anteromedian nucleus (AM), and the oculomotor nucleus (III), and were also present in immediately adjacent regions of the periaqueductal gray and ventral tegmental region. The majority of labelled neurons in EW and AM showed a markedly lower intensity of ChAT-LI than the labelled neurons in III and adjacent regions. To determine the relationship of cells with ChAT-LI to the distribution of ciliary ganglion afferent neurons, a double labelling immunocytochemistry/retrograde transport technique was also used. These experiments showed that many of the cells located outside of III that stained intensely for ChAT-LI project to ciliary ganglion. Very few ciliary ganglion afferent neurons were found in EW or AM itself; instead, the distribution of lightly labelled ChAT-LI-positive neurons in EW and AM more closely matched the known distribution of peptide-containing cells that have descending, central projections.

Afferent Pathways↗

PubMed1987

Cholinergic innervation of the optic tectum in the frog Rana pipiens.

An immunohistochemical method for choline acetyltransferase (ChAT) identifies presumably cholinergic axons in two retino-receptive laminae in the optic tectum of the frog Rana pipiens. Following eye enucleation there is no loss of immunoreactive axons in the optic tectum. Following unilateral ablation of the nucleus isthmi there is a near-total loss of ChAT-positive axons in the superficial cholinergic lamina contralaterally and in the deeper cholinergic lamina ipsilaterally. Thus, the cholinergic innervation of the tectum appears to derive from the nucleus isthmi. However, ChAT-positive staining of the basal optic nucleus does depend upon an intact retinal input and could derive from either retinal axons or some system trophically dependent on them.

Animals↗

PubMed1987

Neostriatal cholinergic neurons receive direct synaptic inputs from dopaminergic axons.

We used an electron microscopic 'mirror technique' to determine whether cholinergic neurons are in direct synaptic contact with dopaminergic axons in the rat neostriatum. Tyrosine hydroxylase-immunoreactive axons make synaptic contacts with the somata and proximal dendrites of large choline acetyltransferase-immunoreactive striatal neurons which are thought to be interneurons. This provides morphological evidence that nigrostriatal dopaminergic neurons can influence monosynaptically the striatal cholinergic neurons.

Adrenergic Fibers↗

PubMed1986

GABAergic input to cholinergic forebrain neurons: an ultrastructural study using retrograde tracing of HRP and double immunolabeling.

Amygdalopetal cholinergic neurons in the ventral pallidum were identified by combining choline acetyltransferase (ChAT) immunohistochemistry with retrograde tracing of horseradish peroxidase (HRP) following injections of the tracer in the basolateral amygdaloid nucleus. Although ChAT-positive terminals were identified in the ventral pallidum, they were never seen in contact with either immunonegative or ChAT-positive amygdalopetal neurons. In material, in which immunostaining against glutamic acid decarboxylase (GAD), the synthesizing enzyme for GABA was combined with retrograde tracing of HRP from the basolateral amygdaloid nucleus, GAD-positive terminals were seen to contact immunonegative amygdalopetal neurons. In addition, when sections of the rostral forebrain were processed, first to preserve and identify the transported HRP, and then were sequentially tested for both ChAT and GAD immunohistochemistry with the immunoperoxidase reaction for both tissue antigens, GAD-immunopositive terminals were seen to make synaptic contacts with cholinergic amygdalopetal neurons. These results suggest that amygdalopetal, presumably cholinergic, neurons receive GAD-positive terminals. In separate experiments using immunoperoxidase for ChAT and ferritin-avidin for GAD labeling, we confirmed the presence of GAD-containing terminals on cholinergic neurons. In addition, cholinergic terminals were seen in synaptic contact with GAD-positive cell bodies. These morphological studies suggest that direct GABAergic-cholinergic and cholinergic-GABAergic interactions take place in the rostral forebrain.

Amygdala↗

PubMed1986

Cholinergic neurons and fibres in the rat visual cortex.

Choline acetyltransferase (ChAT), the acetylcholine synthesizing enzyme, was localized immunocytochemically in neurons and fibres in the rat visual cortex using a monoclonal antibody. ChAT-labelled cells were non-pyramidal neurons, primarily of the bipolar form, distributed in layers II through VI but concentrated in layers II & III. Their perikarya contained a large nucleus and a small amount of perinuclear cytoplasm. The somata and dendrites of all labelled cells received Gray's type I and type II synapses. ChAT-stained axons formed a dense and diffuse network throughout the visual cortex and particularly in layer V. Electron microscopy revealed that the great majority formed type II synaptic contacts with dendrites of various sizes, unlabelled non-pyramidal somata and, on a few occasions, with ChAT-labelled cells. However, a very small number of terminals appeared to form type I synaptic contacts. This study describes the morphological organization of the cholinergic system in the visual cortex, the function of which has been under extensive investigation.

Animals↗

PubMed1986

Morphological characterization of cholinergic neurons in the horizontal limb of the diagonal band of Broca in the basal forebrain of the rat.

A monoclonal antibody against choline acetyltransferase (ChAT), the acetylcholine synthesizing enzyme, was used to identify cholinergic neurons in the nucleus of the horizontal limb of the diagonal band of Broca at the light and electron microscopic levels. ChAT-labelled somata were fusiform, triangular or round in shape and varied considerably in size. Depending on the type of the cell, one to four dendrites emerged from the soma, but an axon could rarely be seen. The nuclei of most cells were round or oval, showed invaginations and displayed prominent nucleoli. The karyoplasm of the larger fusiform and triangular neurons contained abundant organelles including parallel arrays of granular endoplasmic reticulum. The synaptic input to labelled perikarya and proximal dendrites was sparse. It consisted chiefly of asymmetrical synaptic contacts, sometimes with postjunctional densities, but a few symmetrical synapses were also noted. ChAT-positive axon terminals were not identified which suggests that axon collaterals are rare within the nucleus of the horizontal limb of the diagonal band of Broca.

Animals↗

PubMed1986

Inputs to motoneurones in the hypoglossal nucleus of the rat: light and electron microscopic immunocytochemistry for choline acetyltransferase, substance P and enkephalins using monoclonal antibodies.

Light and electron microscopic peroxidase-antiperoxidase immunocytochemistry has been used to localize choline acetyltransferase, substance P and enkephalin in the hypoglossal nucleus of the rat. Choline acetyltransferase immunoreactivity was observed in motoneurone cell bodies and proximal dendrites, in large varicosities in the surrounding neuropil and in nerve terminals in synaptic contact with immunostained motoneurones. Most choline acetyltransferase immunostained terminals which made synaptic contact with motoneurone cell bodies and proximal dendrites possessed prominent subsynaptic cisterns and belong to the terminal type referred to in the literature as C or L. Substance P and enkephalin immunoreactivity did not occur in motoneurones but was seen in fibres and synaptic terminals. Substance P immunoreactive fibres made multiple axosomatic contacts while enkephalin immunoreactive terminals made synaptic contact mainly with large and small dendrites. C terminals were not stained for either substance P or enkephalin. This study provides immunocytochemical support for the classic identification of hypoglossal motoneurones as cholinergic and in addition shows that these neurones are innervated by a number of morphologically and chemically distinct terminal types. C terminals have previously been shown to contain cholinesterase and our demonstration that these terminals contain choline acetyltransferase thus provides additional evidence for their cholinergic nature and for a cholinergic innervation of hypoglossal motoneurones. The origin of the immunoreactive terminals was not identified in this study but possible candidates include the raphe nuclei for substance P. and propriobulbar interneurones for choline acetyltransferase.

Animals↗

PubMed1985

Galanin-like immunoreactivity in cholinergic neurons of the septum-basal forebrain complex projecting to the hippocampus of the rat.

It is now well recognized that there are several groups of cholinergic neurons in the basal forebrain with direct projections to various cortical regions. Immunohistochemical investigations of the distribution of the neuropeptide galanin (GAL) have shown that two of these brain areas, the medial septum and diagonal band, contained large numbers of GAL-immunoreactive neurons. In the present study, double staining techniques using antibodies raised against choline acetyltransferase (ChAT) revealed that GAL- and ChAT-like immunoreactivities are colocalized within a subpopulation of the cholinergic neurons within the medial septum and diagonal band. This colocalization of GAL- and ChAT-immunoreactivities was not seen to occur within other groups of forebrain cholinergic neurons. Immunohistochemistry carried out subsequent to injections of fluorescent retrograde tracers into the hippocampal formation revealed that both ChAT/GAL- and ChAT-containing neurons project to the hippocampal formation. The question of GAL as a modulator of cholinergic transmission in this projection is discussed.

Animals↗

PubMed1985

Cholinergic projections from the midbrain and pons to the thalamus in the rat, identified by combined retrograde tracing and choline acetyltransferase immunohistochemistry.

The distribution of cholinergic neurons in the midbrain and pons which project directly to the thalamus was investigated in the rat using a procedure which allows the simultaneous detection of retrogradely transported horseradish peroxidase (HRP) and immunohistochemical demonstration of choline acetyltransferase (ChAT) in the same neurons. HRP injections were placed in the dorsal half of the anterior third of the thalamus on one side which included the anteroventral nucleus as well as portions of the rostral intralaminar and reticular nuclei. These thalamic nuclei showed the highest density of immunohistochemically detectable cholinergic fibers. Neurons containing both HRP and ChAT, which represented cholinergic neurons projecting directly to the thalamus, were found in the midbrain and pons in the lateral tegmental reticular formation, parabrachial region and lateral dorsal tegmental nucleus. Ipsilateral to the injection site over 91% of the HRP labeled neurons in all of these regions were cholinergic, while an average of 60% of the cholinergic neurons had transported HRP. Contralateral to the injection site 5-6% of the cholinergic neurons in these regions were also retrogradely labeled. These findings demonstrate direct cholinergic projections to the thalamus from neurons in several regions in the tegmentum and suggest that tegmental projections to the thalamus are predominantly cholinergic.

Animals↗

PubMed1985

Nerve growth factor increases choline acetyltransferase but not survival or fiber outgrowth of cultured fetal septal cholinergic neurons.

Neurons dissociated from the septal area of fetal rat brains were grown in culture. Cholinergic neurons were identified by immunocytochemical visualization of choline acetyltransferase and cytochemical demonstration of acetyl cholinesterase. Choline acetyltransferase immunocytochemistry stained cell bodies and proximal processes while acetylcholinesterase cytochemistry visualized the entire neuron. Choline acetyltransferase-positive neurons could only be identified in cultures grown under conditions that produced the maximal choline acetyltransferase activity, measured biochemically. All of the choline acetyltransferase-positive neurons were double stained for acetylcholinesterase while only 6% of the acetylcholinesterase-positive cells were choline acetyltransferase negative in these cultures. These results indicate that acetylcholinesterase is a reliable marker for cholinergic cells in cultures of dissociated septal neurons. Being the more sensitive method, acetylcholinesterase staining was therefore used to identify cholinergic cells in cultures with choline acetyltransferase levels insufficient for immunocytochemical visualization of this enzyme. Addition of nerve growth factor or antibodies to nerve growth factor to the medium did not affect the number of cholinergic neurons surviving in culture. Furthermore, nerve growth factor and anti-nerve growth factor failed to influence the general morphological appearance and the number of processes of these neurons. However, nerve growth factor elevated the biochemically measured activity of choline acetyltransferase up to two-fold. The nerve growth factor-mediated increase in choline acetyltransferase activity was dose dependent with an ED50 of 10 ng/ml (4 X 10(-10) M). The increase was highly specific for nerve growth factor. It was blocked by anti-nerve growth factor, and epidermal growth factor, insulin and other control proteins failed to exert a similar effect. Nerve growth factor had to be present for at least 3 days in the culture medium to increase choline acetyltransferase activity, suggesting that the increase was due to an elevated choline acetyltransferase synthesis rather than to an activation of the enzyme.

Acetylcholinesterase↗

PubMed1984

Immunocytochemical localization of putative cholinergic neurons in the goldfish retina.

The presence of putative cholinergic neurons in goldfish retina was demonstrated by immunocytochemical localization of choline acetyltransferase (ChAT), the synthesizing enzyme for acetylcholine. Four populations of ChAT-immunoreactive neurons were localized: two with cell bodies in the inner nuclear layer and two with cell bodies in the ganglion cell layer. The processes of these neurons ramified in lamina 2 and/or 4 (of 5) in the inner plexiform layer. These cell populations are comparable to populations of putative cholinergic neurons that have been identified by [3H]choline uptake [3, 10].

Acetylcholine↗

PubMed1984

Concurrent visualization of choline acetyltransferase-like immunoreactivity and retrograde transport of neocortically injected markers in basal forebrain neurons of cat and rat.

Magnocellular neurons in the basal forebrain of rats and cats were retrogradely labeled with Fast Blue or horseradish peroxidase injected into the neocortex. Using antisera against choline acetyltransferase (ChAT) a direct double-labeling technique was carried out and it was demonstrated that retrogradely transported markers and ChAT-like immunoreactivity occur within the same neurons. These findings strongly support the cholinergic nature of basal forebrain projection to the neocortex.

Animals↗