Tracing of the entorhinal-hippocampal pathway in vitro

Kluge, Annegret; Hailer, Nils P.; Horváth, Tamas L.; Bechmann, Ingo; Nitsch, Robert

doi:10.1002/(sici)1098-1063(1998)8:1<57::aid-hipo6>3.0.co;2-4

Cited by 41 publications

(27 citation statements)

References 34 publications

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“…Slices were isolated and prepared from C57BL/6 mice (P8 -P11) as described previously (Kluge et al, 1998). In brief, brains were rapidly removed under sterile conditions after decapitation and placed in ice-cold preparation medium (MEM, 1% image glutamine; Invitrogen, Germany) at pH 7.35.…”

Section: T-cell Stimulation and Flow Cytometry Splenocytes From Wt Omentioning

confidence: 99%

Cytotoxic CD8⁺T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death

Meuth¹,

Herrmann²,

Simon³

et al. 2009

J. Neurosci.

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Cytotoxic CD8ϩ T cells are considered important effector cells contributing to neuronal damage in inflammatory and degenerative CNS disorders. Using time-lapse video microscopy and two-photon imaging in combination with whole-cell patch-clamp recordings, we here show that major histocompatibility class I (MHC I)-restricted neuronal antigen presentation and T cell receptor specificity determine CD8 ϩ T-cell locomotion and neuronal damage in culture and hippocampal brain slices. Two separate functional consequences result from a direct cell-cell contact between antigen-presenting neurons and antigen-specific CD8 ϩ T cells. (1) An immediate impairment of electrical signaling in single neurons and neuronal networks occurs as a result of massive shunting of the membrane capacitance after insertion of channel-forming perforin (and probably activation of other transmembrane conductances), which is paralleled by an increase of intracellular Ca 2ϩ levels (within Ͻ10 min). (2) Antigen-dependent neuronal apoptosis may occur independently of perforin and members of the granzyme B cluster (within ϳ1 h), suggesting that extracellular effects can substitute for intracellular delivery of granzymes by perforin. Thus, electrical silencing is an immediate consequence of MHC I-restricted interaction of CD8 ϩ T cells with neurons. This mechanism is clearly perforin-dependent and precedes, but is not causally linked, to neuronal cell death.

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Section: T-cell Stimulation and Flow Cytometry Splenocytes From Wt Omentioning

confidence: 99%

Cytotoxic CD8⁺T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death

Meuth¹,

Herrmann²,

Simon³

et al. 2009

J. Neurosci.

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“…Anyhow, this method may allow to visualize growth of nerve fibers in a co-culture system. Indeed anterograde tracing of the entorhinal-hippocampal pathway in organotypic brain slices has already been demonstrated [9]. In addition to the staining of neurons, many astrocytes were labeled by Mini-ruby, which indicates that the uptake of Mini-ruby was not selective for neurons.…”

Section: Discussionmentioning

confidence: 97%

“…In this study, Mini-ruby, a fluorescently (tetramethylrhodamine) tagged biotinylated dextran amine with a size of 10,000 MW was used to study a tracer-positive cholinergic labeling in organotypic brain slices. Mini-ruby is a widely used neuronal tracer in vitro and in vivo [9,17] and function efficiently as anterograde or retrograde tracers. Mini-ruby is supplied in a crystaline form and is applied onto the slices very carefully at different spots.…”

Section: Discussionmentioning

confidence: 99%

“…Neurotracing techniques [8] are widely used to study axonal transport [9], guidance [10] regeneration [11] or sprouting [11][12][13]. However, no reports are available on the neurotracing of nBM cholinergic neurons.…”

Section: Introductionmentioning

confidence: 99%

“…However, no reports are available on the neurotracing of nBM cholinergic neurons. The present study uses a fluorescently tagged biotinylated dextran amine, Mini-ruby, which is widely used to trace neuronal projections in vitro and in vivo [9,17]. Mini-ruby is a 10,000 MW conjugate of tetramethylrhodamine and biotin with dextran.…”

Section: Introductionmentioning

confidence: 99%

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Mini-Ruby is Rapidly Taken up by Neurons and Astrocytes in Organotypic Brain Slices

Ullrich

Humpel

2011

Neurochem Res

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Cholinergic neurons are intensively studied, because they degenerate in Alzheimer's disease. Although neurotracer techniques are widely used to study axonal transport, guidance, regeneration or sprouting it is not clear if cholinergic neurons can be stained by tracer techniques and studied in brain slices. The aim of the present study was to evaluate the characteristics of the neurotracer Miniruby in organotypic brain slices of the basal nucleus of Meynert (nBM), focusing on cholinergic neurons. Miniruby is a biotinylated dextran amine and is taken up very fast by a variety of cells. When 2-week old nerve growth factor-incubated brain slices of the nBM were treated with Mini-ruby crystals for 1 h, only a few (2-3%) cholinergic neurons were clearly labeled as shown by co-localization with choline acetyltransferase. The staining was found in neuN-positive neurons and microtubule associated protein-2 (MAP-2)-positive nerve fibers. A very rapid dynamic change was observed in these labeled varicosities within seconds. However, Mini-ruby was taken up also by many glutamine synthethase-positive astrocytes. At the site of Mini-ruby application an intense CD11b-positive microglial staining was evident. In conclusion, neurons and astrocytes in organotypic brain slices can be labeled very fast with the fluorescent dye Mini-ruby which undergoes dynamic processes.

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Dose‐dependent short‐ and long‐term effects of ionizing irradiation on neural stem cells in murine hippocampal tissue cultures: neuroprotective potential of resveratrol

Prager¹,

Patties²,

Himmelbach³

et al. 2016

Brain and Behavior

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IntroductionRadiation therapy plays an essential role in the treatment of brain tumors, but neurocognitive deficits remain a significant risk, especially in pediatric patients. In recent trials, hippocampal sparing techniques are applied to reduce these adverse effects. Here, we investigate dose‐dependent effects of ionizing radiation (IR) on juvenile hippocampal neurogenesis. Additionally, we evaluate the radioprotective potential of resveratrol, a plant polyphenol recognized for its bifunctional tumor‐preventive and anticancer effects.MethodsOrganotypic entorhinal–hippocampal slice cultures from transgenic nestin‐CFPnuc C57BL/J6 mice, postnatal days 3–6, were irradiated on a X‐ray machine (4.5, 8, 12, and 16 Gy, single doses) after about 2 weeks. Nestin‐positive neural stem cells were counted at a confocal live imaging microscope 0, 2, 4, 14, 25, and 42 days after IR. Resveratrol (15 μmol/L) was added 2 hr before and 24 hr after IR. Proliferation and cell death were assessed by BrdU pulse label, 48 hr after and by propidium iodide staining 96 hr after IR. GFAP‐ and NeuN‐positive cells were counted 42 days after IR in cryosectioned immunofluorescence‐stained slices.ResultsThe observed age‐related changes of nestin‐positive stem cells in the organotypic slice culture model resembled the reduction of neural stem cells in vivo. IR (4.5–16 Gy) led to a dose‐dependent damage of the neural stem cell pool in the dentate gyrus. No recovery was seen within 42 days after doses from 4.5 Gy onward. The decline of nestin‐positive cells was paralleled by increased cell death and decreased proliferation. The number of GFAP‐positive cells was significantly enhanced. No significant change was detected in the overall NeuN‐positive cell population, whereas the number of newborn, NeuN/BrdU double‐positive neurons was reduced. Resveratrol treatment reversed the irradiation‐induced decline of neural stem cells.ConclusionThe neuroprotective action of resveratrol on irradiated hippocampal tissue warrants further investigation as a possible supplement to hippocampal sparing procedures.

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Tracing of the entorhinal-hippocampal pathway in vitro

Cited by 41 publications

References 34 publications

Cytotoxic CD8⁺T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death

Cytotoxic CD8⁺T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death

Mini-Ruby is Rapidly Taken up by Neurons and Astrocytes in Organotypic Brain Slices

Dose‐dependent short‐ and long‐term effects of ionizing irradiation on neural stem cells in murine hippocampal tissue cultures: neuroprotective potential of resveratrol

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Tracing of the entorhinal-hippocampal pathway in vitro

Cited by 41 publications

References 34 publications

Cytotoxic CD8+T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death

Cytotoxic CD8+T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death

Mini-Ruby is Rapidly Taken up by Neurons and Astrocytes in Organotypic Brain Slices

Dose‐dependent short‐ and long‐term effects of ionizing irradiation on neural stem cells in murine hippocampal tissue cultures: neuroprotective potential of resveratrol

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Product

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Cytotoxic CD8⁺T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death

Cytotoxic CD8⁺T Cell–Neuron Interactions: Perforin-Dependent Electrical Silencing Precedes But Is Not Causally Linked to Neuronal Cell Death