Organotypic Cultures as a Model of Parkinson´s Disease. A Twist to an Old Model

Organotypic cultures from the ventral mesencephalon (VM) are widely used to model Parkinson’s disease (PD). In this method, neurotoxic compounds have traditionally been applied to the media to induce a uniform dopaminergic (DAergic) cell death in the tissue slices, regardless of the variation existing among slices. This study demonstrates a refinement of the toxic induction technique. We show that unilateral application of 6-hydroxydopamine (6-OHDA) at the tissue surface by means of a microelectrode causes a precisely localized cell death that closely resembles an in vivo stereotactic model. This technique introduces an internal control that accounts for variation between slices and enables a precise quantification of the cell loss due to the toxin in use. We characterized organotypic VM cultures in terms of effects of 6-OHDA toxicity and number of DAergic neurons as judged by immunofluorescence and Western blots. Our findings illustrate that this new application technique greatly improves the representativeness of organotypic cultures as a model for PD

Local impact of perivascular plaques on cerebral blood flow dynamics in a transgenic mouse model of Alzheimer's disease.

Cerebrovascular pathology is closely coupled to cognitive function decline, as indicated by numerous studies at the system level. To better understand the mechanisms of this cognitive decline it is important to resolve how pathological changes in the vasculature - such as perivascular plaques - affect local cerebral blood flow dynamics. This issue is ideally studied in the intact brain at very high spatial resolution. Here, we describe initial results obtained by an approach based on in vivo observation by multi-photon microscopy of vascular plaques and local blood flow measurements in a transgenic mouse model engineered to express the human amyloid precursor protein with the Swedish and Arctic mutations. These mice exhibit a striking abundance of perivascular plaques in the cerebral cortex and are well suited to investigate vascular pathology in Alzheimer's disease

Widespread distribution of lymphatic vessels in human dura mater remote from sinus veins

Background and purpose: Previous experimental studies have shown that meningeal lymphatic vessels are located primarily along the walls of the dural sinus veins. Whether they are more widespread throughout human dura mater has presently not been characterized. The present study explored in humans whether meningeal lymphatic vessels may be identified remote from the sinus veins and whether they differ in the various location of dura mater.Methods: We included 15 patients who underwent neurosurgery, in whom dura mater was removed as part of the planned procedure. Tissue was prepared for immunohistochemistry using the lymphatic endothelial cell markers lymphatic vessel endothelial hyaluronan receptor 1 protein (LYVE-1), podoplanin and vascular endothelial growth factor receptor 3 (VEGFR3).Results: Lymphatic endothelial cell positive cells were found in dura mater at the posterior fossa (n = 8), temporal skull base (n = 5), frontal convexity (n = 1), and cranio-cervical junction (n = 1). They were most commonly seen remote from blood vessels, but also occurred along blood vessels, and seemed to be most abundant at the skull base.Conclusion: The present observations show that human lymphatic vessels are widespread in dura mater, not solely lining the dural sinuses

Organotypic Cultures as a Model of Parkinson´s Disease. A Twist to an Old Model

Organotypic cultures from the ventral mesencephalon (VM) are widely used to model Parkinson's disease (PD). In this method, neurotoxic compounds have traditionally been applied to the media to induce a uniform dopaminergic (DAergic) cell death in the tissue slices, regardless of the variation existing among slices. This study demonstrates a refinement of the toxic induction technique. We show that unilateral application of 6-hydroxydopamine (6-OHDA) at the tissue surface by means of a microelectrode causes a precisely localized cell death that closely resembles an in vivo stereotactic model. This technique introduces an internal control that accounts for variation between slices and enables a precise quantification of the cell loss due to the toxin in use. We characterized organotypic VM cultures in terms of effects of 6-OHDA toxicity and number of DAergic neurons as judged by immunofluorescence and Western blots. Our findings illustrate that this new application technique greatly improves the representativeness of organotypic cultures as a model for PD.We characterized organotypic VM cultures in terms of effects of 6-OHDA toxicity and number of DAergic neurons as judged by immunofluorescence and Western blots. Our findings illustrate that this new application technique greatly improves the representativeness of organotypic cultures as a model for PD

Effects of animal manure application on springtails (Collembola) in perennial ley

The density and diversity of springtails (Collembola) in the upper soil layer (0–3.8 cm) were studied in a perennial grass-clover ley in NW Norway during April–June 2012. The study was part of a field experiment comparing yields and soil characteristics after application of non-digested slurry (NS) versus anaerobically digested slurry (DS) from dairy cows. In total for three sampling dates, 39 species of springtails were identified. In the Control plots receiving no manure, the density level was around 30 000 individuals (ind.) m2 throughout the whole season. Three days after slurry application (40 t ha -1), the density of springtails had dropped significantly; from 55 214 to 7410 ind. m2 in the NS treatment and from 41 914 to 10 260 ind. m2 in the DS treatment. After 7 weeks the densities had increased again to 54% and 38% of the initial levels in NS and DS treatments, respectively. The springtails were divided into two ecological groups based on morphology and colour. The epigeic group comprised surface-dwelling species with eye organs and pigmentation. The endogeic group comprised soil-dwelling species lacking eye organs and pigmentation, and generally with shorter extremities than those found in the epigeic group. The negative effect of manure application on density was more severe and long-lasting in the epigeic than in the endogeic group. This effect was similar for both manure types. One species (Parisotoma notabilis) comprised 50% of the epigeic population, while three Mesaphorura spp. and Stanaphorura lubbocki comprised half the endogeic population. In general, the community structure, described by the relative abundance of each species, was more affected by manure application in the epigeic than in the endogeic group. Hence, slurry application seemed to affect surface-dwelling species more negatively than soil dwelling species, even within the small sampling depth used here. The density of endogeic species seemed to recover faster than the density of the epigeic species. A simplified classification of epigeic and endogeic springtails, based on the presence or absence of pigmentation and eyes, may be useful in studies of soil springtails where identification of the actual species is not the primary purpose

Analyzing microglial-associated Aβ in Alzheimer’s disease transgenic mice with a novel mid-domain Aβ-antibody

The mechanisms of amyloid-β (Aβ)-degradation and clearance in Alzheimer’s disease (AD) pathogenesis have been relatively little studied. Short Aβ-fragments form by enzymatic cleavage and alternate amyloid-beta precursor protein (APP)-processing. Here we characterized a novel polyclonal Aβ-antibody raised against an Aβ mid-domain and used it to investigate microglial Aβ-uptake in situ by microscopy at the light- and ultrastructural levels. The rabbit Aβ-mid-domain antibody (ab338), raised against the mid-domain amino acids 21–34 (Aβ21–34), was characterized with biochemical and histological techniques. To identify the epitope in Aβ recognized by ab338, solid phase and solution binding data were compared with peptide folding scores as calculated with the Tango software. The ab338 antibody displayed high average affinity (KD: 6.2 × 10−10 M) and showed preference for C-terminal truncated Aβ-peptides ending at amino acid 34 and Aβ-mid domain peptides with high scores of β-turn structure. In transgenic APP-mouse brain, ab338 labelled amyloid plaques and detected Aβ-fragments in microglia at the ultra- and light microscopic levels. This reinforces a role of microglia/macrophages in Aβ-clearance in vivo. The ab338 antibody might be a valuable tool to study Aβ-clearance by microglial uptake and Aβ-mid-domain peptides generated by enzymatic degradation and alternate production