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A Reichenbach

Publications and source records attributed to A Reichenbach.

At least 19 recordsLinked to original sources
PubMed2007

Downregulation of Kir4.1 inward rectifying potassium channel subunits by RNAi impairs potassium transfer and glutamate uptake by cultured cortical astrocytes.

Glial cell-mediated potassium and glutamate homeostases play important roles in the regulation of neuronal excitability. Diminished potassium and glutamate buffering capabilities of astrocytes result in hyperexcitability of neurons and abnormal synaptic transmission. The role of the different K+ channels in maintaining the membrane potential and buffering capabilities of cortical astrocytes has not yet been definitively determined due to the lack of specific K+ channel blockers. The purpose of the present study was to assess the role of the inward-rectifying K+ channel subunit Kir4.1 on potassium fluxes, glutamate uptake and membrane potential in cultured rat cortical astrocytes using RNAi, whole-cell patch clamp and a colorimetric assay. The membrane potentials of control cortical astrocytes had a bimodal distribution with peaks at -68 and -41 mV. This distribution became unimodal after knockdown of Kir4.1, with the mean membrane potential being shifted in the depolarizing direction (peak at -45 mV). The ability of Kir4.1-suppressed cells to mediate transmembrane potassium flow, as measured by the current response to voltage ramps or sequential application of different extracellular [K+], was dramatically impaired. In addition, glutamate uptake was inhibited by knock-down of Kir4.1-containing channels by RNA interference as well as by blockade of Kir channels with barium (100 microM). Together, these data indicate that Kir4.1 channels are primarily responsible for significant hyperpolarization of cortical astrocytes and are likely to play a major role in potassium buffering. Significant inhibition of glutamate clearance in astrocytes with knock-down of Kir4.1 highlights the role of membrane hyperpolarization in this process.

Animals↗
PubMed2007

Morphometric analysis of the retina from horses infected with the Borna disease virus.

Borna disease (BD) is a fatal disorder of horses, often characterized by blindness. Although degeneration of retinal neurons has been demonstrated in a rat model, there are controversial data concerning whether a similar degeneration occurs in the retina of infected horses. To investigate whether BD may cause degeneration of photoreceptors and possibly of other neuronal cells at least at later stages of the disease, we performed a detailed quantitative morphologic study of retinal tissue from Borna-diseased horses. BD was diagnosed by detection of pathognomonic Joest-Degen inclusion bodies in the postmortem brains. Paraffin sections of paraformaldehyde-fixed retinae were used for histologic and immunohistochemical stainings. Numbers of neurons and Müller glial cells were counted, and neuron-to-Müller cell ratios were calculated. Among tissues from 9 horses with BD, we found retinae with strongly altered histologic appearance as well as retinae with only minor changes. The neuron-to-Müller cell ratio for the whole retina was significantly smaller in diseased animals (8.5 +/- 0.4; P < .01) as compared with controls (17.6 +/- 0.8). It can be concluded that BD in horses causes alterations of the retinal histology of a variable degree. The study provides new data about the pathogenesis of BD concerning the retina and demonstrates that a loss of photoreceptors may explain the observed blindness in infected horses.

Animals↗
PubMed2006

Tandem-pore domain potassium channels are functionally expressed in retinal (Müller) glial cells.

Tandem-pore domain (2P-domain) K+-channels regulate neuronal excitability, but their function in glia, particularly, in retinal glial cells, is unclear. We have previously demonstrated the immunocytochemical localization of the 2P-domain K+ channels TASK-1 and TASK-2 in retinal Müller glial cells of amphibians. The purpose of the present study was to determine whether these channels were functional, by employing whole-cell recording from frog and mammalian (guinea pig, rat and mouse) Müller cells and confocal microscopy to monitor swelling in rat Müller cells. TASK-like immunolabel was localized in these cells. The currents mediated by 2P-domain channels were studied in isolation after blocking Kir, K(A), K(D), and BK channels. The remaining cell conductance was mostly outward and was depressed by acid pH, bupivacaine, methanandamide, quinine, and clofilium, and activated by alkaline pH in a manner consistent with that described for TASK channels. Arachidonic acid (an activator of TREK channels) had no effect on this conductance. Blockade of the conductance with bupivacaine depolarized the Müller cell membrane potential by about 50%. In slices of the rat retina, adenosine inhibited osmotic glial cell swelling via activation of A1 receptors and subsequent opening of 2P-domain K+ channels. The swelling was strongly increased by clofilium and quinine (inhibitors of 2P-domain K+ channels). These data suggest that 2P-domain K+ channels are involved in homeostasis of glial cell volume, in activity-dependent spatial K+ buffering and may play a role in maintenance of a hyperpolarized membrane potential especially in conditions where Kir channels are blocked or downregulated.

Animals↗
PubMed2004

Plasma levels of transforming growth factor-1beta and alpha2-macroglobulin before and after radical prostatectomy: association to clinicopathological parameters.

BACKGROUND: To study the levels of transforming growth factor-1beta (TGF-beta1) and of alpha2-macroglobulin (alpha2-M), a high affinity binding protein of TGF-beta1, in comparison to prostate-specific antigen (PSA) in prostate cancer (PCa) patients before and up to 12 months after prostatectomy, and to correlate the results with clinicopathological parameters. METHODS: Eighty-one patients who underwent radical prostatectomy for PCa were included in this study. Pre- and postoperatively, plasma levels of TGF-beta1, alpha2-M and PSA were measured in the same samples by ELISA, and were correlated with pathological parameters and clinical outcomes. RESULTS: The preoperative TGF-beta1 levels were significantly elevated as compared to the controls; they showed a positive correlation with the Gleason score. Patients with initial androgen-deprivation therapy had lower TGF-beta1 levels than untreated patients. Elevated concentrations of TGF-beta1 levelled off 12 months after prostatectomy approaching values of healthy individuals. Decreased plasma levels of total and transformed alpha2-M (proteinase-complexed form) were observed in PCa. Preoperative levels of TGF-beta1 but not of alpha2-M seem to be influenced by the body mass index (BMI). CONCLUSIONS: Elevated TGF-beta1 and decreased alpha2-M were consistently found in patients with PCa, and may be considered as risk factors for tumor development and progression. In comparison to PSA, the TGF-beta1 levels displayed a slow decline after radical prostatectomy; this indicates that TGF-beta1 is mainly produced outside the prostatic tissue. Since TGF-beta1 levels are influenced by the BMI, this indicates that PCa might be sensitive to diet.

Aged↗
PubMed2004

Somatic and axonal effects of ATP via P2X2 but not P2X7 receptors in rat thoracolumbar sympathetic neurones.

Excitatory ATP responses in rat cultured thoracolumbar sympathetic neurones are mediated by somatic P2X(2) receptors. The present study investigated a possible role of axonal P2X(2) as well as P2X(7) receptors on the same preparation. Confocal laser scanning microscopy demonstrated P2X(2) and P2X(7) immunoreactivity along the axons as well as P2X(7) immunoreactivity surrounding the cell nuclei. P2X(7) mRNA expression was detected in individual neurones using a single-cell RT-PCR approach. Adenosine triphosphate (ATP) caused a significant increase in axonal Ca(2+) concentration which was dependent on external Ca(2+) but insensitive to depletion of the cellular Ca(2+) pools by cyclopiazonic acid. Pyridoxal-phosphate-6-azophenyl-2',4'-disulfonate (PPADS; 30 micro m) virtually abolished the ATP response, whereas brilliant blue G (0.1 micro m), a selective P2X(7) receptor antagonist, had no effect. Dibenzoyl-ATP (BzATP; 100 micro m) induced a much smaller increase in axonal [Ca(2+)] concentration than ATP at equimolar concentrations. The response to BzATP was distinctly reduced by PPADS but not by brilliant blue G. The overall pharmacological profile of the axonal P2X receptors resembled closely that of the somatic P2X(2) receptors. In conclusion, the present data suggest the occurrence of axonal excitatory P2X(2) receptors in thoracolumbar sympathetic neurones. However, the functional significance of axonal and (peri)-nuclear P2X(7) receptors has still to be proven.

Adenosine Triphosphate↗
PubMed2003

Human alpha2-macroglobulin: genotype-phenotype relation.

A pentanucleotide deletion polymorphism in the gene of alpha2-macrolgobulin (alpha2-M) is suggested to be associated with late-onset Alzheimer's disease (AD), though controversial results have been reported. The underlying assumption is that the intronic pentanucleotide deletion may affect the biological function and quantity of the inhibitor and thus contribute to the AD pathology. In the present study we have analyzed the distribution of the deletion polymorphism within a group of 227 healthy Caucasians. In parallel studies, we determined the plasma concentrations of total and transformed alpha2-M. A strong correlation of the total concentration of alpha2-M with age was ascertained (r(s) = -0.54, P < 0.001). However, no significant correlation between age and the genotypes (P = 0.68) was detected, and no statistically significant effect of the genotype on the concentrations of total and transformed alpha2-M was found (P = 0.49 and 0.96, respectively). A significant correlation was observed between total and transformed alpha2-M in the genotype groups Ins/Ins (r(s) = 0.56, P < 0.001) and Ins/Del (r(s) = 0.35, P < 0.004). Furthermore, in the entire data set, a significantly elevated concentration of total alpha2-M was found in females as compared to males (P = 0.003). There was a slight but nonsignificant difference in the genotype distributions between males and females (P = 0.14). To test the proposed existence of genotype-specific alterations of functional properties of alpha2-M, we isolated alpha2-M from the plasma of carriers with different genetic background and analyzed the alpha2-M subunit structure as well as the binding of the inhibitor to growth factors/cytokines, to amyloid-beta and to the receptor. The experiments failed to reveal any genotype-specific functional alterations of the alpha2-M. The absence of abnormalities in alpha2-M mRNA and protein suggests that the alpha2-M deletion polymorphism is probably not associated with functional deficiencies important in AD pathology. However, it can be speculated that the observed general age-related alpha2-M deficiency may lead to accelerated accumulation of amyloid-beta, which might be relevant to AD pathology.

Adult↗
PubMed2003

P2 receptors in satellite glial cells in trigeminal ganglia of mice.

There is strong evidence for the presence of nucleotide (P2) receptors in sensory neurons, which might play a role in the transmission of pain signals. In contrast, virtually nothing is known about P2 receptors in satellite glial cells (SGCs), which are the main glial cells in sensory ganglia. We investigated the possibility that P2 receptors exist in SGCs in murine trigeminal ganglia, using Ca(2+) imaging, patch-clamp recordings, and immunohistochemistry. We found that ATP caused an increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) in SGCs. As adenosine had no effect on [Ca(2+)](i), and the P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid largely blocked the response to ATP we conclude that P1 receptors did not contribute to the responses. We obtained the following evidence that the responses to ATP were mediated by metabotropic P2Y receptors: (i) persistence of the responses in Ca(2+)-free solution, (ii) inhibition of the response by cyclopiazonic acid, (iii) [Ca(2+)](i) increases in response to the P2Y agonists uridine triphosphate, adenosine thiodiphosphate, and 2-methylthio ADP, and (iv) failure of the P2X agonist alpha,beta-methylene ATP to elicit a response. Agonists of P2Y(1) receptors and uridine triphosphate, an agonist at P2Y(2) and P2Y(4) receptors, induced [Ca(2+)](i) increases suggesting that at least these P2Y receptor subtypes are present on SGCs. Using an antibody against the P2Y(4) receptor, we found immunopositive SGCs. Patch-clamp recordings of SGCs did not reveal any inward current due to ATP. Therefore, there was no evidence for the activation of ionotropic P2X receptors under the present conditions. The results indicate the presence of functional nucleotide (P2Y) receptors in SGCs.

Adenosine Triphosphate↗
PubMed2001

Hypoxia: modulation of endothelial cell proliferation by soluble factors released by retinal cells.

A devastating complication of ischemic retinopathies is retinal neovascularization. We studied the impact on retinal endothelial cell proliferation of soluble factors released from cultured retinal glial (Müller) cells and from retinal explant cultures. Hypoxia strongly stimulated VEGF release by all types of cultures but endothelial cell growth was not further increased by the corresponding conditioned media if compared to supernatants obtained under normoxia. When the final concentration of the hypoxia-conditioned media was adjusted to the VEGF level of normoxia-conditioned media, they even inhibited endothelial cell proliferation. Inhibition may be exerted by TGF-beta 2 but TGF-beta 2 mRNA and protein expression in Müller cells were found to be down-regulated under hypoxia. We conclude that retinal endothelial cell proliferation is controlled by the balance of the amount and/or efficacy of several stimulatory and inhibitory factors.

Animals↗
PubMed2001

Role of Muller cells in retinal degenerations.

Muller (radial glial) cells span the entire thickness of the retina, and contact and ensheath every type of neuronal cell body and process. This morphological relationship is reflected by a multitude of functional interactions between retinal neurons and Muller cells, including extracellular ion homeostasis and glutamate recycling by Muller cells. Virtually every disease of the retina is associated with a reactive Muller cell gliosis. Muller cell gliosis may either support the survival of retinal neurons or accelerate the progress of neuronal degeneration. Muller cells are key mediators of nerve cell protection, especially via release of basic fibroblast growth factor, via uptake and degradation of the excitotoxin glutamate, and via secretion of the antioxidant glutathione. Neovascularization during hypoxic conditions is mediated by Muller cells via release of vascular endothelial growth factor and transforming growth factor beta or via direct contact to endothelial cells. Primary Muller cell insufficiency has been suggested to be the cause of different cases of retinal degeneration including hepatic and methanol-induced retinopathy and glaucoma. It is conceivable that, in the future, new therapeutic strategies may utilize Muller cells for, e.g., somatic gene therapy or transdifferentiation of retinal neurons from dedifferentiated Muller cells.

Animals↗
PubMed2001

Kir subfamily in frog retina: specific spatial distribution of Kir 6.1 in glial (Müller) cells.

We show by immunocytochemistry in frog retina that most members of the Kir subfamily are expressed in specific neuronal compartments. However, Kir 6.1, the pore-forming subunit of K(ATP) channels, is expressed exclusively in glial Müller cells. Müller cell endfeet display strong Kir 6.1 immunolabel throughout the retina, whereas the somata are labeled only in the retinal periphery. This spatial pattern is similar to that of Kir 4.1, of the ratio of inward to outward K+ currents, and of spermine/spermidine immunoreactivity. We suggest that the co-expression of Kir 4.1 and Kir 6.1 subunits may enable the cells to maintain their high K+ conductance and hyperpolarized membrane potentials both at high ATP levels (Kir 4.1) and during ATP deficiency (Kir 6.1).

Animals↗
PubMed2001

Electrophysiological properties of rat retinal Müller (glial) cells in postnatally developing and in pathologically altered retinae.

Retinal glial Müller cells are characterized by dominant K(+) conductances. The cells may undergo changes of their membrane currents during ontogeny and gliosis as described in rabbit and man. Although the rat retina is often used in physiological experiments, the electrophysiology of rat Müller cells is less well studied. The aim of the present study was to characterize their membrane currents in postnatal development and in two models of retinal degeneration. Freshly isolated cells were subjected to whole-cell patch clamp recordings. During the first 4 weeks after birth of rats, their Müller cells displayed an increase in all membrane currents, particularly in the inward currents elicited at hyperpolarizing potentials. The decrease of the membrane resistance from more than 760 MOmega to less than 50 MOmega was accompanied by a shift of the zero current potential from about -20 mV to -80 mV, similar as earlier observed in developing rabbit Müller cells. These developmental changes were found in pigmented Brown Norway rats as well as in rats with inherited retinal dystrophy (RCS rats). Moreover, an infection of Lewis rats with the Borna disease virus caused substantial neuroretinal degeneration but did not result in a strong reduction of inward currents and of the zero current potential of the Müller cells. Thus, rat Müller cells fail to change their basic membrane properties in two different models of retinal pathology. This is in contrast to human and rabbit Müller cells, which have been shown to undergo dramatic changes of their membrane physiology in response to retinal diseases and injuries.

Animals↗
PubMed2001

Electrophysiological alterations and upregulation of ATP receptors in retinal glial Müller cells from rats infected with the Borna disease virus.

Infection with the neurotropic Borna disease virus (BDV) causes an immune-mediated neurological disease in a broad range of species. In addition to encephalitis, BDV-infected Lewis rats develop a retinitis histologically characterized by the loss of most retinal neurons. By contrast, the dominating retinal macroglia, the Müller cells, do not degenerate. It is known from several models of neurodegeneration that glial cells may survive but undergo significant alterations of their physiological parameters. This prompted us to study the electrophysiology and ATP-induced changes of intracellular Ca(2+)-concentration ([Ca(2+)](i)) in Müller cells from BDV-infected rat retinae. Freshly isolated cells were used for whole-cell patch-clamp recordings. Whereas neither zero current potentials nor membrane resistances showed significant alterations, the membrane capacitance increased in cells from BDV-infected rats during survival times of up to 8 months. This process was accompanied by a decrease in K(+) current densities. Müller cells from BDV-infected rats were characterized by expression of a prominent fast-inactivating A-type K(+) current which was rarely found in control cells. Moreover, the number of cells displaying Na(+) currents was slightly increased after BDV-infection. ATP evoked increases in [Ca(2+)](i) in Müller cells within retinal wholemounts of both control and BDV-infected animals. However, the number of ATP-responding isolated cells increased from 24% (age-matched controls) to 78% (cells from animals > or =18 weeks after infection). We conclude that in BDV-induced retinopathy, reactive rat Müller cells change their physiological parameters but these changes are different from those in Müller cells during proliferative vitreoretinopathy in man and rabbit.

4-Aminopyridine↗
PubMed2001

Alpha 2-macroglobulin-mediated degradation of amyloid beta 1--42: a mechanism to enhance amyloid beta catabolism.

Peptides derived from proteolytic degradation of the amyloid precursor protein, e.g., amyloid beta (A beta), are considered to be central to the pathology of Alzheimer's disease (AD). Soluble A beta is present in measurable concentrations in cerebrospinal fluid and blood. There are indications that soluble A beta present in circulation can cross the blood-brain barrier via transcytosis mediated by brain capillary endothelial cells. It implies that A beta originating from circulation may contribute to vascular and parenchymal A beta deposition in AD. Enhancing of A beta catabolism mediated by proteolytic degradation or receptor-mediated endocytosis could be a key mechanism to maintain low concentrations of soluble A beta. To launch A beta clearance we have exploited the A beta-degrading activity of diverse alpha 2-macroglobulin (alpha 2-M)-proteinase complexes. Complexes with trypsin, alpha-chymotrypsin, and bromelain strongly degrade (125)I-A beta 1--42 whereas complexes with endogenous proteinases, e.g., plasmin and prostate-specific antigen, were not effective. A beta degradation by the complexes was not inhibited by alpha 1-antichymotrypsin and soybean trypsin inhibitor which normally would inactivate the free serine proteinases. A prerequisite for A beta degradation is its binding to specific binding sites in alpha 2-M that may direct A beta to the active site of the caged proteinase. Ex vivo, enhanced degradation of (125)I-A beta 1--42 in blood could be achieved upon oral administration of high doses of proteinases to volunteers. These results suggest that up-regulation of A beta catabolism could probably reduce the risk of developing AD by preventing A beta accumulation in brain and vasculature.

Alzheimer Disease↗
PubMed2001

Arachidonic acid-induced inhibition of Ca2+ channel currents in retinal glial (Müller) cells.

BACKGROUND: Arachidonic acid is a second messenger that has been implicated in several pathological conditions in nervous tissues. The present study was carried out to determine whether the second messenger arachidonic acid modulates currents through voltage-gated Ca2+ channels in freshly isolated Müller glial cells. METHODS: Whole-cell voltage-clamp recordings were made in human Muller cells to investigate Ba2+ and Na+ currents through high-voltage-activated (HVA) channels, and in rabbit Muller cells to study Na+ currents through low-voltage-activated (LVA) channels. RESULTS: Extracellular application of arachidonic acid reversibly and dose-dependently depressed the amplitude of both LVA (rabbit cells) and HVA currents (human cells). 10 microM arachidonic acid reduced the peak LVA and HVA currents by approximately 70%. A 50% reduction of LVA currents was achieved at 4.7 microM. The block of HVA and LVA currents was not accompanied by alterations in the voltage dependences of current activation and inactivation. A similar reduction of the currents was achieved by 20 microM eicosatetraynoic acid. CONCLUSION: Since eicosatetraynoic acid mimics the effects of arachidonic acid, it is assumed that arachidonic acid itself rather than its degradation products modulates glial Ca2+ channel activity. This Ca2+ channel inhibition may stabilize Muller cell function during pathological conditions in which arachidonic acid levels are elevated and may participate in the cellular action of neurotransmitters.

Animals↗
PubMed2001

Retinal pigment epithelium melanin granules are phagocytozed by Müller glial cells in experimental retinal detachment.

The ability of retinal Müller glial cells to perform phagocytosis in vivo is studied in a rabbit model of experimental retinal detachment where pigment epithelial cells are occasionally detached together with the neural retina. While macrophages and/or microglial cells phagocytoze most of the cellular debris at the sclerad surface of the detached retinae, some Müller cells accumulate melanin granules. The granules are virtually intact at the ultrastructural level, and are surrounded by a membrane. They are often located close to the sclerad end of the cells, but some are distributed throughout the outer stem process up to the soma. It is concluded that rabbit Müller cells in vivo are capable of phagocytosis and of transporting the phagocytozed material within their cytoplasm.

Animals↗
PubMed2001

Experimental retinal detachment causes widespread and multilayered degeneration in rabbit retina.

Retinal detachment remains one of the most frequent causes of visual impairment in humans, even after ophthalmoscopically successful retinal reattachment. This study was aimed at monitoring (ultra-) structural alterations of retinae of rabbits after experimental detachment. A surgical procedure was used to produce local retinal detachments in rabbit eyes similar to the typical lesions in human patients. At various periods after detachment, the detached retinal area as well as neighbouring attached regions were studied by light and electron microscopy. In addition to the well-known degeneration of photoreceptor cells in the detached retina, the following progressive alterations were observed, (i) in both the detached and the attached regions, an incomplete but severe loss of ganglion cell axons occurs; (ii) there is considerable ganglion cell death, particularly in the detached area; (iii) even in the attached retina distant from the detachment, small adherent groups of photoreceptor cells degenerate; (iv) these photoreceptor cells degenerate in an atypical sequence, with severely destructed somata and inner segments but well-maintained outer segments; and (v) the severe loss of retinal neurons is not accompanied by any significant loss of Müller (glial) cells. It is noteworthy that the described progressive (and probably irreparable) retinal destructions occur also in the attached retina, and may account for visual impairment in strikingly large areas of the visual field, even after retinal reattachment.

Animals↗
PubMed2001

P2 receptor-types involved in astrogliosis in vivo.

1. In the nucleus accumbens (NAc) of rats, the involvement of P2X and P2Y receptors in the generation of astrogliosis in vivo, was investigated by local application of their respective ligands. The agonists used had selectivities for P2X1,3 (alpha,beta-methylene adenosine 5'-triphosphate; alpha,beta-meATP), P2Y1,12 (adenosine 5'-O-(2-thiodiphosphate; ADP-beta-S) and P2Y2,4,6 receptors (uridine 5'-O-(3-thiotriphosphate; UTP-gamma-S). Pyridoxalphosphate-6-azophenyl-2,4-disulphonic acid (PPADS) was used as a non-selective antagonist. The astroglial reaction was studied by means of immunocytochemical double-labelling with antibodies to glial fibrillary acidic protein (GFAP) and 5-bromo-2'-deoxyuridine (BrdU). 2. The agonist-induced changes in comparison to the artificial cerebrospinal fluid (aCSF)-treated control side reveal a strong mitogenic potency of ADP-beta-S and alpha,beta-meATP, whereas UTP-gamma-S was ineffective. The P2 receptor antagonist PPADS decreased the injury-induced proliferation when given alone and in addition inhibited all agonist effects. 3. The observed morphogenic changes included hypertrophy of astrocytes, elongation of astrocytic processes and up-regulation of GFAP. A significant increase of both GFAP-immunoreactivity (IR) and GFA-protein content (by using Western blotting) was found after microinfusion of alpha,beta-meATP or ADP-beta-S. In contrast, UTP-gamma-S failed to increase the GFAP-IR. The morphogenic effects were also inhibited by pre-treatment with PPADS. 4. A double immunofluorescence approach with confocal laser scanning microscopy showed the localisation of P2X3 and P2Y1 receptors on the GFAP-labelled astrocytes. 5. In conclusion, the data suggest that P2Y (P2Y1 or P2Y12) receptor subtypes are involved in the generation of astrogliosis in the NAc of rats, with a possible minor contribution of P2X receptor subtypes.

Adenosine Diphosphate↗