Search PubMed⌕ Search

Biomedical subjects

A Reichenbach

Publications and source records attributed to A Reichenbach.

At least 55 records · Page 3Linked to original sources
PubMed2000

Expression of Dp71 in Müller glial cells: a comparison with utrophin- and dystrophin-associated proteins.

PURPOSE: The abnormal retinal electrophysiology observed in patients with Duchenne muscular dystrophy (DMD) has been attributed to an altered expression of C-terminal products of the dystrophin gene. It has been shown that Dp260 is expressed by photoreceptor cells, whereas Dp71 is present in glial cells. The present study was intended to identify all known members of the dystrophin superfamily and their associated proteins expressed in Müller glial cells (MGC). METHODS: The expression of the proteins and of their messengers was studied in MGC cultures from 2-week-old rats, by polymerase chain reaction amplification, Western blot analysis, and immunocytochemistry. An immunocytochemical localization of the proteins was also performed on enzymatically dissociated Müller cells from adult rat retinas. RESULTS: MGCs expressed a spliced isoform of Dp71 called Dp71f, as well as utrophin, beta-dystroglycan, delta and gamma-sarcoglycans, and alpha1-syntrophin. In morphologically preserved differentiated Müller cells, Dp71f was localized in clusters, utrophin was diffusely distributed in the cytoplasm, and dystrophin-associated proteins (DAPs) were membrane-bound. Most of these proteins were preferentially expressed in the vitread portion of the cells. Dp71f and utrophin expression was restricted to MGCs, whereas all DAPs were also present in other retinal cell types. CONCLUSIONS: The exclusive localization of Dp71f and utrophin in MGCs suggests that these proteins, together with DAPs, play a specific role in these cells. Further knowledge of possible interactions of these proteins within a functional complex may provide new insights into the molecular basis of the electroretinogram phenotype in DMD.

Animals↗
PubMed2000

Na(+) currents through Ca(2+) channels in human retinal glial (Müller) cells.

PURPOSE: To detect the presence of voltage-gated Ca(2+) channels in the plasma membranes of freshly isolated Müller glial cells from the human retina and their modulation by GABA(B) receptor agonists. METHODS: Whole cell voltage-clamp recordings were made to study Ca( 2+), Ba(2+), and Na(+) currents through voltage-gated Ca(2+) channels. RESULTS: The vast majority of the investigated cells displayed no resolvable currents through Ca(2+) channels when Ca(2+) ions (2 mM) were present in the extracellular solution. Small-amplitude inwardly directed currents ( approximately 0.6 pA/pF) were detected when Ba(2+) ions (20 mM) were used as charge carrier. However, when Na(+) ions were used as charge carrier in divalent cation-free external solution, currents of large amplitudes ( approximately 7.5 pA/pF) through voltage-gated Ca(2+) channels were detected. Human Müller cells displayed currents through both transient, low voltage-activated Ca(2+) channels and long-lasting, high voltage-activated channels. The Na(+) fluxes through low voltage-activated Ca( 2+) channels were inhibited in a voltage-independent manner in the presence of GABA(B) receptor agonists. CONCLUSIONS: Human Müller glial cells express different kinds of voltage-gated Ca(2+) channels in their plasma membranes that can be activated only under certain physiological or pathophysiological conditions. The record of Na(+) fluxes in divalent cation-free solutions may be a technique to detect the presence of "hidden" voltage-gated Ca(2+) channels in Müller glial cells.

Barium↗
PubMed2000

Age- and disease-related changes of calcium channel-mediated currents in human Müller glial cells.

PURPOSE: To determine whether the expression of voltage-gated Ca2+ channels in human Müller glial cells changes during normal aging and in cells from patients with proliferative vitreoretinopathy (PVR). METHODS: Müller cells were enzymatically isolated from retinas of healthy donors and from excised retinal pieces of patients with PVR, and the whole-cell, voltage-clamp technique was used to characterize the current densities of transient, low-voltage-activated calcium channels and of sustained. high-voltage-activated calcium channels, respectively. To obtain maximal currents through both channel types, Na+ ions were used as the charge carrier. RESULTS: During normal aging, Müller cells developed a hypertrophy, as indicated by an increase of the cell membrane capacitance. The mean membrane capacitance of cells from aged donors (> or = 60 years old) was elevated by 25% compared with cells from younger donors. The hypertrophy was not accompanied by a changed density of low-voltage-activated currents, whereas the density of the high-voltage-activated currents was enhanced by 76%. The density of the high-voltage-activated currents increased in correlation with the increase of the cell membrane capacitance and with the age of the donors. In the case of PVR, Müller cells displayed a strong hypertrophy accompanied by a downregulation of both current types by approximately 65%. CONCLUSIONS: Both normal aging and PVR cause a gliotic reactivity of human Müller cells, as indicated by their hypertrophy. The type of reactivity, however, differs between the two conditions. Normal aging is accompanied by an increased expression of voltage-gated Ca2+ channels, whereas in PVR Ca2+ channel expression is decreased.

Adult↗
PubMed2000

Involvement of calcium-activated potassium channels in the regulation of DNA synthesis in cultured Müller glial cells.

PURPOSE: To determine the involvement of Ca(2+)-activated K(+) channels of big conductance (BK) and of Ca(2+) channels in the regulation of DNA synthesis in cultured guinea pig Müller cells. DNA synthesis was stimulated by elevated extracellular potassium, by serum, or by epidermal growth factor. METHODS: Dissociated retinas from guinea pigs were cultured for 8 days. Just before confluence was achieved, the cultures were treated with the test substances in serum-free or serum-containing media. The rates of DNA synthesis were assessed by a quantitative bromodeoxyuridine immunoassay. The intracellular Ca(2+) concentration was measured by the fura-2 fluorescence technique. RESULTS: Blocking the BK channels with tetraethylammonium or by iberiotoxin had no effect at normal extracellular K(+) (5.8 mM) but decreased the rate of DNA synthesis at higher extracellular K(+) (10 or 25 mM). Epidermal growth factor-induced DNA synthesis was decreased by block of BK channels or by application of the Ca(2+) channel blockers nimodipine and flunarizine. Application of epidermal growth factor elevated the intracellular Ca(2+) concentration of cultured Müller cells. This elevation was diminished by co-application of iberiotoxin or of flunarizine. CONCLUSIONS: The activity of BK channels is necessary for elevated DNA synthesis in Müller cells when their membranes are depolarized and/or when the Ca(2+) influx into Müller cells is increased by growth factors. BK channels may contribute to the maintenance of DNA synthesis by increasing mitogen-induced increase in intracellular Ca(2+) concentration.

Animals↗
PubMed1999

Outwardly rectifying K+ channels display clustering in guinea pig retinal Müller cells.

The cell-attached configuration of the patch-clamp technique was used to characterize the outward currents in acutely isolated Müller cells from the guinea pig retina. Sixty-five of 353 patches displayed macroscopic, outwardly rectifying currents due to depolarizing voltage steps. Single channel transitions were found in only two patches. The remaining patches did not reveal any voltage-dependent currents. Tail current analysis revealed a reversal potential close to the resting membrane potential. The currents disappeared if internal K+ was replaced by Cs+ in inside-out patches. From these results we conclude that guinea pig Müller cells possess voltage-dependent K+ channels that are distributed in clusters.

Animals↗
PubMed1999

Cortical neurons immunoreactive for the potassium channel Kv3.1b subunit are predominantly surrounded by perineuronal nets presumed as a buffering system for cations.

Perineuronal nets (PNs) are known as chondroitin sulphate-rich, lattice-like coatings of the extracellular matrix. In the cortex of mammalian species investigated so far, they were mainly found around GABAergic neurons, but to a lesser degree also around pyramidal cells. Previous investigations in the rat revealed similar distribution patterns of fast-firing neurons expressing both the Kv3.1b subunit of voltage-gated potassium channels and the calcium-binding protein parvalbumin. In the present study, triple fluorescence labelling was applied for the simultaneous demonstration of PNs with the N-acetylgalactosamine-specific Wisteria floribunda agglutinin (WFA), parvalbumin-immunoreactivity (ir) with a monoclonal antibody and of Kv3.1b-ir with several rabbit antibodies. Subsets of non-pyramidal neurons - enwrapped by PNs and expressing parvalbumin and Kv3.1b - were detected in the rat and monkey neocortex and hippocampus. In the rat, faintly stained PNs were additionally found around several layer II/III and V pyramidal cells immunonegative for Kv3.1b, but contacted by Kv3.1b-containing boutons. In the monkey, more intensely labelled PNs frequently occurred around pyramidal cells which themselves appeared to be Kv3. 1b-immunopositive. We also observed minor Kv3.1b-ir and parvalbumin-ir cortical cell populations which were devoid of PNs; occasionally, nets were detected around neurons lacking both immunoreactivities. By confocal laser scanning microscopy, Kv3.1b-ir and WFA-binding sites were found adjoining at the soma and proximal dendritic surface, while lectin-binding sites usually extended on more distal dendritic segments and the axon initial segments which failed to express detectable Kv3.1b-ir. This spatial relationship of both markers was also confirmed by combined WFA-gold labelling and Kv3.1b-immunoperoxidase staining at the electron microscopic level. The data are used for a critical examination of current hypotheses concerning the functional role of PNs. We conclude that PNs may serve as rapid local buffers of excess cation changes in the extracellular space. Somatic membranes of fast-spiking neurons seem to be a main, but not the only source of such changes.

Animals↗
PubMed1999

Protein kinases A and C are opponents in modulating glial Ca2+ -activated K+ channels.

The modulation of the activity of Ca2+ -activated K+ (BK) channels by activators of protein kinases A and C, respectively, was studied in cell-attached patches on isolated Müller (retinal glial) cells from rabbits. The BK channel activity was stimulated by membrane depolarization and by increasing of the intracellular Ca2+ concentration. Extracellular exposure to dibutyryl-cAMP, known to stimulate the protein kinase A, increased the open probability of the channels. Exposure to a phorbol ester, as an activator of protein kinase C, strongly reduced the channel activity whereas exposure to the protein kinase inhibitor, staurosporine, stimulated the channel activity. As glial BK channels are modulated in an opposite manner by protein kinases A and C, they may act as a cellular focus of integration of the inputs from different signaling pathways.

Animals↗
PubMed1999

The pecten oculi of the chicken: a model system for vascular differentiation and barrier maturation.

The pecten oculi is a convolute of blood vessels in the vitreous body of the avian eye. This structure is well known for more than a century, but its functions are still a matter of controversies. One of these functions must be the formation of a blood-retina barrier because there is no diffusion barrier for blood-borne compounds available between the pecten and the retina. Surprisingly, the blood-retina barrier characteristics of this organ have not been studied so far, although the pecten oculi may constitute a fascinating model of vascular differentiation and barrier maturation: Pectinate endothelial cells grow by angiogenesis from the ophthalmotemporal artery into the pecten primordium and consecutively gain barrier properties. The pectinate pigmented cells arise during development from retinal pigment epithelial cells and subsequently lose barrier properties. These inverse transdifferentiation processes may be triggered by the peculiar microenvironment in the vitreous body. In addition, the question is discussed whether the avascularity of the avian retina may be due to the specific metabolic activity of the pecten.

Animals↗
PubMed1999

Microdomains for neuron-glia interaction: parallel fiber signaling to Bergmann glial cells.

Astrocytes are considered a reticulate network of cells, through which calcium signals can spread easily. In Bergmann glia, astrocytic cells of the cerebellum, we identified subcellular compartments termed 'glial microdomains'. These elements have a complex surface consisting of thin membrane sheets, contain few mitochondria and wrap around synapses. To test for neuronal interaction with these structures, we electrically stimulated parallel fibers. This stimulation increased intracellular calcium concentration ([Ca2+]i) in small compartments within Bergmann glial cell processes similar in size to glial microdomains. Thus, a Bergmann glial cell may consist of hundreds of independent compartments capable of autonomous interactions with the particular group of synapses that they ensheath.

Animals↗
PubMed1999

Expression of potassium channels during postnatal differentiation of rabbit Müller glial cells.

The postnatal maturation of Müller glial cells from immature radial glial cells is accompanied by specific changes in the activity of distinct types of K+ channels, as shown by whole-cell and cell-attached records on freshly isolated cells from retinae of young (postnatal days 1-30, P1-P30) and adult rabbits. (i) The density of inwardly rectifying currents, providing the main K+ conductance in adult Müller cells, was very low (0.8 pA/pF) from P1 to P6 but increased rapidly thereafter until a relatively stable level of 11.0 pA/pF was established at P17. (ii) Transient (A-type) K+ currents were expressed in all immature cells at a high density (9.6 pA/pF). After P12, both the percentage of cells with A-type currents and the peak amplitudes of the currents (2.8 pA/pF) declined. (iii) Delayed rectifying K+ currents developed slowly until after P30. (iv) The postnatal maturation of radial glial cells was accompanied by a strong decrease in the activity of large-conductance, Ca2+-activated K+ channels, the open probability of which (measured at the resting membrane potential) decreased from 0.69 at P2-4 to 0.06 at P13-14. The developmental decrease of the activity of Ca2+-activated K+ channels is assumed to be mainly caused by alteration of the resting membrane potential which developed from low values (-49 mV) at P1-6 to high adult values (-84 mV) after P13. The activity of each distinct type of K+ channel investigated is differently modulated by developmental regulation. This may reflect different functional requirements of immature and mature Müller cells.

Animals↗
PubMed1999

Potassium buffering by Müller cells isolated from the center and periphery of the frog retina.

Müller (radial glial) cells span the retina from the outer to the inner limiting membranes. They are the only glial cells found in the amphibian retina. The thickness of the frog (Rana pipiens) retina decreases by a factor of about four from the center to the periphery. Thus, Müller cells were isolated, by enzymatic dissociation, with stalk lengths from 20 to 140 microm. Their ability to transfer K(+) via the stalk between soma and endfoot was studied. Membrane currents were recorded using the whole-cell voltage-clamp technique with the pipette sealed to either the endfoot or the soma. Inward (I(KIN)) or outward (I(KO)) currents were elicited by rapid increases (3 to 10 mM) or decreases (3 to 1 mM) of the extracellular K(+) concentration ([K(+)](o)) either by local application (close or distant to the recording pipette) or around the entire cell (whole cell perfusion). For the long central cells, the ratio I(KIN)/I(KO) was 4.6 +/- 0.6 SE (n = 9) at the endfoot and 1.7 +/- 0.1 SE (n = 8) at the soma. In cells from the retinal periphery, the ratio I(KIN)/I(KO) was higher, 7.0 +/- 0.27 (n = 8) at the endfoot and 3.2 +/- 0.1 (n = 10) at the soma. The results suggest that there is less inward rectification in the somatic than in the endfoot membrane. As expected from previous studies, the sensitivity of the cells to K(+) was higher at the endfoot than at the soma. The amplitude of I(KIN) at the endfoot compared to the soma was about 8-fold for the long central cells but only about 1.5-fold for the short peripheral cells. Currents spread readily from endfoot to soma in the peripheral cells. In the long central Müller cells the soma and endfoot appeared electrotonically isolated. The "functional length constant", lambda, of cell stalk processes was about 70 microm. The relative decrement of large inward currents was stronger than that of smaller outward currents; this difference ("artificial rectification") is explained by a simple model, where larger currents (inward) are attenuated more than smaller (outward) currents. The data support the hypothesis that in the retinal periphery, Müller cells provide extensive spatial K(+) buffering from both plexiform layers into the vitreous body. In the central retina, however, such currents are limited within a short (interlaminar) range.

Animals↗
PubMed1999

Human Müller glial cells: altered potassium channel activity in proliferative vitreoretinopathy.

PURPOSE: To determine differences of K+ channel activity between Müller glial cells obtained from retinas of healthy human donors and of patients with retinal detachment and proliferative vitreoretinopathy. METHODS: Müller cells were enzymatically isolated from retinas of healthy donors and from excised retinal pieces of patients. The whole-cell and the cell-attached configurations of the patch-clamp technique were used to characterize the current densities of different K+ channel types and the activity of single Ca2+ -activated K+ channels of big conductance (BK). RESULTS: Cells from patients displayed a less negative mean membrane potential (-52.8 mV) than cells from healthy donors (-80.6 mV). However, the membrane potentials in cells from patients scattered largely between -6 and -99 mV. The inwardly rectifying K+ permeability in cells from patients was strongly reduced (0.3 pA/pF) when compared with cells from healthy donors (6.0 pA/pF). At the resting membrane potential, single BK channels displayed a higher mean activity (open probability, Po, and channel current amplitude) in cells from patients (Po, 0.30) than in cells from healthy donors (Po: 0.03). The variations of BK current amplitudes were correlated with the variations of the membrane potential. CONCLUSIONS: The dominant expression of inwardly rectifying channels in cells from healthy donors is thought to support important glial cell functions such as the spatial buffering of extracellular K+. The downregulation of these channels and the less negative mean membrane potential in cells from patients should impair spatial buffering currents and neurotransmitter clearance. The increased activity of BK channels may support the proliferative activity of gliotic cells via feedback regulation of Ca2+ entry and membrane potential.

4-Aminopyridine↗
PubMed1999

Cell-cell coupling in cultures of striatal and cortical astrocytes of the monkey Cebus apella.

Astrocytes were cultured from striatum and neocortex of fetal (embryonic day 90) monkeys (Cebus apella). The cultures grew well, and the cells retained viability after freeze-storage and thawing. The cells displayed depolarized membrane potentials (-19 and -33 mV, for striatal and cortical cells, respectively) and the vast majority of cells were dye-coupled to a mean of 7 (1-18) neighbouring cells. Cell coupling was blocked by octanol (0.25 and 0.5 mM) but was independent of high K+ (10 and 50 mM) and glutamate (500 microm). Thus, cultures of fetal primate astrocytic cells are established as a model system for studies on astroglial cell-cell coupling.

Animals↗
PubMed1999

The activity of a transient potassium current in retinal glial (Müller) cells depends on extracellular calcium.

The modulating effects of varying extracellular concentrations of Ca2+ ([Ca2+]e) and of other divalent cations on the fast transient (A-type) K+ current (I(A)) of freshly isolated Muller glial cells from rabbit and human retinae were studied with the whole-cell patch-clamp method. The I(A) of Miller cells was voltage-independently blocked by extracellular 4-aminopyridine (4AP) with a 50 % reduction achieved at 0.94 mM 4AP. The I(A) amplitude was elevated by increased extracellular [K+]. Elevation of the [Ca2+]e had three effects on the glial I(A): (i) it concentration-dependently shifted both the activation and inactivation curves towards less negative membrane potentials, (ii) it increased the peak current amplitude, and (iii) it slowed down the activation and inactivation kinetics. Particularly at depolarized membrane potentials, the I(A) was enlarged and broadened when the [Ca2+]e was increased. Various divalent cations also exerted these effects, although at different concentrations. While Zn2+, Cd2+, Cu2+ and Pb2+ modulated the I(A) in the micromolar range, Mg2+ and Ba2+ had effects in the millimolar range. Extracellular acidification produced a positive shift in the voltage dependence of I(A) gating. However, alterations of the extracellular pH did not abolish the Ca2+ effects on I(A); this indicates that protons and Ca2+ ions mediate their effects on glial K(A) channels by different mechanisms or binding sites, respectively. Physiological (i.e., activity-dependent) changes of the extracellular concentration of divalent cations and of the extracellular pH should influence the retinal excitability via modulation of glial K+ currents. The activation of glial I(A) by divalent cations at depolarized voltages supports a repolarization and, therefore, the maintainance of a hyperpolarized glial membrane potential during periods of increased neuronal activity.

4-Aminopyridine↗
PubMed1998

Cathepsin E immunoreactivity in human ocular tissues: influence of aging and pathological states.

We studied the antigenic expression of the aspartic proteinase cathepsin E in normal and pathologic human ocular tissues obtained from donors of different age. In the retina the enzyme was immunolocalized in neurons of outer and inner plexiform layers and in few ganglionic neurons. Muller cells were also sometimes immunoreactive for cathepsin E. An increase of neuronal enzyme immunoreactivity with age was evident. Immunocompetent blood cells invading the vitreous body were strongly immunostained for the enzyme. The enzyme is possibly involved in the retinal protein metabolism and might play immunological roles in certain pathologic events.

Adolescent↗
PubMed1998

Spermine/spermidine is expressed by retinal glial (Müller) cells and controls distinct K+ channels of their membrane.

There is recent evidence that polyamines such as spermine (spm) and spermidine (spd) may act as endogenous modulators of the activity of inwardly rectifying K+ channels. This type of K+ channels is abundantly expressed by retinal glial (Müller) cells where they are involved in important glial cell functions such as the clearance of excess extracellular K+ ions. This prompted us to study the following questions, i) do mammalian Müller cells contain endogenous spm/spd?; ii) do Müller cells possess the enzymes (e.g., ornithine decarboxylase, ODC) necessary to produce spm/spd?; and iii) does application of exogenous spm/spd exert specific effects onto inwardly rectifying K+ channels of Müller cells? Immunocytochemical studies were performed on histological sections of guinea-pig, rabbit, porcine, and human retinae, and on enzymatically dissociated Müller cells. Whole-cell and patch-clamp recordings were performed on enzymatically dissociated porcine and guinea-pig Müller cells. All above-mentioned questions could be answered with "yes." Specifically, the majority of Müller cells were labeled with antibodies directed to spm/spd, both within retinal sections and enzymatically isolated from retinal tissue. Müller cells in normal retinae express low levels of ODC but increase this expression markedly in cases of retinal pathology such as experimental epiretinal melanoma. Externally applied polyamines (1 mM) reduce (predominantly inward) whole-cell K+ currents, with the efficacies being spm > spd > put. If applied at the inside of membrane patches, spm (1 mM) blocks completely the outward currents through inwardly rectifying K+ channels but fails to affect the activity of large conductance, Ca2+-activated K+ channels. It is concluded that Müller cells contain endogenous channel-active polyamines, the synthesis of which may be up-regulated in pathological situations, and which may be involved in the control of both glial function and cell proliferation.

Adolescent↗
PubMed1998

Retinal gliopathy accompanying thioacetamide-induced liver insufficiency: light and electron microscopic observations.

A recent examination of retinae of patients who had died with symptoms of liver insufficiency (LI) including hepatic encephalopathy (HE) revealed morphological changes in retinal Müller glia similar to the astrocytic changes normally accompanying HE, and the term "hepatic retinopathy" (HR) was coined to define these changes. In the present study, the immunomorphology and ultrastructure of Müller cells were examined in rats in which LI with accompanying HE was induced with a hepatotoxin, thioacetamide (TAA). Light microscopically, retinae of rats with LI were characterized by swelling of the Muller cell cytoplasm. Immunostaining for glia-specific marker proteins in Müller cells from LI rats revealed a strongly enhanced expression of glial fibrillary acidic protein, and a considerable increase in glutamine synthetase immunoreactivity, as compared to control animals. Ultrastructurally, the Müller cells of LI rats showed swelling and vacuolization of cell processes. In particular, the endfeet contained many swollen mitochondria. By contrast, LI produced no morphologically demonstrable changes in retinal neurons and photoreceptor cells. Thus, the retinal changes induced by TAA in the rats strongly resembled those described in human HR, rendering the present rat model suitable for more detailed investigations of the pathomechanism(s) of HR.

Animals↗