Arachidonic acid, which is thought to be involved in pathogenetic mechanisms of the central nervous system, has been shown previously to modulate neuronal ion channels and the glutamate uptake ...carrier of retinal glial (Müller) cells. We have used various configurations of the patch-clamp technique to determine the effects of arachidonic acid on the K
+ currents of freshly isolated Müller glial cells from rabbit and human. Arachidonic acid reduced the peak amplitude of the transient (A-type) outward K
+ currents in a dose-dependent and reversible manner, with a 50% reduction achieved by 4.1
μM arachidonic acid. The inward rectifier-mediated currents remained unchanged after arachidonic acid application. The amplitude of the Ca
2+-activated K
+ outward currents (K
Ca), which were blocked by 1
mM tetraethylammonium chloride and 40
nM iberiotoxin, respectively, was dose-dependently elevated by bath application of arachidonic acid. The activation curve of the K
Ca currents shifted towards more negative membrane potentials. Furthermore, arachidonic acid was found to suppress inwardly directed Na
+ currents. In cell-attached recordings with 3
mM K
+ in the bath and 130
mM K
+ in the pipette, the K
Ca channels of rabbit Müller cells displayed a linear current–voltage relation, with a mean slope conductance of 102
pS. In excised patches, the slope conductance was 220
pS (150
mM K
+
i/130
mM K
+
o). The opening probability of the K
Ca channels increased during membrane depolarization and during elevation of the free Ca
2+ concentration at the intracellular face of the membrane patches. Bath application of arachidonic acid caused a reversible increase of the single-channel opening probability, as well as an increase of the number of open channels. Arachidonic acid did not affect the single-channel conductance. Since arachidonic acid also stimulates the K
Ca channel activity in excised patches, the action of arachidonic acid is assumed to be independent of changes of the intracellular calcium concentration.
Our results demonstrate that arachidonic acid exerts specific effects on distinct types of K
+ channels in retinal glial cells. In pathological cases, elevated arachidonic acid levels may contribute to prolonged Müller cell depolarizations, and to the initiation of reactive glial cell proliferation.
For macular translocation surgery, the native attached retina has to be detached either locally or completely. Although different surgical techniques are used, there is a general search for ...supporting procedures that facilitate and accelerate the retinal detachment.
Pars plana vitrectomies were performed in pigmented rabbits. In all experimental groups, a local retinal detachment was created by infusing the test solution with a thin glass micropipette attached to a glass syringe. In control animals a standard balanced salt solution was used at room temperature, in combination with a standard vitrectomy light source. In two test groups, a calcium- and magnesium-free solution was used for the vitrectomy, under illumination by a standard light source in group I (solution at room temperature) and group II (solution heated up to body temperature). In group III the rabbits were dark-adapted for half an hour, and then, during surgery, a red filter was used in front of the light source (standard balanced salt solution at room temperature). After the rabbits were killed at the end of surgery, the adherence of the retinal pigment epithelium (RPE) to the neural retina in the detachment area was quantified microscopically, and the morphologic integrity of the detached retinal tissue was examined by light and electron microscopy. No electrophysiology was performed.
In all four groups, it was possible to detach the retina. The maximum adherence of the RPE cells to the neural retina was observed in the control group. Virtually no decrease in adherence was found in test group II (36 degrees C solution without calcium and magnesium), whereas a significant decrease was seen in both group I (calcium- and magnesium-free solution at room temperature) and group III (dark adaptation-red light technique; standard balanced salt solution at room temperature). In none of the experimental groups was any obvious damage of the retinal structure observed, even after exposure to the test solutions for 60 minutes.
Both dark adaptation (red illumination) and the use of a calcium chloride- and magnesium chloride-free solution (at room temperature) can facilitate retinal detachment in macular translocation surgery. Both techniques are proposed as a gentle support for the operation, because they protect an intact RPE cell layer and do not cause retinal damage at the ultrastructural level.
In this letter we present a hierarchical unit cell incorporating a nonlinear energy sink (NES) that functions as a low-pass amplitude filter. The hierarchical unit cell is composed of an outer and ...inner mass coupled by essentially nonlinear springs. Other than its nonlinear coupling to the outer mass, the inner mass is otherwise unconstrained and thus acts as an NES. Using numerical simulations, we show that the nonlinear unit cell passively filters incident waves as a function of their amplitude. The resonator is inactive at low amplitudes, permitting waves to pass. At high amplitudes, the NES activates and its resonant behavior reflects incident waves. Consequently, the unit cell effectively operates as an amplitude limiter for specific signal frequency and amplitude ranges. Notably, the passing of low-amplitude incident energy while bypassing the nonlinearity results in very little distortion in transmitted signals. We analytically investigate this behavior further using a harmonic balance method. The analysis supports the transmission filtering behavior as well as the activation of the nonlinear energy sink. Lastly, we design and construct the nonlinear unit cell and perform tests using incident and transmitted waveguides composed of monatomic chains. Both the nonlinear unit cell and monatomic chains are assembled using steel masses and additively-manufactured springs. We test the fabricated filter using low and high amplitude signals and observe amplitude filtering at various excitation frequencies that document strong agreement with numerical simulations and the analytical model. The ability to passively reflect high amplitude waves, and transmit low amplitude waves with minimal distortion, may inspire new devices for hearing protection, or for isolating and protecting sensitive equipment and instruments.
Enzymatically isolated, noncultured Müller glial cells from human organ donor and patient eyes were studied using the whole-cell-voltage-clamp and the patch-clamp technique. The patients suffered ...mainly from choroidal melanomas, retinal detachment due to proliferative vitreorentinopathy, glaucomas, and perforating eye injuries. The organ donor eyes were used as a source of corneas for corneal transplantation. Use of the human retinal tissue was approved by the Ethics Committee of the School of Medicine, University of Leipzig. Most of the patient Müller cells showed a marked or complete loss of inwardly rectifying K+ currents, causing a dramatic increase in the input resistance. The zero current potential of the patient Müller cells, which is equivalent to the membrane potential, was significantly reduced (depolarized) as compared with the donor Müller cells. In contrast to the K+ current loss, the Na+ current density was significantly higher in patient Müller cells than in donor Müller cells; the number of Müller cells depicting Na+ currents increased from 33% (3 pA/pF) in donor Müller cells to 85% (about 12pA/pF) in patient Müller cells. Application of glutamate to the Müller cells generated a glutamate-transporter-mediated current, such as that seen in other species. A highly significant increase was noted for the high-affinity Na+-dependent glutamate-transporter-current density in patient Müller cells compared with donor cells. The application of γ-aminobutyric acid (GABA) evoked, in addition to the GABA transporter currents already known from Müller cells of other mammalian species, GABAA-receptor mediated currents in human Müller cells. We found that GABAA receptors are expressed in human Müller cells, but not in other nonprimate mammals. Whether a difference exists between the GABAA current density in donor and patient Müller cells remains to be seen. The results concerning the disappearance of K+ currents and diminution of the membrane potential may demonstrate early glial changes that may possibly precede pathological neuronal changes, at least in retinas from eyes with choroidal melanomas. In later stages of the diseases, the glial changes may be deleterious for the neurons, because they could diminish glutamate uptake due to the depolarized membrane potential. However, increased extracellular glutamate concentration is toxic for most neurons.
ATP has been shown to be an important extracellular signaling molecule. There are two subgroups of receptors for ATP (and other purines and pyrimidines): the ionotropic P2X and the G-protein-coupled ...P2Y receptors. Different subtypes of these receptors have been identified by molecular biology, but little is known about their functional properties in the nervous system. Here we present data for the existence of P2 receptors in Müller (glial) cells of the human retina. The cells were studied by immunocytochemistry, electrophysiology, Ca(2+)-microfluorimetry, and molecular biology. They displayed both P2Y and P2X receptors. Freshly enzymatically isolated cells were used throughout the study. Although the Ca(2+)(i) response to ATP was dominated by release from intracellular stores, there is multiple evidence that the ATP-induced membrane currents were caused by an activation of P2X(7) receptors. Immunocytochemistry and single-cell RT-PCR revealed the expression of P2X(7) receptors by Müller cells. In patch-clamp studies, we found that (1) benzoyl-benzoyl ATP (BzATP) was the most effective agonist to evoke large inward currents and (2) the currents were abolished by P2X antagonists; however, (3) long-lasting application of BzATP did not cause an opening of large pores in addition to the cationic channels. By microfluorimetry it was shown that the P2X receptors mediated a Ca(2+) influx that contributed a small component to the total Ca(2+)(i) response. Activation of P2X receptors may modulate the uptake of neurotransmitters from the extracellular space by Müller cells in the retina.
Retinal Müller glial cells are known to undergo reactive changes (gliosis) in various retinal diseases. In virtually all cases studied, an upregulation of glial fibrillary acidic protein (GFAP) and a ...hypertrophy can be observed. Physiological alterations, such as a strong downregulation of inwardly rectifying K
+ (Kir) currents, were found after retinal detachment (man, rabbit) and after ischemia/reperfusion (rat) but not in more slowly progressing retinal degenerations (Borna Disease Virus-infected rats, RCS rats). This led us to hypothesize that Müller cells respond with ‘typical’ reactive gliosis only to rapid but not to slow retinal degeneration. To test this hypothesis, we studied Müller cells from
rds mutant mice (
Prph
Rd2
), which show a retinal degeneration of early onset and slow progression, resulting in a complete loss of photoreceptors after 9–12 months. In Müller cells of
rds mice, we found immunoreactivity for GFAP, a marker of gliosis in Müller cells, from postnatal day 21 on, accompanied by a moderately increased membrane capacitance (taken as an indicator of hypertrophy), whereas no change in the expression of the Kir4.1 protein occurred in adult
rds mice. We failed to observe significant changes in the membrane resistance and the membrane potential of cells from
rds mice from first week after birth until 1 year of age. Current densities were decreased in cells from 3- and 5-week old
rds mice. Furthermore, as in control cells from wildtype animals, these cells displayed dominant Kir currents, voltage-dependent Na
+ currents, and glutamate uptake currents. These data support the idea that in mice as well as previously shown in rats, slow retinal degeneration induces an atypical gliosis of Müller cells.
Ischemia–reperfusion of the rat retina causes gliosis of Müller cells that is associated with a decrease of their K
+ conductance. By using quantitative PCR and immunohistochemical staining of ...retinal slices, we investigated the effect of transient ischemia–reperfusion on retinal expression of two inward-rectifying K
+ (Kir) channels, Kir4.1 and Kir2.1. In control retinas, Müller cells prominently expressed both Kir4.1 and Kir2.1 proteins. At 7 days after reperfusion, the expression of Kir4.1 protein was strongly downregulated, while the Kir2.1 protein expression remained unaltered. The expression of Kir4.1 mRNA was reduced by 55% after ischemia while the expression of Kir2.1 mRNA was not altered. The data suggest that the glial expression of distinct Kir channels is differentially regulated after retinal ischemia, with deletarious consequences for K
+ ion and water homeostasis.
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