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At least 775 records · Page 43Linked to original sources
PubMed1985

Extracellular K+ activity changes related to electroretinogram components. I. Amphibian (I-type) retinas.

Electroretinographic (ERG) and extracellular potassium activity measurements were carried out in superfused eyecup preparations of several amphibians. Light-evoked changes in extracellular K+ activity were characterized on the bases of depth profile analysis and latency measurements and through the application of pharmacological agents that have selective actions on the retinal network. Three different extracellular potassium modulations evoked at light onset were identified and characterized according to their phenomenological and pharmacological properties. These modulations include two separable sources of light-evoked increases in extracellular K+: (a) a proximal source that is largely post-bipolar in origin, and (b) a distal source that is primarily or exclusively of depolarizing bipolar cell origin. The pharmacological properties of the distal extracellular potassium increase closely parallel those of the b-wave. A distal light-evoked decrease in extracellular potassium appears to be associated with the slow PIII potential, based on a combination of simultaneous intracellular Müller cell recordings and extracellular ERG and potassium activity measurements before and during pharmacological isolation of the photoreceptor responses. The extracellular potassium activity increases are discussed with respect to the Müller cell theory of b-wave generation.

Ambystoma↗
PubMed1985

Light-evoked increases in extracellular K+ in the plexiform layers of amphibian retinas.

Recordings of light-evoked changes in extracellular K+ concentration (delta[K+]o) were obtained in the retinas of frog and mudpuppy. In eyecup preparations, various recording approaches were used and provided evidence for a K increase near the outer plexiform layer (distal K increase). This distal K increase could be pharmacologically dissociated from the well-known, large K increase in the proximal retina by the application of ethanol and gamma-aminobutyric acid. The distal K increase also often showed surround antagonism. A retinal slice preparation was used to permit electrode placement into the desired retinal layers under direct visual control and without the risk of electrode damage to adjacent layers. In the slice, a distinct distal K increase was found in the outer plexiform layer, in addition to the prominent K increase in the inner plexiform layer. Compared with eyecups, only weak K increases were found in the nuclear layers of the slice. This suggests that the K responses observed in the nuclear layers of eyecups may be generated by K+ diffusing along the electrode track from the plexiform layers. In the context of current models of ERG b-wave generation, the magnitude of the recorded distal K increase, compared with the proximal K increase, seems too small to give rise to the b-wave. However, the distal K increase may be differentially depressed by electrode dead space. It is also possible that if certain aspects of the models of b-wave generation were modified, then the observed distal K increase could give rise to the b-wave.

Animals↗
PubMed1989

B-wave of the electroretinogram. A reflection of ON bipolar cell activity.

Light-evoked intraretinal field potentials (electroretinogram, ERG) have been measured simultaneously with extracellular potassium fluxes in the amphibian retina. The application of highly selective pharmacologic agents permitted us to functionally isolate various classes of retinal neurons. It was found that: (a) application of APB (2-amino-4-phosphonobutyrate), which has previously been shown to selectively abolish the light responsiveness of ON bipolar cells, causes a concomitant loss of the ERG b-wave and ON potassium flux. (b) Conversely, PDA (cis 2,3-piperidine-dicarboxylic acid) or KYN (kynurenic acid), which have been reported to suppress the light responses of OFF bipolar, horizontal, and third-order retinal neurons, causes a loss of the ERG d-wave as well as OFF potassium fluxes. The b-wave and ON potassium fluxes, however, remain undiminished. (c) NMA (N-methyl-DL-aspartate) or GLY (glycine), which have been reported to suppress the responses of third-order neurons, do not diminish the b- or d-waves, nor the potassium fluxes at ON or OFF. This leads to the conclusion that the b-wave of the ERG is a result of the light-evoked depolarization of the ON bipolar neurons. This experimental approach has resulted in two further conclusions: (a) that the d-wave is an expression of OFF bipolar and/or horizontal cell depolarization at the termination of illumination and (b) that light-induced increases in extracellular potassium concentration in both the inner (proximal) and outer (distal) retina are the result of ON bipolar cell depolarization.

Ambystoma↗
PubMed2000

An In vitro assay useful to determine the potency of several bitter compounds.

Gustducin and transducin are guanine nucleotide binding regulatory proteins (G proteins) expressed in taste receptor cells and implicated in transducing taste cell responses to certain compounds that humans consider bitter or sweet. These G proteins can be activated in vitro by taste receptor-containing membranes plus any of several bitter compounds. This activation can be monitored using limited trypsin digestion, sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. Scanning of the autoradiograms enables one to quantitate the level of activation (defined as an activation index), obtain dose-response profiles and estimate the potency of the tastant. This assay may provide a useful substitute for, or adjunct to, the time-consuming human psychophysical analysis and costly animal studies typically used in taste sensory analysis. It may be used to identify and determine the concentration-response function of many bitter components of oral pharmaceuticals and food ingredients. A potential limitation of the assay is that only about half of all bitter compounds tested demonstrated in vitro activity, perhaps due to the presence of multiple transduction pathways. Nevertheless, the rapid throughput and microsample handling capability of this assay make it an ideal method to screen for high-potency bitterness inhibitors.

Animals↗
PubMed2005

Pulse stimulation with odors or IBMX/forskolin potentiates responses in isolated olfactory neurons.

Many odor responses are mediated by the adenosine 3',5'-cyclic monophosphate (cAMP) pathway in which the cAMP-gated current is amplified by Ca2+-dependent Cl- current. In olfactory neurons, prolonged exposure to odors decreases the odor response and is an adaptive effect. Several studies suggest that odor adaptation is linked to elevated intracellular Ca2+. In the present study, using the perforated configuration of the patch clamp technique, we found that repetitive odor stimulation elicits a potentiation of the subsequent responses in olfactory neurons. This potentiation is mimicked by stimulating the cAMP pathway and does not appear to be related to phosphorylation of ion channels since protein kinase inhibitors could not block it. Our data suggest that local increases in [Ca2+]i via activation of the cAMP pathway mediate the pulse-elicited potentiation. In the first odor application, entry of Ca2+ through cyclic nucleotide-gated channels appears to be buffered. Repetitive stimulation allows local increases in [Ca2+]i, recruiting more Ca2+-dependent Cl- channels with each subsequent odor pulse.

1-Methyl-3-isobutylxanthine↗
PubMed1997

Differences in the phosphate oxygen requirements for self-cleavage by the extended and prototypical hammerhead forms.

The hammerhead self-cleaving motif occurs in a variety of RNAs that infect plants and consists of three non-conserved helices connected by a highly conserved central core. A variant hammerhead, called the extended hammerhead, is found in satellite 2 transcripts from a variety of caudate amphibians. The extended hammerhead has the same core as the prototypical hammerhead, but has unusually conserved sequence and structural elements in its peripheral helices. Here we present the results of a thiophosphate substitution interference analysis of the pro-Rp phosphate oxygen requirements in the two hammerhead forms. Five pro-Rp phosphate oxygens, all in the central core, were found to be important for self-cleavage by the prototypical hammerhead. A similar set of core positions were important for self-cleavage by the extended hammerhead, but five non-core positions were also found to be important. Thiosubstitution at one of these positions had the most severe effect on self-cleavage observed in this analysis. Mn2+ did not alleviate this negative effect, indicating that this position was not part of a divalent cation binding site. We propose that novel tertiary interactions in the extended hammerhead help form the same catalytic core structure as that used by the prototypical plant virus hammerhead.

Animals↗
PubMed1999

A computational model of electrical stimulation of the retinal ganglion cell.

Localized retinal electrical stimulation in blind volunteers results in discrete round visual percepts corresponding to the location of the stimulating electrode. The success of such an approach to provide useful vision depends on elucidating the neuronal target of surface electrical stimulation. To determine if electrodes preferentially stimulate ganglion cells directly below them or passing fibers from distant ganglion cells, we developed a compartmental model for electric field stimulation of the retinal ganglion cell (RGC). In this model a RGC is stimulated by extracellular electrical fields with active channels and realistic cell morphology derived directly from a neuronal tracing. Three membrane models were applied: a linear passive model, a Hodgkin-Huxley model with passive dendrites (HH), and a model composed of all active compartments (FCM) with five nonlinear ion channels. Idealized monopolar point and disk stimulating electrodes were positioned above the cell. For the HH and FCM models, the position of lowest cathodal threshold to propagate an action potential was over the soma. Brief (100 microseconds) cathodic stimuli were 20% (HH with disk electrode) to 73% (FCM with point-source) more effective over the soma than over the axon. In the passive model, the axon is preferentially stimulated versus the soma. Although it may be possible to electrically stimulate RGC's near their cell body at lower thresholds than at their axon, these differences are relatively small. Alternative explanations should be sought to explain the focal perceptions observed in previously reported patient trials.

Action Potentials↗
PubMed1985

Computer-based determination of size and shape in living cells.

Measurement of cell volume in living epithelial cells has become an important technique in studies of membrane transport processes that function in cell volume regulation. Planimetry of video images of optical sections enables the measurement of the cross sectional area of each section. Cell volume is calculated from the measured area of each section and the known focus displacements. In the past the measurement of cross section area has been done by manual positioning of a cursor superimposed on the video image. Each experiment generates approximately 200 images in which two or more cells may be analysed. We have developed a computer-based method that uses one image as a template, and allows automated area determination of successive images by template matching and digital image processing. This new method is comparable to the older method in speed and accuracy, but requires much less effort from the experimenter.

Animals↗
PubMed1984

Distribution of glycine, gamma-aminobutyric acid, glutamate decarboxylase, and gamma-aminobutyric acid transaminase in rabbit and mudpuppy retinas.

The distributions of glycine, gamma-aminobutyric acid (GABA), glutamate decarboxylase (EC 4.1.1.15), and GABA transaminase (EC 2.6.1.19) were determined in rabbit and mudpuppy retinas. In both species, peak levels of the amino acids and the enzymes occurred in the inner plexiform layer. Glutamate decarboxylase was almost entirely confined to the inner plexiform layer. Determinations were also made of the GABA content of 107 individual putative amacrine cell somas from mudpuppy retina. About 30% of those somas were found to have high endogenous GABA levels.

4-Aminobutyrate Transaminase↗
PubMed1984

Enzymes of energy metabolism in the mudpuppy retina.

The distributions of glycogen phosphorylase, hexokinase, phosphofructokinase, lactate dehydrogenase, glucose-6-phosphate dehydrogenase, citrate synthase, malate dehydrogenase, beta-hydroxyacyl CoA dehydrogenase, and adenylokinase were determined in the mudpuppy retina. Distinct differences were found in regard to the glycolytic and oxidative capacities of the various layers. In the outer retina, citric acid cycle enzymes were high while glycolytic enzymes were low. Synaptic zones were distinctly enriched in all energy-producing enzymes. Mudpuppy photoreceptors were found to be rich in phosphorylase but poor in glucose-6-phosphate dehydrogenase, suggestive of some evolutionary divergence from mammals in the metabolic machinery which is used to support the visual process.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1992

Millimolar amiloride concentrations block K conductance in proximal tubular cells.

1. Amiloride, applied at millimolar concentrations, results in the blockade of K+ conductance in amphibian proximal convoluted cells (PCT), fused into giant cells. 2. Amiloride results directly in a blockade of K+ conductance that is not related to inhibition of the Na(+)-H+ antiport, which would lower intracellular pH, adversely affecting K+ conductance. On the contrary, high amiloride concentrations promote entry of this lipophilic base in the cell, leading to higher cell pH. 3. Under voltage clamp conditions, control vs. amiloride, current-voltage curves from PCT fused giant cells intersect at -86.2 +/- 3.4 mV, a value close to the equilibrium potential for potassium. 4. Hexamethylene amiloride, 10(-5) M, irreversibly depolarizes the membrane potential. 5. Barium decreased by 50% the initial slope of realkalinization, following removal of a solution containing NH4Cl, as did amiloride. In addition, these blockers reduced membrane conductance by 40%, suggesting that a fraction of the amiloride-suppressible NH4+ efflux may be conductive. 6. Amiloride does not directly inhibit the Na(+)-K+, ATPase in our preparation, contrary to the prevalent belief. 7. In vivo studies show that amiloride interferes with an apical K+ conductance but it does not alter basolateral K+ conductance.

Amiloride↗
PubMed1998

Galanin activates an inwardly rectifying potassium conductance and inhibits a voltage-dependent calcium conductance in mudpuppy parasympathetic neurons.

Galanin-induced activation of an inwardly rectifying membrane potassium (K+) current and inhibition of barium current (IBa) were studied using whole cell voltage clamp recording techniques in parasympathetic neurons dissociated from the mudpuppy cardiac ganglion. Both activation of the K+ current and inhibition of IBa were concentration-dependent with an EC50 (or IC50) of approximately 35 nM and approximately 0.4 nM, respectively. Both actions of galanin were eliminated by pretreatment with pertussis toxin, which suggested involvement of Gi/Go protein activation. Galantide antagonized the galanin-induced activation of K+ current with an IC50 equal to 4 nM. By contrast, galantide, by itself, inhibited IBa with an EC50 equal to 16 nM. Another galanin analog, M40, primarily antagonized the galanin-induced activation of K+ current, but in some cells, M40 also acted as a weak agonist. M40, like galantide, inhibited IBa. The NH2-terminal fragment galanin-(1-16) activated the K+ current and inhibited IBa, indicating that the first 16 amino acids of the galanin peptide were sufficient for both actions. In summary, it is postulated that the effects of galanin on mudpuppy parasympathetic neurons might be mediated by activation of two different subtypes of galanin receptor, one that regulates membrane K+ conductance and a second that modulates calcium conductance.

Animals↗
PubMed1980

Further studies of the chemical sensitivity of the oscillatory potentials of the electroretinogram (ERG) I. GABA- and glycine antagonists.

The oscillatory potentials (OPs) of the mudpuppy ERG were studied to evaluate the effects of GABA and glycine antagonists. Upon exposing the retina to bicuculline and picrotoxin, blocking agents of the putative inhibitory neurotransmitter GABA, all the OPs were selectively abolished. The earlier OPs (01-03) appeared more sensitive to the drug than the latter ones (04-05). There was no appreciable effect on the range of sensitivity and saturation level of the b-wave. Low concentrations of strychnine, blocking the effect of the putative inhibitory neurotransmitter glycine, produced a selective decrease of the amplitude of the OPs. The first (01) appeared less sensitive to the drug than the later ones. Higher concentrations extinguished all the OPs but also decreased the suprathreshold amplitude of the b-wave. In conclusion, the present results in agreement with previous work, suggest that the OPs appear to have a different origin from the b-wave and seem to be generated by inhibitory feed-back circuits within retina. The differential sensitivity of the individual oscillatory peaks indicate that perhaps chemically different synaptic activities might underlie the individual oscillatory potentials.

Action Potentials↗
PubMed1981

Further studies of the chemical sensitivity of the oscillatory potentials of the electroretinogram (ERG). III. Some omega amino acids and ethanol.

The action of some omega amino acids and the influence of ethanol on the oscillatory potentials (OPs) of the ERG were tested. Low doses of beta-alanine, an inhibitory amino acid, selectively suppressed the amplitude of the OPs without affecting the a- and b-waves. The earlier OPs (O1 O2) appeared somewhat more sensitive to the drug than the later ones (O3-O5). Higher doses extinguished all the OPs but also affected the maximum amplitude of the a- and b-wave. There was no noticeable change of the threshold sensitivity of the a- and b-waves. Valine in corresponding concentrations did not affect the OPs. Taurine did not change the OPs in most experiments but in some experiments a concurrent decrease of the OPs, a- and b-waves occurred. Thus, these findings support the view that only inhibitory feed-back circuits initiated by the amacrines give rise to the OPs. Ethanol had a differential and selective effect on the OPs. The later OPs (O3-O5) were sensitive to ethanol. In conclusion, these results support the notion that the individual oscillatory peaks are likely to have different origins and that the later OPs may perhaps be related to the off-components in response to brief stimulus light.

Action Potentials↗
PubMed1986

Spatial characteristics of the oscillatory potentials of the electroretinogram.

The spatial properties of the trans- and intra-retinal oscillatory potentials (OPs), a- and b-waves of the mudpuppy electroretinogram (ERG) were analyzed. A comparison of the intra-retinal potentials was made with the proximal negative response (PNR), an extracellular response, predominantly from amacrine cells. The spatial characteristics of the OPs differed from the a- and b-waves and the PNR. The OPs integrated spatially up to full field illumination. They seem to represent summated neuronal activity over broader areas than that of the PNR and the a- and b-waves. The results indicate that the OPs reflect activity of second order neurons to which visual information converge from the whole retina. These neurons may be the bipolar cells. No evidence that the OPs orginate in the laterally extending amacrines was obtained. The b-wave data further suggested a centre-surround organization for the neurons indirectly involved in the b-wave generation.

Animals↗
PubMed1983

Sustained and transient synaptic inputs to on-off ganglion cells in the mudpuppy retina.

Synaptic inputs to on-off ganglion cells in mudpuppy retina were studied by measuring current-voltage relations in darkness, during different phases of the response to light, and in the presence of 4 mM-Co2+. The addition of Co2+ to the bathing medium usually caused a hyperpolarization of the membrane potential in darkness and an increase in input resistance, indicating that on-off ganglion cells receive tonic excitatory synaptic input in darkness. Other results suggest that an additional synaptic input, with a reversal potential near the dark potential, may also be active in darkness. At the onset of a light stimulus in the receptive field centre all on-off ganglion cells responded with transient excitatory and inhibitory synaptic events, both of which were due to increases in conductance. Similar transient excitatory and inhibitory events occurred at the termination of the light stimulus. In about one-half of the on-off ganglion cells studied the synaptic activity during steady illumination was the same as in darkness. In the remaining cells steady illumination caused an increase in sustained inhibition.

Action Potentials↗