scholarly journals Control of the cyclic GMP phosphodiesterase of frog photoreceptor membranes.

1980 ◽  
Vol 76 (5) ◽  
pp. 631-645 ◽  
Author(s):  
P R Robinson ◽  
S Kawamura ◽  
B Abramson ◽  
M D Bownds
Keyword(s):  
Light Intensity ◽  
Cyclic Gmp ◽  
Outer Segment ◽  
Calcium Concentration ◽  
Light Adaptation ◽  
Light Sensitivity ◽  
Change Mechanism ◽  
Outer Segments ◽  
Labile Component ◽  
Photoreceptor Membranes

The light-activated cyclic GMP phosphodiesterase (PDE) of frog photoreceptor membranes has been assayed in isolated outer segments suspended in a low-calcium Ringer's solution. Activation occurs over a range of light intensity that also causes a decrease in the permeability, cyclic GMP levels, and GTP levels of isolated outer segments. At intermediate intensities, PDE activity assumes constant intermediate values determined by the rate of rhodopsin bleaching. Washing causes an increase in maximal enzyme activity. Increasing light intensity from darkness to a level bleaching 5 x 10(3) rhodopsin molecules per outer segment per second shifts the apparent Michaelis constant (Km) from 100 to 900 microM. Maximum enzyme velocity increases at least 10-fold. The component that normally regulates this light-induced increase in the Km of PDE is removed by the customary sucrose flotation procedures. The presence of 10(-3) M Ca++ increases the light sensitivity of PDE, and maximal activation is caused by illumination bleaching only 5 x 10(2) rhodopsin molecules per outer segment per second. Calcium acts by increasing enzyme velocity while having little influence on Km. The effect of calcium appears to require a labile component, sensitive to aging of the outer segment preparation. The decrease in the light sensitivity of PDE that can be observed upon lowering the calcium concentration may be related to the desensitization of the permeability change mechanism that occurs during light adaptation of rod photoreceptors.

scholarly journals Amplitude, kinetics, and reversibility of a light-induced decrease in guanosine 3',5'-cyclic monophosphate in frog photoreceptor membranes.

1979 ◽  
Vol 73 (5) ◽  
pp. 629-653 ◽  
Author(s):  
M L Woodruff ◽  
M D Bownds
Keyword(s):  
Cyclic Gmp ◽  
Outer Segment ◽  
Calf Serum ◽  
Absolute Number ◽  
Continuous Illumination ◽  
Cyclic Monophosphate ◽  
Outer Segments ◽  
A Value ◽  
Anti Oxidant ◽  
Photoreceptor Membranes

The concentration of guanosine 3',5'-cyclic monophosphate (cyclic GMP) has been examined in suspensions of freshly isolated frog rod outer segments using conditions which previously have been shown to maintain the ability of outer segments to perform a light-induced permeability change (presence of calf serum, anti-oxidant, and low calcium concentration). Illumination causes a rapid decrease in cyclic GMP levels which has a half-time approximately 125 ms. With light exposures that bleach less than 100 rhodopsin molecules in each rod outer segment, at least 10(4)-10(5) molecules of cyclic GMP are hydrolyzed for each rhodopsin molecule bleached. Half of the total cyclic GMP in each outer segment, approximately 2 X 10(7) molecules, is contained in the light-sensitive pool. If outer segments are exposed to continuous illumination, using intensities which bleach between 5.0 X 10(1) and 5.0 X 10(4) rhodopsin molecules/outer segment per second, cyclic GMP levels fall to a value characteristic for the intensity used. This suggests that a balance between synthesis and degradation of cyclic GMP is established. This constant level appears to be regulated by the rate of bleaching rhodopsin molecules (by the intensity of illumination), not the absolute number of rhodopsin molecules bleached...

scholarly journals Guanosine 3',5'-cyclic monophosphate and the in vitro physiology of frog photoreceptor membranes.

1977 ◽  
Vol 69 (5) ◽  
pp. 667-679 ◽  
Author(s):  
M L Woodruff ◽  
D Bownds ◽  
S H Green ◽  
J L Morrisey ◽  
A Shedlovsky
Keyword(s):  
Plasma Membrane ◽  
Light Intensity ◽  
Cyclic Gmp ◽  
Outer Segment ◽  
Phosphodiesterase Inhibitor ◽  
Plasma Membrane Permeability ◽  
Cyclic Monophosphate ◽  
Outer Segments ◽  
In Vitro Physiology

Frog rod outer segments freshly detached from dark-adapted retinas contain approximately 1-2 molecules of guanosine 3',5'-cyclic monophosphate (cyclic GMP) for every 100 molecules of visual pigment present. This cyclic GMP decays to 5'-GMP, and the conversion is accelerated upon illumination of the outer segments. Bleaching one rhodopsin molecule can lead to the hydrolysis of 1,000-2,000 molecules of cyclic GMP within 100-300 ms. The decline in cyclic GMP concentration becomes larger as illumination increases, and varies with the logarithm of light intensity at levels which bleach between 5 X 10(2) and 5 X 10(5) rhodopsin molecules per outer segment-second. Light suppression of plasma membrane permeability, assayed in vitro as light suppression of outer segment swelling in a modified Ringer's solution, occurs over this same range of light intensity. The correlation between cyclic GMP and permeability or swelling is maintained in the presence of two pharmacological perturbations: papaverine, a phosphodiesterase inhibitor, increases both cyclic GMP levels and the dark permeability of the plasma membrane; and beta,gamma-methylene ATP increases the effectiveness of light in suppressing both permeability and cyclic GMP levels.

scholarly journals Effects of cyclic GMP on the kinetics of the photocurrent in rods and in detached rod outer segments.

1987 ◽  
Vol 90 (4) ◽  
pp. 527-551 ◽  
Author(s):  
S Hestrin ◽  
J I Korenbrot
Keyword(s):  
Cyclic Gmp ◽  
Outer Segment ◽  
Time Course ◽  
Intact Cell ◽  
Light Sensitivity ◽  
Biochemical Properties ◽  
Intact Cells ◽  
Outer Segments ◽  
High Concentrations ◽  
Kinetics Of

We investigated the effects of high concentrations of cytoplasmic cyclic GMP on the photocurrent kinetics and light sensitivity of the tiger salamander rod both in intact cells and in detached outer segments. Photoreceptors were internally perfused with cGMP by applying patch pipettes containing cGMP to the inner or outer segment. Large increases in the concentration of cGMP in the outer segment cytoplasm were achieved only when the patch pipette was applied directly to the outer segment. The dark-current amplitude increased with increasing cGMP concentrations up to approximately 1,400 pA. Internal perfusion with 5.0 mM cGMP introduced a delay of 1-3 s in the photocurrent. The magnitude of the delay was inversely proportional to the light intensity. In addition, the photocurrent time course was slowed down and the light sensitivity, measured 1 s after the flash, was decreased approximately 100-fold when compared with that of the intact cell. The observed effects of cGMP were compared with those predicted by a model that assumes that the initial photocurrent time course is determined by the kinetics of the light-activated phosphodiesterase (PDE) and the cGMP dependence of the light-sensitive channels. At high concentrations of cGMP, the experimental data were similar to those predicted by the model and based on the known biochemical properties of the light-activated PDE and cGMP-activated channels.

Cyclic AMP has no effect on the generation, recovery, or background adaptation of light responses in functionally intact rod outer segments: With implications about the function of phosducin

2000 ◽  
Vol 17 (6) ◽  
pp. 887-892 ◽  
Author(s):  
HANA JINDROVA ◽  
PETER B. DETWILER
Keyword(s):  
Outer Segment ◽  
Light Exposure ◽  
Cell Voltage ◽  
Light Sensitivity ◽  
Rod Outer Segments ◽  
Light Responses ◽  
Outer Segments ◽  
Sequence Of Events ◽  
Background Adaptation ◽  
Dependent Processes

In retinal rods, light exposure decreases the total outer segment content of both cGMP and cAMP by about 50%. The functional role of the light-evoked change in cAMP is not known. It is postulated to trigger changes in the phosphorylation state of phosducin, a phosphoprotein that is phosphorylated in the dark by cAMP-dependent protein kinase (PKA) and dephosphorylated by basal phosphatase activity when PKA is inhibited by the light-evoked drop in cAMP. In biochemical studies, dephosphorylated phosducin binds to free βγ dimer of transducin (Tβγ) and prevents the regeneration of heterotrimeric transducin by blocking the re-association of the βγ and α subunits. Phosducin's interaction with Tβγ is blocked when it is phosphorylated on a single residue by PKA. To evaluate the effect of the light-evoked fall in cAMP, functionally intact isolated lizard rod outer segments were dialyzed in whole-cell voltage clamp with a standard internal solution and electrical light responses were recorded with and without adding cAMP to the dialysis solution. Since the total outer segment content of cAMP in darkness is ∼5 μM, internal dialysis with solution containing a much higher concentration (100 μM) of cAMP (or 8-bromo-cAMP) will overcome the effects of a light-evoked decrease in its concentration by keeping cAMP-dependent processes fully activated. Neither cyclic nucleotide had any influence on the generation, light sensitivity, recovery, or background adaptation of the flash response. These results also argue against the participation of phosducin in the sequence of events that are responsible for these aspects of rod function. This does not exclude the possibility of phosducin being involved in adaptation caused by higher light levels than used in the present study, that is, bleaching adaptation, or in light-dependent processes other than phototransduction.

scholarly journals Functional significance of the taper of vertebrate cone photoreceptors

2012 ◽  
Vol 139 (2) ◽  
pp. 159-187 ◽  
Author(s):  
Ferenc I. Hárosi ◽  
Iñigo Novales Flamarique
Keyword(s):  
Outer Segment ◽  
Signal To Noise Ratio ◽  
Light Flux ◽  
Light Sensitivity ◽  
Morphological Data ◽  
Outer Segments ◽  
Primary Determinant ◽  
The One ◽  
Reduced Light ◽  
Theoretical Analyses

Vertebrate photoreceptors are commonly distinguished based on the shape of their outer segments: those of cones taper, whereas the ones from rods do not. The functional advantages of cone taper, a common occurrence in vertebrate retinas, remain elusive. In this study, we investigate this topic using theoretical analyses aimed at revealing structure–function relationships in photoreceptors. Geometrical optics combined with spectrophotometric and morphological data are used to support the analyses and to test predictions. Three functions are considered for correlations between taper and functionality. The first function proposes that outer segment taper serves to compensate for self-screening of the visual pigment contained within. The second function links outer segment taper to compensation for a signal-to-noise ratio decline along the longitudinal dimension. Both functions are supported by the data: real cones taper more than required for these compensatory roles. The third function relates outer segment taper to the optical properties of the inner compartment whereby the primary determinant is the inner segment’s ability to concentrate light via its ellipsoid. In support of this idea, the rod/cone ratios of primarily diurnal animals are predicted based on a principle of equal light flux gathering between photoreceptors. In addition, ellipsoid concentration factor, a measure of ellipsoid ability to concentrate light onto the outer segment, correlates positively with outer segment taper expressed as a ratio of characteristic lengths, where critical taper is the yardstick. Depending on a light-funneling property and the presence of focusing organelles such as oil droplets, cone outer segments can be reduced in size to various degrees. We conclude that outer segment taper is but one component of a miniaturization process that reduces metabolic costs while improving signal detection. Compromise solutions in the various retinas and retinal regions occur between ellipsoid size and acuity, on the one hand, and faster response time and reduced light sensitivity, on the other.

scholarly journals Influence of light and calcium on guanosine 5'-triphosphate in isolated frog rod outer segments.

1979 ◽  
Vol 74 (6) ◽  
pp. 649-669 ◽  
Author(s):  
M S Biernbaum ◽  
M D Bownds
Keyword(s):  
Light Intensity ◽  
Calcium Concentration ◽  
Control Mechanism ◽  
Continuous Light ◽  
Ionophore A23187 ◽  
Rod Outer Segments ◽  
Outer Segments ◽  
Induced Changes ◽  
Hydrolysis Of ◽  
External Calcium

Frog rod outer segments contain approximately 0.25 mol of GTP and 0.25 mol of ATP per mol of rhodopsin 3 min after their isolation from the retina. UTP and CTP are present at 10-fold and 100-fold lower levels, respectively. Concentrations of GTP and ATP decline in parallel over the next 4 min to reach relatively stable levels of 0.1 mol per mol of rhodopsin. Illumination reduces the concentration of endogenous GTP but not ATP. This light-induced decrease in GTP can be as large as 70% and has a half-time of 7 s. GTP is reduced to steady intermediate levels during extended illumination of intermediate intensity, but partially returns to its dark-adapted level after brief illumination. The magnitude of the decrease increases as a linear function of the logarithm of continuous light intensity at levels which bleach between 5 X 10(2) and 5 X 10(6) rhodopsin molecules/outer segment per second. This exceeds the range of intensities over which illumination causes decreases in the cyclic GMP content and permeability of isolated outer segments (Woodruff and Bownds. 1979. J. Gen. Physiol. 73:629-653). Thus, over 4 log units of light intensity, a sensitivity control mechanism functions to make extended illumination less effective in stimulating a GTP decrease. GTP levels in dark-adapted outer segments are sensitive to changes in calcium concentration in the suspending medium. If the external calcium concentration is reduced to 10(-8) M, GTP concentration is lowered to the same level caused by saturating illumination, and the GTP remaining is no longer light-sensitive. Lowering calcium concentration to intermediate levels between 10(-6) and 10(-8) M reduces GTP to stable intermediate levels, and the GTP remaining can be reduced by light. Restoration of millimolar calcium drives synthesis of GTP, but not of ATP, and GTP lability towards illumination is again observed. These calcium-induced changes in GTP are diminished by the addition of the divalent cation ionophore A23187. Lowering or raising magnesium levels does not influence the GTP concentration. These data raise the possibility that light activates either a calcium transport mechanism driven by the hydrolysis of GTP, or some other calcium-sensitive GTPase activity of unknown function. Known light-dependent reactions involving cyclic nucleotide transformations and rhodopsin phosphorylation appear to account for only a small fraction of the light-induced GTP decrease.

Dynamic, spatially nonuniform calcium regulation in frog rods exposed to light

1996 ◽  
Vol 76 (3) ◽  
pp. 1991-2004 ◽  
Author(s):  
S. T. McCarthy ◽  
J. P. Younger ◽  
W. G. Owen
Keyword(s):  
Outer Segment ◽  
Calcium Concentration ◽  
Light Adaptation ◽  
Exchange Current ◽  
Free Calcium ◽  
Test Flash ◽  
Calcium Dynamics ◽  
Fura 2 ◽  
Circulating Current ◽  
Free Calcium Concentration

1. In intact rods of the bullfrog, Rana Catesbeiana, that were loaded with Fura-2 by incubation, we made high-resolution measurements of Na:Ca,K exchange currents and measured cytosolic free calcium concentrations during exposure to steps of illumination. The calcium dynamics we observed are indicative of unmanipulated rods because Fura-2 had little effect on calcium buffering within the outer segment. 2. In the dark, the total concentration of calcium within the outer segment, determined by integrating the exchange current, was near 50 microM. The free calcium concentration in darkness was 200-400 nM, indicating that > of = 99% of the internal calcium was bound to buffer molecules or equivalently sequestered. 3. During saturating illumination, the concentration of free calcium near the membrane (assayed by the exchange current) fell more rapidly than the space-averaged free calcium concentration (measured with Fura-2), but both had time courses that were best described by a sum of three exponential terms. The time constants were the same for each assay, but the weighting factors were different. 4. The relationship between the exchange current and space-averaged calcium concentration is consistent with significant concentration gradients within the outer segment resulting from high buffering power, diffusional restrictions, and the fact that all net gain and loss of calcium occurs at the membrane. The data further indicate that effective buffering, and hence calcium mobility, is not uniform within the outer segment. 5. Calcium kinetics were independent of the calcium concentration, indicating that the dominant buffers effectively have a low affinity for calcium (KD >> [Ca2+]dark free). 6. The dynamics of calcium changes and of exchange currents evoked by saturating and nonsaturating illumination are completely predictable from changes in the circulating current. Calcium and current are related by a linear transformation, indicating that calcium fluxes within the outer segment are passive and that buffers equilibrate rapidly. 7. Although calcium concentrations change slowly with respect to changes in the circulating current, both measured and calculated calcium dynamics are well correlated with changes in light adaptation. Responses to test flashes depended weakly on the detailed time course of the adapting stimulus but strongly on the free cytosolic calcium concentration at the time the test flash was delivered.

scholarly journals Protein Kinase C Activity and Light Sensitivity of Single Amphibian Rods

1997 ◽  
Vol 110 (4) ◽  
pp. 441-452 ◽  
Author(s):  
W.-H. Xiong ◽  
K. Nakatani ◽  
B. Ye ◽  
K.-W. Yau
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Outer Segment ◽  
Light Adaptation ◽  
Light Sensitivity ◽  
Rod Photoreceptors ◽  
Physiological Conditions ◽  
Kinase C ◽  
Protein Kinase C Activity ◽  
Phototransduction Cascade

Biochemical experiments by others have indicated that protein kinase C activity is present in the rod outer segment, with potential or demonstrated targets including rhodopsin, transducin, cGMP-phosphodiesterase (PDE), guanylate cyclase, and arrestin, all of which are components of the phototransduction cascade. In particular, PKC phosphorylations of rhodopsin and the inhibitory subunit of PDE (PDE γ) have been studied in some detail, and suggested to have roles in downregulating the sensitivity of rod photoreceptors to light during illumination. We have examined this question under physiological conditions by recording from a single, dissociated salamander rod with a suction pipette while exposing its outer segment to the PKC activators phorbol-12-myristate,13-acetate (PMA) or phorbol-12,13-dibutyrate (PDBu), or to the PKC-inhibitor GF109203X. No significant effect of any of these agents on rod sensitivity was detected, whether in the absence or presence of a background light, or after a low bleach. These results suggest that PKC probably does not produce any acute downregulation of rod sensitivity as a mechanism of light adaptation, at least for isolated amphibian rods.

The photoreceptors and pigment epithelium of the adult Xenopus retina: morphology and outer segment renewal

1978 ◽  
Vol 201 (1143) ◽  
pp. 131-147 ◽  
Keyword(s):  
Outer Segment ◽  
Light Adaptation ◽  
Pigment Epithelium ◽  
Membrane Topology ◽  
Morphological Evidence ◽  
Experimental Conditions ◽  
Outer Segments ◽  
Pigment Granules ◽  
Discrete Band ◽  
Renewal Time

Outer segment renewal and the fine structure of photoreceptors and pigment epithelium (p. e.) were studied in the adult Xenopus retina by light microscopic autoradiography and electron microscopy. Following the injection of [ 3 H]leucine, the pattern of labelling observed in receptor outer segments was typical of that reported in other adult retinae: only diffuse labelling was found in cones, but in rods a discrete band of label accumulated at the base of the outer segment and migrated sclerally with time. The rate of band displacement and thus disk addition in Xenopus rods was 1.86 μm/day (or 78 disks/day), which is more than twice that reported for red rods in Rana under similar experimental conditions, although these species have similar metabolic rates. Average rod outer segment (r. o. s.) length did not change, demonstrating a balance between disk addition and shedding. R. o. s. renewal time was about 24 days, corresponding to the time when labelled phagosomes were first found in the p. e. Ultrastructurally, one kind of (red) rod and one kind of cone were found whose outer segments differed in membrane topology. Although microfilaments were found in the apical processes of the p. e. and its cytoplasm contained both pigment granules and myeloid bodies, pigment granules did not migrate into these processes during light adaptation. In addition to possible morphological evidence for phago­somes of cone origin, both large and small rod phagosomes were observed in the p. e. The latter appear to represent small stacks of partial disks shed from individual r. o. s. scallops.

Movement of retinal along cone and rod photoreceptors

1994 ◽  
Vol 11 (2) ◽  
pp. 389-399 ◽  
Author(s):  
Jing Jin ◽  
Gregor J. Jones ◽  
M. Carter Cornwall
Keyword(s):  
Cell Body ◽  
Dark Adaptation ◽  
Visual Pigment ◽  
Outer Segment ◽  
Light Adaptation ◽  
Rod Photoreceptors ◽  
Rod Outer Segment ◽  
Rods And Cones ◽  
Outer Segments ◽  
Rod Cell

AbstractSingle isolated photoreceptors can be taken through a visual cycle of light adaptation by bleaching visual pigment, followed by dark adaptation when supplied with 11–cis retinal. Light adaptation after bleaching is manifested by faster response kinetics and a permanent reduction in sensitivity to light flashes, presumed to be due to the presence of bleached visual pigment. The recovery of flash sensitivity during dark adaptation is assumed to be due to regeneration of visual pigment to pre-bleach levels. In previous work, the outer segments of bleached, light-adapted cells were exposed to 11–cis retinal. In the present work, the cell bodies of bleached photoreceptors were exposed. We report a marked difference between rods and cones. Bleached cones recover sensitivity when their cell bodies are exposed to 11–cis retinal. Bleached rods do not. These results imply that retinal can move freely along the cone photoreceptor, but retinal either is not taken up by the rod cell body or retinal cannot move from the rod cell body to the rod outer segment. The free transfer of retinal along cone but not along rod photoreceptors could explain why, during dark adaptation in the retina, cones have access to a store of 11–cis retinal which is not available to rods. Additional experiments investigated the movement of retinal along bleached rod outer segments. The results indicate that retinal can move along the rod outer segment, but that this movement is slow, occurring at about the same rate as the regeneration of visual pigment.

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