scholarly journals A derivative of amiloride blocks both the light-regulated and cyclic GMP-regulated conductances in rod photoreceptors.

1987 ◽  
Vol 90 (5) ◽  
pp. 651-669 ◽  
Author(s):  
G D Nicol ◽  
P P Schnetkamp ◽  
Y Saimi ◽  
E J Cragoe ◽  
M D Bownds
Keyword(s):  
Plasma Membrane ◽  
Cyclic Gmp ◽  
Dark Current ◽  
Outer Segment ◽  
Light Response ◽  
Ionic Channels ◽  
Rod Photoreceptors ◽  
Low Concentrations ◽  
Excised Patches ◽  
Activated Flux

Vertebrate rod photoreceptors in the dark maintain an inward current across the outer segment membrane. The photoresponse results from a light-induced suppression of this dark current. The light-regulated current is not sensitive to either tetrodotoxin or amiloride, potent blockers of Na+ channels. Here, we report that a derivative of amiloride, 3',4'-dichlorobenzamil (DCPA), completely suppresses the dark current and light response recorded from rod photoreceptors. DCPA also blocks a cyclic GMP-activated current in excised patches of rod plasma membrane and a cGMP-induced Ca++ flux from rod disk membranes. These results are consistent with the notion that the Ca++ flux mechanism in the disk membrane and the light-regulated conductance in the plasma membrane are identical. DCPA also inhibits the Na/Ca exchange mechanism in intact rods, but at a 5-10-fold-higher concentration than is required to block the cGMP-activated flux and current. The blocking action of DCPA in 10 nM Ca++ is different from that in 1 mM Ca++, which suggests either that the conductance state of the light-regulated channel may be modified in high and low concentrations of Ca++, or that there may be two ionic channels in the rod outer segment membrane.

scholarly journals Immunocytochemical localization of opsin in the cell membrane of developing rat retinal photoreceptors.

1984 ◽  
Vol 98 (5) ◽  
pp. 1788-1795 ◽  
Author(s):  
I Nir ◽  
D Cohen ◽  
D S Papermaster
Keyword(s):  
Plasma Membrane ◽  
Outer Segment ◽  
Plasma Membranes ◽  
Photoreceptor Cells ◽  
Rod Photoreceptor ◽  
Rod Photoreceptors ◽  
Retinal Photoreceptors ◽  
Retinal Rod ◽  
Ros Formation ◽  
Developing Rat

Mature retinal rod photoreceptors sequester opsin in the disk and plasma membranes of the rod outer segment (ROS). Opsin is synthesized in the inner segment and is transferred to the outer segment along the connecting cilium that joins the two compartments. We have investigated early stages of retinal development during which the polarized distribution of opsin is established in the rod photoreceptor cell. Retinas were isolated from newborn rats, 3-21 d old, and incubated with affinity purified biotinyl-sheep anti-bovine opsin followed by avidin-ferritin. At early postnatal ages prior to the development of the ROS, opsin is labeled by antiopsin on the inner segment plasma membrane. At the fifth postnatal day, as ROS formation begins opsin was detected on the connecting cilium plasma membrane. However, the labeling density of the ciliary plasma membrane was not uniform: the proximal cilium was relatively unlabeled in comparison with the distal cilium and the ROS plasma membrane. In nearly mature rat retinas, opsin was no longer detected on the inner segment plasma membrane. A similar polarized distribution of opsin was also observed in adult human rod photoreceptor cells labeled with the same antibodies. These results suggest that some component(s) of the connecting cilium and its plasma membrane may participate in establishing and maintaining the polarized distribution of opsin.

scholarly journals Transduction persists in rod photoreceptors after depletion of intracellular calcium.

1987 ◽  
Vol 89 (2) ◽  
pp. 297-319 ◽  
Author(s):  
G D Nicol ◽  
U B Kaupp ◽  
M D Bownds
Keyword(s):  
Cyclic Gmp ◽  
Dark Current ◽  
Phosphodiesterase Inhibitor ◽  
Light Regulation ◽  
Response Duration ◽  
Light Sensitivity ◽  
Ionophore A23187 ◽  
Active Suspensions ◽  
Rod Photoreceptors ◽  
Intracellular Ca

We have examined the role of Ca++ in phototransduction by manipulating the intracellular Ca++ concentration in physiologically active suspensions of isolated and purified rod photoreceptors (OS-IS). The results are summarized by the following. Measurement of Ca++ content using arsenazo III spectroscopy demonstrates that incubation of OS-IS in 10 nM Ca++-Ringer's solution containing the Ca++ ionophore A23187 reduces their Ca++ content by 93%, from 1.3 to 0.1 mol Ca++/mol rhodopsin. Virtually the same reduction can be accomplished in 10 nM Ca++-Ringer's without ionophore, presumably via the plasma membrane Na/Ca exchange mechanism. Hundreds of photoresponses can be obtained from the Ca++-depleted OS-IS for at least 1 h in 10 nM Ca++-Ringer's with ionophore. The kinetics and light sensitivity of the photoresponse are essentially the same in the presence or absence of the ionophore in 10 nM Ca++. The addition of A23187 in 1 mM Ca++-Ringer's results in a Ca++ influx that rapidly suppresses the dark current and the photoresponse. This indicates that there is an intracellular site at which Ca++ can modulate the light-regulated conductance. Both the current and photoresponse can be restored if intracellular Ca++ is reduced by lowering the external Ca++ to 10 nM. During the transition from high to low Ca++, the response duration becomes shorter, which suggests that it can be regulated by a Ca++-dependent mechanism. If the dark current and the photoresponse are suppressed by adding A23187 in 1 mM Ca++-Ringer's, the subsequent addition of the cyclic GMP phosphodiesterase inhibitor isobutylmethylxanthine can restore the current and photoresponse. This implies that under conditions where the rod can no longer control its intracellular Ca++, the elevation of cyclic GMP levels can restore light regulation of the channels. The persistence of normal flash responses under conditions where intracellular Ca++ levels are reduced and perturbed suggests that changes in the intracellular Ca++ concentration do not cause the closure of the light-regulated channel.

Cycle GMP-activated channels of rod photoreceptors show neither fast nor slow desensitization

1990 ◽  
Vol 4 (05) ◽  
pp. 481-487 ◽  
Author(s):  
Shu-Ichi Watanabe ◽  
Gary Matthews
Keyword(s):  
Cyclic Gmp ◽  
Outer Segment ◽  
Flow System ◽  
Rod Photoreceptors ◽  
Rapid Flow ◽  
Inside Out

AbstractDesensitization of cGMP-activated channels was examined in excised, inside-out patches obtained from rod photoreceptors. Cyclic GMP was applied using a rapid-flow system in which concentration jumps are complete within 10–50 ms. In outer-segment patches containing many channels, the cGMP-dependent conductance reached a steady plateau that was maintained for tens of seconds in the presence of cGMP; thus, there was no indication of slow desensitization. However, rapid desensitization on the scale of milliseconds could not be ruled out because of limited speed of access of cGMP to the inner face of the patch membrane. To test for rapid desensitization, inner-segment patches containing only a single cGMP-activated channel were used. In these one-channel patches, there was no change in activity of the channel with time from its earliest onset after application of cGMP, indicating that rapid desensitization also did not occur.

scholarly journals Ca2+ fluxes and channel regulation in rods of the albino rat.

1996 ◽  
Vol 107 (5) ◽  
pp. 577-595 ◽  
Author(s):  
A Knopp ◽  
H Rüppel
Keyword(s):  
Plasma Membrane ◽  
Dark Current ◽  
Outer Segment ◽  
Residual Value ◽  
Light Stimulation ◽  
Temperature Dependent ◽  
Albino Rat ◽  
Rat Retina ◽  
Sources And Sinks ◽  
Channel Regulation

By use of microelectrodes, changes in the receptor current and the Ca2+ concentration were measured in the rod layer of the rat retina after stimulation by flashes or steady light. Thereby light induced Ca2+ sources, and sinks along a rod were determined in dependence of time. Thus, the Ca2+ fluxes across the plasma membrane of a mammalian rod could be studied in detail. By light stimulation, Ca2+ sources are evoked along the outer segment only. Immediately after a saturating flash, a maximum of Ca2+ efflux is observed which decays exponentially with tau = 0.3 s at 37 degrees C (4.2 s at 23 degrees C). During regeneration of the dark current, the outer segment acts as a Ca2+ sink, indicating a restoration of the Ca(2+)-depleted outer segment. These findings agree with earlier reports on amphibian rods. Further experiments showed that the peak Ca2+ efflux and tau are temperature dependent. The peak amplitude also depends on the external Ca2+ concentration. In contrast to the reports on amphibian rods, only a part of the Ca2+ ions extruded from the outer segment is directly restored. Surprisingly, during steady light the Ca2+ efflux approaches a permanent residual value. Therefore, in course of a photoresponse, Ca2+ must be liberated irreversibly from internal Ca2+ stores. There is certain evidence that the inner segment acts as a Ca2+ store. Our results show that the Ca2+ fraction of the ions carrying the dark current is proportional to the extracellular Ca2+ concentration. This indicates that the Ca2+ permeability of the plasma membrane of the rod outer segment is independent of the Ca2+ concentration.

Single cyclic GMP-activated channel activity in excised patches of rod outer segment membrane

Nature ◽  
1986 ◽  
Vol 321 (6065) ◽  
pp. 66-70 ◽  
Author(s):  
L. W. Haynes ◽  
A. R. Kay ◽  
K.-W. Yau
Keyword(s):  
Cyclic Gmp ◽  
Outer Segment ◽  
Channel Activity ◽  
Rod Outer Segment ◽  
Excised Patches

Blockers of potassium channels reduce the outward dark current in rod photoreceptor inner segments

1992 ◽  
Vol 8 (5) ◽  
pp. 479-481 ◽  
Author(s):  
Kun Yan ◽  
Gary Matthews
Keyword(s):  
Potassium Channel ◽  
Dark Current ◽  
Outer Segment ◽  
Light Response ◽  
Channel Blockers ◽  
Rod Photoreceptor ◽  
Potassium Channel Blockers ◽  
Internal Ph ◽  
Suction Electrode ◽  
Kinetics Of

AbstractThe dark current of single isolated toad rods was monitored by drawing either the inner segment or the outer segment into a suction electrode. The potassium-channel blockers tetraethylammonium (TEA) and 3,4-diaminopyridine (DAP) reduced the amplitude of the dark current when applied to the inner segment. Both drugs were less effective when applied to the outer segment, suggesting that they act at the inner segment to block part of the outward path for the dark current. In addition, DAP affected the kinetics of the light response, possibly by affecting internal pH.

scholarly journals Calcium flares and compartmentalization in rod photoreceptors

2020 ◽  
Vol 117 (35) ◽  
pp. 21701-21710 ◽  
Author(s):  
Yunzhen Li ◽  
Fabio Falleroni ◽  
Simone Mortal ◽  
Ulisse Bocchero ◽  
Dan Cojoc ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Intracellular Calcium ◽  
Outer Segment ◽  
Near Infrared ◽  
Fundamental Aspect ◽  
Calcium Dynamics ◽  
Rod Photoreceptors ◽  
Functional Gradient ◽  
Bright Flash

Rod photoreceptors are composed of a soma and an inner segment (IS) connected to an outer segment (OS) by a thin cilium. OSs are composed of a stack of ∼800 lipid discs surrounded by the plasma membrane where phototransduction takes place. Intracellular calcium plays a major role in phototransduction and is more concentrated in the discs, where it can be incorporated and released. To study calcium dynamics in rods, we used the fluorescent calcium dye CaSiR-1 AM working in the near-infrared (NIR) (excitation at 650 and emission at 664 nm), an advantage over previously used dyes. In this way, we investigated calcium dynamics with an unprecedented accuracy and most importantly in semidark-adapted conditions. We observed light-induced drops in [Ca2+]iwith kinetics similar to that of photoresponses recorded electrophysiologically. We show three properties of the rods. First, intracellular calcium and key proteins have concentrations that vary from the OS base to tip. At the OS base, [Ca2+]iis ∼80 nM and increases up to ∼200 nM at the OS tip. Second, there are spontaneous calcium flares in healthy and functional rod OSs; these flares are highly localized and are more pronounced at the OS tip. Third, a bright flash of light at 488 nm induces a drop in [Ca2+]iat the OS base but often a flare at the OS tip. Therefore, rod OSs are not homogenous structures but have a structural and functional gradient, which is a fundamental aspect of transduction in vertebrate photoreceptors.

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.

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