Survival factors in retinal degenerations.

Recent experiments on the retina have examined the effectiveness of various factors (e.g. growth factors, neurotrophins and cytokines) for enhancing survival and reducing injury of retinal neurons, such as photoreceptors and ganglion cells, whose death leads to blindness in degenerative retinal diseases. It has also been shown that retinal injury stimulates intrinsic survival mechanisms that promote survival of these neurons.

Effects of hypoxia on potassium homeostasis and pigment epithelial cells in the cat retina.

Intracellular recordings show that light-evoked hyperpolarizations of the apical and basal membranes of the cat retinal pigment epithelium (RPE) are altered by mild hypoxia. RPE cells, like glia, have a high K+ conductance, and measurements with K+-sensitive microelectrodes show that the hypoxic changes in the RPE cell are largely the result of changes in extracellular [K+] in the subretinal space [( K+]o) rather than direct effects on RPE cells. During hypoxia, light-evoked [K+]o responses and membrane responses have longer times to peak, slower and less complete recovery during illumination, and larger amplitudes. In addition to the effects on light-evoked responses, hypoxia causes a depolarization of first the apical and then the basal membranes of RPE cells under dark-adapted conditions. The basal depolarization is accompanied by a decrease in basal membrane resistance. These depolarizations appear to be caused by a rapid increase in [K+]o at the onset of hypoxia, which is maximal in dark adaptation, and smaller if the retina is subjected to maintained illumination. All of the effects are graded with the severity of hypoxia and can be observed at arterial oxygen tensions as high as 65 mmHg, although the threshold may be even higher. We argue that the origin of hypoxic [K+]o changes is probably an inhibition of the photoreceptors' Na+/K+ pump. This work then suggests that photoreceptors are more sensitive to hypoxia than previously believed, and that the high oxygen tension normally provided by the choroidal circulation is necessary for normal photoreceptor function.

Delayed basal hyperpolarization of cat retinal pigment epithelium and its relation to the fast oscillation of the DC electroretinogram.

Previous work has shown that the cat retinal pigment epithelium (RPE) is the source of two potential changes that follow the absorption of light by photoreceptors: a hyperpolarization of the apical membrane, peaking in 2-4 s, which leads to the RPE component of the electroretinogram (ERG) c-wave, and a depolarization of the basal membrane, peaking in 5 min, which leads to the light peak. This paper describes a new basal membrane response of intermediate time course, called the delayed basal hyperpolarization. Isolation of this response from other RPE potentials showed that with maintained illumination the hyperpolarization begins approximately 2 s after light onset, peaks in 20 s, and slowly ends as the membrane repolarizes over the next 60 s. The delayed basal hyperpolarization is very small for stimuli less than 4 s in duration and grows with duration, becoming approximately 15% as large as the preceding apical hyperpolarization with stimuli longer than 20 s. Extracellularly, this response contributes to the transepithelial potential (TEP) across the RPE. In response to light the TEP first rises to a peak, the c-wave, as the apical membrane hyperpolarizes. For stimuli longer than approximately 4 s, the decline of the TEP from the peak of the c-wave results partly from the recovery of apical membrane potential and partly from the delayed basal hyperpolarization. For long periods of illumination (300 s) the delayed basal hyperpolarization leads to a trough in the TEP between the c-wave and light peak. This trough is largely responsible for a corresponding trough in vitreal recordings, which has been called the "fast oscillation." The term "fast oscillation" has also been used to denote the sequence of potential changes resulting from repeated stimuli approximately 1 min in duration. In addition to the delayed basal hyperpolarization, such responses also contain a basal off-response, a delayed depolarization.

Changes in apical [K+] produce delayed basal membrane responses of the retinal pigment epithelium in the gecko.

We describe here a new retinal pigment epithelium (RPE) response, a delayed hyperpolarization of the RPE basal membrane, which is initiated by the light-evoked decrease of [K+]o in the subretinal space. This occurs in addition to an apical hyperpolarization previously described in cat (Steinberg et al., 1970; Schmidt and Steinberg, 1971) and in bullfrog (Oakley et al., 1977; Oakley, 1977). Intracellular and extracellular potentials and measurements of subretinal [K+]o were recorded from an in vitro preparation of neural retina-RPE-choroid from the lizard Gekko gekko in response to light. Extracellularly, the potential across the RPE, the transepithelial potential (TEP), first increased and then decreased during illumination. Whereas the light-evoked decrease in [K+]o predicted the increase in TEP, the subsequent decrease in TEP was greater than predicted by the reaccumulation of [K+]o. Intracellular RPE recordings showed that a delayed hyperpolarization generated at the RPE basal membrane produced the extra TEP decrease. At light offset, the opposite sequence of membrane potential changes occurred. RPE responses to changes in [K+]o were studied directly in the isolated gecko RPE-choroid. Decreasing [K+]o in the apical bathing solution produced first a hyperpolarization of the apical membrane, followed by a delayed hyperpolarization of the basal membrane, a sequence of membrane potential changes identical to those evoked by light. Increasing [K+]o produced the opposite sequence of membrane potential changes. In both preparations, the delayed basal membrane potentials were accompanied by changes in basal membrane conductance. The mechanism by which a change in extracellular [K+] outside the apical membrane leads to a polarization of the basal membrane remains to be determined.

Potassium modulation of taurine transport across the frog retinal pigment epithelium.

Net taurine transport across the frog retinal pigment epithelium-choroid was measured as a function of extracellular potassium concentration, [K+]o. The net rate of retina-to-choroid transport increased monotonically as [K+]o increased from 0.2 mM to 2 mM on the apical (neural retinal) side of the tissue. No further increase was observed when [k+]o was elevated to 5 mM. The [K+]o changes that modulate taurine transport approximate the light-induced [K+]o changes that occur in the extracellular space separating the photoreceptors and the apical membrane of the pigment epithelium. The taurine-potassium interaction was studied by using rubidium as a substitute for potassium and measuring active rubidium transport as a function of extracellular taurine concentration. An increase in apical taurine concentration, from 0.2 mM to 2 mM, produced a threefold increase in active rubidium transport, retina to choroid. Net taurine transport can also be altered by relatively large, 55 mM, changes in [Na+]o. Apical ouabain, 10(-4) M, inhibited active taurine, rubidium, and potassium transport; in the case of taurine, this inhibition is most likely due to a decrease in the sodium electrochemical gradient. In sum, these results suggest that the apical membrane contains a taurine, sodium co-transport mechanism whose rate is modulated, indirectly, through the sodium pump. This pump has previously been shown to be electrogenic and located on the apical membrane, and its rate is modulated, indirectly, by the taurine co-transport mechanism.

Ba2+ unmasks K+ modulation of the Na+-K+ pump in the frog retinal pigment epithelium.

This paper presents electrophysiological evidence that small changes in [K+]o modulate the activity of the Na+-K+ pump on the apical membrane of the frog retinal pigment epithelium (RPE). This membrane also has a large relative K+ conductance so that lowering [K+]o hyperpolarizes it and therefore increases the transepithelial potential (TEP). Ba2+, a K+ channel blocker, eliminated these normal K+-evoked responses; in their place, lowering [K+]o evoked an apical depolarization and TEP decrease that were blocked by apical ouabain or strophanthidin. These data indicate that Ba2+ blocked the major K+ conductance(s) of the RPE apical membrane and unmasked a slowing of the normally hyperpolarizing electrogenic Na+-K+ pump caused by lowering [K+]o. Evidence is also presented that [K+]o modulates the pump in the isolated RPE under physiological conditions (i.e., without Ba2+). In the intact retina, light decreases subretinal [K+]o and produces the vitreal-positive c-wave of the electroretinogram (ERG) that originates primarily in the RPE from a hyperpolarization of the apical membrane and TEP increase. When Ba2+ was present in the retinal perfusate, the apical membrane depolarized in response to light and the TEP decreased so that the ERG c-wave inverted. The retinal component of the c-wave, slow PIII, was abolished by Ba2+. The effects of Ba2+ were completely reversible. We conclude that Ba2+ unmasks a slowing of the RPE Na+-K+ pump by the light-evoked decrease in [K+]o. Such a response would reduce the amplitude of the normal ERG c-wave.

Origin and organization of pigment epithelial apical projections to cones in cat retina.

The apical surface of the retinal pigment epithelial cells (RPE) in the cat extend long sheetlike membranes that wrap concentrically above and around cone outer segments forming the cone sheath. The origin and organization of these sheetlike projections were studied in serial sections by electron microscopy. The apical surface of the RPE cells was found to consist of a thin zone of anastomosing ridges, or microplicae, from which longer projections extend. The lamellar projections forming the cone sheath originate from the microplicae as small cytoplasmic tabs that rapidly expand into broader sheets. Growth of individual sheets to their final size and shape continues by lateral and longitudinal expansion, fusion, and subdivision of the membrane. The small area of connection to the cell body allows the lamellae to overlap and interdigitate in forming the complex organization of the sheath. Microfilaments but not microtubules extend into the apical processes. RPE cilia (9 + 0 microtubules) with associated basal bodies, striated rootlets, and microtubules mark the location of retinal cones. These structures may be part of a microtubule organizing center that participates in morphogenesis of the cone sheath. They also may be involved in anchoring the apical projections forming the sheath, or in the movement of apical projections during the phagocytosis of outer segment discs shed from cone tips.

Disc morphogenesis in vertebrate photoreceptors.

Electron microscopic examination of the bases of adult rod and cone outer segments (rhesus monkey, ground squirrel, and grey squirrel) has led to a new model of disc morphogenesis. In this model the disc surfaces and disc rims develop by separate mechanisms and from separate regions of the membrane of the inner face of the cilium. This membrane is alternately specified into regions that will form either the disc surfaces or the disc rims. The disc surfaces develop by an evagination or outpouching of the ciliary membrane. The two surfaces of an evagination, scleral and vitreal, each form one of the surfaces of adjacent discs. The disc rim is initially specified as a region of ciliary membrane between adjacent disc-surface evaginations. This region grows bilaterally around the circumferences of adjacent discs, zippering together the apposed surfaces to form the rim and completed disc. At the same time it seals the plasma-membrane edges of the evaginations, which have become detached from the surfaces. Incisures form in rod discs by infolding of the rim and surfaces together, and they begin to form before the rim is completed around the disc perimeter. When a number of new discs are developing simultaneously the ciliary membrane at the base of an outer segment consists of a stack of rim forming and surface forming growth points. This model provides, in addition, for the continuous renewal of outer-segment plasma membrane. It also establishes a developmental basis for the structural uniqueness of the disc rim. Finally, it indicates an evolutionary relationship between the discs of vertebrate visual cells and the membrane specializations of invertebrate visual cells.

Contribution from proximal retina to intraretinal pattern ERG: the M-wave.

Intraretinal electroretinographic (ERG) responses were recorded from cat to spatial square wave gratings that were reversed in contrast (pattern ERG, PERG). Maximum 8 Hz PERG amplitudes occurred in proximal retina. Intraretinal ERGs to circular spots (photopic) also were maximal in proximal retina and resembled the M-waves of cold-blooded retinas. The temporal transforms of M-wave responses to 8 Hz flicker (to simulate contrast reversal conditions) imitated the 8 Hz PERG, suggesting that the M-wave may contribute significantly to the PERG.

Mechanisms of hypoxic effects on the cat DC electroretinogram.

Mild hypoxia elevates the standing potential and alters three slow components of the DC electroretinogram in the cat: the c-wave, the fast-oscillation trough, and the light peak. This paper considers the cellular mechanisms of these effects. Elevation of the standing potential results from a depolarization of the basal membrane of retinal pigment epithelial (RPE) cells. The depolarization is indirectly initiated by an elevation of [K+]0 in the subretinal space during hypoxia, and is accompanied by a decrease in basal membrane resistance that leads to an increase in the c-wave. There is also some evidence that hypoxia may alter the standing potential by directly affecting the basal membrane of the RPE. The fast-oscillation trough, which follows the c-wave when illumination is maintained, deepens during hypoxia. This is caused primarily by an increase in the amplitude of the delayed hyperpolarization of the RPE basal membrane that results from a slowing of the rate of recovery of light-evoked [K+]0 during hypoxia. The changes in [K+]0 probably result, in turn, from a decrease in the rate of the photoreceptors' Na+/K+ pump. The light peak's amplitude is reduced during hypoxia and its time-to-peak is lengthened, and this may be related to a change in photoreceptor metabolism that is distinct from the effect on the Na+/K+ pump. Knowledge of these mechanisms may eventually enhance the clinical usefulness of the standing potential and the c-wave, fast-oscillation, and light peak.

Effects of dopamine on the chick retinal pigment epithelium. Membrane potentials and light-evoked responses.

Dopamine, a retinal neurotransmitter, is known to affect electrical measures of retinal pigment epithelial (RPE) function: the standing potential and the DC ERG. To locate the origin of these effects, studies were performed on in vitro preparations of chick retina-RPE-choroid, which were separately perfused on the retinal and choroidal tissue surfaces. Dopamine (250 micrograms) in the retinal bath depolarized the RPE basal membrane, decreased the apparent basal membrane resistance (Rba) and increased the ERG c-wave. At concentrations less than or equal to 100 microM, retinal dopamine often caused a transient basal membrane hyperpolarization, accompanied by an apparent increase in Rba and decrease in c-wave. Surprisingly, 20-100 microM choroidal dopamine induced similar changes in basal membrane potential, resistance and c-wave amplitude, and the transient hyperpolarization and increase in Rba were often more pronounced than at comparable concentrations of retinal dopamine. Experiments in RPE-choroid preparations suggested that the effects of retinal dopamine were not secondary to effects on the neural retina. The effects of retinal and choroidal dopamine in the same tissue often were distinct, suggesting separate receptor populations on the apical and basolateral membranes of the RPE. The c-wave changes could be explained by the changes in Rba, and not by an effect on the light-evoked decrease in subretinal [K+]0. Choroidal perfusion with 50 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS), which appears to block a Cl- conductance in chick RPE, blocked the effects of dopamine perfusion on either side of the tissue. These results suggest that perfusion with either retinal or choroidal dopamine leads to electrical effects on the RPE basal membrane, possibly via a second-messenger system affecting a basal membrane Cl- conductance. Dopamine could suppress the "light-peak" depolarization of the RPE basal membrane. When either retinal or choroidal dopamine induced a large net change in trans-tissue potential (originating as a change in basal membrane potential), the light peak was severely depressed, while smaller changes produced correspondingly smaller decreases in light-peak amplitude. We found, however, that light-peak amplitude was not significantly reduced when there was little net change in the trans-tissue potential, even though dopamine may have produced sizable transient effects. Thus, despite apparent occupation of dopamine receptors on the RPE, the light peak persisted under these conditions. Similar relations between light-peak amplitude and net change in trans-tissue potential have been observed for a variety of different conditions, suggesting that the effect of dopamine on the light peak is nonspecific.

Cobalt increases photoreceptor-dependent responses of the chick retinal pigment epithelium.

While using cobalt (Co2+) to block synaptic transmission in an in vitro preparation of chick retina, the authors observed significant changes in the DC electroretinogram (DC ERG). Cobalt (3.0 mM) increased the amplitudes of all three photoreceptor-dependent responses of the retinal pigment epithelium (RPE): the c-wave, fast-oscillation trough, and light peak. Intracellular recordings from RPE cells revealed that Co2+ increased those light-evoked changes in RPE membrane potentials that contribute to each of these responses. Monitoring of subretinal [K+]o with K(+)-selective microelectrodes showed that Co2+ increased the amplitude of the light-evoked [K+]o decrease, and this must contribute to the observed increase in c-wave and fast-oscillation trough because both are generated by this [K+]o change. Cobalt also increased the initial rate of [K+]o decrease at light-onset, the rate of subretinal [K+]o reaccumulation during maintained illumination, and the amplitude of the [K+]o overshoot at light-offset. The Co(2+)-induced increases in light-evoked RPE responses and subretinal [K+]o changes may arise from a direct effect on photoreceptors because (1) blockade of postphotoreceptoral activity did not block these effects of Co2+; and (2) Co2+ did not alter significantly the electrical properties of isolated RPE-choroid tissues. The authors conclude that, in the cone-dominated chick retina, Co2+ may act directly on the photoreceptors to increase the light response.

Dihydropyridine-sensitive calcium currents in freshly isolated human and monkey retinal pigment epithelial cells.

PURPOSE: The authors previously reported that rat retinal pigment epithelial (RPE) cells exhibit an ionic current through voltage-operated calcium channels that is dihydropyridine sensitive. They attempted to record the same current from freshly isolated fetal or adult primate RPE cells, as well as from cultured cells. METHODS: The whole-cell version of the patch-clamp technique was applied to RPE cells freshly isolated by enzymatic dissociation from fetal human and adult human and monkey eyes, as well as cultured fetal and adult human RPE cells. The cells were loaded with cesium to minimize potassium-related current and were bathed in an extracellular solution containing 40 mM barium to intensify the calcium currents. RESULTS: Freshly isolated cells, both fetal and adult, showed sustained and inward-going barium current through voltage-operated calcium channels with membrane depolarizations from a high holding voltage of -60 mV. This current was affected by dihydropyridine compounds. Cultured human RPE cells showed no sign of a calcium current of this type. CONCLUSIONS: Freshly isolated fetal and adult human RPE cells, as well as adult monkey, exhibit calcium current through voltage-operated, dihydropyridine-sensitive channels, similar to the neuronal L-type, just as in rat RPE cells.

Light peak of cat DC electroretinogram: not generated by a change in [K+]0.

We have inquired whether a change in [K+]0 is associated in time with the light peak of the DC electroretinogram (ERG). If it is, this would implicate a change in [K+]0 as participating in the generation of the light peak. (The same response is usually recorded in humans by the the indirect technique of electro-oculography, giving the electro-oculogram (EOG).) Changes in [K+]0 were recorded with double-barrel, K+-specific microelectrodes from the retina and vitreous of the dark-adapted intact cat eye in response to maintained illumination. The light peak of the DC ERG was recorded from the reference barrel of the microelectrode when it was placed in the vitreous. No correlation was observed between the light peak and the changes in [K+]0 that were recorded in the retina or vitreous. We conclude that a retinal or vitreal change in [K+]0 is not directly involved in the mechanism that generates the light peak of the DC ERG.

Phagosome movement and the diurnal pattern of phagocytosis in the tapetal retinal pigment epithelium of the opossum.

The diurnal pattern of phagocytosis and the movement of phagosomes was studied in the tapetal retinal pigment epithelium (RPE) of the opossum, Didelphis virginiana. The opossum was chosen because of its rod-dominated retina and large tapetal RPE cells (up to 100 micrometers in height), which are packed with reflective granules and contain little melanin. Thus phagosomes and their passage from apical to basal cell border were easily seen. Opossums were maintained on a 12 hr light/12 hr dark cycle and were sacrificed during the day and night. Phagosomes were consulted by light microscopy in sections extending 2 mm along the eye's vertical meridian. The diurnal pattern of rod phagocytosis was generally similar to that reported for other species, although an elevated phagosome content in two animals that in some cases there might be a smaller nighttime peak in addition to the large burst occurring after light onset. To determine the spatial distribution of phagosomes at different times, the RPE cells were divided into apical, middle, and basal thirds, and the phagosomes in each region were counted. A large number of phagosomes was observed in the basal RPE 1 to 2 hr after light onset, after which the number declined, reaching a low level late in the light period. In contrast, there were few phagosomes in the apical and mid-RPE even after light onset. This suggested that phagosomes remained in the apical and mid-RPE for only a short time. To examine its effect on phagosome movement, colchicine was injected intravitreally prior to the burst of phagocytosis. Colchicine blocked the movement of phagosomes producing a row of phagosomes along the apical margin of the RPE. These results suggest that there is a rapid, microtubule-mediated movement of phagosomes from apical to basal border.

Phagosome degradation in the tapetal retinal pigment epithelium of the opossum.

Ultrastructural and cytochemical features of phagosome degradation were examined in the tapetal retinal pigment epithelium (RPE) of the opossum. Didelphis virginiana. The tapetal RPE cells of the opossum measure as much as 100 micrometers in thickness, and the phagosomes traverse these cells so as to occupy a narrow region along the basal border. Both ultrastructural and cytochemical observations showed that degradation of phagosomes by lysosomes occurs only in this basal region. Acid phosphatase activity was present only in the basal RPE, where phagosomes appeared degraded and were observed to interact with each other and with lysosomes. Phagosomes in the apical and mid-RPE always had two membranes surrounding the discs and were acid phosphatase negative. Ultrastructural changes, which may occur in the absence of lysosomal enzymes, were examined in phagosomes that were, on the basis of several criteria, undegraded. These changes were accentuated in phagosomes trapped in the apical RPE by colchicine.

Localized depigmentation of the retinal pigment epithelium and macrophage invasion of the retina in the bullfrog.

An unusual condition of the inferior retinal pigment epithelium (RPE) and neural retina has been observed in essentially all the large bullfrogs examined (Rana catesbeiana, 13 to 20 cm body length, supplied from the U.S. West Coast and Midwest). By ophthalmoscopy the inferior fundus exhibited numerous white spots and lines, which were found by light microscopy to be overlain by smaller black dots and lines. Closer examination revealed that the light areas were regions of depigmented RPE and that the black dots and lines were melanosome-laden macrophages within the adjacent retina. Further examination by light microscopy and electron microscopy allowed the formulation of the following sequence. (1) Monocytes in the choroidal capillaries crossed Bruch's membrane and passed vitreally between adjacent RPE cells. (2) In the subretinal space monocytes transformed into phagocytic macrophages, which became engorged with melanin granules and other RPE inclusions, whereas nearby RPE cells became much thinner and very depigmented. (3) The pigment-laden macrophages then moved vitreally into the avascular neural retina. Although in most areas only the RPE appeared affected by macrophage invasion, occasional localized photoreceptor disruption occurred. The severity of the lesion varied with frog size, being pronounced in large frogs, moderate in medium-sized animals, and absent in small frogs. Because the pigmentary changes were localized to the inferior part of the eye (which receives the most light from the sun overhead) of large bullfrogs (which are likely old), this phenomenon may be due to a change in RPE melanin granules resulting from the cumulative effect of light exposure.

Voltage-operated calcium channels in fresh and cultured rat retinal pigment epithelial cells.

PURPOSE: There is little known about the membrane properties of retinal pigment epithelial (RPE) cells with respect to calcium. The authors attempted to characterize membrane calcium channels from solitary fresh and cultured RPE cells from normal and dystrophic rat retinas. METHODS: RPE cells were enzymatically dissociated from eyes of neonatal rats of several strains, including dystrophic RCS strains. Membrane currents were recorded using the whole-cell version of the patch-clamp technique from either fresh or cultured cells. RESULTS: The authors observed sustained high-voltage-activated calcium channels that were dihydropyridine sensitive and closely resembled neuronal L-type calcium channels. The RCS-rdy+p+ strain was mainly investigated, but high-voltage-activated calcium channels were also recorded from fresh RPE cells of other rats regardless of age or strain, including RCS p+, RCS rdy+, Long Evans, Sprague Dawley, and also cultured RPE cells taken from a neonatal Long Evans strain. Low-voltage-activated calcium channels were not observed in any of these cells. CONCLUSION: Voltage-operated calcium channels of the L-type are the main calcium channels present in rat RPE cells. Cultured cells retained the identical channels. The dystrophic RCS strains (studied until 17 days postnatal) also exhibited these channels.

Mechanisms of effects of small hyperosmotic gradients on the chick RPE.

This paper presents electrophysiological findings of the effects of small trans-tissue osmotic gradients on the chick retinal pigment epithelium (RPE). These gradients are similar to those produced in the human "hyperosmolarity response," a clinical test of RPE integrity. Effects of 25 mOsm osmotic gradients (mannitol) were observed on the electrical parameters of the tissue and on light-evoked responses in a preparation of chick neural retina-RPE-choroid. Making the retinal perfusate hyperosmolar to the choroidal side, (retinal hyperosmolarity), depolarized the RPE basal membrane and increased the amplitude of the light-evoked c-wave of the ERG. Retinal hyperosmolarity also decreased RPE basal membrane resistance as estimated from measurements of resistance parameters (trans-tissue resistance and a, the RPE membrane resistance ratio), and of RPE membrane polarizations during the c-wave. Choroidal hyperosmolarity led to a hyperpolarization of the basal membrane and a decrease in the amplitude of the light-evoked c-wave. Measurements of resistance parameters indicated an increase in basal membrane resistance. In addition, hyperosmolar loads of either direction decreased the amplitude of the light peak of the DC-ERG. The effects on the light-evoked c-wave and light-peak responses did not occur with bilateral hyperosmolarity, indicating that a trans-epithelial osmotic gradient is necessary for the effects on the RPE. We conclude that the basal membrane of the RPE is the principal site of the effects of small hyperosmotic loads of either direction, and that the ERG c-wave is a sensitive measure of the effects on RPE basal membrane resistance.

Effects of current clamp on chick retinal pigment epithelium.

The basal membrane of the retinal pigment epithelium (RPE) is the origin of two components of the electroretinogram, the fast oscillation and the light peak. Both of these responses originate from changes in basal membrane potential (Vba), and both are associated with changes in basal membrane resistance (Rba). In addition, many experimental manipulations that alter Vba also produce apparent changes in Rba. These findings raise the possibility that the basal membrane contains a voltage-sensitive conductance that operates in the physiologic range and is involved causally in light-evoked and other responses. We report the results of current clamp experiments on the isolated retina-RPE-choroid of chick that were designed to test for the presence of such a voltage-sensitive conductance in the basal membrane. Depolarizing Vba by 15 mV with retina-to-choroid current had essentially no effect on either the ratio of membrane resistances (Rap/Rba) or the transtissue resistance (RTotal), indicating no alteration in Rba. In contrast, hyperpolarizing Vba by 15 mV with choroid-to-retina current caused a gradual decrease in RTotal and increase in Rap/Rba. Analysis of accompanying changes in membrane voltages and changes in intracellular c-wave amplitude suggested that the most likely cause of the decrease in RTotal is a decrease in paracellular resistance. Voltage-sensitive conductances of the basal membrane appear to play little or no role in the resistance changes that accompany changes in Vba in the physiologic range. The conductance changes underlying the fast oscillation and light peak probably result from either the modulation of channels by second messengers or changes in intracellular ion concentration.