Adaptation in the Ventral Eye of Limulus is Functionally Independent of the Photochemical Cycle, Membrane Potential, and Membrane Resistance

The early receptor potential (ERP), membrane potential, membrane resistance, and sensitivity were measured during light and/or dark adaptation in the ventral eye of Limulus. After a bright flash, the ERP amplitude recovered with a time constant of 100 ms, whereas the sensitivity recovered with an initial time constant of 20 s. When a strong adapting light was turned off, the recovery of membrane potential and of membrane resistance had time-courses similar to each other, and both recovered more rapidly than the sensitivity. The receptor depolarization was compared during dark adaptation after strong illumination and during light adaptation with weaker illumination; at equal sensitivities the cell was more depolarized during light adaptation than during dark adaptation. Finally, the waveforms of responses to flashes were compared during dark adaptation after strong illumination and during light adaptation with weaker illumination. At equal sensitivities (equal amplitude responses for identical flashes), the responses during light adaptation had faster time-courses than the responses during dark adaptation. Thus neither the photochemical cycle nor the membrane potential nor the membrane resistance is related to sensitivity changes during dark adaptation in the photoreceptors of the ventral eye. By elimination, these results imply that there are (unknown) intermediate process(es) responsible for adaptation interposed between the photochemical cycle and the electrical properties of the photoreceptor.     

Two light-dependent conductances in Lima rhabdomeric photoreceptors

Light-dependent membrane currents were recorded from solitary Lima photoreceptors with the whole-cell clamp technique. Light stimulation from a holding voltage near the cell's resting potential evokes a transient inward current graded with light intensity, accompanied by an increase in membrane conductance. While the photocurrent elicited by dim flashes decays smoothly, at higher stimulus intensities two kinetically distinct components become visible. Superfusion with TEA or intracellular perfusion with Cs do not eliminate this phenomenon, indicating that it is not due to the activation of the Ca-sensitive K channels that are present in these cells. The relative amplitude of the late component vs. the early peak of the light response is significantly more pronounced at -60 mV than at -40 mV. At low light intensities the reversal potential of the photocurrent is around 0 mV, but with brighter lights no single reversal potential is found; rather, a biphasic response with an inward and an outward component can be seen within a certain range of membrane voltages. Light adaptation through repetitive stimulation with bright flashes diminishes the amplitude of the early but not the late phase of the photocurrent. These observations can be accounted for by postulating two separate light-dependent conductances with different ionic selectivity, kinetics, and light sensitivity. The light response is also shown to interact with some of the voltage-sensitive conductances: activation of the Ca current by a brief conditioning prepulse is capable of attenuating the photocurrent evoked by a subsequent test flash. Thus, Ca channels in these cells may not only shape the photoresponse, but also participate in the process of light adaptation.     

Photoreceptor adaptation in intrinsically photosensitive retinal ganglion cells

A rare type of mammalian retinal ganglion cell (RGC) expresses the photopigment melanopsin and is a photoreceptor. These intrinsically photosensitive RGCs (ipRGCs) drive circadian-clock resetting, pupillary constriction, and other non-image-forming photic responses. Both the light responses of ipRGCs and the behaviors they drive are remarkably sustained, raising the possibility that, unlike rods and cones, ipRGCs do not adjust their sensitivity according to lighting conditions ("adaptation"). We found, to the contrary, that ipRGC sensitivity is plastic, strongly influenced by lighting history. When exposed to a constant, bright background, the background-evoked response decayed, and responses to superimposed flashes grew in amplitude, indicating light adaptation. After extinction of a light-adapting background, sensitivity recovered progressively in darkness, indicating dark adaptation. Because these adjustments in sensitivity persisted when synapses were blocked, they constitute "photoreceptor adaptation" rather than "network adaptation." Implications for the mechanisms generating various non-image-forming visual responses are discussed.     

Neural response mechanisms in the photoreceptive pineal organ of goldfish

In order to classify the different cell types involved in signal transmission of the photoreceptive pineal organ of the goldfish, Carassius auratus, intra- and extracellular electrical responses were recorded from photoreceptors and second-order neurons. Photoreceptor responses to light consisted of hyperpolarizing potentials up to 30 mV. The responses were graded with intensity and their voltage-intensity relation followed the hyperbolic function V/Vmax = In/In + sigma n. Latencies varied between 500 msec for responses near threshold and 60 msec for supersaturating flashes. The response duration increased up to 60 sec for flashes 2 log units above the saturation level. Action spectra of individual photoreceptors peaked at lambda max = 530 nm and corresponded to measurements of extracellular slow mass potentials or spike potentials. Slow mass potentials exhibited similar characteristics as intracellular recorded photoreceptor potentials with respect to latency, voltage-intensity curves and spectral sensitivity. Ganglion cells showed maintained discharges under conditions of steady illumination. The discharge rate changed inversely with the logarithm of steady illumination over a range of 8 log units. The response to light flashes was purely achromatic and consisted of inhibition of the maintained discharge. The physiological properties demonstrate that the pineal organ of the goldfish is an effective functional photoreceptor organ operating both in dim and in bright light. The light-induced hyperpolarization of photoreceptors lead to an inhibition of the nervous discharge of ganglion cells. The direct flow of information from photoreceptors to ganglion cells is the basic channel of data processing in the goldfish pineal.     

Pigment migration and illumination of single photoreceptors in a moth

Summary The relation between intensity of illumination of single photoreceptors and position of the screening pigment was studied in the compound eye of the moth Deilephila elpenor. The amplitude of the response to test flashes of white light was measured in 7 dark adapted photoreceptor cells with the screening pigment in the extreme distal and in a proximal position. The experiments demonstrate that the response amplitude of individual photoreceptors varies with the pigment position. The associated variation in screening effect was 1.0 to 1.9 log units.The research reported in this study was supported by Swedish Medical Research Council, Sällskapet för Medicinsk Forskning and Stifteisen Gustaf och Tyra Svenssons Minne.     

The response to monochromatic light flashes of the oxygen consumption of honeybee drone photoreceptors

Local measurements of the fall in oxygen pressure on stimulation of slices of the retina of the honeybee drone by flashes of light were made with oxygen microelectrodes and used to calculate the kinetics of the extra oxygen consumption (delta QO2) induced by each flash. The action spectrum for delta QO2 was obtained from response-intensity curves in response to brief (40 ms) monochromatic light flashes. The action spectrum of receptor potentials was obtained with the same experimental conditions. The two action spectra match closely: they deviate slightly from the photosensitivity spectrum of the drone rhodopsin (R). The deviation is thought to be due to wavelength-dependent light scattering and absorption in the preparation. In these experiments, the visual pigment was first illuminated with orange light, which is known to convert the bistable drone photopigment predominantly to the R state from the metarhodopsin (M) state. When long (300-900 ms) light flashes were used to elicit delta QO2, the responses to different wavelengths could not be matched in time course (as for the short flashes). Flashes producing large R-to-M conversions produced a prolonged delta QO2. The prolongation did not occur after double flashes, which produced both large R-to-M and M-to-R conversions. Similar changes in the length of afterpotentials in the photoreceptor cells and in a long-lasting decrease in photoreceptor intracellular K+ activity were found after long single or double flashes. The results are interpreted to show that the initial event for stimulation by light of metabolism in the drone retina is the same as that for stimulation of electrical responses (i.e., absorption of photons by R). Absorption of photons by M can produce an inhibitory effect on this stimulation.     

Light-induced changes of sensitivity in Limulus ventral photoreceptors

The responses of Limulus ventral photoreceptors to brief test flashes and to longer adapting lights were measured under voltage clamp conditions. When the cell was dark adapted, there was a range of energy of the test flashes over which the peak amplitude of the responses (light-induced currents) was directly proportional to the flash energy. This was also true when test flashes were superposed on adapting stimuli but the proportionality constant (termed peak currently/photon) was reduced. The peak current/photon was attenuated more by brighter adapting stimuli than by less bright adapting stimuli. The peak current/photon is a measure of the sensitivity of the conductance-increase mechanism underlying the light response of the photo-receptor. The response elicited by an adapting stimulus had a large initial transient which declined to a smaller plateau. The peak current/photon decreased sharply during the declining phase of the transient and was relatively stable during the plateau. This indicates that the onset of light adaptation is delayed with respect to the onset of the response to the adapting stimulus. If the adaptational state just before the onset of each of a series of adapting stimuli was constant, the amplitude of the transient was a nearly linear function of intensity. When the total intensity was rapidly doubled (or halved) during a plateau response, the total current approximately doubled (or halved). We argue that the transition from transient to plateau, light-elicited changes of threshold, and the nonlinear function relating the plateau response to stimulus intensity all reflect changes of the responsiveness of the conductance-increase mechanism.     

Basic fibroblast growth factor-induced protection from light damage in the mouse retina in vivo

Basic fibroblast growth factor (bFGF) has proven neuroprotective efficacy in the rodent retina against a diverse array of injurious stimuli. However, there is no consensus to date as to the molecular mechanisms underlying this neuroprotection. The study presented herein demonstrates increased expression of endogenous bFGF in the albino mouse retina in response to acute exposure to sublethal levels of light stress. The increased expression correlates with significant photoreceptor protection from light damage. The neuroprotection is likely to be mediated by bFGF as we demonstrate that a shorter exposure to bright light stress that does not up-regulate bFGF fails to protect photoreceptors from light damage. Furthermore, intravitreal bFGF injection into the retina of mice 3 h prior to light damage affords almost complete photoreceptor protection from light-induced degeneration. In addition, injected bFGF induces the activation of protein kinase B and extracellular signal-regulated kinase 1/2 signalling which correlate directly with the pathways we find to be activated in response to light stress and up-regulated bFGF. Moreover, we demonstrate that both bright light pre-conditioning and intravitreal bFGF injection result in dramatic increases in levels of inactive glycogen synthase kinase 3β and cyclic AMP response element binding protein phosphorylation indicating a potential mechanism by which bFGF promotes survival of photoreceptors in vivo .     

Intracellular responses from the photosensitive pineal organ of the teleost,Phoxinus phoxinus

Summary Intracellular potentials from the isolated dark-adapted pineal organ ofPhoxinus phoxinus were recorded by using glass microelectrodes. The majority of cells had resting potentials of 20 to 35 mV and responded to light with intensitygraded hyperpolarizations. Voltage intensity curves of responses to brief flashes followed the hyperbolic tangent functionV/V _max=Iⁿ/(I ⁿ + ⁿ ).The latency of onset for responses to light stimuli near threshold was 400 ms and decreased with saturating flashes to about 50 ms. The membrane resistance decreased during the hyperpolarization. Spectral sensitivity measurements for these cells exhibited curves with _max=530 nm. Intracellular dye injection unequivocally identified this cell type as a photoreceptor cell.A second cell type with resting potentials between 30 to 40 mV exhibited a biphasic response pattern to light stimulation. The cell depolarized with dim light flashes and hyperpolarized with bright flashes. The amplitude of the hyperpolarizing component showed no saturation over an intensity range of 5 log units. Latencies and rise times were comparable to those of photoreceptor potentials. Spectral sensitivity curves peaked at longer wavelengths ( _max=550 nm) than the action spectra of photoreceptors ( _max=530 nm). It is assumed that this rare cell type represents a small class of pineal interneurons.     

ELECTROPHYSIOLOGY OF EXTRAOCULAR PHOTORECEPTORS IN THE SQUID LOLIGO FORBESI(CEPHALOPODA: LOLIGINIDAE)

The electrophysiology of extraocular photoreception in the myopsidsquid, Loligo forbesi Steenstrup 1856 has been examined. Extracellulargenerator potentials were evoked by white light flashes. Intracellularrecordings from extraocular photoreceptor cells in the parolfactorybodies of the squid demonstrated that they had resting potentialsaround –40 mV, and were depolarised by flashes of white,but not red light (>650 nm). The evoked depolarisation consistedof a transient component, followed by a steady plateau component.The amplitude of depolarisation increased with the logarithmof the light intensity and was maintained for the duration ofthe light stimulus. Action potentials were seen in some recordingsand these increased in frequency during light flash stimulation. (Received 11 February 1997; accepted 10 May 1997)     

Reversal of Photoreceptor Polarity Recorded during the Graded Receptor Potential Response to Light in the Eye of Limulus

Intracellular electrodes were inserted into single photoreceptor units of the excised lateral eye of Limulus, and preparations were selected from which graded receptor potentials of relatively large amplitude could be recorded in response to light stimuli. The experimental data indicated that the graded receptor potential does not arise solely from a collapse of the resting membrane potential of the sensory cells of the eye, since a reversal of polarity of the photoreceptor unit could be demonstrated when the eye was stimulated by light. In the recovery period following stimulation, characteristic changes in the so-called resting potential were recorded. It is suggested that these changes in the so-called resting membrane potential are electrical signs of recovery processes occurring in the photoreceptor, because the potential changes were recorded when the eye was in darkness and because the magnitudes of the potential changes were a predictable function of the intensity and duration parameters of the preceding light stimulus.     

The initial response of Limulus ventral photoreceptors to bright flashes. Released calcium as a synergist to excitation

The leading edge of the response of Limulus ventral photoreceptors to brief flashes was investigated using a voltage clamp. The leading edge of responses increases linearly with flash intensity when dim flashes produce less than one photoisomerization per square micron of cell surface. Brighter flashes accelerate the initial portion of the response, resulting in a fourth-power relationship between the magnitude of the response at brief times after the flash and the flash intensity. The onset of this nonlinearity with increasing flash intensity is determined by the local density of photoisomerizations within the receptor. Responses to bright 10-15-mum-diam spots therefore rise faster than responses to diffuse flashes producing the same number of photoisomerizations within the receptor. Background illumination shortens the response latency and suppresses the initial nonlinearity. These phenomena can be explained by a model of transduction in which light activates two parallel cascades of reactions. Particles released by the first of these cascades open ionic channels, while the second produces an agent that accelerates the rate of production of particles by the first. Injection of the calcium buffer EGTA slows the initial portion of the response to bright flashes and suppresses its nonlinearity, which suggests that the accelerating agent released by the second cascade is calcium.     

Regulation of cGMP levels by guanylate cyclase in truncated frog rod outer segments

Cyclic GMP is the second messenger in phototransduction and regulates the photoreceptor current. In the present work, we tried to understand the regulation mechanism of cytoplasmic cGMP levels in frog photoreceptors by measuring the photoreceptor current using a truncated rod outer segment (tROS) preparation. Since exogenously applied substance diffuses into tROS from the truncated end, we could examine the biochemical reactions relating to the cGMP metabolism by manipulating the cytoplasmic chemical condition. In tROS, exogenously applied GTP produced a dark current whose amplitude was half-maximal at approximately 0.4 mM GTP. The conductance for this current was suppressed by light in a fashion similar to when it is activated by cGMP. In addition, no current was produced in the absence of Mg2+, which is known to be necessary for the guanylate cyclase activity. These results indicate that guanylate cyclase was present in tROS and synthesized cGMP from exogenously applied GTP. The enzyme activity was distributed throughout the rod outer segment. The amount of synthesized cGMP increased as the cytoplasmic Ca2+ concentration of tROS decreased, which indicated the activation of guanylate cyclase at low Ca2+ concentrations. Half-maximal effect of Ca2+ was observed at approximately 100 nM. tROS contained the proteins involved in the phototransduction mechanism and therefore, we could examine the regulation of the light response waveform by Ca2+. At low Ca2+ concentrations, the time course of the light response was speeded up probably because cGMP recovery was facilitated by activation of the cyclase. Then, if the cytoplasmic Ca2+ concentration of a photoreceptor decreases during light stimulation, the Ca2+ decrease may explain the acceleration of the light response during light adaptation. In tROS, however, we did observe an acceleration during repetitive light flashes when the cytoplasmic Ca2+ concentration increased during the stimulation. This result suggests the presence of an additional light-dependent mechanism that is responsible for the acceleration of the light response during light adaptation.     

Physiological roles of Na+/Ca2+ exchange in Limulus ventral photoreceptors

In previous work we have presented evidence for electrogenic Na+/Ca2+ exchange in Limulus ventral photoreceptors (1989. J. Gen. Physiol. 93:473-492). This article assesses the contributions to photoreceptor physiology from Na+/Ca2+ exchange. Four separate physiological processes were considered: maintenance of resting sensitivity, light-induced excitation, light adaptation, and dark adaptation. (a) Resting sensitivity: reduction of [Na+]o caused a [Ca2+]o-dependent reduction in light sensitivity and a speeding of the time courses of the responses to individual test flashes; this effect was dependent on the final value to which [Na+]o was reduced. The desensitization caused by Na+ reduction was dependent on the initial sensitivity of the photoreceptor; in fully dark-adapted conditions no desensitization was observed; in light-adapted conditions, extensive desensitization was observed. (b) Excitation: Na+ reduction in fully dark-adapted conditions caused a Ca2+o-dependent depolarizing phase in the receptor potential that persisted beyond the stimulus duration and was evoked by a bright adapting flash. (c) Light adaptation: the degree of desensitization induced by a bright adapting flash was Na+o dependent, being larger with lower [Na+]o. Na+ reduction enhanced light adaptation only at intensities brighter than 4 x 10(-6) W/cm2. In addition to being Na+o dependent, light adaptation was Ca2+o dependent, being greater at higher [Ca2+]o. (d) Dark adaptation: the recovery of light sensitivity after adapting illumination was Na+o dependent. Dark adaptation after bright illumination in voltage-clamped and in unclamped conditions was faster in normal-Na+ saline than in reduced Na+ saline. The final sensitivity to which photoreceptors recovered was lower in reduced-Na+ saline when bright adapting illumination was used. The results suggest the involvement of Na+/Ca2+ exchange in each of these physiological processes. Na+/Ca2+ exchange may contribute to these processes by counteracting normal elevations in [Ca2+]i.     

Categorical and prolonged potentials are evoked when brief, intermediate-intensity flashes stimulate horseshoe crab lateral eye photoreceptors during octopamine neuromodulation.

Octopamine, a major efferent neurotransmitter in the lateral eye of the horseshoe crab (Limulus polyphemus), has previously been shown to modulate photoreceptor responses evoked by long flashes. Quantification of these data indicates that this modulation produced a genuine increase in sensitivity to light which cannot be entirely due to an increase in optical efficiency consequent on an anatomical alteration. Other previous studies demonstrated that extrinsic current can modulate Limulus lateral eye photoreceptor cells by inducing a bistable membrane potential with two distinct states. The present study was therefore undertaken to find out if octopamine could modulate visual responses by inducing prolonged and bistable polarization shifts similar to those demonstrated in several other neural systems. Intracellular microelectrodes were used to execute an electrophysiological study of the receptor potentials evoked in the lateral eye of Limulus when brief (20-ms) flashes were delivered while 50 microM octopamine perfused dark-adapted photoreceptors. The combined chemical and optical stimuli prolonged photoreceptor responses to light to the degree that they often exceeded the duration of the brief stimulus by hundreds of milliseconds. Moreover, these prolonged potentials were clearly bistable because they were categorical--either a prolongation was perceptually clear-cut and present or it was not, with no intermediate patterns being observed. During seawater control perfusions, such prolongations were absent. This appears to be the first demonstration of such categorical and prolonged potentials in a photoreceptor neuron. This finding particularly suggests that efferent-driven neuromodulation can enable the development of a persisting short-term representation of a brief stimulus, with this representation being retained at the most distal possible neural site.     

Bump amplitude,but not latency and time-course,can depend on position of illumination in Limulus photoreceptor cells

Bumps, the responses evoked by single photons in the ventral photoreceptor of Limulus polyphemus, were measured under voltage clamp conditions. The bumps were evoked by illuminating the photoreceptor either with a global flash or a small light spot (diameter about 5 m) which covers only 0.25% of the light-sensitive part of the cell membrane. The light energy of both flash types was adjusted so that each flash on average evoked one bump. Parameters of bumps evoked by local light spots in various membrane areas were compared with those evoked by light flashes which illuminated the whole photoreceptor. The results show that the bump amplitude depends on the location of the illumination. Membrane areas were found where the average value of the bump amplitude was either smaller or larger for a spot illumination than for a whole cell illumination. The latency and the shape (e.g. width) of the bumps does not depend on the location of the illumination.     

Are two photoreceptors involved in the flowering of a long-day plant?

The effect of daylength extension with narrow spectral bands on the flowering of a long-day plant, Brassica campestris L. cv. Ceres, was investigated to obtain clues to the identity of the photoreceptor involved. Extension of a 9 h photoperiod with 5 h of light pulses at various wavelengths resulted in maximal flowering occurring after irradiation at 710 nm, less at 730 nm, and none at 550, 660 and 750 nm. Flowering at 710 and 730 nm was negated by simultaneous exposures at 550 nm, but not at 660 nm. A short preirradiation at 660 nm enabled a following irradiation at 750 nm to induce flowering. This latter induction was prevented by 550 nm irradiation.
Short flashes of light at 710 nm induced flowering that was negated by a following flash at 550 nm but not at 660 nm. The negation by 550 nm radiation was prevented by subsequent flashes at 710 nm, indicating photoreversibility. A flash at 660 nm enabled subsequent light flashes at 750 nm to initiate flowering that was reversed by a following 550 nm flash.
From the results showing the necessity of red and far-red lights, it is proposed that flowering in this long-day plant is due to two photoreceptors - one is phytochrome and the other an unknown pigment with far-red, green photoreversible properties. By using fluence response data, it is deduced that the unidentified photoreceptor has weak absorption bands in the far-red, but has a strong absorption band in the green. Flowering is induced when effects of red light absorbed by phytochrome interact with effects of far-red light absorbed by the unidentified photoreceptor.     

Responses of retinal rods of the frog to light

To study the effect of the intensity, duration, spectral composition, and diameter of the light spot on the amplitude and shape of the response of single rods of the frog retina, potentials were recorded intracellularly. The rods tested could be divided into two groups on the basis of their responses to light spots of different spectral composition: those with maxima of sensitivity at 507 ± 8 nm and 442 ± 8 nm. With an increase in the intensity of light the response amplitude rose gradually and the time for the response to rise to its maximum was shortened. A bright flash temporarily inhibited the sensitivity of the cell to subsequent test flashes. If light spots of larger diameter (1000–1500 µ) were presented a delayed depolarization wave, due to illumination of the distant surroundings of the receptor, was observed in the course of recovery of the photic response; this effect was maximal for stimulation with red light and it was evidently induced by horizontal cell activity. The possible functional role of the depolarizing effect of illumination of the distant surroundings of the receptor is discussed.M. V. Lomonosov Moscow State University. Translated from Neirofiziologiya, Vol. 7, No. 1, pp. 84–92, January–February, 1975.     

The nature of rhodopsin-triggered photocurrents in Chlamydomonas. II. Influence of monovalent ions.

Chlamydomonas exhibits a sequence of a photoreceptor current and two flagellar currents upon stimulation with bright green flashes. The currents are thought to be a prerequisite for the well-known photophobic responses. In the preceding paper, we analyzed the kinetics of these currents and their dependence on extracellular divalent ions. Here, we show that the photoreceptor current can be carried by monovalent ions (K+ > NH4+ > Na+), provided that the driving force is high enough. The small residual photoreceptor current observed in the absence of Ca2+ is able to evoke flagellar currents at low extracellular pH. This demonstrates that signal transduction from the rhodopsin to the flagella is not inevitably dependent on extracellular Ca2+. Double-flash experiments exclude a contribution of intra-rhodopsin charge movements to the photoreceptor current signal. Evidence will be provided for the existence of nonlocalized K+ outward currents, which counterbalance the localized Ca2+ influx and repolarize the cell after a light flash. A model is presented that explains the different pathways for direction changes and phobic responses.     

Long-term light adaptation in photoreceptors of the housefly, Musca domestica

The photoreceptors of many animals adapt, when illuminated, by reducing their sensitivities to light and improving their response speeds. Light adaptation is usually considered to be rapid and complete within minutes. However, under bright light conditions, I show that functionally significant improvements in impulse response amplitude and speed continue over the course of an hour in photoreceptors of the fly, Musca domestica. After sustained illumination, the average information rate, a measure of signalling performance, improved by 28% in a sample of sixteen photoreceptors. This long-term light adaptation is a robust phenomenon across animals and is repeatable within the same cell when light-adapting sessions are separated by a period of darkness. White-noise analysis of voltage responses to light and current stimuli indicate that much of the long-term changes observed are attributable to an improvement in the reliability with which photoreceptors register the timing of photon absorptions. It is also found that the impedance amplitude of the photoreceptor increases during long-term adaptation, suggesting that the area of the photoreceptor's membrane is reduced.