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.     

Circadian rhythms in Limulus photoreceptors. II. Quantum bumps

The light response of the lateral eye of the horseshoe crab, Limulus polyphemus, increases at night, while the frequency of spontaneous discrete fluctuations of its photoreceptor membrane potential (quantum bumps) decreases. These changes are controlled by a circadian clock in the brain, which transmits activity to the eye via efferent optic nerve fibers (Barlow, R. B., S. J. Bolanski, and M. L Brachman. 1977. Science. 197:86-89). Here we report the results of experiments in which we recorded from single Limulus photoreceptors in vivo for several days and studied in detail changes in their physiological and membrane properties. We found that: (a) The shape of (voltage) quantum bumps changes with the time of day. At night, spontaneous bumps and bumps evoked by dim light are prolonged. The return of the membrane potential to its resting level is delayed, but the rise time of the bump is unaffected. On average, the area under a bump is 2.4 times greater at night than during the day. (b) The rate of spontaneous bumps decreases at night by roughly a factor of 3, but their amplitude distribution remains unchanged. (c) The resting potential and resistance of the photoreceptor membrane do not change with the time of day. (d) the relationship between injected current and impulse rate of the second order neuron, the eccentric cell, also remains unchanged with the time of day. Thus the efferent input from the brain to the retina modulates some of the membrane properties of photoreceptor cells. Our findings suggest that the efferent input acts on ionic channels in the membrane to increase the sensitivity of the photoreceptor to light.     

Development and plasticity of mitochondria and electrical properties of the cell membrane in blowfly photoreceptors

Blowfly photoreceptors are highly energy demanding sensory systems. Their information processing efficiency is enabled by the high temporal resolution of the cell membrane, requiring heavy metabolic support by the mitochondria. We studied the developmental changes of the mitochondrial apparatus and electrical properties of the photoreceptor membrane in the white eyed Calliphora vicina Chalky. Using in vivo microspectrophotometry and Western blot analysis, we found an age-dependent increase in the concentration of mitochondrial pigments. The maximal change occurred during the first week. The age-related changes were smaller in dark-bred than in light-bred flies. The mitochondrial pigment content increased after the switch from dark to light rearing and decreased after the switch from light to dark rearing. The electrical parameters of the photoreceptors were investigated with intracellular recordings. The resting membrane resistance and time constant decreased significantly after eclosion. The decrease was again most significant during the first week of adult life, paralleled with changes in the Na/K pump-dependent hyperpolarizing afterpotential. We conclude that the photoreceptor mitochondria exhibit remarkable ontogenetic and phenotypic plasticity, because the quantity of mitochondrial pigments tightly follows the development of the cell membrane as well as the energy demands of the photoreceptors under different rearing conditions.     

Immunochemical evidence for the light-regulated modulation of phosphatidylinositol-4,5-bisphosphate in rat photoreceptor cells

Immunocytochemical localization of phosphatidylinositol-4,5-bisphosphate (PIP2) in the rat rod photoreceptor outer segments (OS) was investigated with rabbit antiPIP2 antibodies. The OS of the light-adapted rat eye showed little or no staining, whereas the OS of the dark-adapted eye were intensely stained for PIP2. The immunoreactivity of photoreceptor PIP2 in the eye exposed to a brief flash of light was markedly reduced. However, subsequent dark-adaptation of the flash-bleached eye resulted in a rapid recovery of PIP2 immunoreactivity; dark-adaptation for 5 min was sufficient for recovery to the fully dark-adapted level. In dark-adapted eyes exposed to graded light intensities, the PIP2 immunostaining varied with light levels and was correlated with unbleached rhodopsin concentrations. These results suggest that PIP2 in the rat photoreceptor cells is rapidly hydrolyzed upon light exposure and rapidly synthesized in the dark and that the decrease of PIP2 level is triggered by photic bleaching of rhodopsin.     

Electrophysiological organization of the eye of Aplysia

The eye of Aplysia californica was studied by electrophysiological and histological methods. It has a central spheroidal lens which is surrounded by a retina composed of several thousand receptor cells which are replete with clear vesicles, pigmented support cells, neurons which contain secretory granules, and glial cells. The thin optic nerve that connects the eye to the cerebral ganglion gives a simple "on" response of synchronized action potentials. Tonic activity occurs in the optic nerve in the dark and is dependent on previous dark adaptation. Micropipette recordings indicate that the ERG is positive (relative to a bathelectrode) on the outer surface of the eye and negative in the region of the distal segments of the receptors. Intracellular recordings show that receptor cells have resting potentials of 40–50 mv and respond to illumination with graded potentials of up to 55 mv. Dark-adapted receptors exhibit discrete bumps on the graded response to brief light flashes. Other elements in the retina that do not give large graded responses fall into two classes. One class responds to illumination with action potentials that are in synchrony with the extracellularly recorded compound optic nerve potentials. The other class is tonically active and is depolarized or hyperpolarized and inhibited upon illumination. It is apparent that complex excitatory and lateral inhibitory interactions occur among the elements of the retina.     

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.     

Diurnal modulation of photoreceptor potassium conductance in the locust

Single electrode clamp techniques demonstrated diurnal changes in photoreceptor membrane conductance, recorded intracellularly in the intact, dark-adapted retina of the locust Schistocerca gregaria. In the day, locust photoreceptors exhibited the membrane properties of fast cells, as previously defined in rapidly moving diurnal Diptera. Depolarization activated a powerful potassium conductance with two kinetic components, one rapidly activating close to resting potential and the other activating more slowly when further depolarized, giving a pronounced delayed rectification. There was little inactivation. At night, locust photoreceptors resembled slow cells, as defined in weakly flying crepuscular and nocturnal Diptera. Depolarization rapidly activated an outward current which then inactivated over 100 ms to reduce rectification. The change from day to night state was mimicked by applying 10 mM serotonin extracellularly to the retina. We conclude that the potassium conductances of locust photoreceptor membranes are modulated according to a diurnal rhythm, possibly by serotonin. This neuromodulation is used to match photoreceptor membrane properties to photic habitat. Our findings suggest a definite and potentially widespread function for serotonin as a mediator of diurnal changes in the insect visual system.     

Membrane maintenance and electrical properties of photoreceptors of wild-type andrpa (receptor potential absent) mutant blowflies (Calliphora erythrocephala)

Summary The maintenance of photoreceptor cell membranes in the blowfly was investigated in relation to the diurnal cycle, age, and therpa (receptor potential absent) phototransduction mutation. The effect of disturbed membrane assembly on the electrical membrane properties was examined using single-electrode discontinuous current-clamp techniques. In wild-type flies the cross-sectional dimensions of the rhabdomeres were markedly reduced with age, and the quantity of synthetic organelles decreased concurrently, whereas no correlation was found between the diurnal cycle and membrane turnover. Therpa mutation is thought to block the visual transduction cascade in photoreceptor cells and to lead to degeneration of the photoreceptor cell bodies. The volume of rhabdomeres decreased markedly inrpa mutants and the quantity of synthetic organelles was reduced significantly, indicating an imbalance between photoreceptive membrane renewal and degradation. Also, the plasma membrane underwent degenerative changes. The passive electrical properties of photoreceptor cells — resting membrane voltages and input resistances — were only slightly changed from those of wild-type flies, although the photoreceptive membrane did not depolarize in response to light. This indicates no apparent disturbance in the function of the ionic channels in these membranes. Taken together, these results suggest that the photoreceptor cells need a functional phototransduction cascade with its feedback controls to maintain continuous renewal of rhabdomeres, but that the plasma membrane maintains its normal electrochemical properties despite extreme morphological degeneration of photoreceptor cell.     

Limulus ventral eye. Physiological properties of photoreceptor cells in an organ culture medium

Ventral photoreceptor cells bathed in an organ culture medium typically have resting potentials of -85 mV and membrane resistances of 35 Momega and, when dark-adapted, exhibit large potential fluctuations (LPFs) of 60 mV and small potential fluctuations (SPFs) of less than 30 mV. LPFs appear to be regenerative events triggered by SPFs, the well-known quantum bumps. In the dark, SPFs and LPFs occur spontaneously. At intensities near threshold, the rate of occurrence is directly proportional to light intensity, indicating that SPFs and LPFs are elicited by single photon events. At higher intensities, SPFs and LPFs sum to produce a receptor potential that is graded over approximately a 9-log-unit range of light intensity. Amplitude histograms of the discrete potential waves are bimodal, reflecting the SPF and LPF populations. Histograms of current waves are unimodal. SPFs and LPFs are insensitive to 1 microgram tetrodotoxin. I-V characteristics show initial inward currents of approximately 15 nA for voltage clamps to - 40 mV and steady-state outward currents for all clamp potentials. Photoreceptor cells bathed in organ culture medium retain these properties for periods of at least 75 days.     

Hyperpolarization of a barnacle photoreceptor membrane following illumination

Membrane potential changes following illumination of a photoreceptor cell in the lateral ocellus of a barnacle (Balanus eburneus) were studied by means of intracellular recording and polarization techniques. Illumination produces a depolarizing response. When the illumination is terminated, the membrane potential temporarily becomes more negative than the resting potential prior to illumination. Although the amplitude of this postillumination hyperpolarization depends upon the intensity as well as the duration of the light pulse, the time course is fairly constant. The hyperpolarization is not associated with any significant membrane conductance increase and is abolished by 10^-5 M ouabain. It diminishes when the external Na or K ions are removed. An intracellular injection of Na ions produces a hyperpolarization similar to that following illumination. It is suggested that the postillumination hyperpolarization is produced by an electrogenic Na pump which is activated by the Na influx during illumination.     

Lanthanum mimicks the trp photoreceptor mutant of Drosophila in the blowfly Calliphora

The effect of lanthanum on the light response of blowfly (Calliphora erythrocephala) photoreceptors was studied. The electrophysiological behaviour of the photoreceptors in the presence of La can be summarized as follows: 1. Upon long stimulation the photoreceptors responded with a 'transient receptor potential', i.e. the cells depolarized at the onset of the stimulus and then repolarized to (or below) the resting potential. This effect was dependent on stimulus intensity and occurred only at high intensities. During illumination membrane noise was reduced. 2. The light-induced changes in membrane potential were paralleled by changes in membrane resistance. 3. The time course of the receptor response was slowed down. 4. Light adaptation led to an increase in response latency. 5. The recovery of the receptor response after light adaptation was slowed down. 6. The sensitivity of the receptor cells measured by the response to short light stimuli was reduced. In summary, the electrophysiological behaviour of Calliphora photoreceptors in the presence of La was very similar to that of the photoreceptors of the trp (transient receptor potential) mutant of Drosophila melanogaster. This result suggests that La and trp mutation affect the same cellular processes in the photoreceptors.     

Phototaxis in Drosophila: R1–6 input and interaction among ocellar and compound eye receptors

All 3 photoreceptor types in the compound eye of Drosophila can evoke positive phototaxis. Here we describe input from R1–6 receptors which are very sensitive. Previous reports in this series of studies described input from R7 and R8, the other less sensitive receptors. Here we studied fast-walking phototaxis using extremely dim stimuli. We also studied input from the simple ocellar eyes. Thus, we can now summarize a complete synthesis of inputs and interactions among all compound eye and ocellar receptor types. Receptor-deficient mutants were used to establish receptor-specific input. Two other findings are presented: (1) eye colour pigments affect the spectral sensitivity for phototaxis; and (2) the ocelli interact to facilitate input from the compound eye receptor types. Possible mechanisms of receptor interaction are discussed in the light of these findings of positive input from all photoreceptor types in Drosophila.     

Rhabdom changes in the shrimp,Palaemonetes

The fine structure of the principal compound eye of the shrimp, Palaemonetes, was studied under conditions of light and dark adaptation. Ommatidium the situation in other decapod crustaceans. Light and dark adapted eyes differ in that the rhabdom changes its shape; morphological evidence suggests a possible sequence of events involving production, utilization, and degradation of photoreceptor membrane, a discontinuous process occurring only during changes from light to dark and dark to light. A hypothesis of membrane turnover is proposed.     

Hyperpolarizing photoreceptors in the eyes of the giant clamTridacna: physiological evidence for both spiking and nonspiking cell types

Summary Intracellular studies on photoreceptors in the eyes of the giant clamTridacna give evidence for two types of light-sensitive cells, both of which are hyperpolarized by light. These cells are distinguished by the presence or absence of spikes and corresponding characteristics of the receptor potential. In non-spiking (NS) receptors, the average resting potential in the dark is low (-15 mV) and peak receptor potentials are large (to 100 mV) and adapt rapidly to light. Spiking (S) receptors have higher average resting potentials (-45 mV), but receptor potentials do not exceed 20 mV and also do not adapt to light. The spikes in S-receptors are small (3–8 mV), occur spontaneously at low levels of illumination and are inhibited by light. Bursts of spikes arise on the repolarizing off-component of the receptor potential. Light adaptation increases the excitability of S-receptors in terms of a higher frequency and shorter latency of the off response burst. The receptor potential in both cells is due to a light-activated increase in membrane conductance to potassium ions. Membrane conductance decreases in NS-receptors in relation to light adaptation. Unlike the scallop eye, no depolarizing photoreceptors are present.Abbreviations NS non-spiking photoreceptors - S spiking photoreceptors - SW seawater     

Light Adaptation in the Ventral Photoreceptor of Limulus

Light adaptation in both the ventral photoreceptor and the lateral eye photoreceptor is a complex process consisting of at least two phases. One phase, which we call the rapid phase of adaptation, occurs whenever there is temporal overlap of the discrete waves that compose a light response. The recovery from the rapid phase of adaptation follows an exponential time-course with a time constant of approximately 75 ms at 21°C. The rapid phase of adaptation occurs at light intensities barely above discrete wave threshold as well as at substantially higher light intensities with the same recovery time-course at all intensities. It occurs in voltage-clamped and unclamped photoreceptors. The kinetics of the rapid phase of adaptation is closely correlated to the photocurrent which appears to initiate it after a short delay. The rapid phase of adaptation is probably identical to what is called the "adapting bump" process. At light intensities greater than about 10 times discrete wave threshold another phase of light adaptation occurs. It develops slowly over a period of ½ s or so, and decays even more slowly over a period of several seconds. It is graded with light intensity and occurs in both voltage-clamped and unclamped photoreceptors. We call this the slow phase of light adaptation.     

Mechanisms of action of CO2 on the visual response of Galleria mellonella

Carbon dioxide modifies the visual response recorded from the compound eye of the moth Galleria mellonella. Changes brought about include an increase in latency and time course, as well as the loss of a phasic negativegoing transient. Treating the preparation with either organic or inorganic acid vapours affects the time course of the tonic negative receptor potential, initially by increasing it and then decreasing it. Acids have no effect on the latency of the response, but there is a marked increase in amplitude of the phasic negative component. The resting potential in the eye is also sensitive to acids and CO₂. Changes in resting level, however, do not appear to effect the response significantly. Acids and bases have reciprocal effects on the time course of the response. Acids and CO₂ have different effects on the phasic negative transient and CO₂ can reverse amplitude increases caused by acid.We consider the changes in the visual response brought about by the addition of CO₂ of two kinds. Changes in time course are probably due to a shifting of pH in the eye, while the abolition of the phasic negative component is probably due to a CO₂-specific reaction.     

Electrophysiological properties of isolated photoreceptors from the eye of Lima scabra

Photoreceptor cells were enzymatically dissociated from the eye of the file clam, Lima scabra. Micrographs of solitary cells reveal a villous rhabdomeric lobe, a smooth soma, and a heavily pigmented intermediate region. Membrane voltage recordings using patch electrodes show resting potentials around -60 mV. Input resistance ranges from 300 M omega to greater than 1 G omega, while membrane capacitance is of the order of 50-70 pF. In darkness, quantum bumps occur spontaneously and their frequency can be increased by dim continuous illumination in a fashion graded with light intensity. Stimulation with flashes of light produces a depolarizing photoresponse which is usually followed by a transient hyperpolarization if the stimulus is sufficiently intense. Changing the membrane potential with current-clamp causes the early phase to invert around +10 mV, while the hyperpolarizing dip disappears around -80 mV. With bright light, the biphasic response is followed by an additional depolarizing wave, often accompanied by a burst of action potentials. Both Na and Ca ions are required in the extracellular solution for normal photoexcitation: the response to flashes of moderate intensity is greatly degraded either when Na is replaced with Tris, or when Ca is substituted with Mg. By contrast, quantum bumps elicited by dim, sustained light are not affected by Ca removal, but they are markedly suppressed in a reversible way in 0 Na sea water. It was concluded that the generation of the receptor potential is primarily dependent on Na ions, whereas Ca is probably involved in a voltage-dependent process that shapes the photoresponse. Light adaptation by repetitive flashes leads to a decrease of the depolarizing phase and a concomitant enhancement of the hyperpolarizing dip, eventually resulting in a purely hyperpolarizing photoresponse. Dark adaptation restores the original biphasic shape of the photoresponse.     

Intracellular calcium changes in Balanus photoreceptor. A study with calcium ion-selective electrodes and Arsenazo III

Intracellular Ca2+ concentration (Cai) in the dark and during light stimulation, was measured in Balanus photoreceptors with Ca2+ ion-selective electrodes (Ca-ISE) and Arsenazo III absorbance changes (AIII). The average basal Cai of 17 photoreceptors in darkness was 300 +/- 160 nM determined with liquid ion-exchanger (t-HDOPP) Ca-ISE. Ca-ISE measurements indicated that light increased Cai by 700 nM (average), whereas AIII indicated an average change of 450 nM. The time course of AIII absorbance changes matched the time course of changes in the receptor potential more closely than did the Ca-ISE. Changes in Cai were graded with light intensity but the change in Cai was much greater for a decade change in intensity at high light intensity than at low intensity. The peak light induced conductance change of voltage clamped cells had a relationship to light intensity similar to that of the change in Cai. The peak Cai level measured with Ca-ISE was in good agreement with the free Ca2+ concentration of injected buffer solutions. Control Cai levels were usually restored within 5 min following injection of Ca2+ buffers. Injection of Ca2+ buffers with free Ca2+ of 0.6 microM produced a membrane depolarization. Larger increases in Cai (greater than microM) produced by injection of CaCl2 or release of Ca2+ from injected buffers by acidifying the cell, produced a pronounced membrane hyperpolarization. Increasing Cai with all of these techniques reduced the amplitude of the receptor potential. The time course of the receptor potential recovery was usually similar to that of Cai recovery.     

Properties of visual cells in the lateral eye of Limulus in situ: intracellular recordings

Two types of potential fluctuations, large and small, recorded intracellularly from photoreceptors in the dark-adapted Limulus eye in situ underlie the dual properties of the impulse discharge of the optic nerve fibers. The small potential fluctuations (SPFs)--the well-known quantum bumps--were normally less than 20 mV in amplitude. The large potential fluctuations (LPFs) were up to 80 mV in amplitude. LPFs appear to be regenerative events triggered by SPFs that enable single photon absorptions in retinular cells to fire off nerve impulses in the eccentric cell. In the dark, SPFs and LPFs occur spontaneously. At low light intensities, LPFs are the major components of the receptor potential. At high intensities, LPFs are suppressed and SPFs become the major components. SPFs and LPFs together enable single photoreceptor cells to encode approximately a 9-log unit range of light intensity. Excising the eye from the animal or cutting off its blood supply generally abolishes LPFs and thereby reduces the range of light intensity coded in the optic nerve discharge.