Photosensitive Membrane Research Articles

Photosynthetic reaction center functionalized nano-composite films: Effective strategies for probing and exploiting the photo-induced electron transfer of photosensitive membrane protein

Photosynthetic reaction center (RC), a robust transmembrane pigment-protein complex, works as the crucial component participating the primary event of the photo-electrochemical conversion in bacteria. Sparked by the high photo-induced charge separation yield (ca. 100%) of RC, great interests have been aroused to fabricate versatile RC-functionalized nano-composite films for exploring the initial photosynthetic electron transfer (ET) of RC, and thus exploiting well-designed bio-photoelectric converters. In this review, we classify and summarize the current status about the concepts and methods of constructing RC-immobilized nano-composite films or devices for probing the photo-induced ET, and applying to novel bioelectronics if it is possible.

Control of Nanopore Wetting by a Photochromic Spiropyran: A Light-Controlled Valve and Electrical Switch

By modifying the surface of nanoporous alumina membranes using mixtures of a photochromic spiropyran and hydrophobic molecules, it is possible to control the admission of water into the membrane using light. When the spiropyran is in the thermally stable, relatively hydrophobic closed form, the membrane is not wet by an aqueous solution. Upon exposure to UV light, the spiropyran photoisomerizes to the more polar merocyanine form, allowing water to enter the pores and cross the membrane. Thus, the photosensitive membrane acts as a burst valve, allowing the transport of water and ions across the membrane. If the aqueous solution contains ions, then the membrane acts as an electrical switch; photoisomerization leads to a two-order-of-magnitude increase in ionic conductance, allowing a current to flow across the membrane. Exposure to visible light causes photoisomerization of the merocyanine back to the closed, spiro form, but dewetting of the membrane does not occur spontaneously, due to a high activation barrier.

Light-Induced Layer by Layer Thickening in Photosensitive Liquid Crystal Membranes

Photosensitive smectic membranes of pure liquid crystal (LC) were studied under white light illumination. A thickness increase is observed in the illuminated film area. Appropriate light and thermal conditions lead to a thickening layer by layer process. This unusual phenomenon is opposite to the well-known step-by-step thinning transitions under heating. We give a phenomenological interpretation for thickening based on layer transport of LC matter towards the illuminated area.

Holographic Gratings in Photosensitive Acrylic Polymers with High Refractive Index Diphenyl Sulfide

To investigate the effects of polymer matrices on the diffraction efficiency of holographic grating, a series of chiral compounds of bornyl acrylate (BA), bornyl methacrylate (BMA), and their enantiomers derived from borneol were synthesized. Instead of conventional liquid crystals, in this investigation we used a high refractive index diphenyl sulfide (DS, n20D=1.6327) mixed with initiators, bornyl acrylate, bornyl methacrylate, and multifunctional monomers to prepare the photosensitive holographic grating films. Membranes having a large refractive index difference (Δn) between the cured grating arrays and revealing high diffraction efficiency (DE) were created by using the compositions. The dynamic formation of the holographic grating and the dependence of the diffracting efficiency on laser exposure time and intensity were carried out. It was found that the intensity of the writing laser may affect the polymerization rate leading to various results of holographic grating. Bragg grating and Raman–Nath grating depending on the incident writing angles were all obtained. We also investigated the plural recording function of the photosensitive films, the morphologies of the polymer matrices, and laser light grating through the holographic grating membranes. Thermogravimetric analyses of the photosensitive membranes were also estimated.

Moesin contributes an essential structural role in Drosophila photoreceptor morphogenesis.

Ezrin-Radixin-Moesin (ERM) family proteins organize heterogeneous sub-plasma membrane protein scaffolds that shape membranes and their physiology. In Drosophila oocytes and imaginal discs, epithelial organization, fundamental to development and physiology, is devastated by the loss of Moesin. Here, we show that Moesin is crucial for Drosophila photoreceptor morphogenesis. Beyond its requirement for retinal epithelium integrity, Moesin is essential for the proper assembly of the apical membrane skeleton that builds the photosensitive membrane, the rhabdomere. Moesin localizes to the rhabdomere base, a dynamic locus of cytoskeletal reorganization and membrane traffic. Downregulation of Moesin through RNAi or genetic loss of function profoundly disrupts the membrane cytoskeleton and apical membrane organization. We find normal levels and distribution of Moesin in photoreceptors of a Moesin mutant previously regarded as protein null, suggesting alternative interpretations for studies using this allele. Our results show an essential structural role for Moesin in photoreceptor morphology.

Photosensitive Membrane Fabricated by Molecular Imprinting

Photosensitive Membrane Fabricated by Molecular Imprinting

Phototransduction: shedding light on translocation.

Light induces the migration of arrestin to the photosensitive membrane in both vertebrate and invertebrate photoreceptors. New work has identified a phosphoinositide lipid binding domain in Drosophila arrestin and implicates PIP(3) in control of arrestin translocation.

Desensitization of the photoresponse in Limulus ventral photoreceptors by protein kinase C precedes rhabdomere disorganization and endocytosis.

Limulus photoreceptors utilize the phosphoinositide pathway to generate light-induced single photon events (quantum bumps) that sum to form the depolarizing receptor potential. The protein kinase C (PKC) activator, (-)-indolactam V (ILV) rapidly desensitizes the light response in Limulus ventral nerve photoreceptors. Within 10 min of extracellular application, 100 nM (-)-ILV caused a decrease in the mean amplitude of quantum bumps to 38% of control values. PKC activation by (-)-ILV also causes photosensitive membrane disorganization and endocytosis. To investigate whether this precedes desensitization of the electrical response, we fixed cells after 10-min incubation with 25 microM (-)-ILV, a concentration sufficient to cause a 1000-fold desensitization of the receptor potential. The photosensitive microvilli of these photoreceptors remained narrow, densely packed, and well organized. Increasing the incubation time to 60 min did, however, induce disorganization and swelling of the microvilli and endocytosis of the photosensitive membrane, as previously reported. Measurement of membrane capacitance did not indicate a significant reduction in membrane area accompanying desensitization by (-)-ILV. PKC-induced reduction in light sensitivity therefore precedes the detection of ultrastructural changes in the rhabdomeral membrane and is not due to a net loss of membrane.

Role of protein kinase C in light adaptation of molluscan microvillar photoreceptors

The mechanisms by which Ca2+ regulates light adaptation in microvillar photoreceptors remain poorly understood. Protein kinase C (PKC) is a likely candidate, both because some sub-types are activated by Ca2+ and because of its association with the macromolecular 'light-transduction complex' in Drosophila. We investigated the possible role of PKC in the modulation of the light response in molluscan photoreceptors. Western blot analysis with isoform-specific antibodies revealed the presence of PKCalpha in retinal homogenates. Immunocytochemistry in isolated cell preparations confirmed PKCalpha localization in microvillar photoreceptors, preferentially confined to the light-sensing lobe. Light stimulation induced translocation of PKCalpha immunofluorescence to the photosensitive membrane, an effect that provides independent evidence for PKC activation by illumination; a similar outcome was observed after incubation with the phorbol ester PMA. Several chemically distinct activators of PKC, such as phorbol-12-myristate-13-acetate (PMA), (-)indolactam V and 1,2,-dioctanoyl-sn-glycerol (DOG) inhibited the light response of voltage-clamped microvillar photoreceptors, but were ineffective in ciliary photoreceptors, in which light does not activate the G(q)/PLC cascade, nor elevates intracellular Ca2+. Pharmacological inhibition of PKC antagonized the desensitization produced by adapting lights and also caused a small, but consistent enhancement of basal sensitivity. These results strongly support the involvement of PKC activation in the light-dependent regulation of response sensitivity. However, unlike adapting background light or elevation of [Ca2+]i, PKC activators did not speed up the photoresponse, nor did PKC inhibitors antagonize the accelerating effects of background adaptation, suggesting that modulation of photoresponse time course may involve a separate Ca2+-dependent signal.

Central regulation of photosensitive membrane turnover in the lateral eye of Limulus. I. Octopamine primes the retina for daily transient rhabdom shedding.

Limulus lateral eyes shed and renew a portion of their photosensitive membrane (rhabdom) daily. Shedding, in many species including Limulus, is regulated by complex interactions between circadian rhythms and light. Little is known about how circadian clocks and photoreceptors communicate to regulate shedding. Limulus photoreceptors do not contain an endogenous circadian oscillator, but rely upon efferent outflow from a central clock for circadian timing. To investigate whether the putative efferent neurotransmitter octopamine (OA) communicates circadian rhythms that prime the lateral eye for transient rhabdom shedding, we decoupled photoreceptors from the clock by transecting the lateral optic nerve (contains the retinal efferent fibers). Overnight (6 h) intraretinal injections of 40 microM OA restored transient shedding to lateral eyes with transected nerves to levels comparable to those of intact internal control eyes. To determine whether OA acts alone in communicating circadian rhythms that prime the lateral eye for transient shedding, we "primed" eyes with intact nerves for transient shedding with exogenous OA during subjective day. In nature, lateral eyes shed their rhabdoms only once a day at dawn following overnight efferent priming. Eyes in animals placed in darkness during subjective day, when the retinal efferents are quiescent, and injected for 6 h with 40 microM OA shed their rhabdoms in response to a second introduction to light. Untreated control eyes of the same animals did not. The same results were observed in vitro in lateral eyes treated similarly. Octopamine is the only efferent neurotransmitter/messenger required to make lateral eyes competent for transient shedding. Phentolamine, an OA receptor antagonist, reduced the number of photoreceptors primed for transient shedding and the amount of rhabdom shed in those photoreceptors suggesting that OA acts via a specific OA receptor.

Crumbs, the Drosophila homologue of human CRB1/RP12, is essential for photoreceptor morphogenesis.

The apical transmembrane protein Crumbs is a central regulator of epithelial apical-basal polarity in Drosophila. Loss-of-function mutations in the human homologue of Crumbs, CRB1 (RP12), cause recessive retinal dystrophies, including retinitis pigmentosa. Here we show that Crumbs and CRB1 localize to corresponding subdomains of the photoreceptor apical plasma membrane: the stalk of the Drosophila photoreceptor and the inner segment of mammalian photoreceptors. These subdomains support the morphogenesis and orientation of the photosensitive membrane organelles: rhabdomeres and outer segments, respectively. Drosophila Crumbs is required to maintain zonula adherens integrity during the rapid apical membrane expansion that builds the rhabdomere. Crumbs also regulates stalk development by stabilizing the membrane-associated spectrin cytoskeleton, a function mechanistically distinct from its role in epithelial apical-basal polarity. We propose that Crumbs is a central component of a molecular scaffold that controls zonula adherens assembly and defines the stalk as an apical membrane subdomain. Defects in such scaffolds may contribute to human CRB1-related retinal dystrophies.

Eyes closed, a Drosophila p47 homolog, is essential for photoreceptor morphogenesis.

Starting with a mutation impacting photoreceptor morphogenesis, we identify here a Drosophila gene, eyes closed (eyc), as a fly homolog of p47, a protein co-factor of the p97 ATPase implicated in membrane fusion. Temporal misexpression of Eyc during rhabdomere extension early in pupal life results in inappropriate retention of normally transient adhesions between developing rhabdomeres. Later Eyc misexpression results in endoplasmic reticulum proliferation and inhibits rhodopsin transport to the developing photosensitive membrane. Loss of Eyc function results in a lethal failure of nuclear envelope assembly in early zygotic divisions. Phenotypes resulting from eyc mutations provide the first in vivo evidence for a role for p47 in membrane biogenesis.

Protein kinase C‐induced disorganization and endocytosis of photosensitive membrane in Limulus ventral photoreceptors

Protein kinase C (PKC) desensitizes the light response in photoreceptors from the ventral optic nerve of the horseshoe crab Limulus. Photoisomerization of Limulus rhodopsin leads to phosphoinositide hydrolysis, resulting in the production of inositol trisphosphate and diacylglycerol (DAG). Inositol trisphosphate mobilizes intracellular stores of Ca(2+), resulting in photoreceptor excitation in Limulus, while DAG may activate PKC. We investigated whether PKC-mediated desensitization of the photoresponse is accompanied by ultrastructural changes in the rhodopsin-bearing photosensitive membrane (rhabdom) in Limulus ventral photoreceptors. PKC activation by (-)-indolactam V in darkness induces disorganization and swelling of the rhodopsin-containing microvilli and endocytosis of rhabdomeral membrane. The effects of (-)-indolactam V on dark-adapted photoreceptor ultrastructure are reversible, are stereospecific, are blocked by coapplication of PKC inhibitors, and closely match those induced by continuous, bright light. Rhabdom disorganization and endocytosis via PKC activation may, therefore, contribute to desensitization of the light-adapted photoreceptor.

Identification of Novel Molecular Components of the Photoreceptor Connecting Cilium by Immunoscreens

The connecting cilium of photoreceptor cells is the only intracellular link between the morphologically, functionally and biochemically different compartments of the inner and outer segments. The non-motile modified cilium plays an important role in the organization and the function of photoreceptor cells, namely in delivery and turnover of enzymes and substrates of the visual transduction cascade, and the photosensitive membranes of the outer segment. The protein components of the cilium participate in the intracellular transport through the cilium, in the outer segment disk morphogenesis and in the maintenance of discrete membrane domains.In order to identify yet unknown cytoskeletal components of the connecting cilium, a combined biochemical and molecular biological strategy was applied. For this purpose, antibodies were raised against proteins of photoreceptor cell axonemes. Using this AX-4-antiserum, a rat retina cDNA expression library was immunoscreened and clones encoding partial sequences of (i) already known photoreceptor specific proteins; (ii) ubiquitously expressed proteins; (iii) clones with homologies to retinal ESTs; and (iv) clones coding for cytoskeletal proteins were isolated. Further analysis revealed that these candidate clones have homologies to Drosophila flightless I, mouse APC-binding protein EB2, human microtubule associated protein 4 (MAP4), human centrin 3, human cytoplasmic dynein intermediate chain 2C, and human dynamitin.The immunoscreening approach used here was successfully applied to isolate genes encoding yet unknown cytoskeletal proteins of photoreceptor cell axonemes. The obtained information will provide further insight into the role of the connecting cilium in photoreceptor cell function.

Phototoxic process after rapid photosensitive membrane damage of 5,5″-bis(aminomethyl)-2,2′:5′,2″-terthiophene dihydrochloride

We report a new aspect of rapid (<30 s) light-induced cell membrane damage photosensitized by 5,5″-bis(aminomethyl)-2,2′:5′,2″-terthiophene dihydrochloride (BAT), which is a water-soluble α-terthienyl analogue, using a high-power laser (light intensity 1.6 W cm −2). In this paper, we will discuss the relationship between the exposure time of the cells to the photosensitizer and the phototoxic process. Three toxic processes can be identified: first, a non-light-mediated toxicity dependent on BAT-cell incubation; second, a phototoxicity independent of BAT exposure time when the BAT concentration is in the 2–10-μM range; third, a phototoxicity dependent on BAT exposure time when BAT concentration becomes 20 μM. The cytotoxicity decreases when α-tocopherol, an antioxidant, is added to a cell membrane. This pattern of phototoxicity is the typical of a phospholipid peroxidation chain reaction and oxidative damage of membrane proteins triggered by a reactive oxygen species generated by a triplet state of BAT. The BAT exposure time is clearly correlated with the partition of the photosensitizer in the cell membrane and inside the cell.

A direct technique for calculating the profile of aberration of the eye measured by a modified Hartmann–Shack apparatus

We propose a direct numerical method to reconstruct the aberration curve from a set of measured spots, without passing through a wavefront fit. The method is quick and does not require precise knowledge of the position of the pupil center of the human eye. The maximal value of aberration can be estimated even in extreme situations, where the amount of the aberration is large enough that some spots are shifted into areas where other spots are expected. The numerical method also simplifies the discussion of the accuracy and the limitations of the measurement. In addition to the conventional Hartmann–Shack sensor, the direct method is applied to a modified version of this apparatus. In the modified optical setup, losses and noise are reduced, and a well-defined optical pattern is detected so that no image processing — which generally includes subjective decisions — is necessary. The apparatus offers the flexibility of choosing several magnifications of the wavefront reflected by the retina (photosensitive membrane of the eye), without changing any element of the setup. The accuracy and the limitations of the apparatus are discussed, and practical applications are illustrated.

Photo-sensitive lipid membrane perturbation by a single chain lipid having terminal spiropyran group

The synthesis a novel single chain lipid (SP-16A), containing a photo-sensitive spiropyran group at its hydrophobic terminus is described. The photoisomerization behavior of SP-16A has been studied by UV-Vis spectroscopy. The lipid has been incorporated into small unilamellar vesicles (SUVs) in quantities up to 10 mol% by mixing with dipalmitoylphosphatidylcholine (DPPC) and consequent sonication. The photo-induced membrane perturbation of the SP-16A/DPPC mixed SUV liposome was observed by monitoring of leakage of carboxyfluorescein(CF), a water soluble fluorescent dye, from the interior aqueous phase of the SUV. SUVs containing 5–10 mol% of SP-16A showed drastic leakage of CF by UV irradiation at 465 nm, however, neither liposome rupture nor release of the lipid from the SUV was observed after the UV irradiation. On/off switching of the leakage of interior material from the vesicle could also be demonstrated. In order to discuss the mechanism and kinetics of the photo-sensitive membrane perturbation behavior for the SP-16A/DPPC mixed SUV, the isomerization behavior of the SP-16A molecule and the leakage of CF from the SUV over time were investigated using a pulsed excimer laser. These experiments suggested that the main factor involved in membrane perturbation by SP-16A is a conformational change of the lipid in the membrane, which occurs after isomerization of the molecule. This new lipid molecule is thus a novel and useful photo-sensitive switching system, and may have future applications in drug delivery systems.

Circadian rhythms in the horseshoe crab lateral eye: signal transduction and photostasis

The visual system of the horseshoe crab has been an important model system for studies of basic visual processes. It has been particularly useful for understanding the role of efferent feedback and circadian rhythms in vision. Among the many retinal effects of circadian efferent activity is the priming of the daily shedding of photosensitive membrane. This daily event is triggered from the absorption of light by rhodopsin through the phospholipase C/diacylglycerol/protein kinase C cascade also requiring elevated cytosolic calcium released by inositol trisphosphate. The purpose of the daily shedding of photoreceptive rhabdomeral membrane, however, is far from clear. Recently, animals have been discovered at hydrothermal vents in the deep sea that have large photoreceptors and copious amounts of rhabdom but no apparent mechanisms for shedding and synthesizing this membrane array. Since these animals see no cyclic lighting from daylight, the absence of rhabdom shedding is consistent with the suggestion that rhabdom shedding is a homeostatic mechanism for maintaining photostasis (constancy of daily quantum catch). This hypothesis is also consistent with recent experimental data from ongoing studies in the horseshoe crab.

Morphology of the Eye of the Hydrothermal Vent Shrimp,Alvinocaris Markensis

The bresiliid shrimpAlvinocaris markensisis a predator that inhabits the base of sulphide mounds built by the black smoker chimneys of active hydrothermal vents at the Snake Pit site on the Mid-Atlantic Ridge. Casual examination of animals collected with theDSV ‘Alvin’ suggests that, like other biesiliid shrimp from hydrothermal vents, the eyes of this species are adapted for vision in very dim light. However, examination of the structure and ultrastructure of eyes of animals collected and immediately fixed shows that the expected massive array of photoreceptors is partially or completely missing. The eye is enlarged, its dioptric apparatus has disappeared, its screening pigment is essentially gone, and its reflecting pigment cells have formed an enlarged mass of white diffusing cells behind the expected layer of photoreceptors. In half of the animals examined, there were no recognizable photoreceptors in the retina, and in the remaining animals there were only scattered photoreceptors with poorly organized microvillar arrays of photosensitive membrane. We conclude that this species is blind despite some retinal adaptations for vision in very dim light. Apparently, the ambient light of this animal's environment is below the quit point (the minimum level that can be exploited) so that the retina has begun to degenerate by losing its photoreceptors.

Effect of ultraviolet light irradiation on gas permeability in polyimide membranes. 1. Irradiation with low pressure mercury lamp on photosensitive and nonphotosensitive membranes

Effect of ultraviolet light irradiation on gas permeability in polyimide membranes. 1. Irradiation with low pressure mercury lamp on photosensitive and nonphotosensitive membranes