Melanopsin Research Articles

Melanopsin (OPN4) is a novel player in skin homeostasis and attenuates UVA-induced effects.

The presence of melanopsin (OPN4) has been shown in cultured murine melanocytes and was associated with ultraviolet A radiation (UVA) reception. Here we demonstrated the protective role of OPN4 in skin physiology and the increased UVA-induced damage in its absence. Histological analysis showed a thicker dermis and thinner hypodermal white adipose tissue layer in Opn4-/- (KO) mice than in wild-type (WT) animals. Proteomics analyses revealed molecular signatures associated with proteolysis, remodeling chromatin, DNA damage response (DDR), immune response, and oxidative stress coupled with antioxidant responses in the skin of Opn4 KO mice compared to WT. Skin protein variants were found in Opn4 KO mice and Opn2, Opn3, and Opn5 gene expressions were increased in the genotype. We investigated each genotype response to UVA stimulus (100kJ/m2). We found an increase of Opn4 gene expression following stimulus on the skin of WT mice suggesting melanopsin as a UVA sensor. Proteomics findings suggest that UVA decreases DDR pathways associated with ROS accumulation and lipid peroxidation in the skin of Opn4 KO mice. Relative changes in methylation (H3-K79) and acetylation sites of histone between genotypes and differentially modulated by UVA stimulus were also observed. We also identified alterations of molecular traits of the central hypothalamus-pituitary- adrenal (HPA) and the skin HPA-like axes in the absence of OPN4. Higher skin corticosterone levels were detected in UVA-stimulated Opn4 KO compared to irradiated WT mice. Taken altogether, functional proteomics associated with gene expression experiments allowed a high-throughput evaluation that suggests an important protective role of OPN4 in regulating skin physiology in the presence and absence of UVA radiation.

Long-day photoperiods affect expression of OPN5 and the TSH-DIO2/DIO3 pathway in Magang goose ganders

This study sought to understand the regulation mechanism of OPN5 through the TSH-DIO2/DIO3 pathway mediated photoperiod on the breeding activity of short-day breeding birds. In this study, the reproductive activity of Magang goose was regulated by artificial light, and the reproductive activity of the ganders were determined according to the daily laying rate of female geese. The testicular development and the serum reproductive hormone concentrations of ganders were measured during the reproductive period (d 0), the reproductive degeneration period (d 13 and 27) and the resting period (d 45). The mRNA and protein expression patterns of OPN5, the HPG axis reproductive genes, and TSH-DIO2/DIO3 pathway related genes were examined. Results showed that the laying rate of geese and the gonadal indices (GSI) decreased gradually after the photoperiod increased. Histological observation found that the spermatogenic function of the testis was normal on d 0 and 13, while degeneration occurred by d 27 and 45. Serum testosterone, FSH, and LH concentration showed a slight increase on d 13, followed by a sharp decrease on d 27 and 45 (P < 0.01), while PRL concentrations were low on d 0 and 13, and increased rapidly on d 27 and 45 (P < 0.01).The expression pattern of GnRH, FSH, LH, and THRβ mRNA were similar, with high levels on d 0 and 13 and a decreasing trend on d 27 and 45 (P < 0.05 or P < 0.01); and GnRHR mRNA levels were higher on d 13 (P < 0.05), but then had decreased by d 27 and 45 (P < 0.01). The expression pattern of GnIH and GnIHR was similar, which was opposite to that of GnRHR. VIP, PRL, and PRLR increased gradually and peaked on d 45 (P < 0.01). The expression trend of TRH, TSHβ, and DIO2 was similar to that of GnRHR, and the expression abundance increased on d 13, and then decreased on d 27 and 45. GnRH protein expression was significantly higher than during the other 3 periods (P < 0.01) while the GnIH protein levels were extremely low on d 0, had gradually increased by d 13, and significantly increased by d 27 and 45 (P < 0.01). The protein expression trends of THR and DIO2 were similar to that of GNIH. DIO3 protein expression was low on d 0 and 13, and increased by d 27 and 45. These results suggest that when the photoperiod increased, the hypothalamus OPN5 gene and protein were upregulated and the pituitary TSHβ, TSHR, and hypothalamus THRβ, TRH, and DIO2 were downregulated, and thus the reproductive activity of geese was inhibited.

Neuropsin (OPN5): A Non-visual UV Sensitive Photoreceptor Gene Expression Pattern Among Mega and Micro Bats

Bats are nocturnal animals with functional eyes and M/L and S opsin genes in the majority of the species. These genes are prerequisite for daylight, UV and dichromatic colour vision. Several studies suggest that other non-visual light-sensitive pigments are also involved in the UV light perception in animals. Recent behavioural, molecular and immunohistochemical evidence supports that the opsin-like gene, neuropsin (OPN5), is identified in humans, mice and birds, where it serves as a G protein-coupled UV-sensitive photoreceptor. Based on its low sequence homology with other opsin groups, OPN5 is classified as an independent group. While the roles of non-visual light-sensitive pigment OPN5 in bats remain an open question, here we report that bat’s neuropsin (OPN5) encoded by OPN5 gene shares 89-96% amino acid identity and similar domain organization with human and mouse OPN5. By PCR amplification, we confirm that all the mega and micro bats express the OPN5 gene in their genome. The expression of OPN5 is detectable only in the brain, eye and retina and not in the heart, kidney, liver, lungs and testis. This result suggests that OPN5 gene expression is neural specific in bats. OPN5 gene expression level is significantly higher in tree-roosting bats compared to cave-roosting bats. Since, the tree-roosting bats received slightly more sunlight every day when compared to cave-roosting bats. In captive conditions, the expression levels of OPN5 in the neural tissues are significantly lower than those of wild bats. Our preliminary results suggest that the opsin-like gene, neuropsin (OPN5) is involved in UV light perception in bats.

Ontogeny of OPN4, OPN5, GnRH and GnIH mRNA Expression in the Posthatch Male and Female Pekin Duck (Anas platyrhynchos domesticus) Suggests OPN4 May Have Additional Functions beyond Reproduction.

Simple SummaryBirds perceive light differently than mammals. Unlike mammals, birds’ retinas do not participate in the non-image forming, seasonal breeding aspects of light perception. Birds have deep brain photoreceptors that are involved with the seasonal increase in fertility. Until this study, it was unknown how these brain photoreceptors developed after hatching. Our novel finding is that at least one of the photoreceptors, OPN4, is expressed at high levels on the day of hatching, which suggests that this photoreceptor has functions beyond just the reproductive system. These findings could change how we light poultry barns to improve the growth, health and welfare of our poultry species.The hypothalamic–pituitary–gonadal axis (HPG) is known to be regulated by daylength through the deep brain photoreceptor (DBP) system. The post-hatch ontogeny is not known for any of the DBPs. We set out to determine the ontogeny of OPN4 and OPN5 gene expression relative to GnRH and GnIH using qRT-PCR. Brains and serum were collected from five drakes and five hens on the day of hatching (Day 0) and again at 2, 4, 6, 10, 14, 19, 25 and 31 weeks of age and analyzed by qRT-PCR. Hen and drake serum was assayed for circulating levels of estradiol and testosterone, respectively. Data were analyzed between sexes over time using a repeated measures two-way ANOVA. Interestingly, the results show that on the day of hatching (Day 0), ducks showed adult-like levels of relative OPN4, but not OPN5, gene expression. During week 10, DBP levels increased, achieving highest relative expression levels at week 19 that maintained through week 31, typically peak fertility in ducks. GnRH mRNA levels increased following the DBP expression at the onset of puberty, and gonadal steroids increased after GnRH at week 14 while estradiol preceded testosterone. GnIH mRNA levels did not appreciably change during the time course of this experiment. These observations suggest that OPN4 may be active during the peri-hatch period and may have physiological roles beyond puberty and fertility.

Wounding Induces Facultative Opn5-Dependent Circadian Photoreception in the Murine Cornea

PurposeAutonomous molecular circadian clocks are present in the majority of mammalian tissues. These clocks are synchronized to phases appropriate for their physiologic role by internal systemic cues, external environmental cues, or both. The circadian clocks of the in vivo mouse cornea synchronize to the phase of the brain's master clock primarily through systemic cues, but ex vivo corneal clocks entrain to environmental light cycles. We evaluated the underlying mechanisms of this difference.MethodsMolecular circadian clocks of mouse corneas were evaluated in vivo and ex vivo for response to environmental light. The presence of opsins and effect of genetic deletion of opsins were evaluated for influence on circadian photoresponses. Opn5-expressing cells were identified using Opn5Cre;Ai14 mice and RT-PCR, and they were characterized using immunocytochemistry.ResultsMolecular circadian clocks of the cornea remain in phase with behavioral circadian locomotor rhythms in vivo but are photoentrainable in tissue culture. After full-thickness incision or epithelial debridement, expression of the opsin photopigment Opn5 is induced in the cornea in a subset of preexisting epithelial cells adjacent to the wound site. This induction coincides with conferral of direct, short-wavelength light sensitivity to the circadian clocks throughout the cornea.ConclusionsCorneal circadian rhythms become photosensitive after wounding. Opn5 gene function (but not Opn3 or Opn4 function) is necessary for induced photosensitivity. These results demonstrate that opsin-dependent direct light sensitivity can be facultatively induced in the murine cornea.

Unexpected molecular diversity of vertebrate nonvisual opsin Opn5.

Animals depend on light from their external environment to provide information for physiological functions such as vision, photoentrainment of circadian and circannual rhythms, photoperiodism, and background adaptation. Animals have a variety of photoreceptor cells that perform these functions, not only in the retina but also in other tissues, including brain tissue. In these cells, opsins function as universal photoreceptive proteins responsible for both visual and nonvisual photoreception. All opsins identified thus far bind either 11-cis or all-trans retinal as a chromophore and are classified into several groups based on their amino acid sequences. Opn5 forms an independent group that has diversified among vertebrate species. Most mammals only have one Opn5 gene, Opn5m, while nonmammalian vertebrates have two additional Opn5 subtypes, Opn5L1 and Opn5L2. Among these subtypes, Opn5m and Opn5L2 are UV-sensitive pigments in the dark. UV irradiation converts them into the visible light-sensitive active state, which converts back to the dark state by visible light irradiation. Opn5m and Opn5L2 therefore behave as bistable pigments. By contrast, Opn5L1 exclusively binds all-trans retinal to form the active state in the dark. Opn5L1 is converted to the resting state by light irradiation and subsequently reverts to the active state by a thermal process. Thus, Opn5L1 is categorized as a unique reverse photoreceptor whose activity is regulated by its photocyclic reaction. In this review, I introduce the diversity of molecular properties that have been described for vertebrate Opn5 subtypes and their physiological relevance.

Neuropsin (OPN5) Mediates Local Light-Dependent Induction of Circadian Clock Genes and Circadian Photoentrainment in Exposed Murine Skin.

Nearly all mammalian tissues have functional, autonomous circadian clocks, which free-run with non-24h periods and must be synchronized (entrained) to the 24h day. This entrainment mechanism is thought to be hierarchical, with photic input to the retina entraining the master circadian clock in the suprachiasmatic nuclei (SCN) and the SCN in turn synchronizing peripheral tissues via endocrine mechanisms. Here, we assess the function of a population of melanocyte precursor cells in hair and vibrissal follicles that express the photopigment neuropsin (OPN5). Organotypic cultures of murine outer ear and vibrissal skin entrain to a light-dark cycle exvivo, requiring cis-retinal chromophore and Opn5 gene function. Short-wavelength light strongly phase shifts skin circadian rhythms exvivo via an Opn5-dependent mechanism. Invivo, the normal amplitude of Period mRNA expression in outer ear skin is dependent on both the light-dark cycle and Opn5 function. InOpn4-/-; Pde6brd1/rd1 mice that cannot behaviorally entrain to light-dark cycles, the phase of skin-clock gene expression remains synchronized to the light-dark cycle, even as other peripheral clocks remain phase-locked to the free-running behavioral rhythm. Taken together, these results demonstrate the presence of a direct photic circadian entrainment pathway and direct light-response elements for clock genes in murine skin, similar to pathways previously described for invertebrates and certain non-mammalian vertebrates.

Neuropsin (OPN5) Mediates Local Light-Dependent Circadian Responses in Murine Skin

Nearly all mammalian tissues have functional, autonomous circadian clocks, which free-run with non-24-hour periods and must be synchronized (entrained) to the 24-hour day. This entrainment mechanism is thought to be hierarchical, with photic input to the retina entraining the master circadian clock in the suprachiasmatic nuclei (SCN), and the SCN in turn synchronizing peripheral tissues via endocrine mechanisms. A population of LEF1-positive stem cells in hair and vibrissal follicles express the photopigment neuropsin (OPN5). Organotypic cultures of murine outer ear and vibrissal skin entrain to a light-dark cycle ex vivo, requiring cis-retinal chromophore and Opn5 gene function. Short-wavelength light strongly phase shifts skin circadian rhythms ex vivo via an Opn5-dependent mechanism. In vivo, the normal amplitude of Period mRNA expression in outer ear skin is dependent on both the light-dark cycle and on Opn5 function. In Opn4-/-; Pde6brd1/rd1 mice that cannot behaviorally entrain to light-dark cycles, skin clock gene cycling remains phase-entrained to the light-dark cycle even as other peripheral clocks remain phase-locked to the free-running behavioral rhythm. Taken together, these results demonstrate the presence of a direct photic circadian entrainment pathway for murine skin, similar to pathways previously described for invertebrates and certain non-mammalian vertebrates.

The visual basis of the biological clock "sees" the light: intrinsically photosensitive retinal ganglion cells

Mammalian eyes mediate both image-forming and non-image-forming visual functions.Non-image-forming vision provides a measure of the ambient light for the purposes of synchronization of circadian clocks to light/dark cycles and regulation of pupil size, pineal melatonin production and other functions.Traditionally, people used to believe that the classical photoreceptors (rods and cones) regulate both image-forming and non-image-forming visual pathways.However, a small subset of retinal ganglion cells called intrinsically photosensitive RGCs (ipRGCs) has been identified to be a third type of mammalian photoreceptor and determined to be photosensitive, recently.The discovery of ipRGCs has allowed for rapid progress in the past decade toward understanding the non-image-forming visual system, especially about how the circadian clock complete the synchronization with the light/dark cycle.The anatomical and developmental characteristics of ipRGCs, as well as its biological functions and regulation were reviewed in this paper. Key words: Circadian clock; Non-image-forming; Intrinsically photosensitive retinal ganglion cell; Melanopsin

Morphology of intrinsically photosensitive retinal ganglion cells: distinct photoreceptors from rod/cone photoreceptors

Objective This study was to observe the morphology of live intrinsically photosensitive retinal ganglion cells (ipRGCs) at cellular level, and to explore their three-dimensional structure and responses to light in curved retina and the relationship with rod/cone photoreceptors. Methods ipRGCs were identified according to the enhanced green fluorescent potein (EGFP) markers.Two hundred and sixty-three ipRGCs were videoed in mouse whole-mounted retina after strengthening with Lucifer Yellow from patch clamp electrodes.The dendrites and cell bodies were analyzed according to their sublayer-specific localization in inner plexiform layer and ganglion cell layer of curved retina.The relationship with rod/cone system was reconstructed and their functions were speculated.The animal feeding and use was in accordance with the standards set by the ARVO, and the experiment was approved by the Ethic Committee for Experimental Animal of Hubei University of Science and Technology. Results The ipRGCs had strictly sublayer-specific localization in three sublayers of retinal inner plexiform layer.Each sublayer occupies full retina and form photosensitive surface, without any intermediate photosensitive dentric distribution between sublayers.Each ipRGC had randomly dentric distributions among the three sublayer curves, without the specific ON/OFF stratification.The photosensitive sublayers had absolutely perpendicular assignment related to cone/rod photoreceptors.The expression of melanopsin and spike-producing Ca2+ /Na+ channels were randomly distributed in M1, M2 and M3 cells.M4 and M5 cells shared the characteristic properties of conventional ganglion cells. Conclusions In contrast to the rod/cone photoreceptors, ipRGCs form multiple-sublayer photosensitive curved surface, which are perpendicular to rod/cone photoreceptors, their photosensitive melanopsin and intrinsic spike-producing channels randomly occupy these specific sublayers, which suggest their distinct functions from rod/cone photoreceptors. Key words: Intrinsically photosensitive retinal ganglion cells; Three-deminsion morphology; Melanopsin; Retina; Photoreceptor

Measuring melanopsin function in people

Measuring melanopsin function in people

Commentary: Melanopsin Regulates Both Sleep-Promoting and Arousal-Promoting Responses to Light.

Commentary: Melanopsin Regulates Both Sleep-Promoting and Arousal-Promoting Responses to Light.

Immunohistochemical and ultrastructural properties of the larval ciliary band-associated strand in the sea urchin Hemicentrotus pulcherrimus.

BackgroundThe swimming activity of sea urchin larvae is dependent on the ciliary band (CB) on the larval surface and is regulated by several neurotransmitters, including serotonin (5HT), dopamine, and γ-aminobutyric acid (GABA). However, the CB signal transmission mechanism remains unknown. The present study investigated the structural relationship between the CB and external signal receptors by immunohistochemical and transmission electron microscopic analyses of sea urchin, Hemicentrotus pulcherrimus, larvae.ResultsGlutamate decarboxylase (GAD; GABA synthetase) was detected in a strand of multiple cells along the circumoral CB in 6-arm plutei. The GAD-expressing strand was closely associated with the CB on the oral ectoderm side. The ciliary band-associated strand (CBAS) also expressed the 5HT receptor (5HThpr) and encephalopsin (ECPN) throughout the cytoplasm and comprised 1- to 2-μm diameter axon-like long stretched regions and sporadic 6- to 7-μm diameter bulbous nucleated regions (perikarya) that protruded into the oral ectoderm side. Besides the laterally polarized morphology of the CBAS cells, Epith-2, which is the epithelial lateral cell surface-specific protein of the sea urchin embryo and larva, was expressed exclusively by perikarya but not by the axon-like regions. The CBAS exposed its narrow apical surface on the larval epithelium between the CB and squamous cells and formed adherens junctions (AJs) on the apical side between them. Despite the presence of the CBAS axon-like regions, tubulins, such as α-, β-, and acetylated α-tubulins, were not detected. However, the neuroendocrine cell marker protein synaptophysin was detected in the axon-like regions and in bouton-like protrusions that contained numerous small ultrastructural vesicles.ConclusionsThe unique morphology of the CBAS in the sea urchin larva epithelium had not been reported. The CBAS expresses a remarkable number of receptors to environmental stimuli and proteins that are probably involved in signal transmission to the CB. The properties of the CBAS explain previous reports that larval swimming is triggered by environmental stimuli and suggest crosstalk among receptors and potential plural sensory functions of the CBAS.

Deep-brain photoreceptors (DBPs) involved in the photoperiodic gonadal response in an avian species, Gallus gallus

Three primitive photoreceptors [melanopsin (Opn4), neuropsin/opsin5 (Opn5) and vertebrate ancient opsin (VAOpn)] were reported as possible avian deep-brain photoreceptors (DBPs) involved in the perception of photoperiodic information affecting the onset and development of reproduction. The objective of this study was to determine the effect of long-day photostimulation and/or sulfamethazine treatment (SMZ, a compound known to advance light-induced testes development) on gene expression of DBPs and key hypothalamic and pituitary genes involved in avian reproductive function. Two-week old chicks were randomly selected into four experimental groups: short-day control (SC, LD8:16), short-day+SMZ (SS, LD8:16, 0.2% diet SMZ), long-day control (LC, LD16:8), and long-day+SMZ (LS, LD16:8, 0.2% diet SMZ). Birds were sampled on days 3, 7, and 28 after initiation of a long-day photoperiod and/or SMZ dietary treatments. Three brain regions [septal–preoptic, anterior hypothalamic (SepPre/Ant-Hypo) region, mid-hypothalamic (Mid-Hypo) region, posterior-hypothalamic (Post-Hypo) region], and anterior pituitary gland were dissected. Using quantitative real-time RT-PCR, we determined changes of expression levels of genes in distinct brain regions; Opn4 and Opn5 in SepPre/Ant-Hypo and Post-Hypo regions and, VAOpn in the Mid-Hypo region. Long-day treatment resulted in a significantly elevated testes weight on days 7 and 28 compared to controls, and SMZ augmented testes weight in both short- and long-day treatment after day 7 (P<0.05). Long-day photoperiodic treatment on the third day unexpectedly induced a large 8.4-fold increase of VAOpn expression in the Mid-Hypo region, a 15.4-fold increase of Opn4 and a 97.8-fold increase of Opn5 gene expression in the Post-Hypo region compared to SC birds (P<0.01). In contrast, on days 7 and 28, gene expression of the three DBPs was barely detectable. LC group showed a significant increase in GnRH-1 and TRH mRNA in the Mid-Hypo compared to SC on day 3. Pituitary LHβ and FSHβ mRNA were significantly elevated in LC and LS groups compared to SC on days 3 and 7 (P<0.05). On days 3 and 7, TSHβ mRNA level was significantly elevated by long-day treatment compared to the SC groups (P<0.05). Results suggest that long-day photoperiodic activation of DBPs is robust, transient, and temporally related with neuroendocrine genes involved in reproductive function. Additionally, results indicate that two subsets of GnRH-1 neurons exist based upon significantly different gene expression from long-day photostimulation and long-day plus SMZ administration. Taken together, the data indicate that within 3days of a long-day photoperiod, an eminent activation of all three types of DBPs might be involved in priming the neuroendocrine system to activate reproductive function in birds.

Current researches on the circadian rhythm of patients with glaucoma

Like other mammals,human has obvious circadian rhythm,and light is recognized as the principal circadian synchronizer.Rod/cone photoreceptors of the outer retina and the melanopsin-containing retinal ganglion cells(mcRGCs) of the inner retina mediate non-image forming visual responses including entrainment of the circadian clock to the ambient light,the pupillary light reflex (PLR),and light modulation of activity by means of a set of mcRGCs projecting to the suprachiasmatic nucleus (SCN)through retinohypothalamic tract (RHT).Glaucoma is a group of diseases which cause degenerative changes and loss of RGCs including mcRGCs,so the circadian rhythm of patients with glaucoma will be interrupted.The progress in the study on the effect of glaucoma and experimental hypertension on circadian rhythm was reviewed. Key words: Glaucoma; Circadian rhythm; Melanopsin

UV-Sensitive Photoreceptor Protein OPN5 in Humans and Mice

A variety of animal species utilize the ultraviolet (UV) component of sunlight as their environmental cues, whereas physiological roles of UV photoreception in mammals, especially in human beings, remain open questions. Here we report that mouse neuropsin (OPN5) encoded by the Opn5 gene exhibited an absorption maximum (λmax) at 380 nm when reconstituted with 11-cis-retinal. Upon UV-light illumination, OPN5 was converted to a blue-absorbing photoproduct (λmax 470 nm), which was stable in the dark and reverted to the UV-absorbing state by the subsequent orange light illumination, indicating its bistable nature. Human OPN5 also had an absorption maximum at 380 nm with spectral properties similar to mouse OPN5, revealing that OPN5 is the first and hitherto unknown human opsin with peak sensitivity in the UV region. OPN5 was capable of activating heterotrimeric G protein Gi in a UV-dependent manner. Immuno-blotting analyses of mouse tissue extracts identified the retina, the brain and, unexpectedly, the outer ears as the major sites of OPN5 expression. In the tissue sections of mice, OPN5 immuno-reactivities were detected in a subset of non-rod/non-cone retinal neurons as well as in the epidermal and muscle cells of the outer ears. Most of these OPN5-immuno-reactivities in mice were co-localized with positive signals for the alpha-subunit of Gi. These results demonstrate the first example of UV photoreceptor in human beings and strongly suggest that OPN5 triggers a UV-sensitive Gi-mediated signaling pathway in the mammalian tissues.

Progress of the genetic research on asthma

Bronchial asthma is a chronic inflammatory disorder of the airways,caused by multiple genetic and environmental factors.Thtee key methods including candidate gene study,the positional cloning and genomewide association studies.are used for asthma susceptibility gene identification.There are many candidate genes that haye been identified as asthma susceptibility gene, including ADRB2,IL4R,FLG,NPPA,INPP4A,ADAM33, DPP10,OPN3,0RMDL3,PDE4D,CHBLI etc.Replication of genetic associations and demonstration of a functional mechanism for the associated gene variants are needed to COnfirm an asthmasusceptibility gene, this will help us to understand the aetiology of asthma and explore new drugs to treat asthma. Key words: Asthma; Genetics; Susceptibility genes

Cells see the light with melanopsin

Cells see the light with melanopsin

1P243 ニワトリメラノプシンのクローニングと性状解析(光生物 A) 視覚)

1P243 ニワトリメラノプシンのクローニングと性状解析(光生物 A) 視覚)