Rabbit Anti-Human OPN4 Polyclonal Antibody

PURPOSE. To determine the relative contributions of inner and outer retinal photoreception to the pupillary light response METHODS. Wild-type, retinal degenerate (rd/rd), and melanopsin mutant (opn4(-/-)) mice were tested for pupillary light responsiveness by video pupillometry before, during, and after exposure to supersaturating light intensities. Similar lighting protocols were used to probe responses of intrinsically photosensitive retinal ganglion cells (ipRGCs) recorded with multielectrode arrays ex vivo. RESULTS. Both outer retinal photoreceptors (rods and cones) and inner retinal photoreceptors (intrinsically photosensitive retinal ganglion cells [ipRGCs]) are sufficient to drive the pupillary light response in mice. After supersaturating light exposure, rather than bleaching or adapting, rd/rd mice showed paradoxical potentiation of responses to subsaturating light exposure. opn4(-/-) mice, in contrast, could not sustain pupillary constriction under continuous bright illumination, and showed desensitization after bright-light exposure. Both the intensity of light necessary to induce potentiation and the spectral sensitivity for sustained and potentiated responses differed from that necessary to trigger pupillary constriction , suggesting that photopotentiation is dependent on a pigment-state distinct from that triggering the pupillary light response itself. Multielectrode array recordings of ipRGCs from rd/rd retinas demonstrated persistent cell firing under continuous light exposure but did not show potentiation. CONCLUSIONS. Unique photoreceptive properties of intrinsically photosensitive RGCs confer resistance to bleaching and/or adaptation under continuous bright illumination to the pupillary light response and suggest the presence of a photopigment with multiple absorption states.

Purpose: In several species the retinal pigment epithelium (RPE) has the potential to transdifferentiate into retinal cells to regenerate functional retinal tissue after injury. However, this capacity for regeneration is lost in mammals. The synthetic retinoic acid derivative, fenretinide [N(4-hydroxyphenyl) retinamide], induces a neuronal-like phenotype in the human adult retinal pigment epithelial cell line (ARPE-19). These changes are characterized by the appearance of neural-like processes and the expression of neuronal markers not normally associated with RPE cells. Here we assess whether fenretinide can induce a neuroretinal cell phenotype in ARPE-19 cells, by examining retinal cell marker expression. Methods: ARPE-19 cells were treated daily with culture medium containing either 3 mu M fenretinide or dimethyl sulfoxide as a control for 7 days. Cells were processed for immunocytochemistry, western blotting, and for analysis by PCR to examine the expression of a panel of RPE, neural, and retinal-associated cellular markers, including classical and non-canonical opsins. Results: Treatment with fenretinide for 7 days induced the formation of neuronal-like processes in ARPE-19 cells. Fenretinide induced the expression of the cone long wavelength sensitive opsin (OPN1lw) but not rhodopsin (RHO), while decreasing the expression of RPE cell markers. Many of the neuronal and retinal specific markers examined were expressed in both control and fenretinide treated cells, including those involved in photoreceptor cell development and the multipotency of neural retinal progenitor cells. Interestingly, ARPE-19 cells also expressed both photoreceptor specific and non-specific canonical opsins. Conclusions: The expression of retinal-associated markers and loss of RPE cell markers in control ARPE-19 cells suggests that these cells might have dedifferentiated from an RPE cell phenotype under standard culture conditions. The expression of molecules, such as the transcription factors paired box 6 gene (PAX6), sex determining region Y-box 2 (SOX2), cone-rod homeobox (CRX), and neural retina leucine zipper (NRL), further implies that in culture these cells are predisposed toward a retinal progenitor-like state. The fenretinide-induced increase in photoreceptor cell markers, accompanied by a decrease in RPE cell markers, suggests that retinoids may play a role in the transdifferentiation of RPE cells. Importantly, our data show for the first time the expression of a vertebrate ciliary opsin (OPN1lw) and rhabdomeric-like opsin, opsin 4 (OPN4 also known as melanopsin) in a clonal cell line. Together these data suggest that ARPE-19 cells are primed for and possess the capacity to differentiate toward a retinal cell-like lineage.