Addition of human melanopsin renders mammalian cells photoresponsive

Abstract

A small number of mammalian retinal ganglion cells act as photoreceptors for regulating certain non-image forming photoresponses. These intrinsically photosensitive retinal ganglion cells express the putative photopigment melanopsin. Ablation of the melanopsin gene renders these cells insensitive to light; however, the precise role of melanopsin in supporting cellular photosensitivity is unconfirmed. Here we show that heterologous expression of human melanopsin in a mouse paraneuronal cell line (Neuro-2a) is sufficient to render these cells photoreceptive. Under such conditions, melanopsin acts as a sensory photopigment, coupled to a native ion channel via a G-protein signalling cascade, to drive physiological light detection. The melanopsin photoresponse relies on the presence of cis-isoforms of retinaldehyde and is selectively sensitive to short-wavelength light. We also present evidence to show that melanopsin functions as a bistable pigment in this system, having an intrinsic photoisomerase regeneration function that is chromatically shifted to longer wavelengths.

Inner retinal photoreception independent of the visual retinoid cycle

“…In Neuro-2a culture, cells transfected with a human melanopsin cDNA, 11-cis-and 9-cis-retinal were both able to serve as chromophore (15). Interestingly, in these cells, all-trans-retinal could function as chromophore if cells were pretreated with longwavelength light (15).…”

“…Mice with outer retinal dysfunction or degeneration that also lack melanopsin show no nonvisual photoreceptive function (9, 10); no light responses of ipRGCs can be recorded from opn4 Ϫ/Ϫ animals (9, 11, 12); and heterologously expressed melanopsin confers photosensitivity to nonphotoreceptive cells including Xenopus oocytes and immortalized mammalian cell lines (13)(14)(15)(16). Although the requirement for exogenous retinal differs in different heterologous expression systems (e.g., HEK293-TrpC3 cells do not require exogenous retinal; ref.…”

“…Mice with nonfunctional or degenerated rods and cones that also lack melanopsin show no photically influenced behavior or physiology (9,10), and intrinsic photosensitivity of retinal ganglion cells is lost in the absence of melanopsin (9,11,12). Melanopsin forms a functional photopigment when heterologously expressed in COS cells (13), Xenopus oocytes (14), Neuro-2A cells (15), or HEK-293 cells (16). Melanopsin is thus necessary for ipRGC photoreception, and is sufficient to confer photosensitivity on intrinsically insensitive cell types.…”

Effects of Artificial Dawn on Subjective Ratings of Sleep Inertia and Dim Light Melatonin Onset

“…Under broadband natural or artificial light exposure, these two states exist in equilibrium. However, under monochromatic light exposure, it was shown that whereas especially short wavelengths initiated the phototransduction cascade (from R to M), long wavelengths could restore responsiveness by regeneration of the M to the R states (Melyan et al, 2005;Mure et al, 2009). The light intensities used in Mure et al's study to drive the M state back to the R state were quite high.…”

Systems Biology of Mammalian Circadian Clocks

“…In contrast to perturbations achieved by the use of reagents or temperature change, photoperturbation provides an ideal range of adjustability in timing and strength. Although most mammalian cells cannot sense light, recent studies have shown that mammalian cells (Neuro-2a; HEK293) become photoresponsive following the introduction of an exogenous G-protein-coupled photoreceptor, melanopsin (Melyan et al 2005;Qiu et al 2005). It was reported that photostimulation of melanopsin triggers a release of intracellular calcium mediated through the G q -protein signaling pathway, and, importantly, there are several reports that mammalian cellular clocks can be reset by the G q -protein signaling pathway involving a release of intracellular calcium (Balsalobre et al 2000a;Tsuchiya et al 2005).…”