Phototransduction Of Non-image Forming Vision And Spectrum Extension Of Image Vision

Sensing light is one of the most important means for organisms including humans to acquire external information.The input of the light signal not only helps mammals form image vision to perceive shape,color,and motion,but also gives rise to the non-image forming vision to obtain the perception of ambient radiance,thereby regulating a variety of physiological activities.It was recently discovered that the non-image forming vision in mammals is mediated by a rare subpopulation of retinal ganglion cells(termed intrinsically photosensitive retinal ganglion cells,ipRGCs),which were characterized two decades ago.Unlike conventional photoreceptors rods and cones,the kinetics of ipRGC photoresponses are relatively slow and sluggish,with both processes of activation and deactivation being far slower than those of photoresponses by rods and cones.Such kinetic properties are fit for functions of ipRGCs,which require intergration of the environment light over a period of time rather than instant responses.However,details regarding the phototransduction of ipRGCs remain elusive.Previous work demonstrated that melanopsin in ipRGCs activated Gq-PLC?4 signaling pathway,which hydrolyzed PIP2 into IP3 and DAG,and in turn opens up TRPC6/TRPC7 channels.This phototransduction mechanism is similar to the photoransduction in Drosophila(rhodopsin—Gq—PLC?4—TRP/TRPL).For standard GPCRs(G protein-coupled receptors)and phototransduction in rods and cones,the ligand-bound receptors not only can activate the G-protein signaling pathways,but are also able to recruit the arrestin signaling pathway,in which arrestins play an essential role for controlling kinetics of GPCRs.These findings have led us to focus on the role of arrestins during photoresponse of ipRGCs.Previous efforts have been made ?-arrestinl or ?-arrestin2 single knockout mice,and to record photoresponses of ipRGCs using electrophysiology.It was found that there was no deficits in the photoresponses of ipRGCs in either ?-arrestin1 KO or ?-arrestin2 KO mice.However,due to the critical functions of ?-arrestin1 and ?-arrestin2 in numerous GPCRs signal transduction,double knockout of ?-arrestinl and ?-arrestin2 is lethal.In the present study,we employed genetic engineering approach to obtain homozygous mice with ?-arrestin1/2 conditional double knockout(?-arrestin1/2 were cre-dependently knocked out merely in ipRGCs).Through electrophysiological recordings,we found that kinetics and amplitudes of a single photoresponse in?-arrestinl/2 conditionally,double knocked out ipRGCs were not distinguishable from that in WT ipRGCs;but when challenged with repetitive light stimulations,the amplitudes of photoresponses were significantly smaller than that of normal ipRGCs,and mutated ipRGCs needed much longer to recover from inactivation.This indicates that the lack of ?-arrestins leads to defects of ipRGCs in sustaining light responses.Our findings may appear contradict with predictions based on the traditional functions of ?-arrestins,but they imply a new function of P-arrestins in GPCRs signal transduction.For the phototransduction pathway of photoreceptors in Drosophila compound eye,there is a type of skeleton protein named INAD containing multiple PDZ domains which connects the important signal transduction components such as PLC,PKC and TRP channel proteins and plays a significant role in the phototransduction.INAD homologous protein INADL(INAD-like),a skeleton protein containing multiple PDZ domains,is also expressed in ipRGCs,we were interested in the functions of INADL in the phototransduction pathway of ipRGCs in mammal?We began by examining PLC?4,a protein interacting with INADL in the ipRGCs.We mutated the binding sites between PLC?4 and INADL,and then introduced the mutant PL?4 through viral tools into the eyes of Plc?4-/-mice.By recording the photoresponse under various conditions,we found that the dim light response was faster in ipRGCs with mutated PLC?4 than that of the wildtype ones.This suggests INADL in ipRGCS may also function as a skeleton protein to connect signal transduction components and play an important role in keeping sustained photoresposne of ipRGCs.However,mammals cannot see light at a wavelength of over 700 nm.This inability with respect to visual spectrum is due to the inherent physical thermodynamic limitations of opsin,the unique mammalian photon detecting protein on the retinal photoreceptors.The detection of longer wavelength light,such as near infrared(NIR)light,though a highly desirable ability,is a formidable challenge for mammals.This is because detecting such longer wavelength light,with lower energy photons,requires that opsins(e.g.,human red cone opsins)must have much lowered energy barriers.Consequently,this results in unendurable high thermal noise thus making NIR visual pigments impractical.In order to break through this boundary,we invented ocular injectable and photoreceptor-binding upconversion nanoparticles(pbUCNPs)that can be intimately integrated with the mammalian retinal photoreceptors.These nanoparticles can anchor on retinal photoreceptors as miniature covert NIR light transducers in order to create mammalian NIR light image vision.Through extensive physiological examination,from single photoreceptor recordings and electroretinogram(ERG)analyses,to cortical recordings and a wide variety of visual behavior tests,we demonstrated that mice injected with these nanoantennae can,not only perceive NIR light,but also obtain NIR light pattern vision.These data shows that in the phototransduction of ipRGCs,both ?-arrestins and INADL maintain the repeatable and sustained light response through different mechanisms,which is of great significance for the ipRGCs-mediated non-image forming vision.Meanwhile,we can create NIR light image vision of mammals by upconversion nanoparticles,which provides unmatched opportunities for both fundamental vision research and a wide variety of emerging bio-integrated nanodevice designs and applications.