Cui LJ, Chen WH, Liu AL, Han X, Jiang SX, Yuan F, Zhong YM, Yang XL, & Weng SJ. nGnG Amacrine Cells and Brn3b-negative M1 ipRGCs are Specifically Labeled in the ChAT-ChR2-EYFP Mouse. (2020) Invest Ophthalmol Vis Sci 61(2):14. doi: 10.1167/iovs.61.2.14
Summary: The authors speculated that type II cells might be ipRGCs. This was later verified by the strong immunostaining of type II cells in response to the melanopsin antibody UF006 (100%, 141 of 141 cells collected from 5 retinas, Figs. 6B1–B3), which probes multiple ipRGC subtypes.
Dose: Immunostaining 1:10000
Moshirpour M, Nakashima AS, Sehn N, Smith VM, Thackray SE, Dyck RH, Antle MC. Examination of Zinc in the Circadian System. (2020) Neuroscience 432:15-29. doi: 10.1016/j.neuroscience.2020.02.016
Objective: To examine the anatomical and functional aspects of zinc in the circadian system.
Summary: Neither enhancement nor chelation of free zinc at either the SCN or IGL altered circadian responses to phase-shifting light in hamsters.
Dose: Retinal immunohistochemistry included a 20-min wash in 4% PFA prior to initiation of the IHC protocol.
Alarautalahti V, Ragauskas S, Hakkarainen JJ, Uusitalo-Järvinen H, Uusitalo H, Hyttinen J, Kalesnykas G, & Nymark S. Viability of Mouse Retinal Explant Cultures Assessed by Preservation of Functionality and Morphology (2019) Invest Ophthalmol Vis Sci 60(6):1914-1927. doi: 10.1167/iovs.18-25156
Summary: Organotypic retinal explant cultures have been used to study retinal development, retinal diseases and injuries, drug screening, and retinal stem cell therapies.
Dose: The amount of ganglion cells and melanopsin-expressing intrinsically photosensitive RGCs (ipRGCs) were detected by staining of RNA-binding protein. Melanopsin detection by Immunostaining. 1:2500
Lin M-S, Liao P-Y, Chen H-M, Chang C-P, Chen S-K, & Chern Y. Degeneration of ipRGCs in Mouse Models of Huntington’s Disease Disrupts Non-Image-Forming Behaviors Before Motor Impairment. (2019) J Neurosci 39 (8):1505. doi: 10.1523/JNEUROSCI.0571-18.2018
Summary: results show that M1 ipRGCs were susceptible to the toxicity caused by mutant Huntingtin. The resultant impairment of M1 ipRGCs contributed to the early degeneration of the ipRGC–SCN pathway and disrupted circadian regulation during HD progression.
Dose: Immunostaining (1:3000)
Quattrochi LE, Stabio ME, Kim I, Ilardi MC, Michelle Fogerson P, Leyrer ML, & Berson DM. The M6 Cell: A Small-Field Bistratified Photosensitive Retinal Ganglion Cell. (2019) J Comp Neurol 527(1):297-311. doi: 10.1002/cne.24556
Summary: M6 cells express low levels of melanopsin and have correspondingly weak intrinsic light responses.
Dose: In all experiments involving melanopsin immunofluorescence, melanopsin immunodetection was enhanced using tyramide signal amplification coupled with horseradish peroxidase (HRP)-paired with goat anti-rabbit IgG and an Alexa fluorophore. 1:10,000.
Sweeney NT, James KN, Nistorica A, Lorig-Roach RM, & Feldheim DA. Expression of Transcription Factors Divides Retinal Ganglion Cells into Distinct Classes. (2019) J Comp Neurol 527(1):225-235. doi: 10.1002/cne.24172
AB-N38: Melanopsin (Opn4) Immunostaining 1:1000.
Delwig A, Chaney SY, Bertke AS, Verweij JAN, Quirce S, Larsen DD, Yang C, Buhr E, Van Gelder R, Gallar J, Margolis T, & Copenhagen DR. Melanopsin expression in the cornea. (2018). Visual Neuroscience, 35 E004. 01/31.
Objective: To determine melanopsin expression in cornea.
Summary: Found no light-evoked activation of melanopsin-expressing fibers in cornea or in cell bodies in the TG; melanopsin protein might serve other sensory functions in the cornea. One justification for this idea is that melanopsin expressed in Drosophila photoreceptors can serve as a temperature sensor.
Dose: For the primary staining, we used Anti-Melanopsin at 1:5000 for 3 days.
Dhande O, et al. Assembly of functionally antagonistic visual circuits for controlling pupil dynamics (2018) Cell Rep Epub 19 Dec.
Objective: Investigate the mechanisms controlling the specification and establishment of parallel sensory pathways.
Summary: Identification of a novel genetic program that marks and is required for the development of non-image-forming parallel visual pathways.
Dose: IHC: 1:1,000
Grillo SL, & Stella SL, Jr. Melanopsin Retinal Ganglion Cells Are Not Labeled in Thy-1yfp-16 Transgenic Mice. (2018). Neuroreport, 29 (2):118-122. 2017/12/19.
Objective: To determine whether mRGCs are labeled with YFP in Thy-1 YFP-16 transgenic mice. The transgenic Thy-1 YFP mouse line 16 (Thy-1 YFP-16) expresses yellow-fluorescent protein (YFP) in projection neurons, including RGCs.
Summary: The majority of mRGC somas and axons are not labeled with YFP in the transgenic Thy-1 YFP-16 mouse line; therefore, this mouse model may not suitable for research involving mRGC visual pathways.
Dose: Affinity purified rabbit polyclonal antibody raised against melanopsin (1:500) was used to label mRGCs (Retinal ganglion cells expressing melanopsin). Immunolabeled Thy-1 YFP-16 mouse retina with anti-melanopsin and found that majority (∼89%) of mRGCs are not YFP-positive despite numerous other YFP-positive RGCs in the retina.
Ing-Esteves S, Kostadinov D, Marocha J, Sing AD, Joseph KS, Laboulaye MA, Sanes JR, & Lefebvre JL. Combinatorial Effects of Alpha- and Gamma-Protocadherins on Neuronal Survival and Dendritic Self-Avoidance. (2018). The Journal of Neuroscience, 38 (11):2713.
Objective: The clustered protocadherins (Pcdhs) comprise 58 cadherin-related proteins encoded by three tandemly arrayed gene clusters, Pcdh- , Pcdh- , and Pcdh- (Pcdha, Pcdhb, and Pcdhg, respectively). This study sought to determine roles of Pcdhas and Pcdhgs in the retina and cerebellum from mice (both sexes) lacking one or both clusters.
Summary: Study examined two regions of the CNS, the retina and cerebellum and showed that the 14 -Pcdhs and 22 -Pcdhs act synergistically to mediate neuronal survival and dendrite patterning. In retina, Pcdhgs are essential for survival of inner retinal neurons and dendritic self-avoidance of starburst amacrine cells, whereas Pcdhas are dispensable for both processes.
Dose: Anti-Melanopsin (1:5000) used to quantify two mutually exclusive RGC cell types, the Brn3a RGCs and the melanopsin-positive intrinsically photo- sensitive RGCs (Mel ipRGCs).
Jo A, Xu J, Deniz S, Cherian S, DeVries SH, & Zhu Y. Intersectional Strategies for Targeting Amacrine and Ganglion Cell Types in the Mouse Retina. (2018). Frontiers in Neural Circuits, 12 66.
Objective: To obtain unambiguous information about retinal processing.
Summary: Results establish a foundation for future application of intersectional strategies in the retina and retino-recipient regions.
Dose: Immunohistochemistry 1:1000.
Sabbah S, Papendorp C, Koplas E, Beltoja M, Etebari C, Gunesch AN, Carrete L, Kim MT, Manoff G, Bhatia-Lin A, Zhao T, Schreck D, Dowling H, Briggman KL, & Berson DM. Synaptic circuits for irradiance coding by intrinsically photosensitive retinal ganglion cells. (2018) bioRxiv doi: https://doi.org/10.1101/442954
Objective: To explore the synaptic networks responsible for the unique capacity of intrinsically photosensitive retinal ganglion cells (ipRGCs) to encode overall light intensity. This luminance signal is crucial for circadian, pupillary and related reflexive responses light.
Summary: Most ipRGCs sample from all bipolar terminals costratifying with their dendrites, but M1 cells avoid all OFF bipolar input and accept only ectopic ribbon synapses from ON cone bipolar axonal shafts. These monad synapses are equipped with as many as a dozen ribbons and only one postsynaptic process.
Dose: Immunohistochemistry of retina – after recording, retinas were fixed and counterstained with rabbit anti-melanopsin (1:1000) to enhance the fluorescence of the GFP-based GCaMP6f indicator.
Sanchez-Migallon MC, Valiente-Soriano FJ, Nadal-Nicolas FM, Di Pierdomenico J, Vidal-Sanz M, & Agudo-Barriuso M. Survival of melanopsin expressing retinal ganglion cells long term after optic nerve trauma in mice. (2018). Exp Eye Res, 174 93-97. 2018/06/02.
Summary: Melanopsin-positive retinal ganglion cells (RGCs) do not respond to axotomy in the same way than the rest of RGCs, and so while image-forming RGCs die in two exponential phases, non-image forming RGCs die only during the first quick phase.
Dose: Retinas were dissected as flat mounts and double-immunostained against melanopsin (1:5000).
Stabio ME, Sabbah S, Quattrochi LE, Ilardi MC, Fogerson PM, Leyrer ML, Kim MT, Kim I, Schiel M, Renna JM, Briggman KL, & Berson DM. The M5 Cell: A Color-Opponent Intrinsically Photosensitive Retinal Ganglion Cell. (2018) Neuron, 97 (1):150-163.e154.
Objective: To provide a fuller description of the least understood ipRGC type, the M5 cell, and discovered a distinctive functional characteristic— chromatic opponency (ultraviolet excitatory, green inhibitory).
Summary: M5 cells send axons to the dLGN and are thus positioned to provide chromatic signals to visual cortex. These findings under- score that melanopsin’s influence extends beyond unconscious reflex functions to encompass cortical vision, perhaps including the perception of color.
Dose: M5 cells have the weakest melanopsin responses of all known ipRGC types. The intrinsic melanopsin response (~10 pA) in M5 cells is at least an order of magnitude smaller than the extrinsic, synaptically mediated response.
Ueki Y, Shchepetkina V, & Lefcort F. Retina-Specific Loss of lkbkap/Elp1 Causes Mitochondrial Dysfunction That Leads to Selective Retinal Ganglion Cell Degeneration in a Mouse Model of Familial Dysautonomia. (2018). Disease Models & Mechanisms, 10.1242/dmm.033746.
Objective: To determine the pathophysiological mechanisms that are triggered by the absence of IKAP (inhibitor of kappa B kinase complex-associated protein) in the retina.
Summary: The loss of IKAP caused progressive degeneration of retinal ganglion cells (RGCs) by 1 month of age. There was no loss of melanopsin+ intrinsically photosensitive RGCs at 18 months. RGCs were the only cell type that degenerated, with the survival of other retinal neurons unaffected.
Dose: Immunohistochemistry (IHC), RGC Counts, and H&E staining – Anti-melanopsin (1:5000).
Wong JCY, Smyllie NJ, Banks GT, Pothecary CA, Barnard AR, Maywood ES, Jagannath A, Hughes S, van der Horst GTJ, MacLaren RE, Hankins MW, Hastings MH, Nolan PM, Foster RG, & Peirson SN. Differential roles for cryptochromes in the mammalian retinal clock. (2018). The FASEB Journal fj.201701165RR.
Objective: To determine roles of cryptochromes (CRY) in the retinal clock.
Summary: Data suggest that CRY1 is an essential component of the mammalian retinal clock, whereas CRY2 has a more limited role.
Dose: IHC 1:2500.
Bray ER, Noga M, Thakor K, Wang Y, Lemmon VP, Park KK, Tsoulfas P. (2017) 3D Visualization of Individual Regenerating Retinal Ganglion Cell Axons Reveals Surprisingly Complex Growth Paths. eNeuro 4. PMID: 28856242 (read summary)
Huang L, Yuan T, Tan M, Xi Y, Hu Y, Tao Q, Zhao Z, Zheng J, Han Y, Xu F, Luo M, Sollars PJ, Pu M, Pickard GE, So KF, & Ren C. A retinoraphe projection regulates serotonergic activity and looming-evoked defensive behaviour. (2017). Nat Commun, 8 14908. 2017/04/01. PMC5381010
IT-27: Streptavidin-ZAP; IT-44: Melanopsin-SAP; AB-N38: Anti-Melanopsin
Objective: To investigate how the dorsal raphe nucleus (DRN) and superior colliculus work in concert to extract and translate visual threats into defensive behavioural responses.
Summary: A dedicated population of RGCs signals rapidly approaching visual threats and their input to the DRN controls a serotonergic self-gating mechanism that regulates innate defensive responses.
Dose:Mice received bilateral intraocular injection (2 μg per eye) made between Streptavidin-Saporin and a biotinylated CTB antibody, or Anti-Melanopsin-SAP (2 μg per eye). For detection of melanopsin, retinas were incubated for 3 days at 4 °C with anti-melanopsin (1:600).
Joly S, Lamoureux S, & Pernet V. (2017) Nonamyloidogenic Processing of Amyloid Beta Precursor Protein Is Associated with Retinal Function Improvement in Aging Male Appswe/Psde9 Mice. Neurobiol Aging, 53 181-191.
Objective: To determine amyloid beta role in the aging retina in Alzheimer’s Disease.
Summary: Retinal-specific processing of amyloid may confer protection against AD and selectively preserve cone-dependent vision during aging.
Dose: IHC 1:1000
Somasundaram P, Wyrick GR, Fernandez DC, Ghahari A, Pinhal CM, Simmonds Richardson M, Rupp AC, Cui L, Wu Z, Brown RL, Badea TC, Hattar S, Robinson PR. (2017) C-Terminal Phosphorylation Regulates the Kinetics of a Subset of Melanopsin-Mediated Behaviors in Mice. Proc Natl Acad Sci U S A 114(10):2741-46. PMID: 28223508 (Targeting Trends 17q1)
Ueki Y, Ramirez G, Salcedo E, Stabio ME, & Lefcort F. Loss of Ikbkap Causes Slow, Progressive Retinal Degeneration in a Mouse Model of Familial Dysautonomia. (2016). eNeuro, 3 (5):2016/10/05. PMC5037323
Summary: Familial dysautonomia (FD) is an autosomal recessive congenital neuropathy that is caused by a mutation in the gene for inhibitor of kappa B kinase complex-associated protein (IKBKAP). A classic hallmark of the disease is progressive blindness marked by retinal ganglion cell (RGC) loss and optic nerve atrophy. To investigate the consequences of Ikbkap loss in the retina, we generated Ikbkap conditional knockout mice using TUBA1a-Cre. Our data demonstrate that this is a powerful model system that faithfully recapitulates the phenotype and progression of FD blindness.
Berson DM, Castrucci AM, & Provencio I. Morphology and Mosaics of Melanopsin-Expressing Retinal Ganglion Cell Types in Mice. (2010). J Comp Neurol, 518 (13):2405-2422. 2010/05/27. PMC2895505
Objective: To provide a fuller description of murine cell types expressing melanopsin, their contribution to the plexuses of melanopsin dendrites, and mosaics formed by each type.
Summary: M1 cells, corresponding to the originally described ganglion-cell photoreceptors, occupy the ganglion cell or inner nuclear layers. M2 cells ramify in the inner third of the IPL. Rare bistratified cells deploy terminal dendrites within both melanopsin-immunoreactive plexuses within the ON sublayer.
Dose: IHC: the tissue was incubated for 24 hr at 4°C in unpurified primary antiserum diluted 1:2500. Immunoperoxidase labeling of flat mount tissue was incubated for 3 days in the unpurified primary antiserum diluted 1:2500.
Gonzalez-Menendez I, Contreras F, Cernuda-Cernuda R, Provencio I, Garcia-Fernandez JM (2010) Postnatal development and functional adaptations of the melanopsin photoreceptive system in the albino mouse retina. Invest Ophthalmol Vis Sci 51(9):4840-4847. (Targeting Trends 10q4)
Göz D, Studholme K, Lappi DA, Rollag MD, Provencio I, Morin LP (2008) Targeted destruction of photosensitive retinal ganglion cells with a saporin conjugate alters the effects of light on mouse circadian rhythms. PLoS ONE 3(9):e3153. (Targeting Trends 09q4)
Provencio I, Rollag MD, Castrucci AM (2002) Photoreceptive net in the mammalian retina. Nature 415:493.