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Intrinsically Photosensitive Retinal Ganglion Cells: Assessment of Their Influence on Pupillary Responses in Primates

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Circadian Clocks

Part of the book series: Neuromethods ((NM,volume 186))

Abstract

Pupillary responses in primates are driven by melanopsin-containing, intrinsically photosensitive retinal ganglion cells (ipRGCs) that project to the pretectum. These cells also project to the suprachiasmatic nucleus (SCN) to photically entrain circadian rhythms. Given the similarity of the ipRGCs that project to the pretectum and SCN, the study of light-evoked pupillary responses that reflect the activity of these pretectally projecting ipRGCs should provide substantial insights into the ipRGC signals that impinge on the SCN. Here, we describe studies of light-evoked pupillary responses in macaques and humans including the increasingly studied post-illumination pupil response (PIPR). We describe the best practices for conducting these types of experiments and for analyzing the acquired data. We also provide advice about potential pitfalls and confounding variables that can affect the outcome and interpretation of these types of experiments.

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References

  1. Mohawk JA, Green CB, Takahashi JS (2012) Central and peripheral circadian clocks in mammals. Annu Rev Neurosci 35:445–462

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Moore RY, Lenn NJ (1972) A retinohypothalamic projection in the rat. J Comp Neurol 146(1):1–14

    Article  CAS  PubMed  Google Scholar 

  3. Berson DM, Dunn FA, Takao M (2002) Phototransduction by retinal ganglion cells that set the circadian clock. Science 295:1070–1073

    Article  CAS  PubMed  Google Scholar 

  4. Hattar S, Liao HW, Takao M, Berson DM, Yau KW (2002) Melanopsin-containing retinal ganglion cells: architecture, projections, and intrinsic photosensitivity. Science 295:1065–1070

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Lucas RJ, Hattar S, Takao M, Berson DM, Foster RG, Yau KW (2003) Diminished pupillary light reflex at high irradiances in melanopsin-knockout mice. Science 299(5604):245–247

    Article  CAS  PubMed  Google Scholar 

  6. Dacey DM, Liao HW, Peterson BB, Robinson FR, Smith VC, Pokorny J, Yau KW, Gamlin PD (2005) Melanopsin-expressing ganglion cells in primate retina signal colour and irradiance and project to the LGN. Nature 433:749–754

    Article  CAS  PubMed  Google Scholar 

  7. Gamlin PD, McDougal DH, Pokorny J, Smith VC, Yau KW, Dacey DM (2007) Human and macaque pupil responses driven by melanopsin-containing retinal ganglion cells. Vis Res 47(7):946–954

    Article  CAS  PubMed  Google Scholar 

  8. Do MT, Yau KW (2010) Intrinsically photosensitive retinal ganglion cells. Physiol Rev 90(4):1547–1581

    Article  CAS  PubMed  Google Scholar 

  9. Do MTH (2019) Melanopsin and the intrinsically photosensitive retinal ganglion cells: biophysics to behavior. Neuron 104(2):205–226

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Provencio I, Jiang G, De Grip WJ, Hayes WP, Rollag MD (1998) Melanopsin: an opsin in melanophores, brain, and eye. Proc Natl Acad Sci U S A 95(1):340–345

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Provencio I, Rodriguez IR, Jiang G, Hayes WP, Moreira EF, Rollag MD (2000) A novel human opsin in the inner retina. J Neurosci 20(2):600–605

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Qiu X, Kumbalasiri T, Carlson SM, Wong KY, Krishna V, Provencio I, Berson DM (2005) Induction of photosensitivity by heterologous expression of melanopsin. Nature 433(7027):745–749

    Article  CAS  PubMed  Google Scholar 

  13. Perez-Leon JA, Warren EJ, Allen CN, Robinson DW, Brown RL (2006) Synaptic inputs to retinal ganglion cells that set the circadian clock. Eur J Neurosci 24(4):1117–1123

    Article  PubMed  PubMed Central  Google Scholar 

  14. Wong KY, Dunn FA, Graham DM, Berson DM (2007) Synaptic influences on rat ganglion-cell photoreceptors. J Physiol 582(Pt 1):279–296

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Schmidt TM, Kofuji P (2010) Differential cone pathway influence on intrinsically photosensitive retinal ganglion cell subtypes. J Neurosci 30(48):16262–16271

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Liao HW, Ren X, Peterson BB, Marshak DW, Yau KW, Gamlin PD, Dacey DM (2016) Melanopsin-expressing ganglion cells on macaque and human retinas form two morphologically distinct populations. J Comp Neurol 524(14):2845–2872

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Jusuf PR, Lee SC, Hannibal J, Grünert U (2007) Characterization and synaptic connectivity of melanopsin-containing ganglion cells in the primate retina. Eur J Neurosci 26:2906–2921

    Article  PubMed  Google Scholar 

  18. Joo HR, Peterson BB, Dacey DM, Hattar S, Chen SK (2013) Recurrent axon collaterals of intrinsically photosensitive retinal ganglion cells. Vis Neurosci 30:175–182

    Article  PubMed  PubMed Central  Google Scholar 

  19. Hattar S, Lucas RJ, Mrosovsky N, Thompson S, Douglas RH, Hankins MW, Lem J, Biel M, Hofmann F, Foster RG, Yau KW (2003) Melanopsin and rod-cone photoreceptive systems account for all major accessory visual functions in mice. Nature 424(6944):76–81

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Panda S, Provencio I, Tu DC, Pires SS, Rollag MD, Castrucci AM, Pletcher MT, Sato TK, Wiltshire T, Andahazy M, Kay SA, Van Gelder RN, Hogenesch JB (2003) Melanopsin is required for non-image-forming photic responses in blind mice. Science 301(5632):525–527

    Article  CAS  PubMed  Google Scholar 

  21. Fu Y, Zhong H, Wang MH, Luo DG, Liao HW, Maeda H, Hattar S, Frishman LJ, Yau KW (2005) Intrinsically photosensitive retinal ganglion cells detect light with a vitamin A-based photopigment, melanopsin. Proc Natl Acad Sci U S A 102(29):10339–10344

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Hannibal J, Christiansen AT, Heegaard S, Fahrenkrug J, Kiilgaard JF (2017) Melanopsin expressing human retinal ganglion cells: subtypes, distribution, and intraretinal connectivity. J Comp Neurol 525(8):1934–1961

    Article  CAS  PubMed  Google Scholar 

  23. Baver SB, Pickard GE, Sollars PJ, Pickard GE (2008) Two types of melanopsin retinal ganglion cell differentially innervate the hypothalamic suprachiasmatic nucleus and the olivary pretectal nucleus. Eur J Neurosci 27(7):1763–1770

    Article  PubMed  Google Scholar 

  24. Ecker JL, Dumitrescu ON, Wong KY, Alam NM, Chen SK, LeGates T, Renna JM, Prusky GT, Berson DM, Hattar S (2010) Melanopsin-expressing retinal ganglion-cell photoreceptors: cellular diversity and role in pattern vision. Neuron 67(1):49–60

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. McDougal DH, Gamlin PD (2015) Autonomic control of the eye. Compr Physiol 5(1):439–473

    PubMed  PubMed Central  Google Scholar 

  26. Hannibal J, Kankipati L, Strang CE, Peterson BB, Dacey D, Gamlin PD (2014) Central projections of intrinsically photosensitive retinal ganglion cells in the macaque monkey. J Comp Neurol 522(10):2231–2248

    Article  CAS  PubMed  Google Scholar 

  27. McDougal DH, Gamlin PD (2010) The influence of intrinsically-photosensitive retinal ganglion cells on the spectral sensitivity and response dynamics of the human pupillary light reflex. Vis Res 50:72–87

    Article  PubMed  Google Scholar 

  28. Kankipati L, Girkin CA, Gamlin PD (2010) Post-illumination pupil response in subjects without ocular disease. Invest Ophthalmol Vis Sci 51(5):2764–2769

    Article  PubMed  PubMed Central  Google Scholar 

  29. Kankipati L, Girkin CA, Gamlin PD (2011) The post-illumination pupil response is reduced in glaucoma patients. Invest Ophthalmol Vis Sci 52(5):2287–2292

    Article  PubMed  PubMed Central  Google Scholar 

  30. Feigl B, Zele AJ (2014) Melanopsin-expressing intrinsically photosensitive retinal ganglion cells in retinal disease. Optom Vis Sci 91(8):894–903

    Article  PubMed  Google Scholar 

  31. Joyce DS, Feigl B, Cao D, Zele AJ (2015) Temporal characteristics of melanopsin inputs to the human pupil light reflex. Vis Res 107:58–66

    Article  PubMed  Google Scholar 

  32. Cao D, Nicandro N, Barrionuevo PA (2015) A five-primary photostimulator suitable for studying intrinsically photosensitive retinal ganglion cell functions in humans. J Vis 15(1):27

    Article  PubMed  PubMed Central  Google Scholar 

  33. Spitschan M, Jain S, Brainard DH, Aguirre GK (2014) Opponent melanopsin and S-cone signals in the human pupillary light response. Proc Natl Acad Sci U S A 111(43):15568–15572

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Roecklein K, Wong P, Ernecoff N, Miller M, Donofry S, Kamarck M, Wood-Vasey WM, Franzen P (2013) The post illumination pupil response is reduced in seasonal affective disorder. Psychiatry Res 210(1):150–158

    Article  PubMed  PubMed Central  Google Scholar 

  35. Kardon R, Anderson SC, Damarjian TG, Grace EM, Stone E, Kawasaki A (2011) Chromatic pupillometry in patients with retinitis pigmentosa. Ophthalmology 118(2):376–381

    Article  PubMed  Google Scholar 

  36. Park JC, Moura AL, Raza AS, Rhee DW, Kardon RH, Hood DC (2011) Toward a clinical protocol for assessing rod, cone, and melanopsin contributions to the human pupil response. Invest Ophthalmol Vis Sci 52(9):6624–6635

    Article  PubMed  PubMed Central  Google Scholar 

  37. Lucas RJ, Peirson SN, Berson DM, Brown TM, Cooper HM, Czeisler CA, Figueiro MG, Gamlin PD, Lockley SW, O’Hagan JB, Price LL, Provencio I, Skene DJ, Brainard GC (2014) Measuring and using light in the melanopsin age. Trends Neurosci 37(1):1–9

    Article  CAS  PubMed  Google Scholar 

  38. Kelbsch C, Strasser T, Chen Y, Feigl B, Gamlin PD, Kardon R, Peters T, Roecklein KA, Steinhauer SR, Szabadi E, Zele AJ, Wilhelm H, Wilhelm BJ (2019) Standards in pupillography. Front Neurol 10:129

    Article  PubMed  PubMed Central  Google Scholar 

  39. Judge SJ, Richmond BJ, Chu FC (1980) Implantation of magnetic search coils for measurement of eye position: an improved method. Vis Res 20(6):535–538

    Article  CAS  PubMed  Google Scholar 

  40. Clarke RJ, Zhang H, Gamlin PD (2003) Characteristics of the pupillary light reflex in the alert rhesus monkey. J Neurophysiol 89(6):3179–3189

    Article  PubMed  Google Scholar 

  41. Dolan RP, Schiller PH (1994) Effects of ON channel blockade with 2-amino-4-phosphonobutyrate (APB) on brightness and contrast perception in monkeys. Vis Neurosci 11(1):23–32

    Article  CAS  PubMed  Google Scholar 

  42. Sieving PA, Murayama K, Naarendorp F (1994) Push-pull model of the primate photopic electroretinogram: a role for hyperpolarizing neurons in shaping the b-wave. Vis Neurosci 11(3):519–532

    Article  CAS  PubMed  Google Scholar 

  43. Kondo M, Sieving PA (2001) Primate photopic sine-wave flicker ERG: vector modeling analysis of component origins using glutamate analogs. Invest Ophthalmol Vis Sci 42(1):305–312

    CAS  PubMed  Google Scholar 

  44. Binda P, Gamlin PD (2017) Renewed attention on the pupil light reflex. Trends Neurosci 40(8):455–457

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Loewenfeld IE, Lowenstein O (1993) The pupil: anatomy, physiology, and clinical applications. Iowa State University Press, Ames and Wayne State University Press, Detroit

    Google Scholar 

  46. Wilhelm B, Giedke H, Lüdtke H, Bittner E, Hofmann A, Wilhelm H (2001) Daytime variations in central nervous system activation measured by a pupillographic sleepiness test. J Sleep Res 10(1):1–7

    Article  CAS  PubMed  Google Scholar 

  47. McLaren JW, Hauri PJ, Lin SC, Harris CD (2002) Pupillometry in clinically sleepy patients. Sleep Med 3(4):347–352

    Article  PubMed  Google Scholar 

  48. Webster JG, Cohen GH, Boynton RM (1968) Optimizing the use of the criterion response for the pupil light reflex. J Opt Soc Am 58(3):419–424

    Article  CAS  PubMed  Google Scholar 

  49. Melyan Z, Tarttelin EE, Bellingham J, Lucas RJ, Hankins MW (2005) Addition of human melanopsin renders mammalian cells photoresponsive. Nature 433(7027):741–745

    Article  CAS  PubMed  Google Scholar 

  50. Mure LS, Cornut PL, Rieux C, Drouyer E, Denis P, Gronfier C, Cooper HM (2009) Melanopsin bistability: a fly’s eye technology in the human retina. PLoS One 4(6):e5991

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  51. Emanuel AJ, Do MT (2015) Melanopsin tristability for sustained and broadband phototransduction. Neuron 85(5):1043–1055

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Pokorny J, Smith VC, Lutze M (1987) Aging of the human lens. Appl Opt 26(8):1437–1440

    Article  CAS  PubMed  Google Scholar 

  53. Coren S (1987) A rapid method to assess crystalline lens pigment density in vivo. Acta Ophthalmol 65(5):575–578

    Article  CAS  Google Scholar 

  54. Kraft TW, Neitz J, Neitz M (1998) Spectra of human L cones. Vis Res 38(23):3663–3670

    Article  CAS  PubMed  Google Scholar 

  55. Baylor DA, Nunn BJ, Schnapf JL (1984) The photocurrent, noise and spectral sensitivity of rods of the monkey Macaca fascicularis. J Physiol 357:575–607

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Baylor DA, Nunn BJ, Schnapf JL (1987) Spectral sensitivity of cones of the monkey Macaca fascicularis. J Physiol 390:145–160

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Fu Y, Liao HW, Do MT, Yau KW (2005) Non-image-forming ocular photoreception in vertebrates. Curr Opin Neurobiol 15(4):415–422

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Wong KY (2012) A retinal ganglion cell that can signal irradiance continuously for 10 hours. J Neurosci 32(33):11478–11485

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Markwell EL, Feigl B, Zele AJ (2010) Intrinsically photosensitive melanopsin retinal ganglion cell contributions to the pupillary light reflex and circadian rhythm. Clin Exp Optom 93:137–149

    Article  PubMed  Google Scholar 

  60. Adhikari P, Zele AJ, Feigl B (2015) The post-illumination pupil response (PIPR). Invest Ophthalmol Vis Sci 56:3838–3849

    Article  CAS  PubMed  Google Scholar 

  61. Kelbsch C, Maeda F, Lisowska J, Lisowski L, Strasser T, Stingl K, Wilhelm B, Wilhelm H, Peters T (2017) Analysis of retinal function using chromatic pupillography in retinitis pigmentosa and the relationship to electrically evoked phosphene thresholds. Acta Ophthalmol 95(4):e261–e269

    Article  CAS  PubMed  Google Scholar 

  62. Kawasaki A, Herbst K, Sander B, Milea D (2010) Selective wavelength pupillometry in Leber hereditary optic neuropathy. Clin Exp Ophthalmol 38:322–324

    Article  PubMed  Google Scholar 

  63. Herbst K, Sander B, Milea D, Lund-Andersen H, Kawasaki A (2011) Test-retest repeatability of the pupil light response to blue and red light stimuli in normal human eyes using a novel pupillometer. Front Neurol 2:10

    Article  PubMed  PubMed Central  Google Scholar 

  64. Feigl B, Mattes D, Thomas R, Zele AJ (2011) Intrinsically photosensitive (melanopsin) retinal ganglion cell function in glaucoma. Invest Ophthalmol Vis Sci 52:4362–4367

    Article  CAS  PubMed  Google Scholar 

  65. Do MT, Yau KW (2013) Adaptation to steady light by intrinsically photosensitive retinal ganglion cells. Proc Natl Acad Sci U S A 110(18):7470–7475

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Hannibal J, Georg B, Hindersson P, Fahrenkrug J (2005) Light and darkness regulate melanopsin in the retinal ganglion cells of the albino Wistar rat. J Mol Neurosci 27(2):147–155

    Article  CAS  PubMed  Google Scholar 

  67. Sakamoto K, Liu C, Kasamatsu M, Pozdeyev NV, Iuvone PM, Tosini G (2005) Dopamine regulates melanopsin mRNA expression in intrinsically photosensitive retinal ganglion cells. Eur J Neurosci 22(12):3129–3136

    Article  PubMed  Google Scholar 

  68. 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):2741–2746

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. van der Meijden WP, te Lindert BH, Bijlenga D, Coppens JE, Gómez-Herrero G, Bruijel J, Kooij JJ, Cajochen C, Bourgin P, Van Someren EJ (2015) Post-illumination pupil response after blue light: reliability of optimized melanopsin-based phototransduction assessment. Exp Eye Res 139:73–80

    Article  PubMed  CAS  Google Scholar 

  70. Beer RD, Macleod DI, Miller TP (2005) The extended Maxwellian view (BIGMAX): a high-intensity, high-saturation color display for clinical diagnosis and vision research. Behav Res Methods 37(3):513–552

    Article  PubMed  Google Scholar 

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Acknowledgments

Supported by grants from the National Institutes of Health, EY025555 and P30 EY03039. Also supported by Research to Prevent Blindness and the Eyesight Foundation of Alabama. I also thank David McDougal for the use of an unpublished figure.

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Gamlin, P.D. (2022). Intrinsically Photosensitive Retinal Ganglion Cells: Assessment of Their Influence on Pupillary Responses in Primates. In: Hirota, T., Hatori, M., Panda, S. (eds) Circadian Clocks. Neuromethods, vol 186. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2577-4_7

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  • DOI: https://doi.org/10.1007/978-1-0716-2577-4_7

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