Melanopsin photoreceptors

Question 1

Freedman

Answer 1

1999, noted that transgenic rodless/coneless mice maintain circadian photoentrainment responses. These are abolished with removal of the eye, indicating that the murin eye contains additional receptors to regulate the circadian clock

Question 2

Berson

Answer 2

2002, retrogradely labelled SCN-projecting neurons in the rat. Then identified the intrinsic response of these cells by mechanically/pharmacologically isolating them from rods/cones/bipolar cells.
These RGCs were intrinsically photosensitive, with a slow onset, depolarising response to light, maximally sensitie at 480nm (differing from fast, hyperpolarising response of rods/cones).
The sensitivity, spectral tuning, and slow kinetics of their light response matched those obsered in circadian photoentrainment

Question 3

Lucas

Answer 3

2003, the ipRGCs of opn4(-/-) mice were no longer intrinsically photosensitive (though number, morphology and projections were unchanged) indicating that photosensitivity was melanopsin dependent.

Question 4

Do

Answer 4

2009, Generated BAC-transgenic mice expressing tdTomato under the malanopsin promoter. Used perforated-patch recording to make voltage-clamp measurements from pharmacologically/mechanically isolated ipRGCs.
ipRGCs have low sensitivity due to lower photopigment density, but have larger unitary response (shown by single-photon response) which is more prolonged (improving sensitivity by conferring high temporal summation).
ipRGCs have a low threshold and can signal single-photon responses to the brain (as unitary responses are large and prolonged)

Question 5

Hughes

Answer 5

2014, Gq, G11, G14 are highly co-expressed in mouse retina ipRGCs. siRNA experiments showed that melanopsin can signal via all 3 types in vitro, and multiple members of the family could participate in phototransduction in vivo

Question 6

Sekaran

Answer 6

2007, previous studies had shownt hat ipRGCs express TRPC6 and TRPC7. 2-APB (TRPCs and IP3R antagonist) potently inhibits ipRGCs in vivo independent of store-dependent Ca release, indicating 2-APB acts to block the light response by blocking TRPCs

Question 7

Xue

Answer 7

2011, ablating the expression of TRPC6 and TRPC7 in mice eliminated the M1-ipRGC light response

Question 8

Hattar (1)

Answer 8

2006, targetted tau-lacZ to the melanopsin gene locus in mice. Showed that ipRGCs project to multiple brain regions (SCN, intergeniculate leaflet, OPN, vLGN (visual processing), preoptic area (sleep), superior colliculus (motion detection responses) periaqueductal grey, lateral habernula). Thus, ipRGCs may have a role in multiple functions.
Injected anterograde tracer CBT (cholera toxin subunit B) into the retina to label retinal afferents. Co-staining showed that ipRGCS provide most of the retinal input to SCN, IGL and lateral habernula, and much of that to OPN

Question 9

Hattar (2)

Answer 9

2003, showing that only rods, cones and ipRGCs are required for NIF functions.
Opn4(-/-) mice have partially impaired PLR and photoentrainment.
Opn4(-/-) mice with disabled rod and cone function have completely impaired PLR and photoentrainment (son’t show any funciton at all, worse that just Opn4(-/-)), indicating that the rod-cone and ipRGC systems work together.

Question 10

Güler

Answer 10

2008, mice with genetically ablated ipRGCs (the whole cell) show NIF funciton deficits. These deficits are worse that opn4(-/-) mice, and resemble those seen in mice lacking all three photoreceptor classes

Question 11

Panda

Answer 11

2002, opn4(-/-) mice entrain normally to L:D cycles and don’t show circadian rhythm defects in constant darkness.
However, they show severely attenuated phase resetting in response to brief light pulses, indicated impaired phase shifting behaviour.
Thus, melanopsin plays a critical role in photoentrainment

Question 12

Altimus

Answer 12

2010, Found that mice homozygous for the rod transducin locus inactivating mutation (Gnat1(-/-)) failed to photoentrain at scotopic (low) light intensities.
Thus, at low light intensities, cone and ipRGCs do not have enough sensitivity, and photoentrainment relies on rods (which signal through ipRGCs)

Question 13

Lucas

Answer 13

2003, evidence for diminished PLR at high irradiances in opn4(-/-) mice

Question 14

Gamlin

Answer 14

2007, showed that significant PLR persisted in behaving mecaque after pharmacological blockade of rod and cone signals, and that pupillary responses display the characteristics of ipRGC photoresponses

Question 15

Barnard

Answer 15

2006, it's known that ipRGCs exhibit intra-retinal commumination via gap junction coupling. 
Used opn4(-/-) mice to study the role of the inner retinal photoreceptors in diurnal regulation of retinal function (measured by electroretinography). Found that KO abolished the circadian control of the diurnal rhythms of the retinal cone pathway, attenuating the diurnal variation (impaired processing during the day).
Suggests that melanopsin regulates visual processing within the retine and may optimise classical visual pathways according to time of day.

Question 16

Lupi

Answer 16

2008, overal sleep-wake rhythmicity is preserved in the absence of circadian clock, and it remains synchronised to the L:D cycle.
They found that photic regualtion of sleep is predominantely mediated by ipRGCs, as opn4(-/-) mice lacked acute sleep induction

Question 17

Noseda

Answer 17

2010, found that photo-allodynia and exacerbation of migraine headache by light occured in blind individuals (rod/cone degredation).

Question 18

Dacey

Answer 18

2005, found evidence for an anatomically distinct population of ‘giant’ ipRGCs in the primate retina. They were intrinsially photosensitive, and activated by rods and cones, and signalled irradiance over the full range of human vision.
They projected to the LGN

Question 19

Zaidi

Answer 19

2008, found that a blind subject lacking functional rods and cones was able to correctly report a threshold short-wavelength stimulus (approx 480), but not other stimuli

Question 20

Ecker (1)

Answer 20

2010, using a Cre-recombinase reporter mouse line, showed that ipRGCs projected to the superior colliculus and dLGC (both retinotopically organised nuclei mediating object localisation and discrimination).
Mice were trained to distinguish between a sinusoidal grating and a grey screen. Rodless/coneless mice could do this, while triple KO mice could not.

Question 21

Provencio

Answer 21

2002, revealed two networks of melanopsin immunoreactive dendrites using highly sensitive anti-melanopsin ABs

Question 22

Schimdt

Answer 22

2008, used BAC-transgenesis to engineer a mouse line expressing eGFP under the melanopsin promoter and identified three morphological subtypes.
M1: previously characterised, dendrites stratisfying in the outer (OFF) sublamina of the IPL
M2: dendrites stratifying in the inner (ON) sublamina of IPL
M3: dendrites bistratify in the outer and inner sublaminae.
Found that the extrinsic (rod/cone) influence on ipRGCs correlates with their extensive dendritic stratification in IPL.

Question 23

Schmidt and Kofuji (1)

Answer 23

2009, used a transgenic mouse model labelling ipRGCs in vivo to compare the morphology and physiology of M1 and M2 cells.
Morphology: found that M2s had significantly larger dendritic field diameters, more complex and branched dedritic arbors, and larger soma diameter.
Physiology: carried out simulataneous and dual whole-cell recordings, showed that M2 had smaller depolarisation to stimulus, and had a smaller light-evoked current (though it was relatively sustatined compared to the large but rapidly decaying M1 current). M1 had might higher sensitivity (due to much higher melanopsin expression).

Question 24

Schimdt and Kofuji (2)

Answer 24

2011, used a transgenic mouse model labelling ipRGCs with EGFP in vivo to describe the morphology and physiology of M3 cells.
Morphology: M3s were morphologically heterogenous. M3 were more similar to M2 in terms of size.complexity of dendritic arbors, but they differed in stratification.
Physiology: M3s were physiologicall homogenous, and showed unique spiking patterns to M1 and M2.

Question 25

Ecker (2)

Answer 25

2010, crossed a mouse expressing Cre recombinase at opn4 with a cre-dependent EGFP reporter mouse line and identified M4 and M5 cells.
Found that both stratified in the inner sublamina of IPL, but they had unique morphologies. M4s had the largest soma of any identified subtype, and the largest and most complex dendritic arbor. M5s had small and highly branched arbors arranged uniformaly around the soma .
BUT - both had very low levels of melanopsin and weak intrinsic light reponses (especially M5). Perhaps melanopsin was expressed during development but no longer? Or perhaps the identification of M5 was just an anomaly of the tagging system.

Question 26

Ecker (3)

Answer 26

2010, characterised the differential expression of the ipRGCs using an Onp4(Cre/+);Z/AP mouse to trace the axons (better than findings with Opn4(tau-lacZ).
Argues that M1s are the main projections to the SCN and OPN, and non-M1s are the main projections to dLGN, SC, OPN core and others.

Question 27

Chen

Answer 27

2011, found that the Bnr3b transcritpion factor was expressed in all non-M1 cells, but only a fraction of M1 cells.
Brn3b negative M1s innervated the SCN, whilst Brn3b positive M1s projected to all known M1 targets except the SCN.
The ablation of Brn3b positive M1 impaired PRL but didn’t effect photoentrainment. This distinction may provide an explanation for the differential influence of rods and cones on photoentrainment and PRL.