Mark W Hankins

University of Oxford | OX · Nuffield Department of Clinical Neurosciences

Photoreceptor degeneration sufficient to produce severe visual loss often spares the inner retina. This raises hope for vision restoration treatments using optogenetics or electrical stimulation, which generate a replacement light input signal in surviving neurons. The success of these approaches is dependent on the capacity of surviving circuits o...

Photoreceptor degeneration sufficient to produce severe visual loss often spares the inner retina. This raises the hope that treatments using optogenetics or electrical stimulation, which generate a replacement light input signal in surviving neurons, may restore vision. The success of these approaches is dependent on the capacity of surviving circ...

Degenerative retinal disorders are a diverse family of diseases commonly leading to irreversible photoreceptor death, while leaving the inner retina relatively intact. Over recent years, innovative gene replacement therapies aiming to halt the progression of certain inherited retinal disorders have made their way into clinics. By rendering survivin...

During inherited retinal degenerations (IRDs), vision is lost due to photoreceptor cell death, however, a range of optogenetic tools have been shown to restore light responses in animal models. Restored response characteristics vary between tools and the neuronal cell population to which they are delivered: the interplay between these is complex bu...

Mutations in transcription factors often exhibit pleiotropic effects related to their complex expression patterns and multiple regulatory targets. One such mutation in the zinc finger homeobox 3 (ZFHX3) transcription factor, short circuit (Sci, Zfhx3Sci/+), is associated with significant circadian deficits in mice. However, given evidence of its re...

Human opsin-based photopigments have great potential as light-sensitisers, but their requirement for phototransduction cascade-specific second messenger proteins may restrict their functionality in non-native cell types. In this study, eight chimeric human opsins were generated consisting of a backbone of either a rhodopsin (RHO) or long-wavelength...

Purpose Retinal bipolar cells survive even in the later stages of inherited retinal degenerations (IRDs) and so are attractive targets for optogenetic approaches to vision restoration. However, it is not known to what extent the remodelling that these cells undergo during degeneration affects their function. Specifically, it is unclear if they are...

Optogenetic strategies to restore vision in patients blind from end-stage retinal degenerations aim to render remaining retinal neurons light-sensitive. We present an innovative combination of multi-electrode array recordings together with a complex pattern-generating light source as a toolset to determine the extent to which neural retinal respons...

Purpose: To characterize the retinal expression and localization of Kcne2, an ancillary (β) ion-channel subunit with an important role in fine-tuning cellular excitability. Methods: We analyzed available single-cell transcriptome data from tens of thousands of murine retinal cells for cell-type-specific expression of Kcne2 using state-of-the-art...

Melanopsin is a blue light-sensitive opsin photopigment involved in a range of non-image forming behaviours, including circadian photoentrainment and the pupil light response. Many naturally occurring genetic variants exist within the human melanopsin gene (OPN4), yet it remains unclear how these variants affect melanopsin protein function and down...

The discovery of melanopsin-expressing photosensitive retinal ganglion cells (pRGCs) has led to a fundamental change in our understanding of retinal light detection. pRGCs perform a broad range of non-visual functions — most notably mediating circadian entrainment to the environmental light/dark cycle. However, over the last two decades it has beco...

Melanopsin (OPN4) is an opsin photopigment expressed within intrinsically photosensitive retinal ganglion cells (ipRGCs) that mediate non-image forming (NIF) responses to light. Two single nucleotide polymorphisms (SNPs) in human melanopsin (hOPN4), Pro10Leu and Thr394Ile, have recently been associated with abnormal NIF responses to light, includin...

Cryptochromes 1 and 2 (CRY1/2) are key components of the negative limb of the mammalian circadian clock. Like many peripheral tissues, Cry1 and -2 are expressed in the retina, where they are thought to play a role in regulating rhythmic physiology. However, studies differ in consensus as to their localization and function, and CRY1 immunostaining h...

There is much debate on the adeno-associated virus (AAV) serotype that best targets specific retinal cell types and the route of surgical delivery—intravitreal or subretinal. This study compared three of the most efficacious AAV vectors known to date in a mouse model of retinal degeneration (rd1 mouse) and macaque and human retinal explants. GFP dr...

Significance Inherited retinal degenerations may result in blindness due to a progressive loss of photoreceptor cells. We assess subretinal delivery of human melanopsin using an adeno-associated viral vector to remaining retinal cells in a model of end-stage retinal degeneration. Human melanopsin, being already present in the eye, is unlikely to ge...

Two-pore domain (K2P) potassium channels perform essential roles in neuronal function. These channels produce background leak type potassium currents that act to regulate resting membrane potential and levels of cellular excitability. 15 different K2P channels have been identified in mammals and these channels perform important roles in a wide numb...

Circadian rhythms optimize physiology and behavior to the varying demands of the 24 h day. The master circadian clock is located in the suprachiasmatic nuclei (SCN) of the hypothalamus and it regulates circadian oscillators in tissues throughout the body to prevent internal desynchrony. Here, we demonstrate for the first time that, under standard 1...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Electronic Supplementary Material 1: Figures S1–S6, Supplemental Methods, And Supplemental Results And Discussion

A method of treating an eye disease comprising administering an adeno-associated virus (AAV) vector to a mammalian subject by subretinal injection, wherein the AAV vector comprises a nucleotide sequence encoding melanopsin operably linked to an expression control sequence to promote expression of melanopsin in cells of the eye of the subject.

Gene therapy using adeno-associated viral vectors (AAV) for the treatment of retinal degenerations has shown safety and efficacy in clinical trials. However, very high levels of vector expression may be necessary for the treatment of conditions such as Stargardt disease where a dual vector approach is potentially needed, or in optogenetic strategie...

Gnat−/−, Cnga3−/−, Opn4−/− triple knockout (TKO) mice lack essential components of phototransduction signalling pathways present in rods, cones and photosensitive retinal ganglion cells (pRGCs), and are therefore expected to lack all sensitivity to light. However, a number of studies have shown that light responses persist in these mice. In this st...

Light plays a critical role in the regulation of numerous aspects of physiology and behaviour, including the entrainment of circadian rhythms and the regulation of sleep. These responses involve melanopsin (OPN4)-expressing photosensitive retinal ganglion cells (pRGCs) in addition to rods and cones. Nocturnal light exposure in rodents has been show...

Excel spreadsheet containing, in separate sheets, data for Fig 1A–1C and underlying raw values used to generate averages for sleep profiles, sleep latency, and sleep duration in WT mice. (XLSX)

Excel spreadsheet containing, in separate sheets, data for S3A–S3F Fig and underlying raw values to generate averages for white light effect on sleep profiles, sleep latency, and sleep duration in WT- and melanopsin-deficient mice as well as for violet, blue, and green light effect on sleep profiles, sleep latency, and sleep duration in melanopsin-...

Excel spreadsheet containing, in separate sheets, data for Fig 3B–3E and underlying raw values to generate averages for time spent in the hidden zone in the 1st minute and in the following 9 min. (XLSX)

Excel spreadsheet containing, in separate sheets, relative Fos, Per1, and Per2 gene expression; data for Fig 4A–4B. (XLSX)

Excel spreadsheet containing, in separate sheets, data for Fig 5A–5B and underlying raw values of optical density (OD) and corticosterone concentration to generate averages for corticosterone concentration. (XLSX)

Excel spreadsheet containing relative Fos and Galanin gene expression in the SCN and the VLPO; data for Fig 6. (XLSX)

Excel spreadsheet containing, in separate sheets, data to generate S1A Fig (spectral power distribution of LED stimuli used) and S1B Fig (relative photoreceptor activity produced by 405 nm, 470 nm, and 530 nm). (XLSX)

Excel spreadsheet containing data for latency to first entry during behavioural light aversion in WT mice S4 Fig. (XLSX)

Excel spreadsheet containing, in separate sheets, data for S5 Fig and underlying raw values to generate averages for sleep profiles affected by RU-486 and vehicle treatment under blue light exposure. (XLSX)

Different wavelength stimuli used in this study. (A) Spectral power distribution of LED stimuli used. Three different wavelengths were used: 405 nm (violet), 470 nm (blue), and 530 nm (green). All stimuli were confirmed using a calibrated spectrometer. (B) Relative photoreceptor activity produced by 405 nm (violet), 470 nm, (blue), and 530 nm (gree...

Video demonstrates behavioural response to acute green (530 nm) light exposure in WT mouse. The original video of 3 h have been cut down to 1 h and 4 min (2 min darkness before and after 1 h light pulse) and accelerated eight times. The acute light exposure is indicated by text in the upper left corner in the video. WT mouse exposed to green light...

Video demonstrates behavioural response to acute blue (470 nm) light exposure in OPN4-/- mouse. The original video of 3 h have been cut down to 1 h and 4 min (2 min darkness before and after 1 h light pulse) and accelerated eight times. The acute light exposure is indicated by text in the upper left corner in the video. Opn4-/- mouse exposed to blu...

Excel spreadsheet containing, in separate sheets, data for Fig 2A–2E and underlying raw values to generate averages for sleep profiles, sleep latency, and sleep duration in WT- and melanopsin-deficient mice. (XLSX)

Prolonged sleep latency in response to blue light is dependent upon glucocorticoid receptor activation. RU-486 effect on sleep induction during blue light pulse. To investigate the reinforcing effect of blue light-induced plasma corticosterone, the glucocorticoid receptor antagonist RU-486 (100 mg/kg body weight, solid circles) and/or vehicle (cont...

Light available to different retinal photoreceptors under different lighting conditions. (A) Different wavelength stimuli used in this study. (B) Commonly used white light sources incandescent, fluorescent, and cool-white LED, which are designed to be enriched with longer wavelengths for human vision. (C) Daylight versus twilight data, showing the...

Video demonstrates behavioural response to acute blue (470 nm) light exposure in WT mouse. The original video of 3 h have been cut down to 1 h and 4 min (2 min darkness before and after 1 h light pulse) and accelerated 8 times. The acute light exposure is indicated by text in the upper left corner in the video. After acute blue light exposure, WT m...

Excel spreadsheet containing, in separate sheets, data for S2A–S2C Fig and underlying raw values to generate averages for sleep profiles, sleep latency and sleep duration under blue and green light exposure at ZT22. (XLSX)

Excel spreadsheet containing, in separate sheets, data for relative alpha opic lux using white light sources incandescent S6A Fig, fluorescent and cool-white LED S6B Fig as well as daylight versus twilight data S6C Fig. (XLSX)

Effects of different wavelength light on sleep at ZT22. (A) Mice exposed to blue (470 nm) light for 1 hr at ZT22 showed delayed sleep onset compared to green (530 nm) light. (B) Comparable to ZT14, sleep induction was delayed in response to blue light compared with green light. (C) Total sleep duration during the 1 h light pulse was unchanged under...

Effect of white light on sleep induction in Opn4-/- mice. (A) Sleep onset latency induced by acute white light exposure in WT and Opn4-/- mice at ZT14. Opn4-/- deficient as well as WT mice were exposed to 1 hr white light (~250 lux) at ZT14. (B) Opn4-/- mice showed delayed sleep onset under white light exposure compared to wildtype mice (A,C), whic...

Latency to first entry during behavioural light aversion. The latency from placing the mouse into the box until the first entry to the hidden zone was significantly shorter under blue illumination compared to violet, green light and control = dark condition. One-way ANOVA for light condition, F(3.25) = 20.70, p ≤ 0.001. Posthoc Tukey blue versus co...

Video demonstrates behavioural response to acute green (530 nm) light exposure in OPN4-/- mouse. The original video of 3 h have been cut down to 1 h and 4 min (2 min darkness before and after 1 h light pulse) and accelerated eight times. The acute light exposure is indicated by text in the upper left corner in the video. Opn4-/- mouse exposed to gr...

Over the past two decades there have been significant advances in our understanding of both the anatomy and function of the melanopsin system. It has become clear that rather than acting as a simple irradiance detector the melanopsin system is in fact far more complicated. The range of behavioural systems known to be influenced by melanopsin activi...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Acute light exposure exerts various effects on physiology and behaviour. Although the effects of light on brain network activity in humans are well demonstrated, the effects of light on cognitive performance are inconclusive, with the size, as well as direction, of the effect depending on the nature of the task. Similarly, in nocturnal rodents, bri...

Light affects animal physiology and behavior more than simply through classical visual, image-forming pathways. Nonvisual photoreception regulates numerous biological systems, including circadian entrainment, DNA repair, metabolism , and behavior. However, for the majority of these processes, the photoreceptive molecules involved are unknown. Given...

Abstract Purpose: Opsins are light-sensitive G-protein coupled receptor proteins, essential for vision, circadian rhythmicity and eye development. Opsins function by activating G-protein second messenger systems. Rod and cone opsins activate Gαt, while melanopsin activates Gαq/11. If inner retinal neurons, such as bipolar cells, were engineered to...

Melanopsin (OPN4) is a retinal photopigment that mediates a wide range of non-image-forming (NIF) responses to light [1, 2] including circadian entrainment [3], sleep induction [4], the pupillary light response (PLR) [5], and negative masking of locomotor behavior (the acute suppression of activity in response to light) [6]. How these diverse NIF r...

Circadian rhythms in physiology and behavior provide a selective advantage by enabling organisms to anticipate rhythmic changes in their environment. These rhythms are based upon a molecular clock generated via an intracellular transcriptional-translational feedback loop involving a number of key clock genes. However, to be of practical use, circad...

Gene therapy is ideally suited to the treatment of inherited retinal degenerations in order to prevent blindness. The target area of the outer retina is small and relatively immune privileged, which facilitates the delivery of small volumes of vector without generating signifi cant immune reactions. Moreover, most of the currently untreatable forms...

Extraretinal photoreceptors located within the medio-basal hypothalamus regulate the photoperiodic control of seasonal reproduction in birds. An action spectrum for this response describes an opsin photopigment with a λmax of ∼492nm. Beyond this however, the specific identity of the photopigment remains unresolved. Several candidates have emerged i...

Summary A recent study defines a novel role of melanopsin-expressing ipRGCs, showing that these inner retinal photoreceptors function as retinal irradiance detectors and provide a local measure of luminance to regulate functional adaptation in the mammalian retina.

In addition to well-characterised visual systems, many organisms, including the craniates, possess a complex sensory system of non-visual photoreceptors that detect light for a diverse array of non-image-forming tasks. Like the photoreceptors of image-forming systems, the pigments contained within non-visual photoreceptive cells comprise a protein...

In addition to classical image-forming vision, the vertebrates exhibit a range of non-image-forming light detection systems that utilise opsin photopigments. Within the CNS these systems are present in a range of anatomical locations that include both eye and brain. In mammals the eye is both responsible and required for all commonly measured respo...

Melanopsin expressing photosensitive retinal ganglion cells (pRGCs) represent a third class of ocular photoreceptors and mediate a range of non-image forming responses to light. Melanopsin is a G protein coupled receptor (GPCR) and existing data suggest that it employs a membrane bound signalling cascade involving Gnaq/11 type G proteins. However,...

Photopigments are molecules that react to light and mediate a number of processes and behaviours in animals. Visual pigments housed within the photoreceptors of the eye, such as the rods and cones in vertebrates are the best known, however, visual pigments are increasingly being found in other tissues, including other retinal cells, the skin and th...