Melanopsin Activation Research Articles

Chromatic pupillometry as a novel means to assess outer retinal function in health and disease states

AbstractQuantified pupil responses to colored light stimuli is a non‐invasive means to monitor melanopsin function as well as rod and cone activity. Understanding the trivariate contributions to the pupil light reflex has led to renewed interest in using the pupil response to monitor a variety of diseases. The characteristic pupillographic feature of intrinsic, melanopsin‐mediated ganglion cell activity is persistent contraction after light offset. This post‐illumination response is abnormal in patients with glaucoma. In patients with retinal degenerative disease, the retinal sensitivity to blue light as measured by pupillometry is reduced. This correlates with loss of rod function but also hints at reduced melanopsin activity. Clinical examples showing pupil response to red and blue light in patients with neuroretinal visual loss will be discussed. Commercial interest

Circadian and Wake-Dependent Effects on the Pupil Light Reflex in Response to Narrow-Bandwidth Light Pulses

Nonvisual light-dependent functions in humans are conveyed mainly by intrinsically photosensitive retinal ganglion cells, which express melanopsin as photopigment. We aimed to identify the effects of circadian phase and sleepiness across 24 hours on various aspects of the pupil response to light stimulation. We tested 10 healthy adults hourly in two 12-hour sessions covering a 24-hour period. Pupil responses to narrow bandwidth red (635 ± 18 nm) and blue (463 ± 24 nm) light (duration of 1 and 30 seconds) at equal photon fluxes were recorded, and correlated with salivary melatonin concentrations at the same circadian phases and to subjective sleepiness ratings. The magnitude of pupil constriction was determined from minimal pupil size. The post-stimulus pupil response was assessed from the pupil size at 6 seconds following light offset, the area within the redilation curve, and the exponential rate of redilation. Among the measured parameters, the pupil size 6 seconds after light offset correlated with melatonin concentrations (P < 0.05) and showed a significant modulation over 24 hours with maximal values after the nocturnal peak of melatonin secretion. In contrast, the post-stimulus pupil response following red light stimulation correlated with subjective sleepiness (P < 0.05) without significant changes over 24 hours. The post-stimulus pupil response to blue light as a marker of intrinsic melanopsin activity demonstrated a circadian modulation. In contrast, the effect of sleepiness was more apparent in the cone contribution to the pupil response. Thus, pupillary responsiveness to light is under influence of the endogenous circadian clock and subjective sleepiness.

Short wavelength light filtering by the natural human lens and IOLs – implications for entrainment of circadian rhythm

Photoentrainment of circadian rhythm begins with the stimulation of melanopsin containing retinal ganglion cells that respond directly to blue light. With age, the human lens becomes a strong colour filter attenuating transmission of short wavelengths. The purpose of the study was to examine the effect the ageing human lens may have for the photoentrainment of circadian rhythm and to compare with intraocular implant lenses (IOLs) designed to block UV radiation, violet or blue light. The potential for photoentrainment of circadian rhythm was computed for 29 human donor lenses (18-76 years) and five IOLs (one UV, two violet and two blue light blocking) based on the transmission properties of the lenses and the spectral characteristics of melanopsin activation and two of it's physiological outcomes; melanopsin-driven pupillary light reponse and light-induced melatonin suppression. The potential for melanopsin stimulation and melatonin suppression was reduced by 0.6-0.7 percentage point per year of life because of yellowing of the natural lens. The computed effects were small for the IOLs and did not exceed that of a 22.2-year-old natural lens for the blue-blocking IOLs. The results show that photoentrainment of circadian rhythm may be significantly impaired in older subjects because of the colour filtering characteristics of the human lens, whereas the effects were small for all three types of IOLs studied. Consequently, the ageing process of the natural lens is expected to influence the photoentrainment of circadian rhythm, whereas IOLs are not expected to be detrimental to circadian rhythm.

Intrinsic phototransduction persists in melanopsin-expressing ganglion cells lacking diacylglycerol-sensitive TRPC subunits

In mammals, intrinsically photosensitive retinal ganglion cells (ipRGCs) mediate various non-image-forming photic responses, such as circadian photoentrainment, pupillary light reflex and pineal melatonin suppression. ipRGCs directly respond to environmental light by activation of the photopigment melanopsin followed by the opening of an unidentified cation-selective channel. Studies in heterologous expression systems and in the native retina have strongly implicated diacylglycerol-sensitive transient receptor potential channels containing TRPC3, TRPC6 and TRPC7 subunits in melanopsin-evoked depolarization. Here we show that melanopsin-evoked electrical responses largely persist in ipRGCs recorded from early postnatal (P6-P8) and adult (P22-P50) mice lacking expression of functional TRPC3, TRPC6 or TRPC7 subunits. Multielectrode array (MEA) recordings performed at P6-P8 stages under conditions that prevent influences from rod/cone photoreceptors show comparable light sensitivity for the melanopsin-evoked responses in these mutant mouse lines in comparison to wild-type (WT) mice. Patch-clamp recordings from adult mouse ipRGCs lacking TRPC3 or TRPC7 subunits show intrinsic light-evoked responses equivalent to those recorded in WT mice. Persistence of intrinsic light-evoked responses was also noted in ipRGCs lacking TRPC6 subunits, although with significantly smaller magnitudes. These results demonstrate that the melanopsin-evoked depolarization in ipRGCs is not mediated by either TRPC3, TRPC6 or TRPC7 channel subunits alone. They also suggest that the melanopsin signaling pathway includes TRPC6-containing heteromeric channels in mature retinas.

Age-related changes in the transmission properties of the human lens and their relevance to circadian entrainment

To characterize age-related changes in the transmission of light through noncataractous human lenses. Department of Ophthalmology, Glostrup Hospital, Glostrup, Denmark. The spectral transmission of white light was measured along the visual axis in the most central part of the lens in vitro in intact human donor lenses over a wide range of ages. The study evaluated 28 intact human donor lenses of 15 donors aged 18 to 76 years. Increasing age was associated with gradually decreasing transmission at all visible wavelengths, most prominently at shorter wavelengths. Empirical formulas describing the age-related loss of transmission were created for each spectral color. At 480 nm, the absorption peak for melanopsin, transmission decreased by 72% from the age of 10 years to the age of 80 years. The age-related decrease in spectral transmission through the human lens could be modeled by a simple algorithm that may be useful in the design of intraocular lenses that mimic the characteristics of the human lens and in studies of color vision, psychophysics, and melanopsin activation.

Pupillary correlates of light-evoked melanopsin activity in humans

We investigated whether cones are the only photosensitive process mediating the photopic pupillary light reflex. New analyses were performed on previously published recordings, focusing on those evoked by the onset of photopically equated short- and long-wavelength stimuli. Comparisons between responses revealed contraction differences that slowly grew to a peak and gradually declined. The late contraction was associated with short wavelengths and appeared mostly at the higher stimulus intensities. We conclude that cones are not the only photoreception process mediating the photopic ON-reflex and infer that melanopsin is another. Melanopsin contributes to the steady-state pupil size in daylight illumination.

Melanopsin Triggers the Release of Internal Calcium Stores in Response to Light†

Melanopsin is the photopigment that confers photosensitivity upon intrinsically photosensitive retinal ganglion cells (ipRGCs). This subset of retinal ganglion cells comprises less than 2% of all RGCs in the mammalian retina. The paucity of melanopsin-positive cells has made studies on melanopsin signaling difficult to pursue in ipRGCs. To address this issue, we have established several cell lines consisting of a transformed human embryonic kidney cell line (HEK293) stably expressing human melanopsin. With these cell lines, we have investigated the intracellular rise in calcium triggered upon light activation of melanopsin. Our human melanopsin-expressing cells exhibit an irradiance-dependent increase in intracellular calcium. Control cells expressing human melanopsin, where the Schiff-base lysine has been mutated to alanine, show no responses to light. Chelating extracellular calcium has no effect on the light-induced increase in intracellular calcium suggesting that calcium is mobilized from intracellular stores. This involvement of intracellular stores has been confirmed through their depletion by thapsigargin, which inhibits a subsequent light-induced increase in intracellular calcium. Addition of the nonselective cation channel blocker lanthanum does not alter light-induced rises in intracellular calcium, further supporting that melanopsin triggers a release of internal calcium from internal stores. HEK293 cells stably expressing melanopsin have proven to be a useful tool to study melanopsin-initiated signaling.

Chromophore regeneration: Melanopsin does its own thing

Among the most surprising findings in visual science of the last decade has been that photoreception in the mammalian retina is not restricted to the rods and cones, but extends to a small number of cells in the inner retina. These new photoreceptors, which comprise a small subset of retinal ganglion cells (1), are capable of driving a number of “non-image-forming” light responses including circadian photoentrainment and the regulation of pupil size even in the absence of functional rods and cones (2, 3). The so-called intrinsically photosensitive retinal ganglion cells (ipRGCs) absorb light through an opsin/retinaldehyde-based photopigment called melanopsin (4–8). Although the details of melanopsin’s photochemistry in mammals remain only partially defined, the evidence to date supports the hypothesis that, just like other opsin photopigments, the critical first event in melanopsin activation is photoisomerization of the retinaldehyde chromophore from a cis to an all-trans conformation (6–8). An important implication of such a mechanism is that to attain photosensitivity melanopsin requires a steady supply of cis-retinaldehyde. The primary source of chromophore in the vertebrate eye is a multistep enzymatic pathway, known as the retinoid or visual cycle, by which 11-cis retinaldehyde is regenerated from bleached all-trans originating in photoreceptor outer segments (Fig. 1 A). Critical elements of this pathway occur in the retinal pigment epithelium (RPE). As melanopsin is found in the more superficial layers of the retina, distant from the RPE, it would seem poorly placed to obtain cis-retinaldehyde from this source. This raises the question of how melanopsin recovers from bleach. An attractive hypothesis is that it uses an independent, more local, source of cis-retinaldehdye rather than relying on the RPE-based visual cycle. Two articles in this issue of PNAS (9, 10) test this hypothesis …