Melanopsin Ganglion Cells: A Bit of Fly in the Mammalian Eye.
Dustin Graham
[Introduction] [History and discovery] [Melanopsin] [ipRGC form and function] [Phototransduction: a bit of fly in the mammalian eye] [Synaptic connectivity with classic rod/cone photoreceptors] [Central projections] [Behavioral aspects of ipRGC function] [Adaptation] [Intra-retinal signaling] [Conclusions and questions] [References] [Author]
1. Introduction.
For the greater part of 150 years it was assumed that the mammalian retina contained only two types of photoreceptors; rods and cones. However, a flurry of recent evidence has demonstrated the existence of a third type of mammalian photoreceptor that differs greatly from rods and cones. This type utilizes a different photopigment, is much less sensitive to light, and has far less spatial resolution; characteristics that fit perfectly with this photoreceptor's primary function of signaling changes in ambient light levels to the brain throughout the day. Most surprisingly, these photoreceptors are ganglion cells, and thus, have the unique ability to communicate directly with the brain. These intrinsically photosensitive retinal ganglion cells (ipRGCs) are a rare sub-population of ganglion cells (1-3%) whose primary role is to signal light for unconscious visual reflexes, such as pupillary constriction, and regulating a number of daily behavioral and physiological rhythms, collectively called circadian rhythms. This latter process, which adjusts circadian rhythms to the light/dark cycle of an animal's environment, is known as photoentrainment. The visual behaviors under ipRGC control are remarkably tonic, and require long integration times of ambient light levels. The unique properties of ipRGCs, both functionally and anatomically, make them well suited for regulating such behaviors.
2. History and discovery.
As a graduate student in 1923, Clyde Keeler made a number of interesting observations from mice that had severe outer retinal degeneration (Keeler 1928; Van Gelder 2008) . Though they lacked most of their rod and cone photoreceptors, and were considered functionally blind, these mice were still able to generate a number of visual reflexes, including constriction of their pupils in response to light (Keeler 1928). From these observations he deduced that some other photosensitive cell type must be lurking in the retina. Later, using genetically engineered mice with no rods or cones, Foster and colleagues showed that these blind mice also maintained the ability to shift their daily biological rhythms in accordance with shifting light cycles, and suppress pineal activity in response to brief pulses of light (Freedman, Lucas et al. 1999; Lucas, Freedman et al. 1999). These processes disappear, however, when animals have their eyes removed, strongly supporting Keeler's prediction of a novel occular photoreceptor (Klein and Weller 1972). However, it wasn't until the discovery of a new photopigment (or opsin), from an unlikely source, that Keeler's prediction would prove true.
Provencio and colleagues, studying the dermal melanophores of frogs, cloned a novel opsin molecule which they found was responsible for redistribution of skin pigment in direct response to light (Provencio, Jiang et al. 1998). Othologs of this opsin, which they called melanopsin, was also found to be selectively expressed in a small subset of ganglion cells in mouse retina (Fig. 1a) (Provencio, Jiang et al. 1998; Provencio, Rodriguez et al. 2000), and additional work found that these cells send their axons to the suprachiasmatic nucleus (SCN), the site of the mammalian biological clock controlling circadian rhythms (Gooley, Lu et al. 2001; Hannibal, Hindersson et al. 2002). All these findings heavily suggested that these ganglion cells were the mysterious third photoreceptor type Keeler had predicted almost 80 years earlier. They express a putative opsin, potentially allowing them to directly respond to light, and they send axon projections to brain targets known to be involved in visual reflexes that rodless/coneless mice were still capable of performing. However, definitive proof that these melanopsin containing ganglion cells could signal light without the function of rods and cones was still lacking. Combining retrograde tracer injections into the SCN with patch clamp electrophysiology, David Berson and colleagues directly tested the hypothesis that melanopsin ganglion cells are intrinsically photosensitive. They performed targeted patch-clamp recordings and found that the retrolabeled ganglion cells (which stained positive for melanopsin using immuno-hisotchemical techniques) were indeed able to respond to light in the presence of a cocktail of pharmacological blockers that eliminate virtually all rod and cone signaling in the retina (Fig. 1b-c) (Berson, Dunn et al. 2002; Hattar, Liao et al. 2002). Furthermore, even after mechanical isolation, these cells were still intrinsically photosensitive, leaving to rest all doubts that the labeled ganglion cells were true photoreceptors (Berson, Dunn et al. 2002).
3. Melanopsin.
The unique ability of ipRGCs to respond to light is due to their exclusive expression of the photopigment melanopsin. Originally cloned from frog dermal melanophores, the melanopsin gene (OPN4) has orthologs in many mammalian species, including mice, monkeys, and humans (Provencio, Rodriguez et al. 2000). Hydrophobicity analysis of melanospin's amino-acid sequence predicts a 7-transmembrane structure, common to all G-protein coupled receptors (Fig. 2a, 2c) (Provencio, Jiang et al. 1998). Interestingly, melanopsin shares more homology with invertebrate rhabdomeric opsins (r-opsins) than with ciliary opsins of vertebrate species (c-opsins), suggesting that melanopsin may signal light through a different mechanism than that used in vertebrate rods and cones (Fig. 2b) (Provencio, Jiang et al. 1998)
Although the first studies of ipRGCs strongly suggested melanopsin as the photopigment in these cells, a role for a group of blue-light absorbing flavoproteins known as cryptochromes could not be initially ruled out (Berson 2007). Cryptochromes function as circadian photopigments in invertebrates, and early studies favored their functioning as the mammalian circadian photopigments (Kavakli and Sancar 2002; Van Gelder, Gibler et al. 2002). However, there is now overwhelming evidence that melanopsin is the photopigment in ipRGCs, and thus the true circadian photopigment . When the melanopsin gene is deleted via transgenic techniques in mice (knockout mice), retinal ganglion cells labeled from the SCN no longer can signal light (Fig. 3) (Lucas, Hattar et al. 2003). In addition, animals missing the melanopsin gene show deficiencies in multiple visual reflexes such as pupillary constriction and photoentrainment (Panda, Sato et al. 2002; Ruby, Brennan et al. 2002; Lucas, Hattar et al. 2003; Panda, Provencio et al. 2003). As a further test, Tu et. al used a multi-electrode array, which is a planar array of electrodes that allows one to record extracellularly from dozens of retinal ganglion cells at once, to isolate light responses from multiple ipRGCs. Retinas from melanopsin knockout mice showed no intrinsic photoresponses from ganglion cells (Tu, Zhang et al. 2005). Despite this evidence, there were still controversies regarding melanopsin's ability to function as a true photopigment. These were firmly extinguished with a series of elegant experiments whereby the melanopsin gene was expressed in multiple cell types that are normally light insensitive. When made to express the melanopsin gene, these cells are able to robustly respond to light, indicating melanopsin's ability to function as a bonafide photopigment (Fig. 3) (Melyan, Tarttelin et al. 2005; Panda, Nayak et al. 2005; Qiu, Kumbalasiri et al. 2005).
4. ipRGC form and function.
In rodents, ipRGCs make up about 1-3% of the total ganglion cell population, and are distributed throughout the entire retina, at a somewhat higher density superiorly (Fig. 4) (Hattar, Liao et al. 2002). Their dendritic profiles are large, spanning roughly 500 ?m, and form an extensive overlapping plexus within the retinal inner plexiform layer (IPL) (Hattar, Liao et al. 2002; Provencio, Rollag et al. 2002; Berson 2003). Owing to their large dendritic spread, ipRGCs initiate a process of spatial convergence the leads to large receptive fields in target structures like the SCN (Groos and Mason 1980). The dendrites of ipRGCs terminate mainly in the outer-most sublayer of the IPL, corresponding to the main plexus of melanopsin-immunoreactive dendrites (Fig. 4) (Berson, Dunn et al. 2002; Hannibal, Hindersson et al. 2002; Hattar, Liao et al. 2002; Provencio, Rollag et al. 2002), however, in mice there appears to be a second plexus of melanopsin-positive dendrites, suggesting the presence of a possible second population of ipRGCs.
5. Phototransduction: a bit of fly in the mammalian eye.
Animal photoreceptors come in two basic flavors: rhabdomeric (found mainly in invertebrates), and ciliary (vertebrate rods and cones being most notable). A defining feature of rhabdomeric and ciliary photoreceptors is the distinct biochemical cascades both cell types use to transduce light energy into an electrical signal the brain can interpret. Rods and cones use a cascade involving cyclic guanyl monophosphate (cGMP) as a second messenger (Fig. 6) (Arendt 2003; Fu and Yau 2007). The response to light is a decrease of cGMP levels which closes cyclic nucleotide gated channels causing the plasma membrane to hyperpolarize. This is in stark contrast to invertebrate rhabdomeric photoreceptors which use a phosphoinositide signaling cascade involving the enzyme phospholipase C (PLC) and breakdown of the membrane lipid phosphotidylinositol bisphosphate (PIP2) (Fig. 6), leading to the opening of cationic channels and membrane depolarization (Hardie 2001; Hardie and Raghu 2001). In addition, although both phototransduction cascades are G-protein mediated, the specific G-proteins required by ciliary and rhabdomeric photoreceptors are different. Ciliary photoreceptors require transducin, a member of the Gi/o-family of G-proteins, whereas rhabdomeric photoreceptors use a member of the Gq/11-family of G-proteins (Fig. 6).
Melanopsin's strong homology with invertebrate opsins and the depolarizing light response of ipRGCs suggests they may use a rhabdomeric phototransduction cascade. However, early patch clamp and pharmacological studies of ipRGCs could not directly confirm this hypothesis. This was most likely due to the whole-mount retina recording configuration often used in studying ganglion cell function. The combination of photosensitive ipRGC dendrites buried deep within the IPL, and a membrane sheath covering the ganglion cell bodies, can create a significant diffusion barrier for pharmacological agents, especially hydrophobic agents commonly used to study transduction mechanisms. To overcome this hurdle, Graham et al. recorded from disscociated ipRGC cell bodies in culture to study the intracellular phototransduction cascade. Isolated ipRGCs survive remarkably well in culture, generating robust light responses for up to 6 days and allowing for excellent pharmacological manipulation. Using this system, they showed that ipRGC phototransduction follows a rhabdomeric-like phosphoinositide cascade, requiring a member (or possibly members) of the Gq/11 family of G-proteins and the effector enzyme phospholipase C (PLC) (Fig. 7) (Graham, Wong et al. 2008). In addition, the presence of specific Gq/11 and PLC isoforms was confirmed in ipRGCs using single-cell RT-PCR and immunocytochemistry, consistent with the pharmacological findings (Fig. 8) (Graham, Wong et al. 2008).
In rhabdomeric photoreceptors, breakdown of PIP2 by PLC generates two by-products; a membrane-bound component called diacylglycerol (DAG), and inositol-triphosphate (IP3), which is free to move about the cytosol (Hardie 2001). DAG can be broken down within the membrane into polyunsaturated faty acids (PUFA's), which have been shown to directly open Drosophila TRP channels (Chyb, Raghu et al. 1999). IP3, on the other hand, can cause release of intracellular calcium from stores within the cell, which can lead to the opening of so called "store-operated channels". In Drosophila photoreceptors the exact downstream mechanism linking PIP2 breakdown to TRP channel opening is still unknown, although a membrane-associated pathway is heavily favored (Acharya, Jalink et al. 1997). In ipRGCs, data strongly suggest that the cytosolic IP3 pathway is not required for phototransduction, similar to Drosophila photoreceptors (Fig. 9) (Graham, Wong et al. 2008). Intracellular application of high concentrations of IP3 neither induces a current, nor does it block light responses in ipRGCs (Fig. 9), although it does modulate the response properties, suggesting a non-essential secondary role for IP3 and intracellular calcium. Likewise, blocking IP3 receptors with intracellular heparin and intracellular calcium store depletion with thapsigargin, both fail to block phototransduction in ipRGCs (Fig. 9). Only after extended exposure to very high (10mM) concentrations of the general calcium chelator BAPTA, does one see significant blockade of phototransduction, although this is most likely a side effect of clamping resting intracellular calcium levels so low that enzymes such as PLC, are no longer able to function (Hardie 2005; Graham, Wong et al. 2008). The most compelling evidence that ipRGC phototransduction does not require the cytosolic IP3 branch is that excised patches of ipRGC membrane (outside-out and inside-out) remain autonomously photosensitive (Fig. 9), ruling out a vital role for highly diffusible cytosolic components. Based on these findings, all the necessary components for ipRGC phototransduction appear to be within, or tightly coupled to, the plasma membrane.
The exact gating mechanism downstream of PLC activation in ipRGC phototransduction is still not clear. Application of both DAG and PUFA's, either to isolated ipRGC cell bodies, or to excised patches of ipRGC membrane, fail to evoke a current or to block light responses (Fig. 10). An alternative hypothesis, and one in-line with a membrane-associated transduction cascade, is that the breakdown of PIP2 itself (as opposed to production of DAG or PUFA's) is the critical signal that opens the light gated channels in ipRGCs. There is evidence that PIP2 can either open or close a variety of ion channels, including the light-gated channels in Drosophila (Hardie 2003; Suh, Inoue et al. 2006). As a preliminary test of this hypothesis however, Graham et al. pharmacologically interfered with PIP2 synthesis using wortmannin. This drug inhibits phosphoinositide 4-kinase (PI4-K), the synthetic enzyme for phosphatidylinositol 4-phosphate (PIP), which is an essential precursor of PIP2. According to the hypothesis, wortmannin should slow the termination of the photocurrent at light offset by delaying the restoration of resting levels of PIP2 and, thus, the closure of the light gated channels. Indeed, when wortmannin was included in the pipette solution, response shutoff was dramatically delayed (Fig. 10). Further and more direct evidence, however, is still necessary to resolve the gating mechanism of the light sensitive channels in ipRGCs.
The similarity of both the opsin and the signaling cascade in ipRGCs to those in rhabdomeric photoreceptors has encouraged speculation that the light sensitive channels might belong to the TRP family, originally discovered in Drosophila photoreceptors (Montell 2005). Although definitive evidence is still lacking, most studies suggest that a member (or members) of the TRPC family of TRP channels is involved (Warren, Allen et al. 2006; Hartwick, Bramley et al. 2007). Light induces a current that reverses around 0 mV, suggesting a nonspecific cationic channel, and when melanopsin is expressed in heterologous cell systems, co-expression of TRPC channels allows the cells to transduce light (Qiu, Kumbalasiri et al. 2005). Ion substitution experiments suggest that calcium is a significant charge carrier through the light sensitive channels, consistent with findings of Sekaran et al. who used calcium imaging to measure light responses from ipRGCs in retinas lacking rods and cones (Sekaran, Foster et al. 2003; Warren, Allen et al. 2006). Likewise, drugs which are known to block TRPC channels are able to block light response in ipRGCs, although none of the drugs used are specific to TRPC channels (Warren, Allen et al. 2006; Hartwick, Bramley et al. 2007). However, in dissociated ipRGCs, application of DAG analogues (drugs known to activate TRPC channels) neither induce a current nor do they block the light response (Graham, Wong et al. 2008). A stronger case for TRPC channels awaits better pharmacological tools and genetic/molecular manipulation of ipRGCs. It seems safe to say that uncertainty about the identity of the light gated channels in ipRGCs remains the most glaring gap in our understanding of phototransduction mechanisms in these neurons.
6. Synaptic connectivity with classic rod/cone photoreceptors.
Although ipRGCs can function as photoreceptors, they also receive intraretinal synaptic input from rods and cones. Their photosensitive dendrites terminate in the inner plexiform layer (IPL), the layer of synaptic contact for amacrine and bipolar cells to convey rod and cone signals to ganglion cells (Berson 2003). Using electron microscopy, Belenky et al. identified synaptic contacts between amacrine cells and presumed ON-bipolar cells with melanopsin-immunopositive processes in the IPL (Belenky, Smeraski et al. 2003). Also, as demonstrated in both rats and primates, in addition to their intrinsic melanopsin-based light response, ipRGCs respond to light through synaptically mediated input from rods and cones (Dacey, Liao et al. 2005; Perez-Leon, Warren et al. 2006; Wong, Dunn et al. 2007). Dacey et al. showed that at light levels to low to activate cones, ipRGCs responded well, indicating rod-based input. In addition, using light of different wavelengths, they showed that ipRGCs display a color opponency in their responses, indicating that they also receive input from cone photoreceptors.
Using pharmacology and a multielectrode array (MEA) to record extracellularly from multiple ipRGCs simultaneously, one can clearly separate the synaptically driven and intrinsic light responses of ipRGCs (Fig.11) (Wong, Dunn et al. 2007). In addition, the unique response characteristics (insensitive and sluggish) of the intrinsic melanopsin-based signaling becomes quite apparent (Fig. 11)
Using whole-cell voltage clamp recordings combined with pharmacology, investigators demonstrated that ipRGCs have spontaneous inhibitory and excitatory synaptic inputs at rest in the dark , and that they express receptors for glutamate, GABA and glycine (Fig. 12) (Perez-Leon, Warren et al. 2006; Wong, Dunn et al. 2007). In response to light, ipRGCs receive a mix of synaptic input, arising from amacrine cells and both ON and OFF bipolar cells (although the ON signal is much stronger) (Wong, Dunn et al. 2007). Under more physiological conditions (recording with the MEA), the sustained extrinsic response to light appears to be driven by On-bipolar cells, as the drug L-AP4 abolishes the synaptically mediated (rod/cone) driven light response in ipRGCs (Wong, Dunn et al. 2007). Thus, amacrine cells most likely inhibit ipRGCs from firing spontaneously in the dark, while the On-bipolar cell input drives them to spike during light stimulation. This was confirmed by Wong et al. by applying agents to block input from amacrine cells. Under these conditions, ipRGCs begin to fire spontaneously in the dark. Since the dendrites of ipRGCs stratify in the Off-sublamina of the IPL, the exact details of synaptic contacts with other retinal elements driving On and Off responses is not yet known, but a schematic of presumptive ipRGC synaptic connections that can account for the known synaptic physiology is shown in Fig. 12e.
7. Central Projections.
Using a transgenic line of mice where retinal ganglion cells normally expressing melanopsin instead express the marker enzyme _-galactosidase, Hattar et. al characterized the central projections of ipRGCs throughout the brain (Hattar, Kumar et al. 2006). This method provides a very sensitive means to mark cell bodies, and more importantly, axonal projections. As expected for cells that regulate non-visual photic behaviors, such as circadian rhythms and pupillary constriction, they found ipRGCs sent dense projections to the suprachiasmatic nucleus (SCN), the intergeniculate leaflet (IGL), and the olivary pretectal nucleus (OPN) (Fig. 13-14) (Berson 2003; Hattar, Kumar et al. 2006). In addition to these expected sites, a number of other central targets are seen. Within the hypothalamus, ipRGCs send axonal fibers to the ventral portion of the subparaventricular zone (vSPZ), a region controlling the autonomic nervous system (Hattar, Kumar et al. 2006). Rostrolateral to the SCN, a number of scattered fibers reach the lateral and ventrolateral preoptic areas, which influence release of reproductive hormones from the pituitary (Hattar, Kumar et al. 2006). Thus, ipRGCs most likely play a critical role in light regulation of a diverse array of non-image forming visual functions.
In brains optimally stained for _-galactosidase activity, a thin sheet of tortuous fibers with apparent terminal swellings can be detected in the dorsal division of the lateral geniculate nucleus (LGN), the thalamic relay site of retinal input to the visual cortex (Fig.14) (Hattar, Kumar et al. 2006). In addition, the superior colliculus (SC) is a significant target of labeled axons (Hattar, Kumar et al. 2006). These findings suggest that in addition to their known role in regulating visual reflexes, ipRGCs may play an as yet unknown role in shaping conscious visual perception. Interestingly, Dennis Dacey and colleagues found that in primates, ipRGCs could be retrolabeled by injecting fluorescent tracers into the LGN, and that these "giant" melanopsin-expressing ganglion cells display a color-opponent receptive field (Dacey, Liao et al. 2005).
8. Behavioral aspects of ipRGC function.
Daily rhythms in mammalian physiology and behavior, collectively called circadian rhythms, are controlled by a tiny cluster of cells in the suprachiasmatic nuclei (SCN) of the ventral hypothalamus. SCN neurons posses a unique molecular clock that allows them to autonomously regulate activity patterns in near 24-hour rhythms, which ultimately leads to daily changes in animal behaviors such as sleep/wake cycles. However, the clocks are not tuned perfectly to 24 hours, and in order to adjust to changes in the animals environmental light/dark phases, the SCN must be entrained (or reset). Light is by far the most potent entrainment cue, and ipRGCs are the primary cells that carry this daily signal. Melanopsin knockout animals have relatively normal circadian rhythms, and do not display any overt dysfunction in their ability to entrain to light (Fig. 15) (Panda, Sato et al. 2002; Ruby, Brennan et al. 2002). Measured both behaviorally and using cFos staining (a measure of activity in neurons) in the SCN, these animals show no significant difference to wild type animals (Panda, Sato et al. 2002; Ruby, Brennan et al. 2002). However, they display a significantly attenuated phase shifting response (Fig. 16) to brief flashes of light, indicating a crucial role for ipRGCs and melanopsin signaling (Panda, Sato et al. 2002; Ruby, Brennan et al. 2002). The insignificant change in photoentrainment of melanopsin knockout animals is most likely explained by the rod and cone input to ipRGCs, adding a redundant source of photic input to the SCN. This idea was tested directly by doing behavioral analysis on melanopsin knockout mice that lack rods and cones (Fig.17) (Hattar, Lucas et al. 2003; Panda, Provencio et al. 2003). In addition, Samer Hattar and colleagues developed an ingenious mouse line where genes encoding key enzymes in rod and cone signaling, as well as the melanopsin gene, were all deleted, leaving the cellular architecture of the retina completely intact, but devoid of all phototransduciton capabilities (Hattar, Lucas et al. 2003). Much like the melanopsin knockout mice without rods and cones, these "triple knockout" mice could neither phase-shift their circadian rhythms to brief pulses of light, nor could they photoentrain to light/dark cycles (Hattar, Lucas et al. 2003).
In addition to their role in regulating photoentrainment of circadian rhythms, ipRGCs are responsible for constriction of the pupil in response to light (Fig. 18) (Hattar, Lucas et al. 2003; Lucas, Hattar et al. 2003; Panda, Provencio et al. 2003). Projections of ipRGC axons to the olivary pretectal nucleus (OPN), the retino-recipient site responsible for the pupillary light reflex, make them likely candidates for regulating this response. Indeed, melanopsin knockout animals have diminished pupillary light reflexes at high irradiance levels (Fig. 18) (Lucas, Hattar et al. 2003). In addition, melanopsin knockout mice without rods and cones, as well as triple knockout animals, are completely unable to constrict their pupils in response to light (Fig. 18) (Hattar, Lucas et al. 2003; Panda, Provencio et al. 2003).
9. Adaptation.
The mammalian image-forming visual system is capable of operating over a wide spectrum of ambient light intensities covering more than ten orders of magnitude. While much of this range is due to the operation of two separate receptor systems (rods and cones) that operate over different light intensity ranges, some of the capacity is due to the ability of both rods and cones to change their sensitivity to light, a process called adaptation (Dowling 1987; Rodieck 1998). When placed in constant background light, rods and cones can generate an initial peak response which relaxes during the steady illumination, indicating desensitization to the light over time. This desensitization allows the photoreceptors to increase their dynamic range, allowing them to respond to light intensities that would otherwise be saturating. This is termed "light adaptation". Conversely, when placed back into a dark environment, rods and cones are able to regain their sensitivity, a process called "dark adaptation". Both light and dark adaptation are a direct consequence of changes in the biochemical transduction cascades operating in the receptors. While rods have a limited ability to adapt to background illumination, saturating at high light levels, cones exhibit an almost infinite capacity to adapt to light levels (Fain, Matthews et al. 2001; Knox and Solessio 2006).
Due to their role in stable representations of absolute light levels, and unusually tonic response properties, it was not clear whether ipRGCs would display similar adaptation mechanisms as rods and cones. To address this issue, Wong and colleagues performed recordings from ipRGCs under luminance conditions to test for both light and dark adaptation. They found that when exposed to a near saturating background light, ipRGCs gradually became desensitized, and responses to a brighter pulse of light on top of the background light became larger over time, a hallmark of light adaptation (Fig. 19) (Wong, Dunn et al. 2005). Furthermore, when ipRGCs were left in the dark for increasing periods of time, their responses to the same light stimulus intensity increased as well, a clear indication that in addition to adapting to light, ipRGCs are capable of dark adaptation as well Fig. 20 (Wong, Dunn et al. 2005).
10. Intra-retinal signaling.
Dopamine is a neurotransmitter known to play many roles in modulation of neural circuits, and in the retina it is released by a sub-population of amacrine cells (Dowling 1987). These dopaminergic amacrine neurons (DA neurons) play a major role in reshaping the functional properties of retinal circuits according to current lighting conditions, and thus allow the retina to be a more dynamic system. DA neurons come in multiple flavors showing transient, sustained and null responses to light (Zhang, Zhou et al. 2007). Interestingly, only the transient responding DA neurons require On-bipolar cell input, while sustained DA neurons are unaffected by drugs blocking On-bipolar cell input (Fig. 21) (Zhang, Zhou et al. 2007; Zhang, Wong et al. In press). This, along with the properties of their sustained responses, suggests that ipRGCs may be providing the input to these cells in a bizarre centrifugal mechanism (Zhang, Wong et al. In press). The sustained response to light in DA neurons peaks at a wavelength near 480nm, matching very well with a melanopsin-based input.
In addition, staining for cFos, an immediate early gene used to mark activated neurons, showed that roughly the same percentage of DA neurons are activated by sustained light in retinas without rods and cones (rd/rd animals) as retinas with On-bipolar cells pharmacologically blocked using the drug L-AP4 (Fig. 22) (Zhang, Wong et al. In press). Presumably in both situations the only remaining photosensitivity in the retina would be from ipRGCs. Therefore, the evidence suggests that ipRGCs are somehow communicating back to DA neurons through their dendrites, although no evidence of release sites or vesicles in melanopsin-positive dendrites have been seen. However, a number of studies have shown that ipRGC dendrites and DA neurons dendrites co-stratify, providing a potential structural basis for synaptic communication (Belenky, Smeraski et al. 2003; Viney, Balint et al. 2007; Vugler, Redgrave et al. 2007).
11. Conclusions and questions.
ipRGCs are a novel mammalian photoreceptor whose morphological and physiological characteristics seem well suited for their primary role as light detectors for non-image forming visual reflexes. However, many mysteries remain, and an untold number of functions for this rare and special type of ganglion cell should not be overlooked. Their invertebrate-like phototransduction cascade makes them unique among all other known vertebrate photoreceptors, and provide a window into possible mechanisms of the evolution of the retina. In addition to their intrinsic melanopsin-driven photosensitivity, ipRGCs also receive rod and cone synaptic input and thus may provide the brain with different information in series, separated by complex spatial and temporal dynamics. Although they drive a number of tonic behaviors, requiring accurate representation of ambient light levels of long periods of time, ipRGCs have the ability to adapt to both light and darkness, and appear to have an ability to communicate back to the retina, possibly changing the functional properties of retinal circuitry. The functional role these processes have on animal behavior remains to be understood. As the years catch up with this relatively young field of intrinsically photosensitive retinal ganglion cells, new and interesting layers of the onion will no doubt reveal themselves.
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The author Dr. Dustin M. Graham was born and raised in Pleasanton, California and received his Ph.D. in neuroscience from Brown University. He began his research career as an undergraduate student at Santa Clara University in the lab of Dr. David Tauck, studying neural pathways of learning and memory in the pond snail Lymnaea stagnalis. His interests turned to the retina while working with Dr. Ralph Nelson at the National Institutes of Health. There he helped develop a rapid labeling technique to delineate morphological subtypes of retinal neurons in zebrafish retinal slices. As a graduate student Dustin studied mammalian circadian rhythms and the newly discovered melanopsin ganglion cell activity. He joined Dr. David Berson's lab and investigated the role of intrinsically photosensitive retinal ganglion cells (ipRGCs) in generating responses to light in the mammalian biological clock located in the suprachiasmatic nucleus. He focussed on the phototransduction cascade in ipRGCs, and for his thesis work he developed a dissociation and culturing procedure to identify and record light responses from isolated ipRGCs. Dustin is currently a post-doctoral research fellow in the psychology department at the University of Virginia where he studies development and synaptic mechanisms of the gustatory system in rats. |
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[Introduction] [History and discovery] [Melanopsin] [ipRGC form and function] [Phototransduction: a bit of fly in the mammalian eye] [Synaptic connectivity with classic rod/cone photoreceptors] [Central projections] [Behavioral aspects of ipRGC function] [Adaptation] [Intra-retinal signaling] [Conclusions and questions] [References]
August 2008
