Light Affects Mood and Learning through Distinct Retina-Brain Pathways.

Light exerts a range of powerful biological effects beyond image vision, including mood and learning regulation. While the source of photic information affecting mood and cognitive functions is well established, viz. intrinsically photosensitive retinal ganglion cells (ipRGCs), the central mediators are unknown. Here, we reveal that the direct effects of light on learning and mood utilize distinct ipRGC output streams. ipRGCs that project to the suprachiasmatic nucleus (SCN) mediate the effects of light on learning, independently of the SCN's pacemaker function. Mood regulation by light, on the other hand, requires an SCN-independent pathway linking ipRGCs to a previously unrecognized thalamic region, termed perihabenular nucleus (PHb). The PHb is integrated in a distinctive circuitry with mood-regulating centers and is both necessary and sufficient for driving the effects of light on affective behavior. Together, these results provide new insights into the neural basis required for light to influence mood and learning.

Luxotonic signals in human prefrontal cortex as a possible substrate for effects of light on mood and cognition.

Studies with experimental animals have revealed a mood-regulating neural pathway linking intrinsically photosensitive retinal ganglion cells (ipRGCs) and the prefrontal cortex (PFC), involved in the pathophysiology of mood disorders. Since humans also have light-intensity-encoding ipRGCs, we asked whether a similar pathway exists in humans. Here, functional MRI was used to identify PFC regions and other areas exhibiting light-intensity-dependent signals. We report 26 human brain regions having activation that either monotonically decreases or monotonically increases with light intensity. Luxotonic-related activation occurred across the cerebral cortex, in diverse subcortical structures, and in the cerebellum, encompassing regions with functions related to visual image formation, motor control, cognition, and emotion. Light suppressed PFC activation, which monotonically decreased with increasing light intensity. The sustained time course of light-evoked PFC responses and their susceptibility to prior light exposure resembled those of ipRGCs. These findings offer a functional link between light exposure and PFC-mediated cognitive and affective phenomena.

A retinal code for motion along the gravitational and body axes.

Self-motion triggers complementary visual and vestibular reflexes supporting image-stabilization and balance. Translation through space produces one global pattern of retinal image motion (optic flow), rotation another. We examined the direction preferences of direction-sensitive ganglion cells (DSGCs) in flattened mouse retinas in vitro. Here we show that for each subtype of DSGC, direction preference varies topographically so as to align with specific translatory optic flow fields, creating a neural ensemble tuned for a specific direction of motion through space. Four cardinal translatory directions are represented, aligned with two axes of high adaptive relevance: the body and gravitational axes. One subtype maximizes its output when the mouse advances, others when it retreats, rises or falls. Two classes of DSGCs, namely, ON-DSGCs and ON-OFF-DSGCs, share the same spatial geometry but weight the four channels differently. Each subtype ensemble is also tuned for rotation. The relative activation of DSGC channels uniquely encodes every translation and rotation. Although retinal and vestibular systems both encode translatory and rotatory self-motion, their coordinate systems differ.

A high-density narrow-field inhibitory retinal interneuron with direct coupling to Müller glia.

Amacrine cells are interneurons composing the most diverse cell class in the mammalian retina. They help encode visual features such as edges or directed motion by mediating excitatory and inhibitory interactions between input (i.e. bipolar) and output (i.e. ganglion) neurons in the inner plexiform layer (IPL). Like other brain regions, the retina also contains glial cells that contribute to neurotransmitter uptake, metabolic regulation and neurovascular control. Here, we report that in mouse retina (of either sex), an abundant, though previously unstudied inhibitory amacrine cell is coupled directly to Müller glia. Electron microscopic reconstructions of this amacrine type revealed chemical synapses with known retinal cell types and extensive associations with Müller glia, the processes of which often completely ensheathe the neurites of this amacrine cell. Microinjecting small tracer molecules into the somas of these amacrine cells led to selective labelling of nearby Müller glia, leading us to suggest the name "Müller glia-coupled amacrine cell," or MAC. Our data also indicate that MACs release glycine at conventional chemical synapses, and viral retrograde transsynaptic tracing from the dorsal lateral geniculate nucleus (dLGN) showed selective connections between MACs and a subpopulation of RGC types. Visually-evoked responses revealed a strong preference for light increments; these "ON" responses were primarily mediated by excitatory chemical synaptic input and direct electrical coupling with other cells. This initial characterization of the MAC provides the first evidence for neuron-glia coupling in the mammalian retina and identifies the MAC as a potential link between inhibitory processing and glial function.Significance Statement:Gap junctions between pairs of neurons or glial cells are commonly found throughout the nervous system and play multiple roles, including electrical coupling and metabolic exchange. In contrast, gap junctions between neurons and glia cells have rarely been reported and are poorly understood. Here we report the first evidence for neuron-glia coupling in the mammalian retina, specifically between an abundant (but previously unstudied) inhibitory interneuron and Müller glia. Moreover, viral tracing, optogenetics and serial electron microscopy provide new information about the neuron's synaptic partners and physiological responses.

GABA release selectively regulates synapse development at distinct inputs on direction-selective retinal ganglion cells.

Synaptic inhibition controls a neuron's output via functionally distinct inputs at two subcellular compartments, the cell body and the dendrites. It is unclear whether the assembly of these distinct inhibitory inputs can be regulated independently by neurotransmission. In the mammalian retina, γ-aminobutyric acid (GABA) release from starburst amacrine cells (SACs) onto the dendrites of on-off direction-selective ganglion cells (ooDSGCs) is essential for directionally selective responses. We found that ooDSGCs also receive GABAergic input on their somata from other amacrine cells (ACs), including ACs containing the vasoactive intestinal peptide (VIP). When net GABAergic transmission is reduced, somatic, but not dendritic, GABAA receptor clusters on the ooDSGC increased in number and size. Correlative fluorescence imaging and serial electron microscopy revealed that these enlarged somatic receptor clusters are localized to synapses. By contrast, selectively blocking vesicular GABA release from either SACs or VIP ACs did not alter dendritic or somatic receptor distributions on the ooDSGCs, showing that neither SAC nor VIP AC GABA release alone is required for the development of inhibitory synapses in ooDSGCs. Furthermore, a reduction in net GABAergic transmission, but not a selective reduction from SACs, increased excitatory drive onto ooDSGCs. This increased excitation may drive a homeostatic increase in ooDSGC somatic GABAA receptors. Differential regulation of GABAA receptors on the ooDSGC's soma and dendrites could facilitate homeostatic control of the ooDSGC's output while enabling the assembly of the GABAergic connectivity underlying direction selectivity to be indifferent to altered transmission.

Genetic dissection of retinal inputs to brainstem nuclei controlling image stabilization.

When the head rotates, the image of the visual world slips across the retina. A dedicated set of retinal ganglion cells (RGCs) and brainstem visual nuclei termed the "accessory optic system" (AOS) generate slip-compensating eye movements that stabilize visual images on the retina and improve visual performance. Which types of RGCs project to each of the various AOS nuclei remain unresolved. Here we report a new transgenic mouse line, Hoxd10-GFP, in which the RGCs projecting to all the AOS nuclei are fluorescently labeled. Electrophysiological recordings of Hoxd10-GFP RGCs revealed that they include all three subtypes of On direction-selective RGCs (On-DSGCs), responding to upward, downward, or forward motion. Hoxd10-GFP RGCs also include one subtype of On-Off DSGCs tuned for forward motion. Retrograde circuit mapping with modified rabies viruses revealed that the On-DSGCs project to the brainstem centers involved in both horizontal and vertical retinal slip compensation. In contrast, the On-Off DSGCs labeled in Hoxd10-GFP mice projected to AOS nuclei controlling horizontal but not vertical image stabilization. Moreover, the forward tuned On-Off DSGCs appear physiologically and molecularly distinct from all previously genetically identified On-Off DSGCs. These data begin to clarify the cell types and circuits underlying image stabilization during self-motion, and they support an unexpected diversity of DSGC subtypes.

Melanopsin cells are the principal conduits for rod-cone input to non-image-forming vision.

Rod and cone photoreceptors detect light and relay this information through a multisynaptic pathway to the brain by means of retinal ganglion cells (RGCs). These retinal outputs support not only pattern vision but also non-image-forming (NIF) functions, which include circadian photoentrainment and pupillary light reflex (PLR). In mammals, NIF functions are mediated by rods, cones and the melanopsin-containing intrinsically photosensitive retinal ganglion cells (ipRGCs). Rod-cone photoreceptors and ipRGCs are complementary in signalling light intensity for NIF functions. The ipRGCs, in addition to being directly photosensitive, also receive synaptic input from rod-cone networks. To determine how the ipRGCs relay rod-cone light information for both image-forming and non-image-forming functions, we genetically ablated ipRGCs in mice. Here we show that animals lacking ipRGCs retain pattern vision but have deficits in both PLR and circadian photoentrainment that are more extensive than those observed in melanopsin knockouts. The defects in PLR and photoentrainment resemble those observed in animals that lack phototransduction in all three photoreceptor classes. These results indicate that light signals for irradiance detection are dissociated from pattern vision at the retinal ganglion cell level, and animals that cannot detect light for NIF functions are still capable of image formation.

Efficacy and specificity of melanopsin reporters for retinal ganglion cells.

Intrinsically photosensitive retinal ganglion cells (ipRGCs) are specialized retinal output neurons that mediate behavioral, neuroendocrine, and developmental responses to environmental light. There are diverse molecular strategies for marking ipRGCs, especially in mice, making them among the best characterized retinal ganglion cells (RGCs). With the development of more sensitive reporters, new subtypes of ipRGCs have emerged. We therefore tested high-sensitivity reporter systems to see whether we could reveal yet more. Substantial confusion remains about which of the available methods, if any, label all and only ipRGCs. Here, we compared many different methods for labeling of ipRGCs, including anti-melanopsin immunofluorescence, Opn4-GFP BAC transgenic mice, and Opn4cre mice crossed with three different Cre-specific reporters (Z/EG, Ai9, and Ai14) or injected with Cre-dependent (DIO) AAV2. We show that Opn4cre mice, when crossed with sensitive Cre-reporter mice, label numerous ganglion cell types that lack intrinsic photosensitivity. Though other methods label ipRGCs specifically, they do not label the entire population of ipRGCs. We conclude that no existing method labels all and only ipRGCs. We assess the appropriateness of each reporter for particular applications and integrate findings across reporters to estimate that the overall abundance of ipRGCs among mouse RGCs may approach 11%.

The retina's neurovascular unit: Müller glial sheaths and neuronal contacts.

The neurovascular unit (NVU), comprising vascular, glial and neural elements, supports the energetic demands of neural computation, but this aspect of the retina's trilaminar vessel network is poorly understood. Only the innermost vessel layer - the superficial vascular plexus (SVP) - is ensheathed by astrocytes, like brain capillaries, whereas glial ensheathment in other layers derives from radial Müller glia. Using serial electron microscopy reconstructions from mouse and primate retina, we find that Müller processes cover capillaries in a tessellating pattern, mirroring the tiled astrocytic endfeet wrapping brain capillaries. However, gaps in the Müller sheath, found mainly in the intermediate vascular plexus (IVP), permit different neuron types to contact pericytes and the endothelial cells directly. Pericyte somata are a favored target, often at spine-like structures with a reduced or absent vascular basement lamina. Focal application of adenosine triphosphate (ATP) to the vitreal surface evoked Ca2+ signals in Müller sheaths in all three vascular layers. Pharmacological experiments confirmed that Müller sheaths express purinergic receptors that, when activated, trigger intracellular Ca2+ signals that are amplified by IP3-controlled intracellular Ca2+ stores. When rod photoreceptors die in a mouse model of retinitis pigmentosa (rd10), Müller sheaths dissociate from the deep vascular plexus (DVP) but are largely unchanged within the IVP or SVP. Thus, Müller glia interact with retinal vessels in a laminar, compartmentalized manner: glial sheathes are virtually complete in the SVP but fenestrated in the IVP, permitting direct neural-to-vascular contacts. In the DVP, the glial sheath is only modestly fenestrated and is vulnerable to photoreceptor degeneration.

Form and function of the M4 cell, an intrinsically photosensitive retinal ganglion cell type contributing to geniculocortical vision.

The photopigment melanopsin confers photosensitivity upon a minority of retinal output neurons. These intrinsically photosensitive retinal ganglion cells (ipRGCs) are more diverse than once believed, comprising five morphologically distinct types, M1 through M5. Here, in mouse retina, we provide the first in-depth characterization of M4 cells, including their structure, function, and central projections. M4 cells apparently correspond to ON α cells of earlier reports, and are easily distinguished from other ipRGCs by their very large somata. Their dendritic arbors are more radiate and highly branched than those of M1, M2, or M3 cells. The melanopsin-based intrinsic photocurrents of M4 cells are smaller than those of M1 and M2 cells, presumably because melanopsin is more weakly expressed; we can detect it immunohistochemically only with strong amplification. Like M2 cells, M4 cells exhibit robust, sustained, synaptically driven ON responses and dendritic stratification in the ON sublamina of the inner plexiform layer. However, their stratification patterns are subtly different, with M4 dendrites positioned just distal to those of M2 cells and just proximal to the ON cholinergic band. M4 receptive fields are large, with an ON center, antagonistic OFF surround and nonlinear spatial summation. Their synaptically driven photoresponses lack direction selectivity and show higher ultraviolet sensitivity in the ventral retina than in the dorsal retina, echoing the topographic gradient in S- and M-cone opsin expression. M4 cells are readily labeled by retrograde transport from the dorsal lateral geniculate nucleus and thus likely contribute to the pattern vision that persists in mice lacking functional rods and cones.