Reconfiguration of the visual code and retinal cell type complement in closely related diurnal and nocturnal mice.

How does evolution act on neuronal populations to match computational characteristics to functional demands? We address this problem by comparing visual code and retinal cell composition in closely related murid species with different behaviours. Rhabdomys pumilio are diurnal and have substantially thicker inner retina and larger visual thalamus than nocturnal Mus musculus . High-density electrophysiological recordings of visual response features in the dorsal lateral geniculate nucleus (dLGN) reveals that Rhabdomys attains higher spatiotemporal acuity both by denser coverage of the visual scene and a selective expansion of elements of the code characterised by non-linear spatiotemporal summation. Comparative analysis of single cell transcriptomic cell atlases reveals that realignment of the visual code is associated with increased relative abundance of bipolar and ganglion cell types supporting OFF and ON-OFF responses. These findings demonstrate how changes in retinal cell complement can reconfigure the coding of visual information to match changes in visual needs.

The mouse suprachiasmatic nucleus encodes irradiance via a diverse population of neurons monotonically tuned to different ranges of intensity.

Many neurons of the mammalian master circadian oscillator in the suprachiasmatic nuclei (SCN) respond to light pulses with irradiance-dependent changes in firing. Here, we set out to better understand this irradiance coding ability by considering how the SCN tracks more continuous changes in irradiance at both population and single unit level. To this end, we recorded extracellular activity in the SCN of anaesthetised mice presented with up + down irradiance staircase stimuli covering moonlight to daylight conditions and incorporating epochs with steady light or superimposed higher frequency modulations (temporal white noise (WN) and frequency/contrast chirps). Single unit activity was extracted by spike sorting. The population response of SCN units to this stimulus was a progressive increase in firing rate at higher irradiances. This relationship was symmetrical for up vs. down phases of the ramp in the presence of white noise or chirps but exhibited hysteresis for steady light, with firing systematically higher during increasing irradiance. Single units also showed a monotonic relationship between firing and irradiance but exhibited diversity not only in response polarity (increases vs. decreases in firing), but also in the sensitivity (EC50 ) and slope of fitted functions. These data show that individual SCN neurons exhibit monotonic relationships between irradiance and firing rate but differ in the irradiance range over which they respond. This property may help the SCN to encode the large differences in irradiance found in nature using neurons with a constrained range of firing rates. KEY POINTS: Daily changes in environmental light (irradiance) entrain the suprachiasmatic nucleus (SCN) circadian clock. The mouse SCN shows graded increases in neurophysiological activity with light pulses of increasing irradiance. We show that this monotonic relationship between firing rate and irradiance is retained at population and single unit level when probed with more naturalistic staircase increases and decreases in irradiance. The irradiance response is more reliable in the presence of ongoing higher temporal frequency modulations in light intensity than under steady light. Single units varied in sensitivity allowing the population to cover a wide range of irradiances. Irradiance coding in the SCN has characteristics of a sparse code with individual neurons tracking different portions of the natural irradiance range. This property may address the challenge of encoding a 109 -fold day:night difference in irradiance within the constrained range of firing rates available to individual neurons.

Three-dimensional unsupervised probabilistic pose reconstruction (3D-UPPER) for freely moving animals.

A key step in understanding animal behaviour relies in the ability to quantify poses and movements. Methods to track body landmarks in 2D have made great progress over the last few years but accurate 3D reconstruction of freely moving animals still represents a challenge. To address this challenge here we develop the 3D-UPPER algorithm, which is fully automated, requires no a priori knowledge of the properties of the body and can also be applied to 2D data. We find that 3D-UPPER reduces by [Formula: see text] fold the error in 3D reconstruction of mouse body during freely moving behaviour compared with the traditional triangulation of 2D data. To achieve that, 3D-UPPER performs an unsupervised estimation of a Statistical Shape Model (SSM) and uses this model to constrain the viable 3D coordinates. We show, by using simulated data, that our SSM estimator is robust even in datasets containing up to 50% of poses with outliers and/or missing data. In simulated and real data SSM estimation converges rapidly, capturing behaviourally relevant changes in body shape associated with exploratory behaviours (e.g. with rearing and changes in body orientation). Altogether 3D-UPPER represents a simple tool to minimise errors in 3D reconstruction while capturing meaningful behavioural parameters.

Look-up and look-down neurons in the mouse visual thalamus during freely moving exploration.

Visual information reaches cortex via the thalamic dorsal lateral geniculate nucleus (dLGN). dLGN activity is modulated by global sleep/wake states and arousal, indicating that it is not simply a passive relay station. However, its potential for more specific visuomotor integration is largely unexplored. We addressed this question by developing robust 3D video reconstruction of mouse head and body during spontaneous exploration paired with simultaneous neuronal recordings from dLGN. Unbiased evaluation of a wide range of postures and movements revealed a widespread coupling between neuronal activity and few behavioral parameters. In particular, postures associated with the animal looking up/down correlated with activity in >50% neurons, and the extent of this effect was comparable with that induced by full-body movements (typically locomotion). By contrast, thalamic activity was minimally correlated with other postures or movements (e.g., left/right head and body torsions). Importantly, up/down postures and full-body movements were largely independent and jointly coupled to neuronal activity. Thus, although most units were excited during full-body movements, some expressed highest firing when the animal was looking up ("look-up" neurons), whereas others expressed highest firing when the animal was looking down ("look-down" neurons). These results were observed in the dark, thus representing a genuine behavioral modulation, and were amplified in a lit arena. Our results demonstrate that the primary visual thalamus, beyond global modulations by sleep/awake states, is potentially involved in specific visuomotor integration and reveal two distinct couplings between up/down postures and neuronal activity.

Short Wavelengths Contribution to Light-induced Responses and Irradiance Coding in the Rat Dorsal Lateral Geniculate Nucleus - An In vivo Electrophysiological Approach.

The dorsal lateral geniculate nucleus (dLGN) is the main neuronal station en route to higher visual areas. It receives information about environmental light from retinal photoreceptors whose sensitivity peaks are distributed across a visible spectrum. Here, using electrophysiological multichannel recordings in vivo combined with different light stimulations, we investigated short wavelength contribution to the dLGN responses to light and irradiance coding. The results showed that the majority of dLGN cells responded evenly to almost all wavelengths from the 340 to 490 nm spectrum; however, some cells representing extremes of unimodal distribution of Blue-UV index were specialised in the reception of blue or UV light. Moreover, by using alternate yellow and monochromatic light stimuli from blue - UV range, we also assessed the relative spectral contribution to rat dLGN responses to light. Finally, we observed no clear changes in the irradiance coding property of short wavelength-deficient light stimuli, however we noticed a distortion of the coding curves manifested by a significant drop in measure of fit after using short wavelength blocking filter. In conclusion, our data provide the first electrophysiological report on dLGN short wavelength-induced responses under changing light conditions and suggest the presence of colour opponent cells in the rat dLGN.

Using a bistable animal opsin for switchable and scalable optogenetic inhibition of neurons.

There is no consensus on the best inhibitory optogenetic tool. Since Gi/o signalling is a native mechanism of neuronal inhibition, we asked whether Lamprey Parapinopsin ("Lamplight"), a Gi/o-coupled bistable animal opsin, could be used for optogenetic silencing. We show that short (405 nm) and long (525 nm) wavelength pulses repeatedly switch Lamplight between stable signalling active and inactive states, respectively, and that combining these wavelengths can be used to achieve intermediate levels of activity. These properties can be applied to produce switchable neuronal hyperpolarisation and suppression of spontaneous spike firing in the mouse hypothalamic suprachiasmatic nucleus. Expressing Lamplight in (predominantly) ON bipolar cells can photosensitise retinas following advanced photoreceptor degeneration, with 405 and 525 nm stimuli producing responses of opposite sign in the output neurons of the retina. We conclude that bistable animal opsins can co-opt endogenous signalling mechanisms to allow optogenetic inhibition that is scalable, sustained and reversible.

Infra-slow modulation of fast beta/gamma oscillations in the mouse visual system.

KEY POINTS: Neurophysiological activity in the subcortical visual system fluctuates in both infra-slow and fast oscillatory ranges, but the level of co-occurrence and potential functional interaction of these rhythms is unknown. Analysing dark-adapted spontaneous activity in the mouse subcortical visual system, we find that these two types of oscillation interact uniquely through a population of neurons expressing both rhythms. Genetic ablation of rod/cone signalling potentiates infra-slow and abolishes fast beta/gamma oscillations while genetic ablation of melanopsin substantially diminishes the interaction between these two rhythms. Our results indicate that in an intact visual system the phase of infra-slow modulates fast beta/gamma oscillations. Thus one possible impact of infra-slow oscillations in vision is to guide visual processing by interacting with fast narrowband oscillations. ABSTRACT: Infra-slow (<0.02 Hz) and fast beta/gamma (20-100 Hz) oscillations in neurophysiological activity have been widely found in the subcortical visual system. While it is well established that fast beta/gamma oscillations are involved in visual processing, the role (if any) of infra-slow oscillations is currently unknown. One possibility is that infra-slow oscillations exert influence by modulating the amplitude of fast oscillations, yet the extent to which these different oscillations arise independently and interact remains unknown. We addressed these questions by recording in vivo spontaneous activity from the subcortical visual system of visually intact mice, and animals whose retinal network was disrupted by advanced rod/cone degeneration (rd/rd cl) or melanopsin loss (Opn4-/- ). We found many neurons expressing only one type of oscillation, and indeed fast oscillations were absent in rd/rd cl. Conversely, neurons co-expressing the two oscillations were also common, and were encountered more often than expected by chance in visually intact but not Opn4-/- mice. Finally, where they co-occurred we found that beta/gamma amplitude was modulated by the infra-slow rhythm. Our data thus reveal that: (1) infra-slow and beta-gamma oscillations are separable phenomena; and (2) that they actively co-occur in a subset of neurones in which the phase of infra-slow oscillations defines beta-gamma oscillations amplitude. These findings suggest that infra-slow oscillations could influence vision by modulating beta-gamma oscillations, and raise the possibility that disruptions in these oscillatory behaviours contribute to vision dysfunction in retinal dystrophy.

Melanopsin: From a small molecule to brain functions.

Melanopsin, a G family coupled receptor, found within the ganglion cell layer in the retina, plays an important role in non-image-forming visual functions, including hormone secretion, entrainment of circadian rhythms, cognitive and affective processes. Diffuse projections of melanopsin-containing cells to many brain areas suggest that different responses may involve different neural projections, thus different melanopsin cells. Considering the complexity of the melanopsin system, its contribution to so many different biological functions is not surprising. In this review, we summarize the current knowledge about melanopsin in terms of its photophysics, photochemistry, mechanisms of activation, cell signaling, morphology, and physiology. In the last part, the role of melanopsin in image and non-image forming processes and cognitive and affective functioning of animals and humans, are discussed. Although in recent years considerable insight has been gained into the melanopsin system, it still remains an open question of how one protein expressed by several thousand cells in the retina, could be responsible for so many diverse functions and what activation mechanism(s) it uses.

Neuronal Responses to Short Wavelength Light Deficiency in the Rat Subcortical Visual System.

The amount and spectral composition of light changes considerably during the day, with dawn and dusk being the most crucial moments when light is within the mesopic range and short wavelength enriched. It was recently shown that animals use both cues to adjust their internal circadian clock, thereby their behavior and physiology, with the solar cycle. The role of blue light in circadian processes and neuronal responses is well established, however, an unanswered question remains: how do changes in the spectral composition of light (short wavelengths blocking) influence neuronal activity? In this study we addressed this question by performing electrophysiological recordings in image (dorsal lateral geniculate nucleus; dLGN) and non-image (the olivary pretectal nucleus; OPN, the suprachiasmatic nucleus; SCN) visual structures to determine neuronal responses to spectrally varied light stimuli. We found that removing short-wavelength from the polychromatic light (cut off at 525 nm) attenuates the most transient ON and sustained cells in the dLGN and OPN, respectively. Moreover, we compared the ability of different types of sustained OPN neurons (either changing or not their response profile to filtered polychromatic light) to irradiance coding, and show that both groups achieve it with equal efficacy. On the other hand, even very dim monochromatic UV light (360 nm; log 9.95 photons/cm2/s) evokes neuronal responses in the dLGN and SCN. To our knowledge, this is the first electrophysiological experiment supporting previous behavioral findings showing visual and circadian functions disruptions under short wavelength blocking environment. The current results confirm that neuronal activity in response to polychromatic light in retinorecipient structures is affected by removing short wavelengths, however, with type and structure - specific action. Moreover, they show that rats are sensitive to even very dim UV light.

Orexin A as a modulator of dorsal lateral geniculate neuronal activity: a comprehensive electrophysiological study on adult rats.

Orexins (OXA, OXB) are hypothalamic peptides playing crucial roles in arousal, feeding, social and reward-related behaviours. A recent study on juvenile rats suggested their involvement in vision modulation due to their direct action on dorsal lateral geniculate (dLGN) neurons. The present study aimed to verify whether a similar action of OXA can be observed in adulthood. Thus, in vivo and in vitro electrophysiological recordings on adult Wistar rats across light-dark and cortical cycles were conducted under urethane anaesthesia. OXA influenced ~28% of dLGN neurons recorded in vivo by either excitation or suppression of neuronal firing. OXA-responsive neurons did not show any spatial distribution nor represent a coherent group of dLGN cells, and responded to OXA similarly across the light-dark cycle. Interestingly, some OXA-responsive neurons worked in a cortical state-dependent manner, especially during the dark phase, and 'preferred' cortical activation over slow-wave activity induced by urethane. The corresponding patch clamp study confirmed these results by showing that < 20% of dLGN neurons were excited by OXA under both light regimes. The results suggest that OXA is involved in the development of the visual system rather than in visual processes and further implicate OXA in the mediation of circadian and arousal-related activity.

Modulation of Spontaneous and Light-Induced Activity in the Rat Dorsal Lateral Geniculate Nucleus by General Brain State Alterations under Urethane Anesthesia.

The thalamic dorsal lateral geniculate nucleus (dLGN) serves as a gating station for the transfer of light information en route to the primary visual cortex (V1). Although the modulatory input arising from the V1 and several brainstem nuclei to the dLGN is well characterised in higher mammals, little is known about its influence on dLGN activity in rodents. Using simultaneous recordings of electrocorticogram (ECoG) and single unit neuronal activity under urethane anesthesia in Long Evans rats, we managed to show that cyclic changes in the general brain state strongly affect spontaneous activity and light encoding properties of dLGN neurons. First, we characterised several groups of dLGN cells: neurons led by ECoG, neurons in which the spike rate preceded ECoG changes and neurons co-occurring or not correlated with ECoG signal. Secondly, we verified that although the general light responsiveness of the dLGN is not influenced by the state of the brain, modulation of types of photoresponses and differences in ability to encode ambient light levels were observed. Cells responding to light in a sustained manner encoded light intensity more accurately during the cortical activation phase of urethane anesthesia. On the other hand, isoflurane anesthesia does not induce such rhythmic changes in ECoG and shuts down the spontaneous neuronal activity in the dLGN. Together, these data suggest a greater modulation of spontaneous activity and dLGN neurons function, than it was previously reported for the rodent dLGN and highlight the role of anesthesia in interpretations of findings from ongoing acute experiments.

Retinal gap junctions are involved in rhythmogenesis of neuronal activity at remote locations - Study on infra-slow oscillations in the rat olivary pretectal nucleus.

A subpopulation of olivary pretectal nucleus (OPN) neurons fire action potentials in a rhythmic manner with an eruption of activity occurring approximately every two minutes. These infra-slow oscillations depend critically on functional retinal input and are subject to modulation by light. Interestingly, the activity of photoreceptors is necessary for the emergence of the rhythm and while classic photoreceptors (rods and cones) are necessary in darkness and dim light, melanopsin photoreceptors are indispensable in bright light. Using pharmacological and electrophysiological approaches in vivo, we show that also blocking retinal gap junctions (GJs), which are expressed by multitude of retinal cells, leads to the disruption of oscillatory activity in the rat OPN. Intravitreal injection of carbenoxolone (CBX) quenched oscillations in a concentration-dependent manner with 1mM being ineffective, 5mM showing partial and 20mM showing complete effectiveness in disrupting oscillations. Moreover, the most effective CBX concentration depressed cone-mediated light-induced responses of oscillatory neurons suggesting that CBX is also acting on targets other than GJs. In contrast, intravitreal injection of meclofenamic acid (MFA, 20mM) led to disruption of the rhythm but did not interfere with cone-mediated light-induced responses of oscillatory neurons, implying that MFA is more specific toward GJs than CBX, as suggested before. We conclude that electrical coupling between various types of retinal cells and resultant synchronous firing of retinal ganglion cells is necessary for the generation of infra-slow oscillations in the rat OPN.

Genomic Response to Selection for Predatory Behavior in a Mammalian Model of Adaptive Radiation.

If genetic architectures of various quantitative traits are similar, as studies on model organisms suggest, comparable selection pressures should produce similar molecular patterns for various traits. To test this prediction, we used a laboratory model of vertebrate adaptive radiation to investigate the genetic basis of the response to selection for predatory behavior and compare it with evolution of aerobic capacity reported in an earlier work. After 13 generations of selection, the proportion of bank voles (Myodes [=Clethrionomys] glareolus) showing predatory behavior was five times higher in selected lines than in controls. We analyzed the hippocampus and liver transcriptomes and found repeatable changes in allele frequencies and gene expression. Genes with the largest differences between predatory and control lines are associated with hunger, aggression, biological rhythms, and functioning of the nervous system. Evolution of predatory behavior could be meaningfully compared with evolution of high aerobic capacity, because the experiments and analyses were performed in the same methodological framework. The number of genes that changed expression was much smaller in predatory lines, and allele frequencies changed repeatably in predatory but not in aerobic lines. This suggests that more variants of smaller effects underlie variation in aerobic performance, whereas fewer variants of larger effects underlie variation in predatory behavior. Our results thus contradict the view that comparable selection pressures for different quantitative traits produce similar molecular patterns. Therefore, to gain knowledge about molecular-level response to selection for complex traits, we need to investigate not only multiple replicate populations but also multiple quantitative traits.

The contribution of inner and outer retinal photoreceptors to infra-slow oscillations in the rat olivary pretectal nucleus.

A subpopulation of olivary pretectal nucleus (OPN) neurons discharges action potentials in an oscillatory manner, with a period of approximately two minutes. This 'infra-slow' oscillatory activity depends on synaptic excitation originating in the retina. Signals from rod-cone photoreceptors reach the OPN via the axons of either classic retinal ganglion cells or intrinsically photosensitive retinal ganglion cells (ipRGCs), which use melanopsin for photon capturing. Although both cell types convey light information, their physiological functions differ considerably. The aim of the present study was to disentangle how rod-cone and melanopsin photoresponses contribute to generation of oscillatory activity. Pharmacological manipulations of specific phototransduction cascades were used whilst recording extracellular single-unit activity in the OPN of anaesthetized rats. The results show that under photopic conditions (bright light), ipRGCs play a major role in driving infra-slow oscillations, as blocking melanopsin phototransmission abolishes or transiently disturbs oscillatory firing of the OPN neurons. On the other hand, blocking rod-cone phototransmission does not change firing patterns in photopic conditions. However, under mesopic conditions (moderate light), when melanopsin phototransmission is absent, blocking rod-cone signalling causes disturbances or even the disappearance of oscillations implying that classic photoreceptors are of greater importance under moderate light. Evidence is provided that all photoreceptors are required for the generation of oscillations in the OPN, although their roles in driving the rhythm are determined by the lighting conditions, consistent with their relative sensitivities. The results further suggest that maintained retinal activity is crucial to observe infra-slow oscillatory activity in the OPN.

Melanopsin-driven increases in maintained activity enhance thalamic visual response reliability across a simulated dawn.

Twice a day, at dawn and dusk, we experience gradual but very high amplitude changes in background light intensity (irradiance). Although we perceive the associated change in environmental brightness, the representation of such very slow alterations in irradiance by the early visual system has been little studied. Here, we addressed this deficit by recording electrophysiological activity in the mouse dorsal lateral geniculate nucleus under exposure to a simulated dawn. As irradiance increased we found a widespread enhancement in baseline firing that extended to units with ON as well as OFF responses to fast luminance increments. This change in baseline firing was equally apparent when the slow irradiance ramp appeared alone or when a variety of higher-frequency artificial or natural visual stimuli were superimposed upon it. Using a combination of conventional knockout, chemogenetic, and receptor-silent substitution manipulations, we continued to show that, over higher irradiances, this increase in firing originates with inner-retinal melanopsin photoreception. At the single-unit level, irradiance-dependent increases in baseline firing were strongly correlated with improvements in the amplitude of responses to higher-frequency visual stimuli. This in turn results in an up to threefold increase in single-trial reliability of fast visual responses. In this way, our data indicate that melanopsin drives a generalized increase in dorsal lateral geniculate nucleus excitability as dawn progresses that both conveys information about changing background light intensity and increases the signal:noise for fast visual responses.