This study investigated the anatomical consequences of a photoreceptor toxin, iodoacetic acid (IAA), in the rabbit retina. Retinae were examined 2 weeks, 1, 3, and 6 months after systemic IAA injection. The retinae were processed using standard histological methods to assess the gross morphology and topographical distribution of damage, and by immunohistochemistry to examine specific cell populations in the retina. Degeneration was restricted to the photoreceptors and was most common in the ventral retina and visual streak. In damaged regions, the outer nuclear layer was reduced in thickness or eliminated entirely, with a concomitant loss of immunoreactivity for rhodopsin. However, the magnitude of the effect varied between animals with the same IAA dose and survival time, suggesting individual differences in the bioavailability of the toxin. In all eyes, the inner retina remained intact, as judged by the thickness of the inner nuclear layer, and by the pattern of immunoreactivity for protein kinase C-α (rod bipolar cells) and calbindin D-28 (horizontal cells). Müller cell stalks became immunoreactive for glial fibrillary acidic protein (GFAP) even in IAA-treated retinae that had no signs of cell loss, indicating a response of the retina to the toxin. However, no marked hypertrophy or proliferation of Müller cells was observed with either GFAP or vimentin immunohistochemistry. Thus the selective, long lasting damage to the photoreceptors produced by this toxin did not lead to a reorganization of the surviving cells, at least with survival as long as 6 months, in contrast to the remodeling of the inner retina that is observed in inherited retinal degenerations such as retinitis pigmentosa and retinal injuries such as retinal detachment.

We studied the development of sensitivity to complex motion using plaid patterns. We hypothesized, based on neurophysiological data showing a dearth of pattern direction–selective (PDS) cells in area medial temporal (MT) of infant macaques, that sensitivity to pattern motion would develop later than other forms of global motion sensitivity. We tested 10 macaque monkeys (Macaca nemestrina) ranging in age from 7 weeks to 109–160 weeks (adult). The monkeys discriminated horizontal from vertical pattern motion; sensitivity for one-dimensional (1D) direction discrimination and detection were tested as control tasks. The results show that pattern motion discrimination ability develops relatively late, between 10 and 18 weeks, while performance on the 1D control tasks was excellent at the earliest test ages. Plaid discrimination performance depends on both the speed and spatial scale of the underlying patterns. However, development is not limited by contrast sensitivity. These results support the idea that pattern motion perception depends on a different mechanism than other forms of global motion perception and are consistent with the idea that the representation of PDS neurons in MT may limit the development of complex motion perception.

The ventral lateral geniculate nucleus (vLGN), the pretectal nuclear complex (PNC) and the superior colliculus (SC) are structures that all receive retinal input. All three structures are important relay stations of the subcortical visual system. They are strongly connected with each other and involved in circadian and/or visuomotor processes. However, the information transferred along these pathways is unknown and their possible functions are, therefore, not well understood. Here, we characterized multiple pathways between the vLGN, the PNC, and the SC electrophysiologically and anatomically in an in vitro study using acute rat brain slices. Using orthodromic and antidromic electrical stimulation, we first characterized vLGN neurons that receive pretectal input and those that project to the PNC. Morphological reconstructions of cells labeled after patch clamp recordings identified these neurons as geniculo-tectal neurons and as medium-sized multipolar neurons. We identified inhibitory connections in both pathways and we could show that inhibitory postsynaptic currents (IPSCs) evoked from the PNC in vLGN neurons are mediated only by GABAA receptors, while IPSCs evoked in PNC neurons by vLGN stimulation are either mediated by both, GABAA and GABAC receptors or by a GABA receptor with mixed GABAA and GABAC receptor-like pharmacology. Finally, retrograde double labeling experiments with two different fluorescent dextran amines indicated that pretectal neurons which project to the ipsilateral vLGN also project to the ipsilateral SC.

The purpose of this study was to specify isoluminance at different retinal eccentricities and to characterize macular pigment optical density (MPOD) and distribution using the steady-state visual evoked potential (VEP). Red–green (R/G) and blue–green (B/G) gratings were generated within two circular stimulus fields (radius = 0.55 or 1.1 deg) and within four annular fields (maximum mean radius = 6.0 deg) on a color monitor. Temporal frequency was 15 Hz. Isoluminance was determined for each stimulus using minimum flicker photometry. Steady-state onset–offset VEPs were recorded to the same annular stimuli as the luminance ratio between adjacent chromatic components was changed from 0.25 to 0.85 in 11 automated steps (0.5 representing photometric isoluminance). Fourier analysis showed that the power of the first harmonic was minimized at the isoluminant ratio specific to each subject. Relative OD was computed by comparing the isoluminant ratio at any location with that for the most eccentric annulus. To compensate for the broadband characteristics of the monitor, OD values were corrected according to minimum flicker measurements made through known concentrations of carotenoid solution. MPOD was additionally measured using minimum motion photometry.

There was high correlation between the isoluminant ratios determined by minimum flicker and VEPs for both R/G and B/G stimulation (r = 0.91, P < 0.005, slope = 1). Calibrated OD values computed from VEP estimates of B/G isoluminance correlated with those derived from minimum flicker (r = 0.96, P < 0.0005, slope = 0.85) and motion photometry (r = 0.94, P < 0.0005, slope = 0.88). OD values derived from B/G VEPs increased toward the fovea and corresponded closely with minimum flicker and minimum motion assessment of MP distribution profiles. The steady-state VEP can be used to determine isoluminance at different retinal eccentricities. MPOD and distribution can be measured by steady-state VEPs to B/G stimuli.

We have determined the sequence and genomic organization of the genes encoding the cone visual pigment of the platypus (Ornithorhynchus anatinus) and the echidna (Tachyglossus aculeatus), and inferred their spectral properties and evolutionary pathways. We prepared platypus and echidna retinal RNA and used primers of the middle-wave-sensitive (MWS), long-wave-sensitive (LWS), and short-wave sensitive (SWS1) pigments corresponding to coding sequences that are highly conserved among mammals; to PCR amplify the corresponding pigment sequences. Amplification from the retinal RNA revealed the expression of LWS pigment mRNA that is homologous in sequence and spectral properties to the primate LWS visual pigments. However, we were unable to amplify the mammalian SWS1 pigment from these two species, indicating this gene was lost prior to the echidna-platypus divergence (∼21 MYA). Subsequently, when the platypus genome sequence became available, we found an LWS pigment gene in a conserved genomic arrangement that resembles the primate pigment, but, surprisingly we found an adjacent (∼20 kb) SWS2 pigment gene within this conserved genomic arrangement. We obtained the same result after sequencing the echidna genes. The encoded SWS2 pigment is predicted to have a wavelength of maximal absorption of about 440 nm, and is paralogous to SWS pigments typically found in reptiles, birds, and fish but not in mammals. This study suggests the locus control region (LCR) has played an important role in the conservation of photo receptor gene arrays and the control of their spatial and temporal expression in the retina in all mammals. In conclusion, a duplication event of an ancestral cone visual pigment gene, followed by sequence divergence and selection gave rise to the LWS and SWS2 visual pigments. So far, the echidna and platypus are the only mammals that share the gene structure of the LWS-SWS2 pigment gene complex with reptiles, birds and fishes.

Wide-field cone bipolar cells with sparse dendritic branching and proposed connectivity to blue cones were first identified in rabbit and cat. In rabbit, these were subdivided into type a (wa) and type b (wb), with axonal branching in sublamina a, and sublamina b, respectively, of the inner plexiform layer (IPL). Recent studies in rabbit support the earlier hypothesis of exclusive blue/short wavelength cone connectivity for both types. The homologues of wb cells (but not wa cells) have been identified in other mammals. The axonal branching of wa cone bipolar cells is shown to co-stratify with the dendrites of the “fiducial,” type a starburst amacrine cell, although a few branches extend into sublamina b. The axon terminal of wb cone bipolar cells is shown to be narrowly stratified in stratum 5α, deep to the dendrites of the type b starburst amacrine cell. Rabbit ganglion cells postsynaptic to wa cells are unknown, but may include class III.2a cells, similarly stratified in the IPL. The wb axon terminal is shown here to co-stratify with and to make close, likely synaptic, contacts with the dendrites of a recently described morphological subtype of class II ganglion cell in rabbit retina, IIb2. Recent morpho-physiological correlation indicates that class IIb2 cells correspond to the blue-ON-center-X or ON-brisk-sustained ganglion cells, defined physiologically in rabbit. In contrast, the wb cell in cat retina must innervate the physiologically identified blue-ON-center-sluggish-sustained ganglion cell. In monkey retina, the wb-like bipolar cells apparently innervate a small, partly bi-stratified ganglion cell. Mammals share a common pathway from short-wavelength-sensitive (S/blue) cone photoreceptors to ON-center ganglion cells in sublamina b of the IPL, in the form of wb or wb-like cone bipolar cells, but the type of ganglion cell innervated appears to be particular, and may serve different functional roles in different mammalian orders.

In support of the long-held idea that cone ratio is genetically determined by variation linked to the X-chromosome opsin gene locus, the present study identified nucleotide differences in DNA segments containing regulatory regions of the L and M opsin genes that are associated with significant differences in the relative number of L versus M cones. Specific haplotypes (combinations of genetic differences) were identified that correlated with high versus low L:M cone ratio. These findings are consistent with the biological principle that DNA sequence variations affect binding affinities for protein components of complexes that influence the relative probability that an L versus M opsin gene will be silenced during development, and in turn, produce variation in the proportion of L to M cones.

The aim of this study was to use non-invasive magneto-encephalographic techniques, together with visual stimulus paradigms that can psychophysically separate the M- and P-pathways, to examine the physiological relations of the pathways at the fovea with (1) the magneto-encephalography components M70 and M100 (in latency and amplitude), and (2) the cortical oscillatory activities (alpha, beta, and gamma), respectively. The checkerboard stimuli accompanied with different spatial frequencies (SFs) (0.5 or 4 cycles per degree) were presented (within 2° of the retinal center) to six healthy subjects by using steady-pedestal and pulse paradigms, which could activate distinct populations of M- and P-neurons. SF analyzed brain responses in each paradigm. The results show a consistent trend in M70 and M100 with increased latencies and amplitudes in response to the high SF. Mean while, the beta to gamma activities are apparently enhanced by the stimulus of high SF, especially under pulse paradigm (p = 0.03). In this study, we suggest that M70 can be a good clue to characterize the P-pathway. Moreover, in the frequency analysis, the beta oscillations may serve for more detailed visual information, while the gamma oscillations seem to reflect the signal processing in the P-pathway and with sensitivity to the fovea.

Recent work has revealed multiple pathways for cross-orientation suppression in cat and human vision. In particular, ipsiocular and interocular pathways appear to assert their influence before binocular summation in human but have different (1) spatial tuning, (2) temporal dependencies, and (3) adaptation after-effects. Here we use mask components that fall outside the excitatory passband of the detecting mechanism to investigate the rules for pooling multiple mask components within these pathways. We measured psychophysical contrast masking functions for vertical 1 cycle/deg sine-wave gratings in the presence of left or right oblique (±45 deg) 3 cycles/deg mask gratings with contrast C%, or a plaid made from their sum, where each component (i) had contrast 0.5Ci%. Masks and targets were presented to two eyes (binocular), one eye (monoptic), or different eyes (dichoptic). Binocular-masking functions superimposed when plotted against C, but in the monoptic and dichoptic conditions, the grating produced slightly more suppression than the plaid when Ci ≥ 16%. We tested contrast gain control models involving two types of contrast combination on the denominator: (1) spatial pooling of the mask after a local nonlinearity (to calculate either root mean square contrast or energy) and (2) “linear suppression” (Holmes & Meese, 2004, Journal of Vision4, 1080–1089), involving the linear sum of the mask component contrasts. Monoptic and dichoptic masking were typically better fit by the spatial pooling models, but binocular masking was not: it demanded strict linear summation of the Michelson contrast across mask orientation. Another scheme, in which suppressive pooling followed compressive contrast responses to the mask components (e.g., oriented cortical cells), was ruled out by all of our data. We conclude that the different processes that underlie monoptic and dichoptic masking use the same type of contrast pooling within their respective suppressive fields, but the effects do not sum to predict the binocular case.

Most studies on people with age-related macular degeneration (AMD) have been focused on investigations of low-level processes with simple stimuli like gratings, letters, and in perception of isolated faces or objects. We investigated the ability of people with low vision to analyze more complex stimuli like photographs of natural scenes. Fifteen participants with AMD and low vision (acuity on the better eye <20/200) and 11 normally sighted age-matched controls took part in the study. They were presented with photographs of either colored or achromatic gray level scenes in one condition and with photographs of natural scenes versus isolated objects extracted from these scenes in another condition. The photographs were centrally displayed for 300 ms. In both conditions, observers were instructed to press a key when they saw a predefined target (a face or an animal). The target was present in half of the trials. Color facilitated performance in people with low vision, while equivalent performance was found for colored and achromatic pictures in normally sighted participants. Isolated objects were categorized more accurately than objects in scenes in people with low vision. No difference was found for normally sighted observers. The results suggest that spatial properties that facilitate image segmentation (e.g., color and reduced crowding) help object perception in people with low vision.

Retinal cones are depolarized in darkness, keeping voltage-gated Ca2+ channels open and sustaining exocytosis of synaptic vesicles. Light hyperpolarizes the membrane potential, closing Ca2+ channels and suppressing exocytosis. Here, we quantify the Ca2+ concentration in cone terminals, with Ca2+ indicator dyes. Two-photon ratiometric imaging of fura-2 shows that global Ca2+ averages ~360 nM in darkness and falls to ~190 nM in bright light. Depolarizing cones from their light to their dark membrane potential reveals hot spots of Ca2+ that co-label with a fluorescent probe for the synaptic ribbon protein ribeye, consistent with tight localization of Ca2+ channels near ribbons. Measurements with a low-affinity Ca2+ indicator show that the local Ca2+ concentration near the ribbon exceeds 4 μM in darkness. The high level of Ca2+ near the ribbon combined with previous estimates of the Ca2+ sensitivity of release leads to a predicted dark release rate that is much faster than observed, suggesting that the cone synapse operates in a maintained state of synaptic depression in darkness.

Different types of retinal ganglion cell show differences in their response properties. Here we investigated the question of whether these differences are related to the distribution of the synaptic input to the dendritic tree. We measured the distribution and density of synaptic input to the dendrites of midget and parasol ganglion cells in the retina of a New World monkey, the marmoset, Callithrix jacchus. Ganglion cells were retrogradely labeled by dye injection into parvocellular or magnocellular regions of the lateral geniculate nucleus and subsequently photo-filled. Presumed bipolar cell synapses were identified immunocytochemically using antibodies against the ribbon protein CtBP2 or the GluR4 subunit of the AMPA receptor. For all cells, colocalized immunoreactive puncta were distributed across the entire dendritic tree. The density of the presumed bipolar input to midget ganglion cells was comparable for both synaptic markers, suggesting that the AMPA receptor GluR4 subunit is expressed at all synapses between midget bipolar and midget ganglion cells. Midget ganglion cells had an average of nine colocalized immunoreactive puncta per 100 μm2 dendritic surface, and parasol cells had an average of seven colocalized immunoreactive puncta per 100 μm2 dendritic surface. The densities were comparable in different regions of the dendritic tree and were not influenced by the location of the cells with respect to the fovea. Our findings suggest that the differences in the response characteristics of midget and parasol cells are not due to differences in the density of synaptic input to their dendritic tree.

Motion transparency occurs when multiple object velocities are present within a local region of retinotopic space. Transparent signals can carry information useful in the segmentation of moving objects and in the extraction of three-dimensional structure from relative motion cues. However, the physiological substrate underlying the detection of motion transparency is poorly understood. Direction tuned neurons in area MT are suppressed by transparent stimuli, suggesting that other motion sensitive areas may be needed to represent this signal robustly. Recent neuroimaging evidence implicated two such areas in the macaque superior temporal sulcus. We studied one of these, FST, with electrophysiological methods and found that a large fraction of the neurons responded well to two opposite directions of motion and to transparent stimuli containing those same directions. A linear combination of MT-like responses qualitatively reproduces this behavior and predicts that FST neurons can be tuned for transparent motion containing specific direction and depth components. We suggest that FST plays a role in motion segmentation based on transparent signals.

Age-related macular degeneration (AMD) is a major cause of visual impairment in people older than 50 years in Western countries, affecting essential tasks such as reading and face recognition. Here we investigated the mechanisms underlying the deficit in recognition of facial expressions in an AMD population with low vision. Pictures of faces displaying different emotions with the mouth open or closed were centrally displayed for 300 ms. Participants with AMD with low acuity (mean 20/200) and normally sighted age-matched controls performed one of two emotion tasks: detecting whether a face had an expression or not (expressive/non expressive (EXNEX) task) or categorizing the facial emotion as happy, angry, or neutral (categorization of expression (CATEX) task). Previous research has shown that healthy observers are mainly using high spatial frequencies in an EXNEX task while performance at a CATEX task was preferentially based on low spatial frequencies. Due to impaired processing of high spatial frequencies in central vision, we expected and observed that AMD participants failed at deciding whether a face was expressive or not but categorized normally the emotion of the face (e.g., happy, angry, neutral). Moreover, we observed that AMD participants mostly identified emotions using the lower part of the face (mouth). Accuracy did not differ between the two tasks for normally sighted observers. The results indicate that AMD participants are able to identify facial emotion but must base their decision mainly on the low spatial frequencies, as they lack the perception of finer details.

Delivery of foreign opsin genes to cone photoreceptors using recombinant adeno-associated virus (rAAV) is a potential tool for studying the basic mechanisms underlying cone based vision and for treating vision disorders. We used an in vivo retinal imaging system to monitor, over time, expression of virally-delivered genes targeted to cone photoreceptors in the Mongolian gerbil (Meriones unguiculatus). Gerbils have a well-developed photopic visual system, with 11–14% of their photoreceptors being cones. We used replication deficient serotype 5 rAAV to deliver a gene for green fluorescent protein (GFP). In an effort to direct expression of the gene specifically to either S or M cones, the transgene was under the control of either the human X-chromosome opsin gene regulatory elements, i.e., an enhancer termed the locus control region (LCR) and L promoter, or the human S-opsin promoter. Longitudinal fluorescence images reveal that gene expression is first detectable about 14 days post-injection, reaches a peak after about 3 months, and is observed more than a year post-injection if the initial viral concentration is sufficiently high. The regulatory elements are able to direct expression to a subpopulation of cones while excluding expression in rods and non-photoreceptor retinal cells. When the same viral constructs are used to deliver a human long-wavelength opsin gene to gerbil cones, stimulation of the introduced human photopigment with long-wavelength light produces robust cone responses.

Calcium-binding protein expression has proven useful in delineating neural pathways. For example, in birds, calbindin is strongly expressed in the tectofugal pathway, whereas parvalbumin (PV) is strongly expressed in the thalamofugal pathway. Whether neurons within other visual regions also differentially express calcium-binding proteins, however, has not been extensively studied. The nucleus of the basal optic root (nBOR) is a retinal-recipient nucleus that is critical for the generation of the optokinetic response. The nBOR projects to the cerebellum both directly and indirectly via the inferior olive (IO). The cerebellar and IO projections originate from different neurons within the nBOR, but whether they can also be differentiated based on calcium-binding protein expression is unknown. In this study, we combined retrograde neuronal tracing from the cerebellum and IO with fluorescent immunohistochemistry for PV and calretinin (CR) in the nBOR of pigeons. We found that about half (52.3%) of the cerebellar-projecting neurons were CR+ve, and about one-third (33.6%) were PV+ve. Most (90%) of these PV+ve cells were also labeled for CR. In contrast, very few of the IO-projecting neurons expressed CR or PV (≤2%). Thus, the direct nBOR–cerebellar and indirect nBOR–olivocerebellar pathways to the cerebellum can be distinguished based on the differential expression of CR and PV.

Transcranial direct current stimulation (tDCS) is a non-invasive method of modulating levels of cortical excitability. In this study, data gathered over a number of previously conducted experiments before and after tDCS, has been re-analyzed to investigate correlations between sex differences with respect to neuroplastic effects. Visual evoked potentials (VEPs), phosphene thresholds (PTs), and contrast sensitivity measurements (CSs) are used as indicators of the excitability of the primary visual cortex. The data revealed that cathodally induced excitability effects 10 min post stimulation with tDCS, showed no significant difference between genders. However, stimulation in the anodal direction revealed sex-specific effects: in women, anodal stimulation heightened cortical excitability significantly when compared to the age-matched male subject group. There was no significant difference between male and female subjects immediately after stimulation. These results indicate that sex differences exist within the visual cortex of humans, and may be subject to the influences of modulatory neurotransmitters or gonadal hormones which mirror short-term neuroplastic effects.

Endothelin-1 (ET-1), a potent vasoconstrictor peptide, has been implicated in the development of normal- and high-tension glaucoma. We investigated the effects of unoprostone on extracellular signal-regulated kinase (ERK) in ET-1-induced retinal ganglion cell (RGC) death and optic nerve injury. Our morphometric study showed that intravitreal injection of ET-1 led to cell loss in the RGC layer (RGCL) in 28 days. Western blot analysis showed decreased neurofilament (NF) protein in the optic nerve 28 days after ET-1 injection. In this in vivo model, increased phosphorylated ERK (p-ERK) was observed in the retina on 1 day and subsequently in the optic nerve from 7 days after ET-1 injection. Simultaneous injection of M1, as a metabolite of unoprostone, showed further increased p-ERK levels compared with ET-1 injection alone. Our morphometric study of flat-mount preparations stained with cresyl violet or retrograde labeling with a neuro-tracer and Western blot analysis of NF showed that inhibition of ERK phosphorylation led to acceleration of ET-1-induced RGC death and optic nerve damage. In addition, M1 significantly attenuated both RGC loss and the decrease in NF protein induced by ET-1. The protective effects of M1 were significantly inhibited by U0126, an ERK inhibitor. These results suggest that unoprostone has neuroprotective effects against ET-1-induced neuronal injury through ERK phosphorylation.

Many attempts have been made over the years to distinguish human and primate L (long-wavelength sensitive) from M (middle-wavelength sensitive) cone photoreceptors using either immunohistochemistry or in situ hybridization. These attempts have been unsuccessful due to the very high degree of identity between the sequences of the L and M proteins and encoding mRNAs. The recent development of chemically modified oligonucleotide probes, referred to as locked nucleic acid (LNA) probes, has shown that they hybridize with much greater affinity and specificity to the target nucleic acid. This has greatly increased the potential for differentiating L from M cones by in situ hybridization. We have designed LNA oligonucleotide probes that are complementary to either the L or M coding sequences located in exon 5 of the Macaca nemestrina L and M pigment genes. We have shown that the LNA-M and LNA-L probes hybridize specifically to their respective target nucleic acid sequences in vitro. This result strongly suggests that these probes would be instrumental in rapidly distinguishing L from M cone in the entire retina, and in defining the cone mosaic during development and in adults.