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Aging, a universal process that affects all cells in an organism, is a major risk factor for a group of neuropathies called glaucoma, where elevated intraocular pressure is one of the known stresses affecting the tissue. Our understanding of molecular impact of aging on response to stress in retina is very limited; therefore, we developed a new mouse model to approach this question experimentally. Here we show that susceptibility to response to stress increases with age and is primed on chromatin level. We demonstrate that ocular hypertension activates a stress response that is similar to natural aging and involves activation of inflammation and senescence. We show that multiple instances of pressure elevation cause aging of young retina as measured on transcriptional and DNA methylation level and are accompanied by local histone modification changes. Our data show that repeated stress accelerates appearance of aging features in tissues and suggest chromatin modifications as the key molecular components of aging. Lastly, our work further emphasizes the importance of early diagnosis and prevention as well as age-specific management of age-related diseases, including glaucoma.

Aging, a universal process that affects all cells in an organism, is a major risk factor for a group of neuropathies called glaucoma, where elevated intraocular pressure is one of the known stresses affecting the tissue. Our understanding of molecular impact of aging on response to stress in retina is very limited, therefore we developed a new mouse model to approach this question experimentally. Here we show that susceptibility to response to stress increases with age and is primed on epigenetic level. We demonstrate that program activated by hypertension is similar to natural aging, and that one of the earliest pathways activated upon stress is senescence. Finally, we show that multiple instances of pressure elevation cause accelerated aging of young retina as measured on transcriptional and epigenetic level. Our work emphasizes the importance of early diagnosis and prevention as well as age-specific management of age-related eye-diseases, including glaucoma.

Cytosine base editors and adenine base editors (ABEs) can correct point mutations predictably and independent of Cas9-induced double-stranded DNA breaks (which causes substantial indel formation) and homology-directed repair (which typically leads to low editing efficiency). Here, we show, in adult mice, that a subretinal injection of a lentivirus expressing an ABE and a single-guide RNA targeting a de novo nonsense mutation in the Rpe65 gene corrects the pathogenic mutation with up to 29% efficiency and with minimal formation of indel and off-target mutations, despite the absence of the canonical NGG sequence as a protospacer-adjacent motif. The ABE-treated mice displayed restored RPE65 expression and retinoid isomerase activity, and near-normal levels of retinal and visual functions. Our findings motivate the further testing of ABEs for the treatment of inherited retinal diseases and for the correction of pathological mutations with non-canonical protospacer-adjacent motifs.

Inherited retinal degenerative diseases are a prominent cause of blindness. Even though mutations causing death of photoreceptors are mostly known, the pathophysiology downstream in the inner retina and along the visual pathway is incompletely characterized in the earliest disease stages. Here we investigated retinal, midbrain and cortical visual function using electroretinography (ERG), the optomotor response (OMR), visual evoked potentials (VEPs), respectively, and single unit electrophysiology at the primary visual cortex (V1) in light-adapted juvenile (∼ 1-month-old) and young adult (3-month-old)RhoP23H/WTmice, representative of early-stage retinitis pigmentosa (RP). Photopic ERG revealed up to ∼ 30 % hypersensitivity to light inRhoP23H/WTmice, as measured by the light intensity required to generate half-maximal b-wave (I50parameter).RhoP23H/WTmice also showed increased optomotor responses towards low spatial frequency drifting gratings, indicative of visual overexcitation at the midbrain level. At the V1 level, VEPs and single-cell recordings revealed prominent hyperexcitability in the juvenileRhoP23H/WTmice. Mean VEP amplitudes for light ON stimuli were nearly doubled in 1-month-oldRhoP23H/WTmice compared to controls, and more than doubled for light OFF. Single-cell recordings showed a significantly increased spontaneous V1 neuron firing in theRhoP23H/WTmice, and persistent contrast and temporal sensitivities. In contrast, direction selectivity was severely compromised. Our data suggest that during early RP, the visual pathway becomes hyperexcited. This could have both compensatory and deleterious consequences for visual behavior. Further studies on the mechanisms of hyperexcitability are warranted as this could lead to therapeutic interventions for RP.Significance statementLost retinal function in many blinding retinal degenerative disorders could soon be alleviated by advanced therapies that restore photoreception. However, it is unknown whether a visual system rewired downstream of the photoreceptors can process signals adequately. We studied the functional consequences of early rod death along the visual pathway in young retinitis pigmentosa (RP) mice. Photopic inner retina responses were moderately hypersensitized in the electroretinograms of RP mice. Reflex-based visual behavior and visual cortex electrophysiology showed hyperexcitability. Some aspects of complex visual processing were remarkably resistant to degeneration, whereas others were severely impacted. We conclude that the visual system adapts to lost photoreception by increasing sensitivity, but simultaneously becomes detrimentally hyperexcited. Mechanistic understanding could lead to therapeutic preservation and restoration of vision.

Primary sensory areas of the mammalian neocortex have a remarkable degree of plasticity, allowing neural circuits to adapt to dynamic environments. However, little is known about the effects of traumatic brain injury on visual circuit function. Here we used anatomy and in vivo electrophysiological recordings in adult mice to quantify neuron responses to visual stimuli two weeks and three months after mild controlled cortical impact injury to primary visual cortex (V1). We found that, although V1 remained largely intact in brain-injured mice, there was ~35% reduction in the number of neurons that affected inhibitory cells more broadly than excitatory neurons. V1 neurons showed dramatically reduced activity, impaired responses to visual stimuli and weaker size selectivity and orientation tuning in vivo. Our results show a single, mild contusion injury produces profound and long-lasting impairments in the way V1 neurons encode visual input. These findings provide initial insight into cortical circuit dysfunction following central visual system neurotrauma., Jan Frankowski and Andrzej Foik et al. characterized neuroanatomical, electrophysiological, and functional deficits in the primary visual cortex of mice after traumatic brain injury (TBI). Their results suggest that TBI produces long-lasting impairments to V1 neurons, and provide further insight into circuit dysfunction following injury of the visual cortex.

Despite the fundamental importance of understanding the brain’s wiring diagram, our knowledge of how neuronal connectivity is rewired by traumatic brain injury remains remarkably incomplete. Here we use cellular resolution whole-brain imaging to generate brain-wide maps of the input to inhibitory neurons in a mouse model of traumatic brain injury. We find that somatostatin interneurons are converted into hyperconnected hubs in multiple brain regions, with rich local network connections but diminished long-range inputs, even at areas not directly damaged. The loss of long-range input does not correlate with cell loss in distant brain regions. Interneurons transplanted into the injury site receive orthotopic local and long-range input, suggesting the machinery for establishing distant connections remains intact even after a severe injury. Our results uncover a potential strategy to sustain and optimize inhibition after traumatic brain injury that involves spatial reorganization of the direct inputs to inhibitory neurons across the brain.

Adiponectin receptor 1 (ADIPOR1) is a lipid and glucose metabolism regulator that possesses intrinsic ceramidase activity. Mutations of the ADIPOR1 gene have been associated with nonsyndromic and syndromic retinitis pigmentosa. Here, we show that the absence of AdipoR1 in mice leads to progressive photoreceptor degeneration, significant reduction of electroretinogram amplitudes, decreased retinoid content in the retina, and reduced cone opsin expression. Single-cell RNA-Seq results indicate that ADIPOR1 encoded the most abundantly expressed ceramidase in mice and one of the 2 most highly expressed ceramidases in the human retina, next to acid ceramidase ASAH1. We discovered an accumulation of ceramides in the AdipoR1-/- retina, likely due to insufficient ceramidase activity for healthy retina function, resulting in photoreceptor death. Combined treatment with desipramine/L-cycloserine (DC) lowered ceramide levels and exerted a protective effect on photoreceptors in AdipoR1-/- mice. Moreover, we observed improvement in cone-mediated retinal function in the DC-treated animals. Lastly, we found that prolonged DC treatment corrected the electrical responses of the primary visual cortex to visual stimuli, approaching near-normal levels for some parameters. These results highlight the importance of ADIPOR1 ceramidase in the retina and show that pharmacological inhibition of ceramide generation can provide a therapeutic strategy for ADIPOR1-related retinopathy.

Leber congenital amaurosis (LCA) is the most common cause of inherited retinal degeneration in children. LCA patients with RPE65 mutations show accelerated cone photoreceptor dysfunction and death, resulting in early visual impairment. It is therefore crucial to develop a robust therapy that not only compensates for lost RPE65 function but also protects photoreceptors from further degeneration. Here, we show that in vivo correction of an Rpe65 mutation by adenine base editor (ABE) prolongs the survival of cones in an LCA mouse model. In vitro screening of ABEs and sgRNAs enables the identification of a variant that enhances in vivo correction efficiency. Subretinal delivery of ABE and sgRNA corrects up to 40% of Rpe65 transcripts, restores cone-mediated visual function, and preserves cones in LCA mice. Single-cell RNA-seq reveals upregulation of genes associated with cone phototransduction and survival. Our findings demonstrate base editing as a potential gene therapy that confers long-lasting retinal protection.

The extrageniculate visual pathway, which carries visual information from the retina through the superficial layers of the superior colliculus and the pulvinar, is poorly understood. The pulvinar is thought to modulate information flow between cortical areas, and has been implicated in cognitive tasks like directing visually guided actions. In order to better understand the underlying circuitry, we performed retrograde injections of modified rabies virus in the visual cortex and pulvinar of the Long-Evans rat. We found a relatively small population of cells projecting to primary visual cortex (V1), compared to a much larger population projecting to higher visual cortex. Reciprocal corticothalamic projections showed a similar result, implying that pulvinar does not play as big a role in directly modulating rodent V1 activity as previously thought.

SummaryPrimary sensory areas of the mammalian neocortex have a remarkable degree of plasticity, allowing neural circuits to adapt to dynamic environments. However, little is known about the effect of traumatic brain injury on visual system function. Here we applied a mild focal contusion injury to primary visual cortex (V1) in adult mice. We found that, although V1 was largely intact in brain-injured mice, there was a reduction in the number of inhibitory interneurons that extended into deep cortical layers. In general, we found a preferential reduction of interneurons located in superficial layers, near the impact site, while interneurons positioned in deeper layers were better preserved. Three months after injury, V1 neurons showed dramatically reduced responses to visual stimuli and weaker orientation selectivity and tuning, consistent with the loss of cortical inhibition. Our results demonstrate that V1 neurons no longer robustly and stably encode visual input following a mild traumatic injury.HighlightsInhibitory neurons are lost throughout brain injured visual cortexVisually-evoked potentials are severely degraded after injuryInjured V1 neurons show weaker selectivity and tuning consistent with reduced interneurons

The pulvinar is a higher-order thalamic relay and a central component of the extrageniculate visual pathway, with input from the superior colliculus and visual cortex and output to all of visual cortex. Rodent pulvinar, more commonly called the lateral posterior nucleus (LP), consists of three highly-conserved subdivisions, and offers the advantage of simplicity in its study compared to more subdivided primate pulvinar. Little is known about receptive field properties of LP, let alone whether functional differences exist between different LP subdivisions, making it difficult to understand what visual information is relayed and what kinds of computations the pulvinar might support. Here, we characterized single-cell response properties in two V1 recipient subdivisions of rat pulvinar, the rostromedial (LPrm) and lateral (LPl), and found that a fourth of the cells were selective for orientation, compared to half in V1, and that LP tuning widths were significantly broader. Response latencies were also significantly longer and preferred size more than three times larger on average than in V1; the latter suggesting pulvinar as a source of spatial context to V1. Between subdivisons, LPl cells preferred higher temporal frequencies, whereas LPrm showed a greater degree of direction selectivity and pattern motion detection. Taken together with known differences in connectivity patterns, these results suggest two separate visual feature processing channels in the pulvinar, one in LPl related to higher speed processing which likely derives from superior colliculus input, and the other in LPrm for motion processing derived through input from visual cortex.Significance StatementThe pulvinar has a perplexing role in visual cognition as no clear link has been found between the functional properties of its neurons and behavioral deficits that arise when it is damaged. The pulvinar, called the lateral posterior nucleus (LP) in rats, is a higher order thalamic relay with input from the superior colliculus and visual cortex and output to all of visual cortex. By characterizing single-cell response properties in anatomically distinct subdivisions we found two separate visual feature processing channels in the pulvinar, one in lateral LP related to higher speed processing which likely derives from superior colliculus input, and the other in rostromedial LP for motion processing derived through input from visual cortex.

Optogenetic tools have become of great utility in the causal analysis of systems in the brain. However, current optogenetic techniques do not reliably support both excitation and suppression of the same cells in vivo, limiting analysis and slowing research. Here we developed a novel glycoprotein-deleted rabies virus expressing two channelrhodopsin proteins, GtACR2 and Chrimson, in order to independently manipulate excitatory and inhibitory transmembrane potentials, respectively. Using this approach, we demonstrated that rodent pulvinar neurons modulate cortical size tuning and suppress flash responses, but do not drive activity in visual cortex. While our goal was primarily to develop this novel method to study the structure-function organization of thalamocortical circuits, this technique is readily applicable to study any brain region.

The extrageniculate visual pathway, which carries visual information from the retina through the superficial layers of the superior colliculus and the pulvinar nucleus, is poorly understood. The pulvinar is thought to modulate information flow between cortical areas, and has been implicated in cognitive tasks like directing visually guided actions. In order to better understand the underlying circuitry, we performed retrograde injections of modified rabies virus in the visual cortex and pulvinar of the Long-Evans rat. We found a relatively small population of cells projecting to primary visual cortex (V1), compared to a much larger population projecting to higher visual cortex. Reciprocal corticothalamic projections showed a similar result, implying that pulvinar does not play as big a role in directly modulating V1 activity as previously thought.

To combat retinal degeneration, healthy fetal retinal sheets have been successfully transplanted into both rodent models and humans, with synaptic connectivity between transplant and degenerated host retina having been confirmed. In rodent studies, transplants have been shown to restore responses to flashes of light in a region of the superior colliculus corresponding to the location of the transplant in the host retina. To determine the quality and detail of visual information provided by the transplant, visual responsivity was studied here at the level of visual cortex where higher visual perception is processed. For our model, we used the transgenicRho-S334ter line-3rat (both sexes), which loses photoreceptors at an early age and is effectively blind at postnatal day 30. These rats received fetal retinal sheet transplants in one eye between 24 and 40 d of age. Three to 10 months following surgery, visually responsive neurons were found in regions of primary visual cortex matching the transplanted region of the retina that were as highly selective as normal rat to stimulus orientation, size, contrast, and spatial and temporal frequencies. Conversely, we found that selective response properties were largely absent in nontransplantedline-3rats. Our data show that fetal retinal sheet transplants can result in remarkably normal visual function in visual cortex of rats with a degenerated host retina and represents a critical step toward developing an effective remedy for the visually impaired human population.SIGNIFICANCE STATEMENTAge-related macular degeneration and retinitis pigmentosa lead to profound vision loss in millions of people worldwide. Many patients lose both retinal pigment epithelium and photoreceptors. Hence, there is a great demand for the development of efficient techniques that allow for long-term vision restoration. In this study, we transplanted dissected fetal retinal sheets, which can differentiate into photoreceptors and integrate with the host retina of rats with severe retinal degeneration. Remarkably, we show that transplants generated visual responses in cortex similar in quality to normal rats. Furthermore, transplants preserved connectivity within visual cortex and the retinal relay from the lateral geniculate nucleus to visual cortex, supporting their potential application in curing vision loss associated with retinal degeneration.

The basal forebrain provides cholinergic inputs to primary visual cortex (V1) that play a key modulatory role on visual function. While basal forebrain afferents terminate in the infragranular layers of V1, acetylcholine is delivered to more superficial layers through volume transmission. Nevertheless, direct synaptic contact in deep layers 5 and 6 may provide a more immediate effect on V1 modulation. Using helper viruses with cell type specific promoters to target retrograde infection of pseudotyped and genetically modified rabies virus evidence was found for direct synaptic input onto V1 inhibitory neurons. These inputs were similar in number to geniculocortical inputs and, therefore, considered robust. In contrast, while clear evidence for dorsal lateral geniculate nucleus input to V1 excitatory neurons was found, there was no evidence of direct synaptic input from the basal forebrain. These results suggest a direct and more immediate influence of the basal forebrain on local V1 inhibition.

Key points The process of orientation tuning is an important and well-characterized feature of neurons in primary visual cortex. The combination of ascending and descending circuits involved is not only relevant to understanding visual processing but the function of neocortex in general. The classic feed-forward model of orientation tuning predicts a broadening effect due to increasing contrast; yet, experimental results consistently report contrast invariance. We show here that contrast invariance actually depends on stimulus size such that large stimuli extending beyond the neuron's receptive field engage circuits that promote invariance, whereas optimally sized, smaller stimuli result in contrast variance that is more in line with the classical orientation tuning model. These results illustrate the importance of optimizing stimulus parameters to best reflect the sensory pathways under study and provide new clues about different circuits that may be involved in variant and invariant response properties. Abstract Selective response to stimulus orientation is a key feature of neurons in primary visual cortex, yet the underlying mechanisms generating orientation tuning are not fully understood. The combination of feed-forward and cortical mechanisms involved is not only relevant to understanding visual processing but the function of neocortex in general. The classic feed-forward model predicts that orientation tuning should broaden considerably with increasing contrast; however, experimental results consistently report contrast invariance. We show here, in primary visual cortex of anaesthetized cats under neuromuscular blockade, that contrast invariance occurs when visual stimuli are large enough to include the extraclassical surround (ECS), which is likely to involve circuits of suppression that may not be entirely feed-forward in origin. On the other hand, when stimulus size is optimized to the classical receptive field of each neuron, the population average shows a statistically significant 40% increase in tuning width at high contrast, demonstrating that contrast variance of orientation tuning can occur. Conversely, our results also suggest that the phenomenon of contrast invariance relies in part on the presence of the ECS. Moreover, these results illustrate the importance of optimizing stimulus parameters to best reflect the neural pathways under study.

Item does not contain fulltext For studies of visual cortex organization, mouse is becoming an increasingly more often used model. In addition to its genetic tractability, the relatively small area of cortical surface devoted to visual processing simplifies efforts in relating the structure of visual cortex to visual function. However, the nature of this compact organization can make some comparisons to the much larger non-human primate visual cortex difficult. The squirrel, as a highly visual rodent offers a useful means for better understanding how mouse and monkey cortical organization compares. More in line with primates than their nocturnal rodent cousin, squirrels rely much more on sight and have evolved a larger expanse of cortex devoted to visual processing. To reveal the detailed organization of visual cortex in squirrels, we injected a highly sensitive monosynaptic retrograde tracer (glycoprotein deleted rabies virus) into several locations of primary visual cortex (V1) in California ground squirrels. The resulting pattern of connectivity revealed an organizational scheme in the squirrel that retains some of the basic features of the mouse visual cortex along the medial and posterior borders of V1, but unlike mouse has an elaborate and extensive pattern laterally that is more similar to the early visual cortex organization found in monkeys. In this way, we show that the squirrel can serve as a useful model for comparison to both mouse and primate visual systems, and may help facilitate comparisons between these two very different yet widely used animal models of visual processing. 13 p.

Background: Cortical inhibition plays a critical role in controlling and modulating cortical excitation, and a more detailed understanding of the neuronal circuits contributing to each will provide more insight into their roles in complex cortical computations. Traditional neuronal tracers lack a means for easily distinguishing between circuits of inhibitory and excitatory neurons. To overcome this limitation, we have developed a technique for retrogradely labeling inputs to local clusters of inhibitory or excitatory neurons, but not both, using neurotropic adenoassociated and lentiviral vectors, cell-type-specific promoters, and a modified rabies virus. Results: Applied to primary visual cortex (V1) in mouse, the cell-type-specific tracing technique labeled thousands of presynaptically connected neurons and revealed that the dominant source of input to inhibitory and excitatory neurons is local in origin. Neurons in other visual areas are also labeled; the percentage of these intercortical inputs to excitatory neurons is somewhat higher (∼20%) than to inhibitory neurons (

The intrinsic functional architecture of early cortical areas in highly visual mammals is characterized by the presence of domains and pinwheels, with orientation preference of the inputs to these regions being more and less selective, respectively. We exploited this organizational feature to investigate mechanisms supporting extraclassical surround suppression, a process thought to be critical for figure ground segregation and form vision. Combining intrinsic signal optical imaging and single-unit recording in V1 of anesthetized cats, we show for the first time that the orientation tuning of the suppressive surround is sharper for domain than for pinwheel neurons. This difference depends on high center gain and is more pronounced in superficial cortex. In addition, when we remove the near component of the surround stimulus, the strength of suppression induced by the iso-oriented surround is significantly reduced for domain neurons but is unchanged for orthogonal oriented surrounds. This leads to broader orientation tuning of suppression that renders domain cells indistinguishable from pinwheel cells. Because the limited receptive field of the near surround can be accounted for by the lateral spread of long-range connections in V1, our findings suggest that intrinsic V1 circuits play a key role in the orientation tuning of extraclassical surround suppression.

When somatosensory cortex (S1) is deprived of some of its inputs after section of ascending afferents in the dorsal columns of the spinal cord, it reorganizes to overrepresent the surviving inputs. As somatosensory cortex provides guiding sensory information to motor cortex, such sensory loss and representational reorganization could affect the development of the motor map in primary motor cortex (M1), especially if the sensory loss occurs early in development. To address this possibility, the dorsal columns of the spinal cord were sectioned between cervical levels (C3–5) 3–12 days after birth in five macaque monkeys. After 3–5 years of maturation (young adults), we determined how movements were represented in M1 contralateral to the lesion by using microelectrodes to electrically stimulate sites in M1 to evoke movements. Although the details of the motor maps in these five monkeys varied, the forelimb motor maps were abnormal. The representations of digit movements were reduced and abnormally arranged. Current levels for evoking movements from the forelimb region of M1 were in the normal range, but the lowest mean stimulation thresholds were for wrist or elbow instead of digit movements. Incomplete lesions and bilateral lesions produced fewer abnormalities. The results suggest that the development of normal motor cortex maps in M1 depends on sensory feedback from somatosensory maps.

SummaryThe superior colliculus (SC) is the first station in a subcortical relay of retinal information to extrastriate visual cortex. Ascending SC projections pass through pulvinar and LGN on their way to cortex, but it is not clear how many synapses are required to complete these circuits or which cortical areas are involved. To examine this relay directly, we injected transynaptic rabies virus into several extrastriate visual areas. We observed disynaptically labeled cells in superficial, retino-recipient SC layers from injections in dorsal stream areas MT and V3, but not the earliest extrastriate area, V2, nor ventral stream area V4. This robust SC-dorsal stream pathway is most likely relayed through the inferior pulvinar and can provide magnocellular-like sensory inputs necessary for motion perception and the computation of orienting movements. Furthermore, by circumventing primary visual cortex, this pathway may also underlie the remaining visual capacities associated with blindsight.

[Abstract] In V1, local circuitry depends on the position in the orientation map: close to pinwheel centers, recurrent inputs show variable orientation preferences; within iso-orientation domains, inputs are relatively uniformly tuned. Physiological properties such as cell's membrane potentials, spike outputs, and temporal characteristics change systematically with map location. We investigate in a firing rate and a Hodgkin–Huxley network model what constraints these tuning characteristics of V1 neurons impose on the cortical operating regime. Systematically varying the strength of both recurrent excitation and inhibition, we test a wide range of model classes and find the likely models to account for the experimental observations. We show that recent intracellular and extracellular recordings from cat V1 provide the strongest evidence for a regime where excitatory and inhibitory recurrent inputs are balanced and dominate the feed-forward input. Our results are robust against changes in model assumptions such as spatial extent and strength of lateral inhibition. Intriguingly, the most likely recurrent regime is in a region of parameter space where small changes have large effects on the network dynamics, and it is close to a regime of “runaway excitation,” where the network shows strong self-sustained activity. This could make the cortical response particularly sensitive to modulation.

Christina M Cerkevich,1 David C Lyon,2 Pooja Balaram,3 Jon H Kaas3 1Department of Neurobiology, University of Pittsburgh School of Medicine, Systems Neuroscience Institute, Pittsburgh, PA, USA; 2Department of Anatomy and Neurobiology, University of California, Irvine, CA, USA; 3Department of Psychology, Vanderbilt University, Nashville, TN, USA Abstract: To better reveal the pattern of corticotectal projections to the superficial layers of the superior colliculus (SC), we made a total of ten retrograde tracer injections into the SC of three macaque monkeys (Macaca mulatta). The majority of these injections were in the superficial layers of the SC, which process visual information. To isolate inputs to the purely visual layers in the superficial SC from those inputs to the motor and multisensory layers deeper in the SC, two injections were placed to include the intermediate and deep layers of the SC. In another case, an injection was placed in the medial pulvinar, a nucleus not known to be strongly connected with visual cortex, to identify possible projections from tracer spread past the lateral boundary of the SC. Four conclusions are supported by the results: 1) all early visual areas of cortex, including V1, V2, V3, and the middle temporal area, project to the superficial layers of the SC; 2) with the possible exception of the frontal eye field, few areas of cortex outside of the early visual areas project to the superficial SC, although many do, however, project to the intermediate and deep layers of the SC; 3) roughly matching retinotopy is conserved in the projections of visual areas to the SC; and 4) the projections from different visual areas are similarly dense, although projections from early visual areas appear somewhat denser than those of higher order visual areas in macaque cortex. Keywords: visual cortex, superior colliculus, frontal eye field, posterior parietal cortex, visual system

In primates, the cortex adjoining the rostral border of V2 has been variously interpreted as belonging to a single visual area, V3, with dorsal V3 (V3d) representing the lower visual quadrant and ventral V3 (V3v) representing the upper visual quadrant, V3d and V3v constituting separate, incomplete visual areas, V3d and ventral posterior (VP), or V3d being divided into several visual areas, including a dorsomedial (DM) visual area, a medial visual area (M), and dorsal extension of VP (or VLP). In our view, the evidence from V1 connections strongly supports the contention that V3v and V3d are parts of a single visual area, V3, and that DM is a separate visual area along the rostral border of V3d. In addition, the retinotopy revealed by V1 connection patterns, microelectrode mapping, optical imaging mapping, and functional magnetic resonance imaging (fmri) mapping indicates that much of the proposed territory of V3d corresponds to V3. Yet, other evidence from microelectrode mapping and anatomical connection patterns supports the possibility of an upper quadrant representation along the rostral border of the middle of dorsal V2 (V2d), interpreted as part of DM or DM plus DI, and along the midline end of V2d, interpreted as the visual area M. While the data supporting these different interpretations appear contradictory, they also seem, to some extent, valid. We suggest that V3d may have a gap in its middle, possibly representing part of the upper visual quadrant that is not part of DM. In addition, another visual area, M, is likely located at the DM tip of V3d. There is no evidence for a similar disruption of V3v. For the present, we favor continuing the traditional concept of V3 with the possible modification of a gap in V3d in at least some primates.

[Abstract] Cortical computations critically involve local neuronal circuits. The computations are often invariant across a cortical area yet are carried out by networks that can vary widely within an area according to its functional architecture. Here we demonstrate a mechanism by which orientation selectivity is computed invariantly in cat primary visual cortex across an orientation preference map that provides a wide diversity of local circuits. Visually evoked excitatory and inhibitory synaptic conductances are balanced exquisitely in cortical neurons and thus keep the spike response sharply tuned at all map locations. This functional balance derives from spatially isotropic local connectivity of both excitatory and inhibitory cells. Modeling results demonstrate that such covariation is a signature of recurrent rather than purely feed-forward processing and that the observed isotropic local circuit is sufficient to generate invariant spike tuning.

Patterns of thalamocortical connections were related to architectonically defined subdivisions of the pulvinar complex and the dorsolateral geniculate nucleus (LGN) in tree shrews (Tupaia belangeri). Tree shrews are of special interest because they are considered close relatives of primates, and they have a highly developed visual system. Several distinguishable tracers were injected within and across cortical visual areas in individual tree shrews in order to reveal retinotopic patterns and cortical targets of subdivisions of the pulvinar. The results indicate that each of the three architectonic regions of the pulvinar has a distinctive pattern of cortical connections and that one of these divisions is further divided into two regions with different patterns of connections. Two of the pulvinar nuclei have similar retinotopic patterns of projections to caudal visual cortex. The large central nucleus of the pulvinar (Pc) projects to the first and second visual areas, V1 and V2, and an adjoining temporal dorsal area (TD) in retinotopic patterns indicating that the upper visual quadrant is represented dorsal to the lower quadrant in Pc. The smaller ventral nucleus (Pv) which stains darkly for the Cat-301 antigen, projects to these same cortical areas, with a retinotopic pattern. Pv also projects to a temporal anterior area, TA. The dorsal nucleus (Pd), which densely expresses AChE, projects to posterior and ventral areas of temporal extrastriate cortex, areas TP and TPI. A posterior nucleus, Pp, projects to anterior areas TAL and TI, of the temporal lobe, as well as TPI. Injections in different cortical areas as much as 6 mm apart labeled overlapping zones in Pp and double-labeled some cells. These results indicate that the visual pulvinar of tree shrews contains at least four functionally distinct subdivisions, or nuclei. In addition, the cortical injections revealed that the LGN projects topographically and densely to V1 and that a significant number of LGN neurons project to V2 and TD.

We used patterns of connections of primary visual cortex (V1) to reevaluate differing proposals on the organization of extrastriate cortex in three species of New World monkeys. Several fluorescent tracers and the bidirectional tracer cholera toxin B subunit (CTB) were injected into dorsal V1 (representing the lower visual quadrant) and ventral V1 (representing the upper visual quadrant) of titi, squirrel, and owl monkeys. Labeled cells and terminals were plotted on brain sections cut parallel to the surface of flattened cortex and were related to architectonic boundaries. The results provided compelling evidence for both dorsal V3 with dorsal V1 connections and ventral V3 with ventral V1 connections. The connection pattern indicated that V3 represents the visual hemifield as a mirror image of V2. In addition, V3 could be recognized by a weak banding pattern in brain sections processed for cytochrome oxidase. V1 has connections with at least 12 subdivisions of visual cortex, with half of the connections involving V2 and 20% V3. Comparable results were obtained from all three species, suggesting that visual cortex is similarly organized.

Previously we described patterns of connections that support the concept of V3 in small New World marmoset monkeys, three species of larger New World monkeys, and two species of Old World macaque monkeys. Here we describe a pattern of V1 connections with extrastriate visual cortex in Galago garnetti (also known as Otolemur garnetti) that demonstrates the existence of a V3 in a strepsirhine (prosimian) primate. Injections of fluorochromes or cholera toxin subunit-B (CTB) in V1 labeled cells and terminals in retinotopically matched regions in V2, V3, DL (V4), and MT. Labeled axon terminations were more focused primarily in middle layers of cortex, likely representing ‘feedforward’ input from V1, whereas labeled cells were more widespread and found in both superficial and deeper cortical layers, indicative of feedback projections. Averaged across injections, V3 had the third largest percentage of labeled cells (11%), following only V2 (47%) and the middle temporal area (MT; 19%). The dorsolateral area (DL, or V4; 9%) also contained a relatively large number of retrogradely labeled cells. These results indicate that V2, V3, DL (V4), and MT are retinotopically connected with V1, and provide major sources of feedback. Other extrastriate areas were less densely connected to V1, and there was no clear indication of labeled terminals. Inferotemporal cortex (IT) provided nearly 7% of feedback connections, whereas the dorsomedial area (DM) contributed about 3%. The remaining areas that have been proposed for galago extrastriate cortex, MTc, MST, FST, LPP and VPP, each accounted for about 1% or less of the total number of labeled cells. Thus, six extrastriate areas, V2, MT, V3, DL (V4), IT, and DM provide over 96% of visual cortex projections to V1. These areas also provide most of the projections to V1 in New and Old World monkeys.

Through more than 30 years of research, the nature of the third visual area, V3, and even its existence have been in question. Here, we used injections of up to five distinguishable tracers into both dorsal and ventral portions of V1 of macaque monkeys (representing the lower and upper visual quadrant, respectively) to provide compelling evidence for a V3 that is smaller than V2. This V3 includes both dorsal and ventral halves mirroring dorsal and ventral V2 in retinotopic organization. Of the approximately ten areas with V1 connections, V3 appears to account for about 20%.

Intrinsic signal optical imaging reveals a highly modular map of orientation preference in the primary visual cortex (V1) of several species. This orientation map is characterized by domains and pinwheels where local circuitry is either more or less orientation selective, respectively. It has now been repeatedly demonstrated that neurons in pinwheels tend to be more broadly tuned to orientation, likely due to the broad range of orientation preference of the neighboring neurons forming pinwheels. However, certain stimulus conditions, such as a decrease in contrast or an increase in size, significantly sharpen tuning widths of V1 neurons. Here, we find that pinwheel neuron tuning widths are broader than domain neurons only for high contrast, optimally sized stimuli, conditions that maximize excitation through feedforward, and local cortical processing. When contrast was lowered or size increased, orientation tuning width sharpened and became equal. These latter conditions are conducive to less local excitation either through lower feedforward drive or by surround suppression arising from long-range cortical circuits. Tuning width differences between pinwheel and domain neurons likely arise through more local circuitry and are overcome through recruitment of longer-range cortical circuits.

Yong-Jun Liu, Miguel Arreola, Cassandra M Coleman, David C Lyon Department of Anatomy and Neurobiology, School of Medicine, University of California, Irvine, CA, USA Abstract: Neurons in primary visual cortex (V1) integrate across the representation of the visual field through networks of long-range projecting pyramidal neurons. These projections, which originate from within V1 and through feedback from higher visual areas, are likely to play a key role in such visual processes as low contrast facilitation and extraclassical surround suppression. The extent of the visual field representation covered by feedback is generally much larger than that covered through monosynaptic horizontal connections within V1, and, although it may be possible that multisynaptic horizontal connections across V1 could also lead to more widespread spatial integration, nothing is known regarding such circuits. In this study, we used injections of the CVS-11 strain of rabies virus to examine disynaptic long-range horizontal connections within macaque monkey V1. Injections were made around the representation of 5° eccentricity in the lower visual field. Along the opercular surface of V1, we found that the majority of connected neurons extended up to 8 mm in most layers, consistent with twice the typically reported distances of monosynaptic connections. In addition, mainly in layer 6, a steady presence of connected neurons within V1 was observed up to 16 mm away. A relatively high percentage of these connected neurons had large-diameter somata characteristic of Meynert cells, which are known to project as far as 8 mm individually. Several neurons, predominantly in layer 6, were also found deep within the calcarine sulcus, reaching as far as 20° of eccentricity, based on estimates, and extending well into the upper visual field representation. Thus, our anatomical results provide evidence for a wide-ranging disynaptic circuit within V1, mediated largely through layer 6, that accounts for integration across a large region of the visual field. Keywords: visual cortex, horizontal connections, long-range lateral connections, Meynert cells, surround suppression, feedback

The existence of a third visual area, V3, along the outer margin of V2 was originally proposed for primates on the basis of projections from V1. The evidence for V3 was never totally convincing because investigators failed to demonstrate V1 projections to ventral V3, and projections to dorsal V3 could be attributed to the dorsomedial visual area (DM). We have reexamined the issue by placing large injections into both dorsal and ventral portions of V1 and subsequently processing flattened cortex for myelin and cytochrome oxidase so that borders of V1 and V2 could be determined accurately. The injections were in small-brained marmosets, where ventral V1 was most accessible and cortex could be flattened easily. The results indicate that dorsal V1 (representing the lower visual quadrant) projects to a narrow “dorsal V3” located between DM and dorsal V2, whereas ventral V1 (representing the upper visual quadrant) projects to a narrow “ventral V3.” Architectonic borders for these dorsal and ventral strips were clearly apparent. In addition, all parts of V1 project to DM, whereas ventral V1 connections indicate that the dorsolateral area (DL) extends more ventral than has been established previously. We also placed injections within dorsal V2, dorsal and ventral DM, and dorsal, central, and ventral middle temporal (MT) area. Results from these injections were consistent with the proposed retinotopic organizations of V3, DM, and DL. We provide compelling evidence for the existence of areas V3, DM, and DL in marmosets and suggest that these areas are likely to be found in all primates.

Cortical organization was examined in five shrew species. In three species, Blarina brevicauda, Cryptotis parva, and Sorex palustris, microelectrode recordings were made in cortex to determine the organization of sensory areas. Cortical recordings were then related to flattened sections of cortex processed for cytochrome oxidase or myelin to reveal architectural borders. An additional two species (Sorex cinereus and Sorex longirostris) with visible cortical subdivisions based on histology alone were analyzed without electrophysiological mapping. A single basic plan of cortical organization was found in shrews, consisting of a few clearly defined sensory areas located caudally in cortex. Two somatosensory areas contained complete representations of the contralateral body, corresponding to primary somatosensory cortex (S1) and secondary somatosensory cortex (S2). A small primary visual cortex (V1) was located closely adjacent to S1, whereas auditory cortex (A1) was located in extreme caudolateral cortex, partially encircled by S2. Areas did not overlap and had sharp, histochemically apparent and electrophysiologically defined borders. The adjacency of these areas suggests a complete absence of intervening higher level or association areas. Based on a previous study of corticospinal connections, a presumptive primary motor cortex (M1) was identified directly rostral to S1. Apparently, in shrews, the solution to having extremely little neocortex is to have only a few small cortical subdivisions. However, the small areas remain discrete, well organized, and functional. This cortical organization in shrews is likely a derived condition, because a wide range of extant mammals have a greater number of cortical subdivisions.

The ipsilateral and contralateral cortical connections of visual cortex of tree shrews (Tupaia belangeri) were investigated by placing restricted injections of fluorochrome tracers, wheat germ agglutinin-horseradish peroxidase, or biotinylated dextran amine into area 17 (V1), area 18 (V2), or the adjoining temporal dorsal area (TD). As previously reported, V1 was characterized by a widespread, patchy pattern of intrinsic connections; ipsilateral connections with V2, TD, and to a lesser extent, other areas of the temporal cortex; and contralateral connections with V1, V2, and TD. A surface-view of the myelin pattern in V1 revealed a patchwork of light and dark module-like regions. The ipsilateral connections with V2 and TD were roughly topographic, whereas heterotopic locations in V1 were callosally connected. Injections in V2 labeled as much as one third of V2 in a patchy pattern, and portions of ipsilateral V1 and TD in roughly topographic patterns. In addition, connections with several other visual areas in the temporal lobe were revealed. Contralaterally, most of the label was in V2, with some in V1 and TD. Injections in TD demonstrated connections within the region, and with adjoining portions of the temporal cortex, V2, and V1. There were sparse connections with an oval of densely myelinated cortex, which we have termed the temporal inferior area (TI). Callosal connections were concentrated in TD, but also included V2. The results provide further evidence for modular organizations within V1 and V2, and reveal for the first time the complete patterns of cortical connections of V2 and TD. The results are consistent with the proposal that at least three visual areas, the temporal anterior area, TA, the temporal dorsal area, TD, and the temporal posterior area, TP, exist along the rostrolateral border of V2 in tree shrews; suggest visual involvement of at least three other areas, the temporal inferior area, TI, the temporal anterior lateral area, and the temporal posterior inferior area located more ventrally in the temporal cortex; and fortify the conclusion that TD is the likely homologue of the middle temporal visual area of primates. Because tree shrews are considered close relatives of primates, the evidence for several visual areas along the border of V2 is more compatible with theories that propose a series of visual areas along V2 in primates, rather than a single visual area, V3.

The commentary by Rosa et al. , argues that while a V3-like area (VLP), termed VLP, is present in the V3v region immediately adjacent to the ventral half of V2; dorsally, a V3d is not found along the dorsal half of V2 in any primate species. Instead, the dorsal region (which represents the lower

Neuronal responses in primary visual cortex (V1) can be suppressed by a stimulus presented to the extraclassical surround, and such interactions are thought to be critical for figure ground segregation and form perception. While surround suppression likely originates from both feedforward afferents and multiple cortical circuits, it is unclear what role each circuit plays in the surround’s orientation tuning. To investigate this we recorded from single units in V1 of anesthetized cat and analyzed the orientation tuning of the suppressive-surround over time. In addition, based on orientation maps derived through optical imaging prior to recording, neurons were classified as being located in domains or pinwheels. For both types of neurons, shortly after response onset (10 ms) the suppressive-surround is broadly tuned to orientation, but this is followed by a steep improvement in tuning over the next ∼30 ms. While the tuning of the pinwheel cells plateaus at this point, tuning is enhanced further for domain cells, especially those located superficially in the cortex, reaching a peak at 80 ms from response onset. This relatively slow evolution of the orientation tuning of the suppressive surround suggests that fast-arriving feedforward circuits (10 ms) likely only provide broadly tuned suppression, but that feedback from higher visual areas which is likely to arrive over the next 30 ms and can cover both the receptive field center and the extraclassical surround contributes to the initial steep rise in tuning for both cell types. Moreover, we speculate that the even later enhancement in tuning for domain neurons could mean the involvement of inputs from relatively long-range lateral connections, which not only propagate slowly but also link like-oriented domains corresponding to the receptive field of only the extraclassical surround.

In primates, retinal inputs are relayed through the magno- and parvocells of the lateral geniculate nucleus (LGN) indirectly to extrastriate visual cortex. The most direct pathway identified to the extrastriate cortex is a disynaptic one that provides robust magno- and parvocellular inputs to the middle temporal area (MT). The inclusion of parvocells in this projection is somewhat surprising because of their importance for color and form vision, whereas MT is more strictly tuned to velocity. This raises the question of whether areas more involved in color and form processing, such as V4, receive similar projections. We report here on experiments that use rabies virus injections into V4 to retrogradely label mono- and disynaptic inputs. We find only a small number of labeled neurons in the LGN in a pattern consistent with monosynaptic labeling of koniocells, rather than disynaptic labeling of magno- and parvocells. The lack of robust magno- and parvocellular label was not due to ineffective viral transport because in the same cases we find hundreds of neurons labeled in the thalamic reticular nucleus, a structure that can only be labeled disynaptically from the cortex. We also find a complete absence of neurons labeled in V1, but thousands in adjacent areas V2 and V3. This result helps explain the absence of labeled magno- and parvocells in the LGN because disynaptic transport from an extrastriate visual area should require a relay through V1. Taken together, these results suggest that ascending magno/parvocellular inputs to V4 are more hierarchically organized than the relatively direct inputs to MT. J. Comp. Neurol. 520:2500–2511, 2012. © 2012 Wiley Periodicals, Inc.

The visual system in primates is represented by a remarkably large expanse of the cerebral cortex. While more precise investigative studies that can be performed in non-human primates contribute towards understanding the organization of the human brain, there are several issues of visual cortex organization in monkey species that remain unresolved. In all, more than 20 areas comprise the primate visual cortex, yet there is little agreement as to the exact number, size and visual field representation of all but three. A case in point is the third visual area, V3. It is found relatively early in the visual system hierarchy, yet over the last 40 years its organization and even its very existence have been a matter of debate among prominent neuroscientists. In this review, we discuss a large body of recent work that provides straightforward evidence for the existence of V3. In light of this, we then re-examine results from several seminal reports and provide parsimonious re-interpretations in favour of V3. We conclude with analysis of human and monkey functional magnetic resonance imaging literature to make the case that a complete V3 is an organizational feature of all primate species and may play a greater role in the dorsal stream of visual processing.

The visual cortex of cats is highly evolved. Analogously to the brains of primates, large numbers of visual areas are arranged hierarchically and can be parsed into separate dorsal and ventral streams for object recognition and visuospatial representation. Within early primate visual areas, V1 and V2, and to a lesser extent V3, the two streams are relatively segregated and relayed in parallel to higher order cortex, although there is some evidence suggesting an alignment of V2 and V3 to one stream over the other. For cats, there is no evidence of anatomical segregation in areas 18 and 19, the analogs to V2 and V3. However, previous work was only qualitative in nature. Here we re-examined the feedback connectivity patterns of areas 18/19 in quantitative detail. To accomplish this, we used a genetically modified rabies virus that acts as a retrograde tracer and fills neurons with fluorescent protein. After injections into area 19, many more neurons were labeled in putative ventral stream area 21a than in putative dorsal stream region posterolateral suprasylvian complex of areas (PLS), and the dendrites of neurons in 21a were significantly more complex. Conversely, area 18 injections labeled more neurons in PLS, and these were more complex than neurons in 21a. We infer from our results that area 19 in cat is more aligned to the ventral stream and area 18 to the dorsal stream. Based on the success of our approach, we suggest that this method could be applied to resolve similar issues related to primate V3. J. Comp. Neurol. 520:988–1004, 2012. © 2011 Wiley Periodicals, Inc.

Visual stimuli outside of the classical receptive field (CRF) can influence the response of neurons in primary visual cortex (V1). While recording single units in cat, we presented drifting sinusoidal gratings in circular apertures of different sizes to investigate this extraclassical surround modulation over time. For the full 2-s stimulus time course, three types of neurons were found: 1) 68% of the cells were “suppressive,” 2) 25% were “plateau” cells that showed response saturation with no suppression, and 3) the remaining 6% of cells were “facilitative.” Analysis of the response dynamics revealed that at response onset, activity of one-half of facilitative cells, 70% of plateau cells, and all suppressive cells is suppressed by the surround. However, over the next 20–30 ms, surround modulation changes to stronger suppression for suppressive cells, substantial facilitation for facilitative cells, and weak facilitation for plateau cells. For all three cell types, these modulatory effects then stabilize between 100 and 200 ms from stimulus onset. Thus our findings illustrate two stages of surround modulation. Early modulation is mainly suppressive regardless of cell type and, because of rapid onset, may rely on feedforward mechanisms. Surround modulation that evolves later in time is not always suppressive, depending on cell type, and may be generated through different combinations of cortical circuits. Additional analysis of modulation throughout the cortical column suggests the possibility that the larger excitatory fields of facilitative cells, primarily found in infragranular layers, may contribute to the second stage of suppression through intracolumnar circuitry.

The visual pulvinar is part of the dorsal thalamus, and in primates it is especially well developed. Recently, our understanding of how the visual pulvinar is subdivided into nuclei has greatly improved as a number of histological procedures have revealed marked architectonic differences within the pulvinar complex. At the same time, there have been unparalleled advances in understanding of how visual cortex of primates is subdivided into areas and how these areas interconnect. In addition, considerable evidence supports the view that the hierarchy of interconnected visual areas is divided into two major processing streams, a ventral stream for object vision, and a dorsal stream for visually guided actions. In this review, we present evidence that a subset of medial nuclei in the inferior pulvinar function predominantly as a subcortical component of the dorsal stream while the most lateral nucleus of the inferior pulvinar and the adjoining ventrolateral nucleus of the lateral pulvinar are more devoted to the ventral stream of cortical processing. These nuclei provide cortico-pulvinar-cortical interactions that spread information across areas within streams, as well as information relayed from the superior colliculus via inferior pulvinar nuclei to largely dorsal stream areas.