Recombinant rabies viral vectors have proven useful for applications including retrograde targeting of projection neurons and monosynaptic tracing, but their cytotoxicity has limited their use to short-term experiments. Here we introduce a new class of double-deletion-mutant rabies viral vectors that left transduced cells alive and healthy indefinitely. Deletion of the viral polymerase gene abolished cytotoxicity and reduced transgene expression to trace levels but left vectors still able to retrogradely infect projection neurons and express recombinases, allowing downstream expression of other transgene products such as fluorophores and calcium indicators. The morphology of retrogradely targeted cells appeared unperturbed at 1 year postinjection. Whole-cell patch-clamp recordings showed no physiological abnormalities at 8 weeks. Longitudinal two-photon structural and functional imaging in vivo, tracking thousands of individual neurons for up to 4 months, showed that transduced neurons did not die but retained stable visual response properties even at the longest time points imaged.

The trichromatic primate retina parses the colour content of a visual scene into 'red/green' and 'blue/yellow' representations. Cortical circuits must combine the information encoded in these colour-opponent signals to reconstruct the full range of perceived colours. Red/green and blue/yellow inputs are relayed by the lateral geniculate nucleus (LGN) of thalamus to primary visual cortex (V1), so understanding how cortical circuits transform these signals requires understanding how LGN inputs to V1 are organized. Here we report direct recordings from LGN afferent axons in muscimol-inactivated V1. We found that blue/yellow afferents terminated exclusively in superficial cortical layers 3B and 4A, whereas red/green afferents were encountered only in deeper cortex, in lower layer 4C. We also describe a distinct cortical target for 'blue-OFF' cells, whose afferents terminated in layer 4A and seemed patchy in organization. The more common 'blue-ON' afferents were found in 4A as well as lower layer 2/3. Chromatic information is thus conveyed to V1 by parallel, anatomically segregated colour-opponent systems, to be combined at a later stage of the colour circuit.

Color has become a premier model system for understanding how information is processed by neural circuits, and for investigating the relationships among genes, neural circuits, and perception. Both the physical stimulus for color and the perceptual output experienced as color are quite well characterized, but the neural mechanisms that underlie the transformation from stimulus to perception are incompletely understood. The past several years have seen important scientific and technical advances that are changing our understanding of these mechanisms. Here, and in the accompanying minisymposium, we review the latest findings and hypotheses regarding color computations in the retina, primary visual cortex, and higher-order visual areas, focusing on non-human primates, a model of human color vision.In trichromatic primates, including humans and Old World monkeys, there are three types of cone photoreceptors that are responsible for color vision ( Fig. 1 A, B). The cone classes are called L, M, and S because of their spectral-sensitivity peaks, which lie in the long-, middle-, and short-wavelength regions of the visible spectrum. These labels replace the misleading terms "red," "green," and "blue." Two physically distinct stimuli appear as different colors only if they produce different relative activations in at least two cone types; conversely, any pair of physically distinct stimuli that activate the cone types in the same relative amount appear the same, like the two yellows shown in Figure 1C. While photoreceptor responses are easily computed from the spectral distribution of the stimulus, there is no straightforward relationship between photoreceptor response and color (Hofer et al., 2005a;Shevell and Kingdom, 2008). The multitude of color phenomena, including color afterimages, color assimilation, neon-color spreading, color constancy, and colored shadows, is compelling because in many cases two physically identical stimuli are made to appear different colors, or two physically different stimuli are made to appear the same simply by changing the spatial or temporal context (Fig. 2). A full description of the neural machinery for color should account for these observations, as well as more cognitive phenomena involving the relationship between experience, language, memory, emotion and color. The neural basis of color has been reviewed previously from a range of perspectives (Gegenfurtner, 2003;Gegenfurtner and Kiper, 2003;Lennie and Movshon, 2005;Sincich and Horton, 2005;Solomon and Lennie, 2007;Conway, 2009;Dobkins, 2009;Jacobs and Nathans, 2009;Stockman and Brainard, 2010). Here we focus on advances and pressing questions regarding the mechanisms of color in retina, striate cortex, and extrastriate cortex of non-human primates, although we note that other species are emerging as excellent model systems of color processing (Lotto and Chittka, 2005;Van Hooser and Nelson, 2006;Osorio and Vorobyev, 2008;Borst, 2009;Johnson et al., 2010;Srinivasan, 2010). Retinal mechanismsA single cone by itself is color blind b...

The magnocellular visual pathway is believed to receive input from long (L) and middle (M), but not short (S), wavelength-sensitive cones. Recording from neurons in magnocellular layers of lateral geniculate nucleus (LGN) in macaque monkeys, we found that magnocellular neurons were unequivocally responsive to S cone-isolating stimuli. A quantitative analysis suggests that S cones provided about 10% of the input to these cells, on average, while L:M ratios were far more variable. S cone signals influenced responses with the same sign as L and M cone inputs (i.e., no color opponency). Magnocellular afferent recordings following inactivation of primary visual cortex demonstrated that S cone signals were feedforward in nature and did not arise from cortical feedback to LGN

The spatial summation properties of visual signals were analyzed for geniculocortical afferents in the primary visual cortex (V1) of anesthetized paralyzed macaque monkeys. Afferent input responses were recorded extracellularly during cortical inactivation through superfusion of the cortex with muscimol, allowing investigation of lateral geniculate nucleus of the thalamus (LGN) cell properties in the absence of cortical feedback. Responses from afferent inputs were classified as magno-, parvo-, or koniocellular based on anatomical organization within the cortex, established through histological reconstructions, and visual response wavelength sensitivity. More than 80% of afferents showed strong surround suppression [suppression index (SI) >0.5] and 14% showed negligible surround suppression (SI < 0.2). Afferent responses with weak and strong surround suppression were found throughout cortical input layers 4C and 4A. High-contrast estimates of the spatial extent of the classical surround were similar to the nonclassical surround. The classical and nonclassical surrounds were, on average, 1.5-fold larger than the excitatory center. Unlike neurons within V1, the spatial extent of excitatory summation for geniculocortical afferents was contrast invariant. Nonclassical surround suppression showed slight contrast dependency with estimates larger (20%) at lower contrasts and stronger at higher contrasts (13%). Surround suppression is inherent in cortical input responses and likely derives from lateral inhibition in either the LGN or retina. Although surround suppression within afferent responses increases slightly with contrast, the spatial spread of excitation remains fixed with contrast. This argues for distinct mechanisms of action for contrast-dependent modulation in cortical and subcortical responses.

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Brain circuits comprise vast numbers of intricately interconnected neurons with diverse molecular, anatomical and physiological properties. To allow “user-defined” targeting of individual neurons for structural and functional studies, we created light-inducible site-specific DNA recombinases (SSRs) based on Cre, Dre and Flp (RecVs). RecVs can induce genomic modifications by one-photon or two-photon light induction in vivo . They can produce targeted, sparse and strong labeling of individual neurons by modifying multiple loci within mouse and zebrafish genomes. In combination with other genetic strategies, they allow intersectional targeting of different neuronal classes. In the mouse cortex they enable sparse labeling and whole-brain morphological reconstructions of individual neurons. Furthermore, these enzymes allow single-cell two-photon targeted genetic modifications and can be used in combination with functional optical indicators with minimal interference. In summary, RecVs enable spatiotemporally-precise optogenomic modifications that can facilitate detailed single-cell analysis of neural circuits by linking genetic identity, morphology, connectivity and function.

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