Synaptic organization of visual space in primary visual cortex

Abstract

How a sensory stimulus is processed and perceived depends on the surrounding sensory scene. In the visual cortex, contextual signals can be conveyed by an extensive network of intra- and inter-areal excitatory connections that link neurons representing stimulus features separated in visual space. However, the connectional logic of visual contextual inputs remains unknown; it is not clear what information individual neurons receive from different parts of the visual field, nor how this input relates to the visual features that a neuron encodes, defined by its spatial receptive field. Here we determine the organization of excitatory synaptic inputs responding to different locations in the visual scene by mapping spatial receptive fields in dendritic spines of mouse visual cortex neurons using two-photon calcium imaging. We find that neurons receive functionally diverse inputs from extended regions of visual space. Inputs representing similar visual features from the same location in visual space are more likely to cluster on neighbouring spines. Inputs from visual field regions beyond the receptive field of the postsynaptic neuron often synapse on higher-order dendritic branches. These putative long-range inputs are more frequent and more likely to share the preference for oriented edges with the postsynaptic neuron when the receptive field of the input is spatially displaced along the axis of the receptive field orientation of the postsynaptic neuron. Therefore, the connectivity between neurons with displaced receptive fields obeys a specific rule, whereby they connect preferentially when their receptive fields are co-oriented and co-axially aligned. This organization of synaptic connectivity is ideally suited for the amplification of elongated edges, which are enriched in the visual environment, and thus provides a potential substrate for contour integration and object grouping.

Extended Data Figure 1. Isolation of spine-specific signals using robust regression

a, Calcium signal in the spine as a function of the signal in the corresponding dendritic shaft for one example spine. The slope of the robust fit (red dashed line), which indicates the contribution of dendritic activity to the spine signal, was used as a scaling factor. The scaled dendrite signal was then subtracted from the spine signal. b, Example traces of the calcium signal in the dendrite (top), the signal in the spine and the estimated dendritic component (scaled dendrite signal, middle) and the isolated spine-specific signal after subtraction (bottom).

Extended Data Figure 2. The relationship between orientation preference derived from spine RFs and drifting grating responses.

a, Smoothed RFs (top), and orientation preference extracted from the RFs (bottom) for three example spines. b, Example orientation tuning curves obtained using sinusoidal gratings for the same spines as in a. Normalized responses were fitted with the sum of two Gaussians (See Methods). Error bars indicate SEM. c, Polar plots of the grating responses above in b. d, Correspondence of orientation preference derived from responses to drifting gratings and from the RF Gabor fit of individual spines. Correlation coefficient and p-value from circular correlation, n = 89 spines. e, The frequency of spines as a function of the difference in their orientation preference derived from RFs and grating responses (ΔOrientation). The majority of spines show similar orientation preferences for the two methods.

Extended Data Figure 3. The relationship between spine pair distance and different visual response properties

a-g, Dendritic separation of spines pairs versus RF similarity (a, spatial RF correlation coefficients), ON subfield correlation coefficient (b), OFF subfield correlation coefficient (c), ON + OFF RF overlap (d, see Methods), RF centre distance (e), difference in orientation preference (f, ΔOrientation), and correlation coefficient of calcium signals (g, total correlation). N = 3966 spine pairs, 74 dendrites, 21 mice. Blue shading represents the 95% confidence interval of the mean.

Extended Data Figure 4. Simultaneous imaging of dendritic and somatic calcium signals.

a, Two imaging planes separated by 10 μm comprising the soma and dendrites of a V1 layer 2/3 neuron expressing GCaMP6s. Dashed red lines indicate 13 dendritic ROIs from the same neuron. b, RFs calculated from calcium signals in the cell body and in the dendritic ROIs indicated in a. Numbers in the upper right corner of the dendritic RF maps indicate correlation with the somatic RF map. c, The frequency of dendrite ROIs as a function of the similarity of their RF with that of the soma (pixel-by-pixel RF map correlation). The majority of dendrites show similar RFs to that of the soma.

Extended Data Figure 5. Relationship between the physical distance of somata and the distance of their RFs

a, Example imaging region with layer 2/3 neurons expressing GCaMP6s. b, Median physical cell body distance of all cell pairs as a function of the distance in visual space of their RFs. Shading indicates 95% confidence interval. c, Likelihood of encountering cell pairs with overlapping (< 15 deg distance, red) and displaced (> 30 deg distance, blue) RFs for different physical cell body distances.

Extended Data Figure 6. Anatomical location of spines with retinotopically displaced RFs

a-d, Distance in visual space of the RFs of spines from that of the parent neuron as a function of the physical distance between spine and soma measured along the dendritic tree (a), of the dendritic branch order of the dendrite (b), of the depth of the soma beneath the cortical surface (c), and of the depth of the imaged dendrite (d).

Extended Data Figure 7. Retinotopic organization of visual inputs.

a, Position of colored dots indicates the cortical position of spines relative to the cell body on a plane parallel to the cortical surface. Dots are color-coded according to the spines’ RF position in visual field elevation (left) and visual field azimuth (right) relative to the parent neuron’s RF. Spines from all cells are combined, aligned to the cell body position shown by the black dot. Arrows indicate axes of cortical space that correlate best with changes in receptive field elevation (left) or azimuth (right). b, Relationship between RF distance in elevation (left) and azimuth (right) and cortical distance of spines and soma in the direction of the best fit as indicated by arrows in a. c, Relationship between RF distance in elevation (left) and azimuth (right) and cortical distance of dendrites and soma in the direction of the best fit as indicated by arrows in a, after averaging the position and RF elevation or RF azimuth of all spines on the same dendritic branch. M: medial, A: anterior. N = 32 dendrites, 15 mice (all dendrites for which the cell body position was recovered).

Extended Data Figure 8. Transformation of dendrite and spine RFs

Transformation of RFs of dendrites and their corresponding spines for pooling of all spine RFs in Figure 3b,c. a, To combine the position and orientation of all spine RFs relative to dendritic RFs in a common coordinate framework, we rotated the dendritic RFs such that their orientation was vertical and then translated them such that their centres were aligned at the same position. The parameters of this transformation were then used to transform the RFs of all spines to maintain the spatial relationship of their RF to that of their parent dendrite. b, RFs of two example dendrites and two of their corresponding spines before (top) and after transformation (bottom) as described in a. The visual space was defined as co-axial (green) or orthogonal (purple) relative to the centre and orientation of the dendrite RF.

Extended Data Figure 9. Control analysis for potential artefacts caused by global dendritic signals

a-e, Main analyses repeated after including only spines with responses not significantly correlated with the activity of their corresponding dendrite (see Methods, N = 522 spines, 26% of spines removed). a, corresponds to Figure 2e. b, corresponds to Figure 3d. c, corresponds to Figure 3e. d, corresponds to Figure 1g. e, corresponds to Figure 1h. f-i, Analyses of displaced spine RFs repeated after excluding all stimulus presentation trials in which the dendrite showed a calcium transient. f, Example RFs computed including all trials (top) or only trials in which the dendrite was not active (bottom). Numbers in the upper right corner indicate spatial RF correlation between the two RFs. g, Frequency of spines as a function of the similarity of their RF maps (spatial RF correlation) computed with and without trials with dendrite activity. h, corresponds to Figure 3d. i, corresponds to Figure 3e.

Figure 1. Dendritic clustering of synaptic inputs with similar receptive fields

a, Z-projection of a layer 2/3 neuron expressing GCaMP6s in mouse V1. b,c Schematic of receptive field (RF) mapping stimuli and a representative calcium signal (b) of the dendritic segment (c) indicated in a. d, Raw (top), smoothed (middle) and combined (bottom) ON and OFF RF subfield maps from calcium signals extracted from the ROI over the dendrite shown in c. a.u., arbitrary units. Color intensity denotes response strength to light increments (ON, red) and light decrements (OFF, blue). e, Spine calcium signals after removal of the dendritic component (top row), smoothed RFs (middle row), and orientation preference derived from the RFs (bottom row) of the example spines in c. f, The distribution of pairwise spatial RF correlation coefficients for all imaged spine pairs (N=3966 spine pairs, 74 dendrites, 21 mice). Triangle indicates median. Inset, example matrix of correlation coefficients of RFs from the spines in c and e. g,h, Relationship between the dendritic distance separating pairs of spines and their spatial RF correlation coefficients (g) and between spine-pair distances and the difference in their orientation preference (h, ΔOrientation). Shadings represent SEM. P-values from permutation test. Inset, the distribution of correlation coefficients between spine pair distance and spatial RF correlation (g), or difference in orientation preference (h) for individual dendrites. P-values from Wilcoxon signed-rank test, N= 3728 spine pairs, 39 dendrites, 18 mice.

Figure 2. Organisation of synaptic inputs from extended regions of visual space

a, Example RFs for two spine-dendrite pairs with either overlapping (top) or displaced RFs (bottom). Dashed lines indicate the dendrite RF Gabor fit outline. b, Distribution of distances in visual space between the RFs of spines and their corresponding dendrite. (N=62 dendrites, 21 mice). c, Mean spine - dendrite RF distance as a function of branch order of the imaged dendritic segment. Error bars represent SEM. d, Mean spine - dendrite RF distance as a function of the physical distance between spine and soma along the dendritic tree. e-f, The frequency of spines as a function of the difference between their preferred orientation (ΔOrientation) and that of the corresponding dendrite, for spine-dendrite pairs with retinotopically overlapping RFs (e), and for retinotopically displaced inputs (f). The numbers above bars indicate the number of spine - dendrite pairs. P-values are derived from permutation tests (see Methods).

Figure 3. Preferential synaptic input from neurons with co-oriented and co-axially aligned receptive fields.

a, Two example dendrite RFs with two retinotopically displaced spine RFs each. The visual field was divided into two sectors relative to the orientation of the dendrite RF: the co-axial visual space refers to the region around the orientation axis of the RF (< ± 45 deg) running through its centre, the orthogonal region occupies the remainder of visual space. b-c, Position in visual space and orientation difference relative to the dendrite RF of spines with displaced RFs located in co-axial (b, 97 spines) or orthogonal (c, 62 spines) visual space. Circles indicate individual spines. Colour denotes the difference in orientation preference (ΔOrientation) between the spine and dendrite. d-e, The frequency of spines with displaced RFs as a function of the difference in their preferred orientation from that of the corresponding dendrite for spines with RFs located in co-axial (d) or orthogonal (e) visual space. Schematics above illustrate the relationship between spine and dendrite RFs for each bin. Spine numbers are indicated above bars. P-values from permutation tests, N = 44 dendrites, 17 mice. f, Left: representative natural image. Green and purple squares represent co-axially and orthogonally displaced image sub-regions from a reference sub-region (red square). Right: probability of co-occurrence of features with similar orientations (ΔOrientation < 30 deg) in natural images for pairs of image features spatially displaced co-axially or orthogonally according to their orientation. The two distributions are significantly different for all displacements beyond 2 degrees (2 to 50 degrees, bin size 2 degrees, P< 0.01, Wilcoxon rank sum tests, Bonferroni correction for multiple comparisons).