Very-long-range disynaptic V1 connections through layer 6 pyramidal neurons revealed by transneuronal tracing with rabies virus

Figures & data

Figure 1 Rabies virus injections in macaque monkey primary visual cortex.

Notes: (A) Locations of the V1 injection sites (white dots within the black oval) are shown on a drawing of the dorsal lateral view of the right hemisphere of the adult male macaque monkey. The approximate location of the V1/V2 border is indicated by the curved dashed line just posterior to the LuS. (B) An image from a representative brain shows a dorsal view of the posterior end of cortex. The representations of the HM and VM and iso-eccentricities of 1°, 5°, and 10° are based on Lyon and Kaas,⁸³ and were derived from retinotopic maps provided in Van Essen et al⁵⁹ and Weller and Kaas.⁶⁰ The digital image in (C) shows one of the V1 injection sites. The image was taken from a coronal section through the plane indicated by the vertical dashed line in (A). Rabies-infected neurons are stained black from immunohistochemistry of the rabies nucleocapsid protein. The arrowhead points to a perturbation on the cortical surface caused by insertion of the injection pipette. The diagonal dashed line estimates the path of the injection pipette. Cortical layers were determined from cytochrome oxidase staining patterns (see also [D]), and are indicated at the bottom of the image. (D) A digital image of a cytochrome oxidase-stained coronal section shows the V1 lamination in greater detail. Gray text to the left shows the terminology of Hassler,⁸⁵ as an alternative to the terminology of Brodmann,⁵⁶ which is shown in black. The scale bars in (A) and (B) are 5 mm. The scale bar in (D) is 1 mm and also applies to (C).
Abbreviations: CalcS, calcarine sulcus; CS, central sulcus; HM, horizontal meridian; IOS, inferior occipital sulcus; IPS, intraparietal sulcus; LatS, lateral sulcus; LuS, lunate sulcus; STS, superior temporal sulcus; V1, primary visual cortex; V2, secondary visual cortex; VM, vertical meridian; WM, white matter.

Figure 2 The distribution of rabies-infected neurons in two coronal sections of V1 of the right hemisphere.

Notes: (A) A reconstruction of section 97 shows portions of two rabies virus injection sites (black arrowheads). Neurons retrogradely labeled from these injections are shown as small black dots or large gray triangles depending on whether or not they were larger than 30 μm in diameter. The parallel blue lines represent the upper and lower border of layer 4A (Hassler’s layer 3B⁸⁴); parallel red lines represent the upper and lower border of layer 4C (Hassler’s layer 4⁸⁴); and the green line represents the layer 5/6 border. The thick black line indicates the cortical surface, and the thin black line represents the border between white matter and layer 6. The digital image in (B) shows layers 4C-2/3 and is from the region in (A) outlined by the yellow rectangle. The digital image in (C) shows layers 5 and 6 and is from the region in (A) outlined by the orange rectangle. (D) A reconstruction of section 121 shows portions of two rabies virus injection sites indicated by black arrowheads. The digital image in (E) shows rabies-infected neurons throughout layers 2/3–6, and is from the inset region outlined in (D) by the orange rectangle. Enlarged views of the layer 4B and layer 2/3 regions outlined by left and right black squares, respectively, in (E) are shown in (F) and (G). The scale bar in (A) is 2 mm and also applies to (D). The scale bar in (E) is 1 mm. Scale bars in (B) and (G) correspond to 100 µm and also apply to (C) and (F).
Abbreviations: CalcS, calcarine sulcus; V1, primary visual cortex; WM, white matter.

Figure 3 The distribution of rabies-infected neurons in a single cross-section of V1 of the right hemisphere.

Notes: (A) A reconstruction of section 109 shows portions of three rabies virus injection sites (black arrowheads). Neurons retrogradely labeled from these injections are shown as small black dots or large gray triangles. The two solid black lines adjacent to the medial and lateral injection sites represent the starting point (0 mm) for the calculations of all connected neurons along the opercular surface, laterally, and the medial wall, medially. Distances of up to 18 mm are indicated. The digital image in (B) shows rabies-infected neurons in layers 2/3–6 in V1 and is from the inset region outlined in (A) by the black rectangle. An enlarged view of a cluster of small layer 4A and 4B rabies-infected neurons is shown in (C), and is taken from the area outlined by the left square in (B); An enlarged view of a cluster of large layer 6 neurons, all with cell body diameters >30 μm, is shown in (D), and is taken from the area outlined by the right square in (B). The scale bars in (A), (B), and (D) are 2 mm, 1 mm, and 50 µm, respectively. The scale bar in (D) also applies to (C).
Abbreviations: V1, primary visual cortex; WM, white matter.

Figure 4 Serial reconstructions show the pattern of mono- and disynaptically connected neurons in visual cortex following rabies virus injections in V1.

Notes: Ten coronal sections are shown in order by rows, ranging from the most posterior (top left section) to the most anterior (bottom right section). Section numbers are listed to the lower left of each section and range from section 97 to section 371. Retrogradely infected neurons (black dots and gray triangles) were found in striate (V1) and extrastriate visual cortex. Red parallel lines representing layer 4C are used to delineate V1. Black dots = small neurons and gray triangles = large neurons. The relevant sulci are given in gray lettering. The scale bar shown for section 97 equals 2 mm and applies to all sections.
Abbreviations: CalcS, calcarine sulcus; IOS, inferior occipital sulcus; IPS, intraparietal sulcus; LuS, lunate sulcus; STS, superior temporal sulcus; V1, primary visual cortex.

Figure 5 The numbers of retrogradely infected neurons found across several millimeters of the opercular surface of primary visual cortex.

Note: Small and large cells are graphed together in the left-hand panels (A–F), whereas large cells only are graphed in the right-hand panels (G–L).

Figure 6 The numbers of retrogradely infected V1 neurons present in the calcarine sulcus.

Note: The proportions of large cells, which were present in layer 4B (5%) and layer 6 (26%), are indicated by gray shading.
Abbreviation: VI, primary visual cortex.

Figure 7 The region of visual space covered by the very-long-range disynaptic connections is shown on an unfolded, two-dimensional view of the V1 surface.

Notes: Iso-eccentricity contours from 2.5° to 80° and the HM are shown, and are based on the summary of V1 retinotopy provided in Van Essen et al.⁵⁹ The light gray oval represents the injection sites. The dark gray shading represents the spread of rabies-labeled neurons found for layer 6. The spread for layer 5 and for layers superficial to layer 4C were less extensive and are not shown. Upper and lower visual fields are indicated by + and −, respectively. The scale bar is 1 cm.
Abbreviations: HM, horizontal meridian; V1, primary visual cortex.

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