Neuronal projections from V1 to V2 in amblyopia

Abstract

The mechanism of amblyopia in children with congenital cataract is not understood fully, but studies in macaques have shown that geniculate synapses are lost in striate cortex (V1). To search for other projection abnormalities in amblyopia, the pathway from V1 to V2 was examined using a triple-label technique in three animals raised with monocular suture. [(3)H]proline was injected into one eye to label the ocular dominance columns. Cholera toxin B subunit conjugated to gold (CTB-Au) was injected into V2 to label V1 projection neurons. Alternate sections were processed for cytochrome oxidase (CO) and CTB-Au, or dipped for autoradiography. Eight fields of CTB-Au-labeled cells in V1 opposite injection sites were plotted in layers 2/3 or 4B. After thin stripe injection, labeled cells were concentrated in CO patches. Despite column shrinkage, cells in deprived and normal columns were equal in size and density in both layers 2/3 and 4B. After pale or thick stripe injection, labeled cells were concentrated in interpatches. Only 23% of projection neurons originated from deprived columns. This reduction exceeded the degree of column shrinkage, a result explained by the fact that column shrinkage causes disproportionate loss of interpatch territory. These data indicate that early monocular form deprivation does not alter the segregation of patch and interpatch pathways to V2 stripes or cause selective loss or atrophy of V1 projection neurons. The effect of shrinkage of geniculocortical afferents in layer 4C following visual deprivation is not amplified further by attenuation of the amblyopic eye's projections from V1 to V2.

Figure 1.

Projections from V1 to V2 in amblyopia. Visual deprivation of the right eye causes shrinkage of ocular dominance (OD) columns and reduction in the size of CO patches. Although the CO patches are smaller, they occupy a greater percentage of the deprived eye's columns. The appearance of the thin, pale, and thick stripes in V2 remains normal in amblyopia. In this study, the layer 3 projections were compared from normal eye columns (blue arrows) and deprived eye columns (red arrows) to V2 stripes, as well as those from layer 4B (not illustrated).

Figure 2.

Shrinkage of ocular dominance columns. Autoradiograph of a single tangential section cut through a flatmount of the right striate cortex from a monkey that underwent right eye suture at age 20 d. [³H]proline was injected into the right eye when the animal was adult to label the ocular dominance columns in layer 4C. They appear bright in this darkfield image, and are shrunken relative to the unlabeled ocular dominance columns serving the normal left eye. The CTB-Au injections (arrows) in V2 are visible near the dorsal V1 border of the operculum, because the D-19 developer produced some silver enhancement. Rectangle is shown at higher magnification in Figure 5a. mc, Monocular crescent. The asterisk represents the left eye's blind spot.

Figure 3.

CTB-Au tracer injection into a thin stripe. Layer 3 CO section after silver enhancement, showing four CTB-Au injections in V2, close to the V1 border. The rectangle indicates the field of retrogradely labeled V1 cells analyzed in Figures 4 and 5. The inset at upper right shows the CTB-Au injection site for this field before silver enhancement. The injection is small and well centered in a thin stripe. The white arrows denote rows of small, pale CO patches, aligned with the deprived eye's ocular dominance columns in layer 4C. Black arrows show thin stripes, brackets show thick stripes.

Figure 4.

Thin stripe CTB-Au injection labels cells in patches. a, Darkfield image showing clusters of retrogradely filled cells. The field is enclosed by the rectangle in Figure 3. b, Identical section in brightfield, showing the CO patches in layer 3, coinciding with the clusters of labeled cells in a. c, Density contours, superimposed on b, to divide CO activity into six zones of equal area. d, Plot of CTB-Au-filled cells in a, showing their relationship to the CO patches, which correspond to the darkest two zones (33%) in c.

Figure 5.

Identification of normal eye and deprived eye patches. a, Ocular dominance columns from the box in Figure 2, at higher magnification. b, Boundaries of the ocular dominance columns in a, with the cells in Figure 4d plotted on the CO section in Figure 4b. c, Cells in b, plotted in blue for normal eye columns and in red for deprived eye columns. The boundary surrounding the cells was used to define areas for cell density measurements (see Materials and Methods).

Figure 6.

Comparison of layer 3 projection neuron soma size. Examples of two fields, from a normal eye column and a deprived eye column, where cell soma areas were measured by tracing outlines of CTB-Au-filled cells at 1000×.

Figure 7.

Histograms of soma size. Plot showing cell body areas for projection neurons sampled in patches serving the normal eye and the deprived eye. There was no significant difference between the two populations (p > 0.05).

Figure 8.

Layer 4B labeling. a, Darkfield image of cell labeling in layer 4B from the same region shown in Figures 4 and 5. b, Plot of CTB-filled cells, showing their relationship to the patches in layer 3 defined by red contour lines. c, Cells in b, plotted with respect to the ocular dominance columns (blue, open eye; red, deprived eye).

Figure 9.

Effect of column shrinkage on projection cell density after tracer injection into a pale or thick stripe. In this simulation, each blue dot represents a cell, with 84% distributed randomly in the interpatches. The oval patches run down the middle of the ocular dominance columns, with the open eye's columns (gray stripes) expanded to fill 70% of the cortex. The density of projection neurons is 23% lower in the deprived columns (white stripes), simply because they have lost interpatch tissue.