Distribution of non-phosphorylated neurofilament in squirrel monkey V1 is complementary to the pattern of cytochrome-oxidase blobs

Abstract

The geniculo-recipient zones of the primate primary visual cortex (V1) stain more strongly for cytochrome oxidase (CO) than other regions. Labeling V1 with an antibody (SMI-32) against neurofilament protein produces a laminar pattern that is largely complementary to that of CO: the layers that receive the strongest geniculate input react weakly for SMI-32. We evaluated whether the complementary laminar relationship extends throughout the superficial layers where there are regularly spaced blobs of dark CO staining that are known to receive geniculate input. In all hemispheres, neurofilament labeling in the superficial layers was indeed complementary to the CO pattern. The density of SMI-32 labeled neurons was quantified and found to be greater within the CO interblobs than in the blobs. These results demonstrate that blobs and interblobs can be distinguished by examining the pattern of neurofilament expression in V1. That neurofilament expression is highest within interblobs raises the possibility that the distribution of cell types may be non-uniform across blobs and interblobs.

Figure 1

Adjacent coronal sections of squirrel monkey V1 labeled for CO (A) and SMI-32 (B). As demonstrated in previous studies, the layers richest in CO (IVA, IVCα and IVCβ) were weakly labeled for SMI-32. Scale bar = 500 μm.

Figure 2

Tangentially cut sections of V1 labeled for CO (A) and SMI-32 (B). Adjacent sections were aligned using the pattern of radial blood vessels. CO blobs were identified in an automated way and contours were drawn around the border of each blob (C). The blob contours were then superimposed onto the section labeled for SMI-32. The heaviest SMI-32 labeling was found outside blob centers (D). Each SMI-32 positive cell was identified under a microscope and a dot was placed at its location in the adjacent CO section (E). The distribution of blob contours and SMI-32 positive cells was then examined (F). This analysis revealed a tendency for SMI-32 positive neurons to be located within interblobs. Scale bar = 500 μm.

Figure 3

High magnification photographs of SMI-32 immunoreactivity (A and C) and CO staining (B and D). The pattern of SMI-32 was comprised of dark neuronal somata and dendritic processes, as well as apical dendrites from deeper neurons that created a punctate pattern outside of blobs (arrows in B). At this magnification, individual SMI-32 positive cells could be seen to lie primarily outside of blobs. Scale bars = 250 μm (A and B) and 100 μm (C and D).

Figure 4

Another example of the relationship between CO blobs (A) and SMI-32 labeling (B) in V1 from a different monkey. The regions defined by contours generated around CO blobs (C) were within regions that were light for SMI-32 labeling (D). Individual SMI-32 positive neurons were plotted as black dots superimposed onto the adjacent CO section (E). Comparison of these two patterns shows that SMI-32 labeled cells fall preferentially within inter-blob regions (F). Scale bar = 500 μm.

Figure 5

A sequential series of sections reacted for CO (left column) or SMI-32 (right column). The sections start at the top of layer I (row A), move through layers II/III (rows B, C and D) and end in layer IVA (row F). SMI-32 labeling was apparent in the most superficial sections examined and consisted of apical dendrite shafts (row A). Sections taken from layers II/III (rows B, C and D) revealed a distribution of SMI-32 labeled cells that clustered outside of CO blobs. Sections sampled from deep layer III and layer IVA show weak SMI-32 labeling with the distribution of cells not showing a strong correlation with the CO pattern (E and F). Scale bar = 200 μm.

Figure 6

Quantification of the spatial relationship between SMI-32 labeling and CO blobs. The distribution of SMI-32 labeled cells was plotted from a 5 mm² area of V1 for each of the five hemispheres we examined. In this graph, the density of labeled cells inside of blobs and interblobs is plotted. The density of SMI-32 positive neurons is significantly greater within interblobs (P < 0.001; t-test). Error bars represent the standard error of the mean.