The shrinkage of ocular dominance columns following early visual deprivation provides a striking example of how neuronal connections in the brain can be altered by abnormal sensory experience (Hubel et al., 1977
; Wiesel, 1982
). Since this landmark discovery the field has made only modest advances in describing the anatomical changes that occur in amblyopia. It is clear that major physiological deficits are present beyond V1, in extrastriate areas such as V2 and MT (Jones et al., 1984
; Imamura et al., 1997
; El-Shamayleh et al., 2010
; Hess et al., 2010
; Bi et al., 2011
). It is possible that these abnormalities are due entirely to loss of geniculocortical synapses in layer 4C serving the deprived eye. It seems more likely, however, that synaptic changes at multiple levels, from the lateral geniculate body to higher association areas, combine in series to impair visual function (Kiorpes, 2006
In monkeys raised with eyelid suture, about 20% of cells outside layer 4C still respond to visual stimulation via the deprived eye. To early investigators, these units seemed to have normal orientation selectivity (Hubel et al., 1977
; Blakemore et al., 1978
). However, optical imaging in kittens has shown convincingly that orientation tuning is degraded by monocular deprivation (Kim and Bonhoeffer, 1994
; Crair et al., 1997
). It is unknown how projections from layer 4C to other layers are affected by form deprivation. A precise description of intracortical wiring abnormalities might explain why orientation selectivity is affected, and shed further light on the anatomical changes responsible for amblyopia. Unfortunately, this objective is not attainable with currently available neuroanatomical methods. Probing intracortical connections by extracellular injection of retrograde tracers, even into single layers, is fraught by uptake into dendrites, fibers of passage, and local axon collaterals (Fitzpatrick et al., 1985
; Lachica et al., 1992
). These problems make it difficult to explore vertical cortical projections, although this approach has been used for horizontal projections in strabismic monkeys to demonstrate selective loss of connections between cells in ocular dominance columns serving opposite eyes (Tychsen et al., 2004
). In the future, retrograde viral tracers that cross several synapses may facilitate the examination of wiring abnormalities in amblyopia (DeFalco et al., 2001
; Campbell and Herbison, 2007
; Callaway, 2008
). A powerful experiment would be to infect a single cell in a layer such as 4B, and to compare the number of labeled cells in each set of layer 4C ocular dominance columns, the laminae of the lateral geniculate body, and even the two eyes.
We made retrograde tracer injections in area V2 to test for intercortical wiring anomalies in amblyopia. This approach had the advantage of cleanly labeling projection neurons from V1, to compare their numbers in ocular dominance columns serving either the normal eye or the deprived eye. Tracer injections into V2 always label neurons in both sets of ocular dominance columns (Sincich and Horton, 2005
; Sincich et al., 2010
). Labeling is never confined to one eye’s columns, no matter how small the tracer injection. This observation means that V1 axon terminals preferentially serving the right eye and the left eye are intermingled intimately in V2. This arrangement should maximize the potential for competition between the eyes, resulting in selective loss of V1 connections serving the deprived eye, according to Hebbian principles (Butz et al., 2009
In fact, we found little evidence for such an effect. The patches corresponding to the deprived eye stained lightly for CO (). This finding indicated that their cells were less active physiologically. Nonetheless, the projection neurons in deprived CO patches did not fail to label after tracer injection into thin stripes. In fact, analysis showed that the clusters of labeled projection neurons had the same density in normal columns and deprived columns (). In terms of absolute numbers, only 33% of cells projecting to thin stripes were located in deprived columns. This reduction was commensurate with the amount of lost cortical territory. The effect of column shrinkage was not amplified further by selective dropout of cell projections from deprived patches to V2. One might wonder why shrinkage of ocular dominance columns caused any loss of projection neurons to thin stripes, given that patches are located in the center of ocular dominance columns. The reason is that about 20% of neurons that project to thin stripes are actually located in interpatch zones. In addition, neurons are exchanged along bridges between patches in adjacent ocular dominance columns ().
After pale or thick stripe injection, only 23% of projection neurons were located in deprived columns and their density was reduced by 24% (). A model of the impact of visual deprivation showed that these changes could be accounted for by column shrinkage, without postulating selective dropout of projection neurons in deprived columns ().
Of course, connections to V2 might be lost without any reduction in the number of retrogradely labeled cells in V1. The geniculocortical projection provides an instructive precedent: visual deprivation causes shrinkage of axon terminals in layer 4C but no loss of cells in the lateral geniculate body. The only sign of visual deprivation at the level of the geniculate body is a change in cell size (Wiesel and Hubel, 1963a
; Guillery and Stelzner, 1970
; Headon et al., 1985
). This motivated us to measure the size of projection neurons in normal columns and deprived columns. There was no difference in either layer 4B or 2/3. This suggests that the eyes’ projection neurons supported axon terminal arbors of equal size in V2. This conclusion was supported by the observation that deprived eye patches were labeled as intensely as normal eye patches (). If their projection neurons had less extensive axon terminal arbors, one would expect them to imbibe less retrograde tracer at injection sites. Had this occurred, deprived patches would have appeared fainter in comparison with normal patches. Ultimately, the definitive way to approach this problem would be to label the axons of individual cells projecting to V2, and reconstruct their terminal arbors (LeVay and Ferster, 1977
; Rockland and Virga, 1990
; Friedlander et al., 1991
). By comparing a large inventory of axons emanating from normal columns and deprived columns, one could determine whether visual deprivation has any impact on axon terminal morphology or synapse number.
The projection from V1 to V2 is adult-like in macaques by age one week, and probably even at birth (Kiorpes et al., 2003
). The lack of any obvious impact from eyelid suture, even at age 8 days, suggests that either this projection is impervious to the effects of sensory deprivation or its critical period comes at an earlier age. The latter possibility seems unlikely, because it would push the critical period into fetal life. Perhaps structural changes in the V1 to V2 projection are induced by monocular deprivation, but they are too subtle to detect with our experimental approach. For example, synaptic connections in V2 could be corrupted in strength, location, specificity, and chemistry by visual deprivation, without necessarily reducing the uptake of a retrograde tracer. These possibilities aside, it is clear that dramatic contraction of afferents, as seen in the geniculate projection to layer 4C, does not occur. The shrinkage of geniculate arbors in layer 4C may represent an exceptional response to sensory deprivation, rather than a common phenomenon throughout the nervous system.
As mentioned previously, column shrinkage involves surrender of more interpatch tissue than patch tissue. Therefore, in deprived columns, compared with normal columns, a greater proportion of cells projects to thin stripes than to pale or thick stripes. It is interesting to speculate what consequences this difference might have for visual function in amblyopia. There is evidence that patches are specialized for the processing of color information (Livingstone and Hubel, 1984
; Lu and Roe, 2008
). Relative sparing of patches could explain why color perception seems to be impaired less severely than spatial resolution in patients with amblyopia (Mangelschots et al., 1996
; Almog and Nemet, 2010
). However, there is some doubt that color and form are handled by largely segregated cell populations in V1, implying that color processing may not be limited to patches (Gegenfurtner and Kiper, 2003
; Shapley and Hawken, 2011
). Moreover, at low spatial frequencies, color and luminance discrimination are similarly affected in most amblyopes (Bradley et al., 1986
; Mullen et al., 1996
). Finally, it is worth recalling that in most forms of amblyopia there is no shrinkage of ocular dominance columns (Horton and Stryker, 1993
; Horton and Hocking, 1996b
). Therefore, in the majority of patients, amblyopia does not have a greater impact on interpatch projections to V2.
We have examined the impact of early form deprivation – which produces the most severe type of amblyopia – on the projections between V1 and V2. To our surprise, the effect predicted from the shrinkage of ocular dominance columns in V1 was not found. There was no evidence for loss or atrophy of V2 projection neurons in deprived ocular dominance columns. Instead, the changes in projection neuron density were commensurate with column shrinkage. This finding is encouraging from a clinical standpoint. If one could devise a treatment that restored a normal geniculocortical projection in layer 4C, the downstream pathways at the disposal of the deprived eye might still be quite functional.