Blockade of retinal or cortical activity does not prevent the development of callosal patches normally associated with ocular dominance columns in primary visual cortex

Abstract

Callosal patches in primary visual cortex of Long Evans rats, normally associated with ocular dominance columns, emerge by postnatal day 10 (P10), but they do not form in rats monocularly enucleated a few days before P10. We investigated whether we could replicate the results of monocular enucleation by using tetrodotoxin (TTX) to block neural activity in one eye, or in primary visual cortex. Animals received daily intravitreal (P6-P9) or intracortical (P7-P9) injections of TTX, and our physiological evaluation of the efficacy of these injections indicated that the blockade induced by a single injection lasted at least 24 h. Four weeks later, the patterns of callosal connections in one hemisphere were revealed after multiple injections of horseradish peroxidase in the other hemisphere. We found that in rats receiving either intravitreal or cortical injections of TTX, the patterns of callosal patches analyzed in tangential sections from the flattened cortex were not significantly different from the pattern in normal rats. Our findings, therefore, suggest that the effects of monocular enucleation on the distribution of callosal connections are not due to the resulting imbalance of afferent ganglion cell activity, and that factors other than neural activity are likely involved.

Keywords: Long Evans rats; columnar organization; interhemispheric connections; segregation; tetrodotoxin.

Fig. 1.

Assessment of the effectiveness of intravitreal and intracortical injections of TTX. (A) Intravitreal injections of TTX. Upper panel illustrates activity recorded before the TTX injection. It shows bursts of activity correlated with periods when the light stimulus was turned on (horizontal line segment underneath activity trace). Lower panel shows that 15 min after the TTX injection, only spontaneous activity uncorrelated to the light stimuli was recorded. (B) Intracortical TTX injection. The arrow points to the location of the TTX injection (asterisk), located 0.5 mm anterior to the lambda suture, and 3.5 mm lateral to the brain midline. The black contour corresponds to the border of V1. Lateral to the right, posterior is down. The numbers indicate eight recording sites separated by about 1.0 mm from each other. Scale bar = 1.0 mm. Activity was recorded by stimulating the contralateral eye. (C) Activity recorded before the intracortical TTX injection. The tracings show activity recorded at the sites indicated. (D) Activity recorded 3 h after the injection. The tracings show recordings at the sites indicated. Line segments under the recordings indicate light on.

Fig. 2.

(A) Pattern of HRP-labeled callosal connections in normal adult rat (modified from Olavarria et al., 2021). The thin line delineates the central segment of V1 and illustrates the region used for analysis in all cases. This region does not include the band of callosal labeling at the 17/18a border. (B,C) Patterns of callosal connections ipsilateral to the remaining eye (B) and ipsilateral to the enucleation (C) in rats monocularly enucleated at P8. (D,E) Patterns of callosal connections in the hemisphere ipsilateral to the intravitreal injection of TTX. (F,G) Patterns of callosal connections in the hemisphere contralateral to the intravitreal injection of TTX. (H,I) Patterns of callosal connections in hemispheres injected with TTX. In A–I, note that callosal connections segregate into distinct patches in the central segment in all cases. Lateral is to the right, posterior is down. (J) Magnified view of cells in V1 labeled retrogradely with HRP in the case shown in (B). (K) HRP labeling pattern in the LGN of a normal adult rat showing ipsilateral (left) and contralateral (right) eye projections (modified from Olavarria et al., 2021). (L) HRP labeling in the LGN ipsilateral (left) and contralateral (right) to the intravitreal injections of TTX followed by injections of HRP. Arrows in K and L indicate dorsomedial region of the LGN ipsilateral to the HRP injections that remains unlabeled in both normal and TTX injected rats, suggesting that the intravitreal injection of TTX did not produce an expansion of the ipsilateral retino-geniculate projection into this region, as it occurs for the ipsilateral projection from the remaining eye in rats monocularly enucleated at P7 (Olavarria et al., 2021). Scale bar in J = 100 μm, other scale bars = 1.0 mm.

Fig. 3.

(A) Comparison of patch indices in the central segment in MEP8, normal adult, in the hemisphere ipsilateral (Ipsi TTX) and contralateral (Contra TTX) to the intravitreal injection of TTX, and in the hemisphere injected with TTX (Cortex TTX). (B) Comparison of the %V1 occupied by callosal patches in the central segment in MEP8, normal adult, in the hemisphere ipsilateral (Ipsi TTX) and contralateral (Contra TTX) to the intravitreal injection of TTX, and in the hemisphere injected with TTX (Cortex TTX). Numbers above the bars indicate number of animals in each group. Error bars indicate s.e.m. MEP8, monocular enucleation at P8.