Long-term usability and bio-integration of polyimide-based intra-neural stimulating electrodes.

Stimulation of peripheral nerves has transiently restored lost sensation and has the potential to alleviate motor deficits. However, incomplete characterization of the long-term usability and bio-integration of intra-neural implants has restricted their use for clinical applications. Here, we conducted a longitudinal assessment of the selectivity, stability, functionality, and biocompatibility of polyimide-based intra-neural implants that were inserted in the sciatic nerve of twenty-three healthy adult rats for up to six months. We found that the stimulation threshold and impedance of the electrodes increased moderately during the first four weeks after implantation, and then remained stable over the following five months. The time course of these adaptations correlated with the progressive development of a fibrotic capsule around the implants. The selectivity of the electrodes enabled the preferential recruitment of extensor and flexor muscles of the ankle. Despite the foreign body reaction, this selectivity remained stable over time. These functional properties supported the development of control algorithms that modulated the forces produced by ankle extensor and flexor muscles with high precision. The comprehensive characterization of the implant encapsulation revealed hyper-cellularity, increased microvascular density, Wallerian degeneration, and infiltration of macrophages within the endoneurial space early after implantation. Over time, the amount of macrophages markedly decreased, and a layer of multinucleated giant cells surrounded by a capsule of fibrotic tissue developed around the implant, causing an enlargement of the diameter of the nerve. However, the density of nerve fibers above and below the inserted implant remained unaffected. Upon removal of the implant, we did not detect alteration of skilled leg movements and only observed mild tissue reaction. Our study characterized the interplay between the development of foreign body responses and changes in the electrical properties of actively used intra-neural electrodes, highlighting functional stability of polyimide-based implants over more than six months. These results are essential for refining and validating these implants and open a realistic pathway for long-term clinical applications in humans.

[LAMINAR LOCATION OF NEURONS PROVIDING INTERHEMISPHERIC CONNECTIONS IN THE VISUAL CORTEX IN CATS WITH IMPAIRMENTS OF BINOCULAR VISION].

The distributio of cells in the visual cortical layers of intact cats (n=7) and cats with experimentally induced strabismus (n=10) and monocular deprivation (n=5) was studied after microiontophoretic injection of horseradish peroxidase into the ocular-dominance columns in areas 17, 18 and the transition zone 17/18. It was found that in cats with impaired binocular vision, the callosal cells were located deeper in layers of I/II, and higher - in layer IV, as compared to those in intact cats. Also in cats with impaired binocular vision, the proportion of callosal cells in layer IV was increased, while in layers II/III it was reduced as compared to intact cats. The most pronounced changes were noted in monocular deprived animals. These findings suggest an important role of sensory input in the formation of the callosal neurons layer distribution.

[The influence of strabismus and monocular deprivation on the size of callosal cells in cortical areas 17 and 18 of the cat].

To reveal the changes in visual cortex structure following impaired early binocular experience, the size (somatic area) of callosal cells in areas 17, 18 ofmonocularly deprived and convergent strabismic cats was measured. Horseradish peroxidase was injected into the single ocular dominance columns of areas 17, 18 and the transition zone 17/18. In both groups of impaired cats the mean size of callosal cells in area 17 was increased in comparison to intact cats. In area 18, the similar difference was found in monocularly deprived cats only. It was shown that the differences in the mean sizes of cells are due to the increase of the number of large cells. In strabismic cats, the portion of large cells (soma > 200 mkm2) in area 17 was 58% and in area 18 was 8%. The relative share of large cells in areas 17 and 18 of monocularly deprived cats was similar (28 and 26 % correspondingly). These data show that early binocular vision impairments may lead to the changes in cytoarchitecture of cortical layers where the interhemispheric connections originate.

[The size of cells providing interhemispheric and intrahemispheric connections in the visual cortex of binocular vision-impaired cats].

The size (somatic area) of 658 cells located in layers 2/3 of cortical areas 17, 18 of both hemispheres in intact monocularly deprived and bilateral strabismic cats was measured. These cells were retrogradely labelled after injections of horseradish peroxidase into ocular dominance columns in areas 17, 18. In all groups of cats, the mean somatic area of callosal cells was significantly larger than the mean somatic area of intrahemispheric cells. It was found that the mean somatic area of callosal cells was increased by 26.6% in monocularly deprived cats and by 20.2% in strabismic cats in relation to the mean somatic area of callosal cells in intact cats. In addition, the mean somatic area of intrahemispheric cells in monocularly deprived cats was indistinguishable from the mean somatic area of intrahemispheric cells in strabismic cats and in intact cats. It is concluded that early binocular vision impairments produce enlargement of callosal cells' size in the visual cortex.

Interhemisphere connections of eye dominance columns in the cat visual cortex in conditions of impaired binocular vision.

Data from studies of interhemisphere connections in fields 17 and 18 of cats reared in conditions of impaired binocular vision (monocular deprivation, uni- and bilateral strabismus) are presented. Monosynaptic connections between neurons were studied by microiontophoretic application of horseradish peroxidase into cortical eye dominance columns and the distributions of retrograde labeled callosal cells were analyzed. Spatial asymmetry and eye-specific interhemisphere neuron connections persisted in conditions of monocular deprivation and strabismus. Quantitative changes in connections were less marked in monocular deprivation than strabismus. In cats with impaired binocular vision, as in intact animals, the widths of callosal-receiving zones were greater than the widths of the callosal cell zones, which is evidence for the non-reciprocity of interhemisphere connections in cortical areas distant from the projection of the vertical meridian. Morphofunctional differences between cells mediating connections in the opposite directions are proposed.

Neuronal connections of eye-dominance columns in the cat cerebral cortex after monocular deprivation.

Plastic changes in intrahemisphere neuronal connections of the eye-dominance columns of cortical fields 17 and 18 were studied in monocularly deprived cats. The methodology consisted of microintophoretic administration of horseradish peroxidase into cortical columns and three-dimensional reconstruction of the areas of retrograde labeled cells. The eye dominance of columns was established, as were their coordinates in the projection of the visual field. In field 17, the horizontal connections of columns receiving inputs from the non-deprived eye via the crossed-over visual tracts were longer than the connections of the "non-crossed" columns of this eye and were longer than in normal conditions; the connections of the columns of the deprived eye were significantly reduced. Changes in the spatial organization of horizontal connections in field 17 were seen for the columns of the non-deprived eye (areas of labeled cells were rounder and the density of labeled cells in these areas were non-uniform). The longest horizontal connections in deprived cats were no longer than the lengths of these connections in cats with strabismus. It is suggested that the axon length of cells giving rise to the horizontal connections of cortical columns has a limit which is independent of visual stimulation during the critical period of development of the visual system.

[Interhemispheric connections of ocular-dominance columns in cats with binocular vision impairment].

We have investigated the interhemispheric connections of areas 17 and 18 in cats with impaired binocular vision (monocular deprivation, uni- and bilateral strabismus). Monosynaptic neuronal connections were studied using microionophoretic injections of horseradish peroxidase in the single cortical columns and analsys of spatial distribution of retrogradely labelled callosal cells was performed. In the cases of monocular deprivation and strabismus, the spatial asymmetry and eye-specificity of interhemispheric connections are retained. Quantitative changes of connections are more pronounced in strabismic cats. In cats with binocular vision impairments, as well as in control ones, the width of callosal-recipient zone is larger than of the callosal cells zone. This may indicate that interhemispheric connections are non-reciprocal in the areas of cortex that are more distant from the projection of vertical meridian of visual field. We expect that there should be morpho-functional in the cells that are providing connections in opposite directions.

[Neuronal connections of ocular dominance columns in the cortex of monocularly deprived cats].

We have investigated the developmental changes of intrahemispheric neuronal connections of the areas 17 & 18 ocular dominance columns in monocularly deprived cats. Single cortical columns were microiontophoretically injected with horseradish peroxidase and 3D reconstruction of retrogradely labelled cells' region was done. Ocular dominance of injected columns and their coordinates in the visual field map were determined. In area 17 it was shown that for non-deprived eye the connections of columns that are driven via the crossed pathways were longer than connections of columns driven via uncrossed ones, and in both cases they were longer than connections in intact cats. The connections of deprived eye columns are significantly reduced. We have observed some changes in the spatial organization of long-range connections in area 17 for columns driven by the non-deprived eye (more rounded shape of regions of labeled cells, non-uniform distribution of cells within it). Maximal length of such connections did not exceed the length of connections in strabismic cats. We speculate that the length of cell axons providing for the horizontal connections of cortical columns has some intrinsic limit that does not depend on visual stimulation during the critical period of development.

Interhemisphere connections of the visual cortex in cats with bilateral strabismus.

The distribution of retrograde labeled callosal cells after microiontophoretic application of horseradish peroxidase into individual cortical columns in fields 17 and 18 was studied in cats reared with bilateral strabismus (with an angle of eye deviation of 10-35 degrees ). The area containing labeled cells was located asymmetrically in relation to the position of the injected column in the opposite hemisphere. Some of the cells were located in those parts of the transitional zone between fields 17 and 18 whose retinotopic coordinates corresponded to the column coordinates (as in intact cats). Other labeled cells were located in fields 17 and 18 and were grouped into clusters located at distances of about 1000 microm from the marginal clusters of the transitional zone. The locations of labeled cells in the lateral geniculate body showed that most columns receive inputs from the ipsilateral eye. Evidence for eye specificity of these monosynaptic interhemisphere connections is presented. The functional significance of changes in these connections in bilateral strabismus is discussed.

Changes in the structure of neuronal connections in the visual cortex of cats with experimentally induced bilateral strabismus.

The spatial distribution of neuronal connections in cortical field 17 was studied in cats with experimentally induced bilateral convergent strabismus on postnatal days 10-14. Horseradish peroxidase was applied microiontophoretically to individual columns of neurons in fields 17 and 18 and retrograde-labeled cells were identified in both hemispheres. Increases and decreases in the extent of intra-hemisphere connections were seen in the mediolateral direction (projections of the horizontal meridian of the visual field). Most columns showed increases in inter-hemisphere connections in this same direction, which may support the more reliable unification of the two visual hemifields. In addition, some columns showed increases in intra-and inter-hemisphere connections in the rostrocaudal direction (projections of the vertical meridian). Thus, bilateral strabismus induced during the critical period of development leads to changes in the structure of both intra-hemisphere and inter-hemisphere connections of individual cortical columns in fields 17 and 18.

Eye-rotation-induced spatial reorganization of horizontal connections in field 17 of the cat cortex.

Six cats with rotation of one or both eyes (strabismus) produced surgically in the early postnatal period demonstrated torsional deviation of the eyes by 10-20 degrees in addition to the rotation. The spatial distribution of retrograde labeled neurons in field 17 was studied by microiontophoretic administration of horseradish peroxidase into individual cortical columns in fields 17 and 18. These studies showed that rotation of the eyes increased the extent of horizontal neuronal connections in field 17 along the projection of the vertical meridian of the field of vision. It is suggested that this reorganization of neuronal connections may support functional changes compensating for eye rotation, as described in the literature.

[Interhemispheric connections in the visual cortex of cats reared with bilateral strabismus].

We have determined the spatial distribution of retrograde labelled callosal cells after microiontophoretic horseradish peroxidase injections into the single cortical columns of area 17, 18 in cats reared with bilateral convergent strabismus. The obtained strabismus angle was in the range 10-35 degrees. The zone of labelled cells was located asymmetrically in respect to location of injected column in opposite hemisphere. Some cells were revealed in the transition zone 17/18 and their retinotopic coordinates corresponded to the injected column, as was shown in intact cats. Other labelled cells were located in areas 17, 18, in clusters approximately in 1000 mkm from marginal clusters of transition zone. Analysis of labeling in lateral geniculate nucleus has shown that most of the injected columns were driven by ipsilateral eye. The data obtained may be interpreted as evidence of eye-specificity of monosynaptic callosal connections. The functional role in such connections changes in cats with bilateral strabismus is discussed.

[Spatial reorganization of horizontal connections of cat cortical area 17 induced by eye rotation].

In six cats with experimental unilateral or bilateral strabismus, surgically induced early in postnatal life, torsion eye rotation with a deviation angle of 10 to 20 degrees was also detected. Spatial distribution of retrogradely labeled neurons in area 17 was studied following microiontophoretic injection of horseradish peroxidase in area 17 or 18 cortical columns. Eye rotation was shown to cause the increase of the length of horizontal neuronal connections in area 17 along the projection of visual field vertical meridian. The reorganization of neuronal connections, detected in this work, may promote the functional changes, described in the literature, that compensate for the eye rotation.

[Changes of the structure of neuronal connections in visual cortex of cats with experimentally induced bilateral strabismus].

The spatial distribution of neuronal connections in cortical area 17 in cats with bilateral convergent strabismus, surgically induced on postnatal days 10-14, was investigated. Single cortical columns of areas 17 and 18 were microiontophoretically injected with horseradish peroxidase and retrogradely labeled cells in both hemispheres were detected. The intrahemispheric connections in area 17 along the mediolateral direction (the projection of visual field horizontal meridian) were found to be either dilated or reduced. For most columns, the interhemispheric connections in this direction were increased, which may provide for more reliable unification of two visual hemifields. Moreover, the extension of intra- and interhemispheric connections in rostrocaudal direction (along the visual field vertical meridian projection) was detected in some cortical columns. Thus bilateral strabismus induced during the developmental critical period, modifies the structure of both of intra- and inter-hemispheric spatial neuronal connections of individual cortical columns in areas 17 and 18.

[Interhemispheric connections of areas 17 and 18 in the cortical columns of cats with unilateral strabismus]. / Mezhpolusharnye sviazi korkovykh kolonok polei 17 i 18 u koshek s unilateral'nym kosoglaziem.

The interhemispheric connections of areas 17 and 18 of the cerebral cortex were investigated in cats with experimental unilateral strabismus. Single cortical columns were microiontophoretically injected with horseradish peroxidase, and retrogradelly labeled cells were demonstrated in the opposite brain hemisphere. After tracer injection in area 18 columns, the labeled callosal cells were located in area 17/18 transitional zone, similar to what was found in normal cats. However, in some cases the expansion of the region of callosally labeled cells distribution, was found. It is proposed that the extent of the region of callosally-connected cells may vary depending on whether the cells receive their input from intact or strabismic eye.