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Neural pathfinding on uni- and multidirectional photopolymerized micropatterns.

Tuft BW, Xu L, White SP, Seline AE, Erwood AM, Hansen MR, Guymon CA - ACS Appl Mater Interfaces (2014)

Bottom Line: SGN neurites orient randomly on unpatterned photopolymer controls, align and consistently track unidirectional patterns, and are substantially influenced by, but do not consistently track, multidirectional turning cues.Neurite lengths are 20% shorter on multidirectional substrates compared to unidirectional patterns while neurite branching and microfeature crossing events are significantly higher.Developing methods to understand neural pathfinding and to guide de novo neurite growth to specific stimulatory elements will enable design of innovative biomaterials that improve functional outcomes of devices that interface with the nervous system.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical and Biochemical Engineering, University of Iowa , Iowa City, Iowa 52242, United States, United States.

ABSTRACT
Overcoming signal resolution barriers of neural prostheses, such as the commercially available cochlear impant (CI) or the developing retinal implant, will likely require spatial control of regenerative neural elements. To rationally design materials that direct nerve growth, it is first necessary to determine pathfinding behavior of de novo neurite growth from prosthesis-relevant cells such as spiral ganglion neurons (SGNs) in the inner ear. Accordingly, in this work, repeating 90° turns were fabricated as multidirectional micropatterns to determine SGN neurite turning capability and pathfinding. Unidirectional micropatterns and unpatterned substrates are used as comparisons. Spiral ganglion Schwann cell alignment (SGSC) is also examined on each surface type. Micropatterns are fabricated using the spatial reaction control inherent to photopolymerization with photomasks that have either parallel line spacing gratings for unidirectional patterns or repeating 90° angle steps for multidirectional patterns. Feature depth is controlled by modulating UV exposure time by shuttering the light source at given time increments. Substrate topography is characterized by white light interferometry and scanning electron microscopy (SEM). Both pattern types exhibit features that are 25 μm in width and 7.4 ± 0.7 μm in depth. SGN neurites orient randomly on unpatterned photopolymer controls, align and consistently track unidirectional patterns, and are substantially influenced by, but do not consistently track, multidirectional turning cues. Neurite lengths are 20% shorter on multidirectional substrates compared to unidirectional patterns while neurite branching and microfeature crossing events are significantly higher. For both pattern types, the majority of the neurite length is located in depressed surface features. Developing methods to understand neural pathfinding and to guide de novo neurite growth to specific stimulatory elements will enable design of innovative biomaterials that improve functional outcomes of devices that interface with the nervous system.

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SGN percent neurite length in depressedmicrofeatures and feature crossing per neurite length on uni- andmultidirectional topographic cues. (A) The majority of SGN neuritelength on both parallel and 90° angle patterns is located inthe grooves (*p < 0.05, Mann–Whitney RankSum test). (B,C) Immunofluorescent images of SGN neurite growth ingroove microfeatures. (D) SGN neurites crossed ridge-groove transitionssignificantly more on multidirectional patterns compared to unidirectionalsubstrates (*p < 0.05, Mann–Whitney RankSum test). (E,F) Immunofluorescent images of SGN neurites crossingridge-groove transitions on various micropatterns. Dissociated cultureswere stained with anti-NF200 antibodies. Micropatterned substrateshave a channel amplitude of 7 μm.
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fig7: SGN percent neurite length in depressedmicrofeatures and feature crossing per neurite length on uni- andmultidirectional topographic cues. (A) The majority of SGN neuritelength on both parallel and 90° angle patterns is located inthe grooves (*p < 0.05, Mann–Whitney RankSum test). (B,C) Immunofluorescent images of SGN neurite growth ingroove microfeatures. (D) SGN neurites crossed ridge-groove transitionssignificantly more on multidirectional patterns compared to unidirectionalsubstrates (*p < 0.05, Mann–Whitney RankSum test). (E,F) Immunofluorescent images of SGN neurites crossingridge-groove transitions on various micropatterns. Dissociated cultureswere stained with anti-NF200 antibodies. Micropatterned substrateshave a channel amplitude of 7 μm.

Mentions: The percentage of neurite length in microfeature grooves and thenumber of feature crossings per neurite length were measured to furthercharacterize differences in neural pathfinding on uni- versus multidirectionalsurface cues (Figure 7). The majority of regenerativeSGN neurite length is found in depressed microfeatures (i.e., grooves)on both parallel and repeating angle micropatterns. Nearly 75% ofall neurite length tracks surface depressions when SGNs are culturedon parallel feature micropatterns. A few neurites were even observedto turn 180° while remaining sequestered within microgrooves(Figure 7B). The quantified preference fordepressed features is in contrast to other research, which observedneural processes preferentially growing on elevated features; however,the width of the features used were much narrower (i.e., < 1 μm)than the photopolymerized patterns used here.14 While the majority of neurite length on 90° angle patternsis also found in surface depressions, the percent length in the depressionsis still significantly less than that of neurites on unidirectionalfeatures. The difference is likely due to the presentation of multipledirectional cues to the advancing growth cone, which increases thenumber of potential guidance points, and ultimately leads to increasedtopographic feature crossings.


Neural pathfinding on uni- and multidirectional photopolymerized micropatterns.

Tuft BW, Xu L, White SP, Seline AE, Erwood AM, Hansen MR, Guymon CA - ACS Appl Mater Interfaces (2014)

SGN percent neurite length in depressedmicrofeatures and feature crossing per neurite length on uni- andmultidirectional topographic cues. (A) The majority of SGN neuritelength on both parallel and 90° angle patterns is located inthe grooves (*p < 0.05, Mann–Whitney RankSum test). (B,C) Immunofluorescent images of SGN neurite growth ingroove microfeatures. (D) SGN neurites crossed ridge-groove transitionssignificantly more on multidirectional patterns compared to unidirectionalsubstrates (*p < 0.05, Mann–Whitney RankSum test). (E,F) Immunofluorescent images of SGN neurites crossingridge-groove transitions on various micropatterns. Dissociated cultureswere stained with anti-NF200 antibodies. Micropatterned substrateshave a channel amplitude of 7 μm.
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Related In: Results  -  Collection

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fig7: SGN percent neurite length in depressedmicrofeatures and feature crossing per neurite length on uni- andmultidirectional topographic cues. (A) The majority of SGN neuritelength on both parallel and 90° angle patterns is located inthe grooves (*p < 0.05, Mann–Whitney RankSum test). (B,C) Immunofluorescent images of SGN neurite growth ingroove microfeatures. (D) SGN neurites crossed ridge-groove transitionssignificantly more on multidirectional patterns compared to unidirectionalsubstrates (*p < 0.05, Mann–Whitney RankSum test). (E,F) Immunofluorescent images of SGN neurites crossingridge-groove transitions on various micropatterns. Dissociated cultureswere stained with anti-NF200 antibodies. Micropatterned substrateshave a channel amplitude of 7 μm.
Mentions: The percentage of neurite length in microfeature grooves and thenumber of feature crossings per neurite length were measured to furthercharacterize differences in neural pathfinding on uni- versus multidirectionalsurface cues (Figure 7). The majority of regenerativeSGN neurite length is found in depressed microfeatures (i.e., grooves)on both parallel and repeating angle micropatterns. Nearly 75% ofall neurite length tracks surface depressions when SGNs are culturedon parallel feature micropatterns. A few neurites were even observedto turn 180° while remaining sequestered within microgrooves(Figure 7B). The quantified preference fordepressed features is in contrast to other research, which observedneural processes preferentially growing on elevated features; however,the width of the features used were much narrower (i.e., < 1 μm)than the photopolymerized patterns used here.14 While the majority of neurite length on 90° angle patternsis also found in surface depressions, the percent length in the depressionsis still significantly less than that of neurites on unidirectionalfeatures. The difference is likely due to the presentation of multipledirectional cues to the advancing growth cone, which increases thenumber of potential guidance points, and ultimately leads to increasedtopographic feature crossings.

Bottom Line: SGN neurites orient randomly on unpatterned photopolymer controls, align and consistently track unidirectional patterns, and are substantially influenced by, but do not consistently track, multidirectional turning cues.Neurite lengths are 20% shorter on multidirectional substrates compared to unidirectional patterns while neurite branching and microfeature crossing events are significantly higher.Developing methods to understand neural pathfinding and to guide de novo neurite growth to specific stimulatory elements will enable design of innovative biomaterials that improve functional outcomes of devices that interface with the nervous system.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical and Biochemical Engineering, University of Iowa , Iowa City, Iowa 52242, United States, United States.

ABSTRACT
Overcoming signal resolution barriers of neural prostheses, such as the commercially available cochlear impant (CI) or the developing retinal implant, will likely require spatial control of regenerative neural elements. To rationally design materials that direct nerve growth, it is first necessary to determine pathfinding behavior of de novo neurite growth from prosthesis-relevant cells such as spiral ganglion neurons (SGNs) in the inner ear. Accordingly, in this work, repeating 90° turns were fabricated as multidirectional micropatterns to determine SGN neurite turning capability and pathfinding. Unidirectional micropatterns and unpatterned substrates are used as comparisons. Spiral ganglion Schwann cell alignment (SGSC) is also examined on each surface type. Micropatterns are fabricated using the spatial reaction control inherent to photopolymerization with photomasks that have either parallel line spacing gratings for unidirectional patterns or repeating 90° angle steps for multidirectional patterns. Feature depth is controlled by modulating UV exposure time by shuttering the light source at given time increments. Substrate topography is characterized by white light interferometry and scanning electron microscopy (SEM). Both pattern types exhibit features that are 25 μm in width and 7.4 ± 0.7 μm in depth. SGN neurites orient randomly on unpatterned photopolymer controls, align and consistently track unidirectional patterns, and are substantially influenced by, but do not consistently track, multidirectional turning cues. Neurite lengths are 20% shorter on multidirectional substrates compared to unidirectional patterns while neurite branching and microfeature crossing events are significantly higher. For both pattern types, the majority of the neurite length is located in depressed surface features. Developing methods to understand neural pathfinding and to guide de novo neurite growth to specific stimulatory elements will enable design of innovative biomaterials that improve functional outcomes of devices that interface with the nervous system.

Show MeSH
Related in: MedlinePlus