Adhesion to Carbon Nanotube Conductive Scaffolds Forces Action-Potential Appearance in Immature Rat Spinal Neurons
et al. (2013) Adhesion to Carbon Nanotube Conductive Scaffolds Forces Action-Potential
Appearance in Immature Rat Spinal Neurons. PLoS ONE 8(8): e73621. doi:10.1371/journal.pone.0073621
Adhesion to Carbon Nanotube Conductive Scaffolds Forces Action-Potential Appearance in Immature Rat Spinal Neurons
Alessandra Fabbro
Antonietta Sucapane
Francesca Maria Toma
Enrica Calura
Lisa Rizzetto
Claudia Carrieri
Paola Roncaglia
Valentina Martinelli
Denis Scaini
Lara Masten
Antonio Turco
Stefano Gustincich
Maurizio Prato (MP)
Laura Ballerini
Masaya Yamamoto, Institute for Frontier Medical Sciences, Kyoto University, Japan
In the last decade, carbon nanotube growth substrates have been used to investigate neurons and neuronal networks formation in vitro when guided by artificial nano-scaled cues. Besides, nanotube-based interfaces are being developed, such as prosthesis for monitoring brain activity. We recently described how carbon nanotube substrates alter the electrophysiological and synaptic responses of hippocampal neurons in culture. This observation highlighted the exceptional ability of this material in interfering with nerve tissue growth. Here we test the hypothesis that carbon nanotube scaffolds promote the development of immature neurons isolated from the neonatal rat spinal cord, and maintained in vitro. To address this issue we performed electrophysiological studies associated to gene expression analysis. Our results indicate that spinal neurons plated on electro-conductive carbon nanotubes show a facilitated development. Spinal neurons anticipate the expression of functional markers of maturation, such as the generation of voltage dependent currents or action potentials. These changes are accompanied by a selective modulation of gene expression, involving neuronal and non-neuronal components. Our microarray experiments suggest that carbon nanotube platforms trigger reparative activities involving microglia, in the absence of reactive gliosis. Hence, future tissue scaffolds blended with conductive nanotubes may be exploited to promote cell differentiation and reparative pathways in neural regeneration strategies.
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Funding: Funding provided by NEURONANO-NMP4-CT-2006-031847 CARBONANOBRIDGE- ERC-2008-227135 http://erc.europa.eu/. The funders had
no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
These authors contributed equally to this work.
Nanomaterials are increasingly used for organ engineering
purposes [1,2]. Scaffolds with manufactured three-dimensional
properties may promote cells reorganization into functional
tissue. This possibility has driven a growing interest in studying
physical-chemical features of scaffolds at the nano-scale, to
activate cell-specific molecular machineries [14].
Scaffolds blended with various materials have been
constructed for the repair of different tissues, such as bones,
liver and other organs [1,2,57]. However, attempts to
construct scaffolds for the repair of the central nervous system
(CNS) have had limited success, because of its intrinsic
complexity, low regenerative potential and anatomically
restrictive nature, which pose a unique set of challenges
[813]. Despite this fact, an increasing amount of studies in
modern neuroscience addresses the ability of growth substrate
topography or physical features in driving neuronal networks
reconstruction. In cultured systems, the interaction of neurons
with their growth substrate may influence neuron differentiation,
morphology, adhesion and outgrowth [1319].
Our approach to address this issue was to incorporate
neuronal cultures to artificial conductive nanostructures,
namely carbon nanotubes. Recently, carbon nanotubes have
attracted tremendous attention for the development of nano-bio
hybrid systems able to govern cell-specific behaviors in
cultured neuronal networks and explants [2027] and have
been shown to promote proliferation of neonatal cardiac
myocytes [28].
Carbon nanotubes are cylindrically shaped nanostructures,
made of one or more concentric rolled-up graphene sheets,
which possess peculiar properties including high surface area,
high mechanical strength, ultra-light weight, rich electronic
properties, and excellent chemical and thermal stability [29,30].
In vivo, carbon nanotubes have been shown to be a blood
compatible and a suitable scaffold for bone regeneration
[31,32] or, in vitro, for cultured synaptic network formation
[2224,26,27,33] and neonatal cardiomyocyte maturation [28].
In the present work we investigate the interaction between
carbon nanotube scaffolds and immature spinal cord neurons.
Here we show that in vitro spinal neurons adherent to carbon
nanotube substrates undergo a functional maturation
characterized by an earlier appearance of voltage dependent
currents and of action potentials. To address the mechanistic
pathways between the enhan (...truncated)