Biomedical Engineering Reference
In-Depth Information
system. Nonetheless, these differences should not detract from the fact that (un)
functionalized CNTs can serve as permissive substrates/scaffolds for neuronal
growth and that the neurite outgrowth can be infl uenced by scaffold properties.
Deposition of AP-MWCNTs to glass coverslips can be problematic, so it may
result in detachment of MWCNT mats which then fl oat to the surface of the culture
media (Lovat et al.
2005
). To improve on MWCNT substrate adhesion to glass cov-
erslips, Lovat et al. (Lovat et al.
2005
) developed an alternative MWCNT deposition
procedure. First, they subjected AP-MWCNTs to 1,3-dipolar cycloaddition of
azomethine ylides, which generates pyrrolidine (PYR) groups on the MWCNT ends
and sidewalls. This reaction confers MWCNT-PYR with increased solubility in
organic solvents. Hence, the MWCNT-PYR material was dissolved in dimethylfor-
mamide and applied to glass coverslips. Following evaporation of the solvent, cov-
erslips containing MWCNT-PYR were baked to remove PYR groups. The resulting
glass coverslips contained a retainable fi lm of bare/defunctionalized MWCNTs,
which was used as a substrate for growth of hippocampal neurons. Since morpho-
logical parameters of neuronal growth were not the subject of this study, the com-
parison of growth and neurite outgrowth on this substrate to those described above
using various scaffolds is not possible. However, the neurons grown on top of
MWCNT fi lms survived and formed functional networks displaying spontaneous
synaptic transmission within 8-10 days in culture. The detailed description of this
approach and the effects that interactions between neurons and MWCNTs can have
on synaptic transmission is available elsewhere [Chap. 9 in this volume; also see
(Lovat et al.
2005
; Mazzatenta et al.
2007
; Cellot et al.
2009
) ].
Thus far, we only discussed the usage of MWCNTs as planar substrates to support
neuronal growth. Galvan-Garcia et al.(Galvan-Garcia et al.
2007
) presented evidence
that nonfunctionalized MWCNTs not only in the form of sheets, but also as yarns, can
serve as permissive substrates/scaffolds for neuronal attachment and growth. They
generated MWCNT sheets using the catalytic CVD. MWCNT yarns were made by a
subsequent spinning of these sheets. MWCNTs are directionally oriented in both
forms. Galvan-Garcia et al. evaluated permissiveness of bare MWCNT sheets and
yarns as a substrate for neuronal growth in culture. Both materials were biocompatible
since they promoted cell attachment, differentiation, growth, and long-term survival
of various cell types, including dissociated cortical and cerebellar neurons from rat
pups (postnatal day 0-10) and neonatal mice DRG explants containing neurons. Using
bright and fl uorescence light microscopy, they demonstrated that DRG neurons sur-
vived over 2 weeks in culture. These neurons extended their axons from explants onto
MWCNT sheets; their neurites could attach and extend along MWCNT yarns.
Dissociated cortical and cerebellar neurons displayed neurite outgrowth to a similar
extent as when they were seeded onto MWCNT sheets or onto the commonly used
polycationic substrate for neural cell growth, polyornithine (PO), which was precoated
on glass coverslips. Interestingly, cortical and cerebellar neurons cultured on MWCNT
sheets showed larger growth cones than when grown on PO while the length of neu-
rites was not signifi cantly different. Based on electrostatic charge alone, this is an
unexpected fi nding since MWCNT sheets should be neutrally charged in DMEM
while PO should be positively charged. However, besides the charge difference
