Biomedical Engineering Reference
In-Depth Information
PEI had a signifi cantly higher number of growth cones than those grown on 10- and
30-nm SWCNT fi lms, but not higher than those grown on 60-nm SWCNT fi lms
(Fig.
3f
). Since positively charged PEI has a much smoother surface than SWCNT
fi lms, it appears that perhaps a combination of charge, roughness, and/or conductiv-
ity could cause this effect. Taken together, these results indicate that a SWCNT-PEG
substrate can modulate neuronal growth and neurite outgrowth within a narrow
range of its conductivity. The studies discussed above showed that various qualities
of CNT substrates/scaffold, most notably their charge, roughness/topology, and
conductivity, can modulate neuronal growth and neurite outgrowth. Consequently,
these physical properties of CNTs should be taken into consideration when design-
ing CNTs as scaffolds for neuronal growth.
3
Water-Dispersible CNTs as Modulators of Neuronal Growth
Experimental approaches discussed thus far aimed to explore the effects of CNTs as
biocompatible substrate/scaffolds. Of course, an additional strategy is to generate
CNTs that are soluble/dispersible in aqueous media of extracellular space of the
brain so that they could be delivered as diffusive agents to affect neurite outgrowth.
Ni et al. (Ni et al.
2005
) addressed this issue by treating neurons grown on PEI sub-
strate with water-soluble SWCNTs that were functionalized with either PEG or
PABS via the SWCNT-COCl intermediate. At physiological pH, these graft copoly-
mers are neutral or zwitterionic, respectively. Using calcein-loaded hippocampal
neurons, prepared from 0- to 2-day-old rats, and fl uorescence microscopy (Fig.
4a-c
),
they demonstrated that neurons treated with either form of soluble SWCNTs showed
a reduced number of neurites and growth cones when compared to control (sham
treated with the vehicle) neurons. Coincidentally, neurons treated with water-solu-
ble SWCNTs also exhibited longer neurites than controls (Fig.
4d
); these effects
were concentration dependent. Ni et al. investigated a possible mechanism underly-
ing actions of water-soluble SWCNTs on neurite outgrowth. It appears that SWCNT-
PEG reduced Ca
2+
infl ux from the extracellular space through voltage-dependent
Ca
2+
channels (VDCCs), known regulators of neurite elongation (Mattson and Kater
1987
; Kater et al.
1988
; Mattson et al.
1988
). This was investigated using the intra-
cellular calcium indicator fl uo-3. When compared to control, neurons treated with
SWCNT-PEG had reduced cytosolic Ca
2+
accumulation after depolarizing them
with high extracellular potassium (50 mM) to open VDCCs; the depolarization-
dependent Ca
2+
entry was sensitive to the VDCC blocker Cd
2+
. An increase in neu-
ronal intracellular Ca
2+
levels can regulate plasma membrane/vesicular recycling,
which has been implicated to play a role in the rate of neurite elongation (Zakharenko
and Popov
2000
). Consequently, Malarkey at al. examined whether SWCNTs could
affect membrane recycling (Malarkey et al.
2008
). Reasoning that the fl uorescent
dye
N
-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styrl)pyridinium dibromide
(FM 1-43) is plasma membrane impermeable and is taken up by cells through endo-
cytosis, they compared the effect of SWCNT-PEG exposure on FM 1-43 load in
