Electronic Detection Techniques - Sensor Technology in Neuroscience - page 91

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Figure 3.1 Examples of topographical features on silicon and plastic surfaces.
(A) Silicon nanoscale roughness (scale bar ¼ 20 nm). (B) Silicon nanoscale
pillars (scale bar ¼ 1000 nm). (C) Grooves on poly(dimethylsiloxane)

(PDMS) (scale bar ¼ 5 nm). 15
(Reprinted by kind permission of Wiley.)

Several techniques have been developed to direct the growth of neurons and
their network structure. Designed in vitro network platforms can employ
topographical and chemical modification of surfaces (Figure 3.1). In large part
these techniques try to mimic or amplify the natural topographical effects of the
cell's base membranes and to artificially create a designed substrate with
nanostructures that stimulate directed growth of neurons. Such modifications
illustrated in Figure 3.1 are hypothesized to involve the creation of
submicrometer features by careful design such as laser etching, photo-
lithography and other etching techniques or by applying bulk surface modifi-
cations such as roughening of the surface. 11,12 Alternative methods employed
separately and in addition to topographical changes can involve changing the
chemical composition of a surface by binding cell adhesive molecules to
promote directed growth. 13 In order to mimic the spatial structure of the brain,
3D matrices consisting of polyethylene glycol (PEG) hydrogel have been
employed. 14 In these platforms the hydrogel is supplemented with poly- L -lysine
(PLL), which has proved not to be cytotoxic and allows cell culture growth.
Such systems can be useful for analyzing the cell interactions in vitro in a more
natural 3D system.
Glial cells play an essential role in supporting and nourishing the neurons.
They provide natural physical cues to migrating neurons. Thus to gain further
insight into the study of neurobiology, it is important to consider carefully the
contribution of glial cells. 16,17 Figure 3.2 shows the complicated protocol used
to obtain a low density dissociated cell culture of hippocampal neurons from
embryonic rats or mice. The neurons are cultured on PLL-treated coverslips,
which are suspended above an astrocyte feeder layer and maintained in serum
free medium. When cultured according to this protocol, hippocampal neurons
are appropriately polarized and develop extensive axonal and dendritic arbors
to form numerous, functional synaptic connections with one another.
Hippocampal cultures have been widely used to visualize the subcellular
localization of endogenous or expressed proteins. This allows for imaging of
protein tracking and the defining of molecular mechanisms underlying the
development of neuronal polarity, dendritic growth and synapse formation. 18
Protocols such as these may be adapted to generate growth of neurons from

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