CELLS OF THE NERVOUS SYSTEM AND ELECTRICAL STIMULATION - Advanced Biomaterials: Fundamentals, Processing, and Applications - page 616

of the media will be changed when using a constant current, so one must design a
system that prevents cells from experiencing change in pH to use DC. Solving this
problem can often be diffi cult. Most often media perfusion is a good solution, but
this increases the risk of contamination.
One other solution is the use of agar or agarose salt bridges to prevent elec-
trochemical byproducts and changes in pH caused by the electrodes to enter the
media. However, the saline in the salt bridges will diffuse out and media will
diffuse into the salt bridge when stimulating cells for an extended period and
temperatures close to the agar gelation temperature. The concentration differ-
ence between the media and agar-saline bridge drives the diffusion of salt into the
media and of media into the salt bridge. Diffusion will be slower at room tem-
perature than at incubator temperature which is 37 °C. At temperatures close to
37 °C agar is close or at its gelation temperature allowing diffusion in and out of
the agar. Additionally, agar has been shown to cause changes in the genetic mate-
rial of cells when an electromagnetic fi eld is applied. This may not be related to
DC stimulation but should be taken as a precaution (Cohen et al. 1988). There-
fore, using agar salt bridges is an appropriate solution for experiments on cold-
blooded animal cells, but for mammalian cells, agarose gel bridges and media
perfusion is better.
18.5.7 Cellular Response to Electric Field In Vitro
The effect of any electrical stimulation is not predictable or the same for all cell
types. There is no individual signal that results in the same cellular response in all
cell types, although a strong electric signal that passes high enough current
through the cell will cause cell death. A thorough review, written by Nuccitelli
et al., where the effects of in vitro electrical stimulation on several cells types
are listed (Nuccitelli 2003), shows that no single response to an identical stimulus
is the same for all cell types. The cells mentioned in the list by Nuccitelli et al.
were stimulated with DC EFs. Cells types that have been stimulated with other
methodologies are: PC12 cells (Kimura et al. 1998; Schmidt et al. 1997), astroglial
cells (Koyama et al. 1997), HeLa cancer cells (Manabe et al. 2004), and epithelial
cells (Zhao et al. 1996), again, with no identical response to a particular
stimulus.
The focus of this chapter is on electrical stimulation related to the nervous
system; therefore, studies with PC12 and astroglial cells are considered here in
greater detail. PC12 cells were stimulated with rectangular impulses of 200 mV
and 400 mV, peak - to - peak, with frequencies of 50 Hz, 100 Hz, 500 Hz and 1 kHz
for 96 hours. The result was that PC12 cells matured and extended neurites
without the use of NGF. In other words, electrical stimulation caused the cells to
differentiate into more mature neurons, which normally only occurs by incorpo-
rating NGF into the media (Kimura et al. 1998).
In the latter study, astroglial cells were shown to secrete NGF when an elec-
trical stimulation was applied. A 10 Hz, sine wave with a potential difference of
+0.3 V was shown to maximize the amount of NGF secreted into the media. These
two studies are related since they both show that electrical stimulation affects the