Biology Reference
In-Depth Information
3.3. Alternative
Confi gurations
Many alternative confi gurations of recording can be made based
on the whole-cell recording for different purposes of studies.
Inside-out recording . This recording confi guration is achieved by
pulling off the patch of membrane shortly after gigaseal formation.
In this confi guration, the inside of the cell membrane is facing the
bath solution, hence the name. The ease of changing the cytoso-
lic solutions of the patch makes this confi guration very useful to
study the role of cytosolic signalling molecules on channel func-
tions at the single channel level. We used this technique to study
the BK channel function at the single channel level in a dog model
of SAH ( 3 ).
Outside-out recording . This confi guration is made by pulling off
the patch of membrane from the cell after the formation of the
whole-cell confi guration. The ease of changing the extracellular
solutions of the patch makes this confi guration very useful to study
functions of receptor-gated ion channels at the single channel level.
Perforated patch-clamp recording . In the whole cell confi guration,
cell contents will slowly leak out into the patch pipette and thus
alter the intracellular milieu. When the loss of cytosolic constituents
is a concern, the perforated patch-clamp confi guration can be
performed. This could be achieved by including channel forming
substances in the pipette solution, such as nystatin and amphotericin B.
The small pores formed by these substances are large enough for
ions to pass but small enough to reduce the loss of cytosolic contents.
We used this technique to assess the resting membrane potential of
smooth muscle cells ( 7 ).
4. When and What
to Study with
Electrophysio-
logical Techniques
After SAH
Patch-clamp techniques can be used to study cells at any time after
SAH. Since it is still quite diffi cult to molecularly alter ion chan-
nels in smooth muscle cells and then determine what effect these
changes have on vasospasm, other methods to determine whether
changes in ion channels cause vasospasm are used. For example,
the researcher can study the same channel function over a series of
time points to have a general picture of what happens to a particular
ion channel after SAH ( 1 ).
Patch-clamping has been used to characterize functional
changes in K + channels ( 1, 3, 5, 7, 13, 15 ), Ca 2+ channels ( 4, 8, 9 )
and TRP proteins ( 6 ) of vasospastic arteries from SAH models.
Many aspects of ion channel properties and functions can be studied,
including resting membrane potential of the cells, as well as voltage,
Ca 2+ and time-dependent features of the channels. Single channel
recordings allow the investigator to understand more about the
biophysical properties of the channel. Using similar parameters to
those in the aforementioned publications as a starting point, other
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