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and calcium release-activated channels (CRAC) that mediates store-operated calci-
um entry (SOCE) across the plasma membrane (
Hille, 2001; Hogan et al.,2010
). All
of these proteins provide channels that allow calcium to di
use into the cytosol when
the channel opens. Each open channel protein has a unique unitary conductance for
calcium, ranging from approximately 0.1 to several hundred picosiemens (pS), but
the proteins spontaneously cycle between open and closed conformations on a time
scale of milliseconds. It is the rates of these transitions rather than the conductances
which are regulated by physiological events to control the amplitude of calcium
fluxes. Such unitary currents are often di
V
cult tomeasure because they are small and
individual openings last less than a millisecond. However, the total amount of
current crossing the entire surface membrane of a cell at any time is the product of
the number of channels (N), the fraction of time they spend in the open state (P
o
),
their unitary conductance (g), and the electrochemical driving force (
D
V) measured
as the di
Y
V
erence between the voltage across the membrane (V
m
) and the ion's Nernst
potential.
Even small currents produce physiologically significant increases in intracellular
calcium. For example, a current of only 0.1 pA (pA
10
12
A), which corresponds
ΒΌ
to 0.1 pC of charge per second, or
300,000 divalent ions per second, will transfer
300 calcium ions each millisecond the channel is open. Such a current, 0.1 pA
lasting 1 ms, is just below the current technology of detection with the patch clamp
technique. Nevertheless, it would produce a physiologically significant change in
intracellular calcium. The calcium ions cannot di
use on average much farther
than a micrometer in a millisecond, so the concentration under the membrane will
rise transiently to 0.5
m
M, more than double the resting level of calcium. If there
was only one such channel that opened for 1 ms in every square micrometer of
membrane of a spherical cell with a diameter of 10
m
m (volume
V
0.5 pL; area -
300
m
m
2
), then the resulting 30 pA current would almost double intracellular
calcium concentration throughout the cell. Action potentials that depolarize cells
for tens of milliseconds will have correspondingly larger e
V
ects. Thus, millisecond
di
erences in calcium channel kinetics have profound consequences for cell physi-
ology and human health (
Erxleben et al., 2006
).
This calculation also illustrates the danger of expressing recombinant channels
in mammalian fibroblasts. Investigators routinely report currents of a few
nanoamperes, which even inexperienced investigators can measure with the
patch clamp technique. However, in the scenario outlined above, a 3-nA current
would represent a 100
V
larger
increases in calcium, which might lead to cytotoxic reactions. In most cases, such
recordings are made with exogenous calcium bu
higher density of channels and produce 100
ers in the cytosol, which not only
prevent cytotoxicity but also preclude analysis of physiological regulation of
calcium channels by calcium-dependent signaling. In addition, because most calci-
um channels have a low probability of opening (P
o
) less than 0.1, the larger current
density reflects at least 1000 channel proteins per square micrometer, or more than
10% of the space available with close packing. At this density, there might not be
room for each channel protein to be associated with its normal penumbra of
V
