454 pH and Closed-State Inactivation
depolarizations at normal and acidic pH. These data sug-
gest the possibility that channels are closed-inactivated
at low pH, and are able to activate directly into Na
conductive depolarization-induced inactivated states
without actually opening. This idea is consistent with the
present uorescence and single channel experiments,
which show directly that a population of channels be-
comes unavailable during acidic pH (Figs. 5, 6, and 10)
due to the stabilization of inactivation from resting
closed channel states (Figs. 10–13), making them unable
to open. A kinetic model to describe the Na
ation experiments suggested that channels accumulated
in the open-inactivated state during depolarization and
made the transition to closed-inactivated states via an
intermediate “R” state in the recovery pathway (Zhang
et al., 2003). Channels could not reopen from resting
closed-inactivated states in the model (Zhang et al., 2003),
but it is not clear whether or not Na
as the permeant ion
causes other structural changes within the selectivity lter
that do not allow it to accurately model activation of
closed-inactivated channels with K
as the charge carrier
(Panyi and Deutsch, 2006).
It has previously been shown that Shaker channels may
undergo a type of closed-state inactivation called U-type
inactivation (Klemic et al., 2001) and it is helpful to
consider whether this is the closed state stabilized at low
pH. In U-type inactivation, maximal current reduction
occurs at intermediate potentials, where the dwell time
in intermediate states is longer, and less inactivation oc-
curs at more depolarized potentials when the open
probability is high, resulting in a characteristic U-shaped
inactivation–voltage relationship. However, although
the uorescence data strongly support closed-state inac-
tivation, the effects of raising external K
are opposite to those expected for U-type inactivation.
U-type inactivation is enhanced by raising external K
(Klemic et al., 2001), whereas we show in Fig. 10 that
high external K
rescues channels from closed-state in-
activation, which suggests that the pH-induced closed-
state inactivation is unlikely to be U-type in nature.
This work was supported by grants from the Heart and Stroke
Foundation of British Columbia and Yukon and the Canadian In-
stitutes of Health Research to D. Fedida and S.J. Kehl. D. Fedida
is supported by a Career Investigator award from the Heart and
Stroke Foundation of British Columbia and Yukon. T.W. Claydon
was supported by a postdoctoral research fellowship funded by a
Focus on Stroke strategic initiative from The Canadian Stroke
Network, the Heart and Stroke Foundation, the CIHR Institute of
Circulatory and Respiratory Health, and the CIHR/Rx&D Pro-
gram along with AstraZeneca Canada. M. Vaid was supported by a
Michael Smith Foundation for Health Research Studentship. S.
Rezazadeh was supported by a Heart and Stroke Foundation of
British Columbia and Yukon Studentship and a University of Brit-
ish Columbia Graduate Fellowship.
Olaf S. Andersen served as editor.
Submitted: 2 March 2007
Accepted: 6 April 2007
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