From the Department of Biology, Georgia State University, Atlanta, Georgia 30302-4010
Received for publication, November 11, 2002, and in revised form, December 13, 2002
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ABSTRACT |
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G-protein-coupled inward rectification
K+ (GIRK) channels play an important role in
modulation of synaptic transmission and cellular excitability. The GIRK
channels are regulated by diverse intra- and extracellular signaling
molecules. Previously, we have shown that GIRK1/GIRK4 channels are
activated by extracellular protons. The channel activation depends on a
histidine residue in the M1-H5 linker and may play a role in
neurotransmission. Here, we show evidence that the heteromeric
GIRK1/GIRK4 channels are inhibited by intracellular acidification. This
inhibition was produced by selective decrease in the channel open
probability with a modest drop in the single-channel conductance. The
inhibition does not seem to require G-proteins as it was seen in two
G-protein coupling-defective GIRK mutants and in excised patches in the absence of exogenous G-proteins. Three histidine residues in
intracellular domains were critical for the inhibition. Individual
mutation of His-64, His-228, or His-352 in GIRK4 abolished or greatly
diminished the inhibition in homomeric GIRK4. Mutations of any of these
histidine residues in GIRK4 or their counterparts in GIRK1 were
sufficient to eliminate the pHi sensitivity of the
heteromeric GIRK1/GIRK4 channels. Thus, the molecular and biophysical
bases for the inhibition of GIRK channels by intracellular protons are
illustrated. Because of the inequality of the pHi and
pHo in most cells and their relatively independent controls by
cellular versus systemic mechanisms, such pHi
sensitivity may allow these channels to regulate cellular excitability
in certain physiological and pathophysiological conditions when
intracellular acidosis occurs.
G-protein-coupled inward rectifier K+
(GIRK)1 channels play an
important role in regulating cellular excitability and synaptic transmission (1, 2). Activation of GIRK channels leads to inhibition of
neurons, cardiac myocytes, and endocrine cells (3). It is known that
the activation of GIRK channels depends on G-protein One important signaling molecule is the hydrogen ion. Our recent
studies have shown that the heteromeric GIRK1/GIRK4 channels are
stimulated by a drop in extracellular pH (pHo) (20). The
pHo sensitivity depends on a histidine residue located in the
M1-H5 linker and may play a role in enhancing the GIRK-mediated synaptic transmission.
Increasing evidence indicates that intracellular protons can serve as a
second messenger in regulating multiple cellular functions (21, 22).
Whereas pHo is controlled by several buffer systems, allowing
its change in a rather narrow range, intracellular pH (pHi) can
vary widely under certain physiological and pathophysiological
conditions. In the central nervous system, intracellular protons are
involved in modulations of synaptic transmission, membrane
excitability, and neuronal plasticity (23) in which GIRK channels are
ubiquitously involved (3). In the brainstem, pH-dependent
regulation of G-protein-coupled K+ channels are known to be
important for the generation and control of central respiratory
activity and CO2 central chemoreception (24-27). There is
evidence suggesting that certain types of epilepsy are related to
dysfunction of GIRK channels and the decrease in pHi (28, 29).
Furthermore, pHi shifts seem to be a mechanism for several
local anesthetics (30, 31) that are likely to be produced by changing
GIRK channel activity as well (32). Therefore, the modulation of GIRK
channels by intracellular protons is an important cellular mechanism in
regulating membrane excitability under these physiological and
pathophysiological conditions.
GIRK channels belong to a subfamily of inward rectifying K+
channels, several members of which have been demonstrated to be pHi-sensitive (30-36). To test the hypothesis that the heteromeric GIRK1/GIRK4 channels are sensitive to intracellular protons, we performed these experiments in a heterologous expression system. Our data showed that these channels were inhibited by intracellular pH at near physiological levels. The pHi sensitivity does not seem to require G-proteins and relies on a few
histidine residues in the intracellular C and N termini.
Oocyte Preparation and Injection--
Experiments were performed
as we described previously (34-37). In brief, oocytes from
Xenopus laevis were used in the present study.
Frogs were anesthetized by bathing them in 0.3% 3-aminobenzoic acid
ethyl ester. A few lobes of ovaries were removed after a small
abdominal incision (~5 mm). Then, the surgical incision was closed,
and the frogs were allowed to recover from the anesthesia. Xenopus oocytes were treated with 2 mg/ml of
collagenase (Type IA, Sigma) in the OR2 solution: 82 mM NaCl, 2 mM KCl, 1 mM
MgCl2, and 5 mM HEPES, pH 7.4 for 60 min at
room temperature. After three washes (10 min each) of the oocytes with
the OR2 solution, cDNAs (25-50 ng in 50 nl of water) were injected
into the oocytes. The oocytes were then incubated at 18 °C in the
ND-96 solution containing 96 mM NaCl, 2 mM KCl, 1 mM MgCl2, 1.8 mM CaCl2, 5 mM HEPES, and 2.5 mM sodium pyruvate with 100 mg/liter geneticin added (pH
7.4).
Molecular Biology--
Rat GIRK1 (Kir3.1) cDNA
(GenBankTM accession number U01071) and rat GIRK4 (Kir3.4)
cDNA (GenBankTM accession number X83584) are gifts from
Dr. Norman Davidson at Caltech, Pasadena, CA. These cDNAs
were subcloned into a eukaryotic expression vector pcDNA3.1
(Invitrogen) and used for Xenopus oocyte expression without
cRNA synthesis. Homomeric expression of GIRK4 was attempted using cRNA
that was synthesized with an in vitro transcription kit
(Riboprobe, Promega Inc., Madison, WI) by the T7 promoter. PCR was used
to generate GIRK1/GIRK4 dimer. Two PCR fragments encoding the
entire length of GIRK1 and GIRK4 were joined with an XbaI
restriction site created in the primers with the GIRK1 at the 5' end
and then cloned to the pcDNA3.1. Five extra amino acids (RCQQQ)
were created between the C terminus of GIRK1 and the N terminus of
GIRK4. We did not find any detectable effect of these additional
residues on channel expression, current profile, and pH sensitivity.
Site-specific mutations were made using a site-directed mutagenesis kit
(Stratagene, La Jolla, CA). Correct constructions and mutations were
confirmed with DNA sequencing.
Electrophysiology--
Whole-cell currents were studied on the
oocytes 2-4 days after injection. The two-electrode voltage
clamp test was performed using an amplifier (Geneclamp 500, Axon
Instruments Inc., Foster City, CA) at room temperature (~24 °C).
The extracellular solution contained 90 mM KCl, 3 mM MgCl2, and 5 mM HEPES (pH 7.4).
Experiments were performed using a perfusion chamber with a total
volume of 1 ml (RC-3Z, Warner Instruments, New Haven, CT).
Intracellular acidification was produced using an extracellular
solution in which all KCl (90 mM) was replaced with 90 mM KHCO3. This solution was titrated to pH 7.4 immediately before experiments. Intracellular pH was measured in our
previous studies, indicating that exposure of the Xenopus
oocytes to this solution produces a rapid drop in intracellular pH to
6.6 (35, 37). Leak currents were constantly monitored using a series of
command pulses. The current-voltage (I-V) relationship was plotted. The
I-V plots were then compared between the test record and baseline
control at the lowest conductance region. There was no detectable
leakage in most cells. In a small number of cells (10-20%), the
conductance changes were evident. In the cells, the off-line leak
subtraction was performed to remove the additional linear current
component using Clampfit 6.0 (Axon Instruments, Inc.).
Patch Clamp Studies--
Single-channel currents were studied in
an inside-out patch configuration. The bath solution contained 140 mM KCl, 10 mM
Na2H2P2O7, 5 mM NaF, 0.1 mM Na3VO3,
0.2 mM ATP, 0.2 mM GTP, 10 mM EGTA,
and 10 mM HEPES (pH = 7.4). The pipette was filled
with the same solution (37, 38). The open-state probability
(Po) was calculated as we described previously
(37, 38). The number of active channels was determined at the maximal
channel activity and used for all records obtained from a given patch.
The single-channel conductance was measured using a slope command
potential from Data Analysis--
Data are presented as means ± S.E. (standard error). Analysis of variance or Student's t
test was used. Differences of pH effects before versus
during exposures were considered to be statistically significant if
p Heteromeric GIRK1/GIRK4 channels were expressed in
Xenopus oocytes and studied using the two-electrode voltage
clamp test. Whole-cell currents were recorded using a solution
containing 90 mM K+. The currents showed strong
inward rectification (Fig. 1A)
and were blockable with micromolar concentrations of Ba2+
(data not shown). To study the effect of low pHi on GIRK
channels, intracellular acidification was produced by perfusing the
oocytes for 10 min with a solution in which all KCl was replaced with
90 mM KHCO3 (pH 7.4). This solution has been
previously shown to selectively reduce the pHi to 6.6 without
changing pHo (33, 36, 37). Under this condition, GIRK1/GIRK4 currents were inhibited by 37.5 ± 3.3% (n = 11).
In comparison, currents recorded from oocytes injected with the
expression vector alone or the Kir2.1 cDNA showed no significant
changes (
INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
subunits (G
) that are released from the G
heterotrimer when G-protein-coupled receptors are activated by
extracellular hormones or neurotransmitters. The
G
-dependent activation can be eliminated by
disrupting certain protein domains and amino acid residues that may be
involved in channel gating or the interaction with the G
(4-8).
In addition to the G
dependence, GIRK channels are modulated by a
number of cellular signaling molecules, such as phospholipids (9, 10),
arachidonic acid (11, 12), Na+ (13, 14), Mg2+
(15), redox agents (16, 17), and protein kinases (18, 19). Such complex
regulations suggest that GIRK channels are capable of integrating
extra- and intracellular signals and coupling the information to
cellular excitability.
MATERIALS AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
100 to 100 mV or an all-time histogram fitted with
Gaussian distribution. To change pHi, the internal surface of
patch membranes was perfused with solutions in different pH levels with
no dead space.
0.05.
RESULTS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
1.9 ± 1.7%, n = 4, and 2.8 ± 1.1%, n = 4), indicating that the GIRK1/GIRK4 channels
but not Xenopus oocyte-endogenous currents are inhibited. This inhibition occurred at 3 min and reached plateau in 6-7 min (Fig.
1, A and B). Washout led to complete recovery in
most cells studied. The current inhibition did not show voltage
dependence (Fig. 1, C-E). Similar current inhibition was
observed in a GIRK1-GIRK4 tandem-dimeric channel (32.4 ± 2.2%,
n = 7).
View larger version (25K):
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Fig. 1.
Inhibition of
heteromeric GIRK1/GIRK4 by intracellular
acidification. Whole-cell currents were studied in a two-electrode
voltage clamp test using an extracellular solution containing 90 mM K+. As shown in A, inward
rectifying currents were recorded from an oocyte 3 days after
co-injection of the GIRK1 and GIRK4 cDNAs. Membrane potential
(Vm) was held at 0 mV. A series of command pulse
potentials from 160 mV to 100 mV with a 20-mV increment was applied
to the cell. Intracellular acidification was produced using 90 mM KHCO3 to replace 90 mM KCl in
the perfusion solution, yielding a pHi of 6.6 (35, 37). The
perfusate was titrated to pH 7.4 immediately before use. In such a
condition, the GIRK currents were reversibly inhibited. As shown in
B, the current amplitude measured at
160 mV was plotted
against time. The profile of current amplitude changes shows that the
currents decreased rapidly when perfusate was switched to 90 mM KHCO3 and recovered with washout. At maximal
inhibition, the current amplitude was inhibited by ~40%.
Arrows indicate where the data are obtained from in
A from left to right, respectively. As
shown in C and D, the voltage dependence was
studied in whole-cell recording before (C) and during
(D) KHCO3 exposure. As shown in E,
when these currents were scaled to the same amplitude, the I-V
relationship of D (triangle) is similar to that
of C (circle) at negative membrane
potentials.
The biophysical mechanism underlying the GIRK1/GIRK4 inhibition was
studied in inside-out patches. In these experiments, oocytes expressing
GIRK currents were first identified in whole-cell recording. In the
cell-attached patch configuration, large inward rectifying currents
were seen. Channel activity rapidly declined after patch excision. In
about two-thirds of the patches, these currents remained active in
inside-out patches with single-channel conductance of 31.0 ± 0.3 picosiemens (n = 8). Exposure of the
internal surface of patch membranes to perfusates with various pH
levels produced concentration-dependent inhibitions of
the Po (Fig.
2A). The inhibition was
reversible and dependent on pH levels. The pH-current relationship showed a sigmoid shape with the 50% current inhibition (pK)
at pH 6.95 (Fig. 2D). Acidic pH also had modest inhibitory
effect on single-channel conductance. At pH 6.2, the single-channel
conductance was slightly inhibited by ~10% (27.3 ± 0.4 picosiemens, n = 7) (Fig. 2, B and
C). Thus, these results indicate that the inhibition of
whole-cell GIRK currents at acidic pHo is mainly produced by
the suppression of the Po.
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Since GIRK channels are activated by G (4, 39) and inhibited by
G
(40), it is possible that intracellular protons exert effects by
interference of the interaction of the GIRK channels with G-proteins.
To address this possibility, we took advantage of two mutants that are
defective in G-protein-dependent channel gating
(i.e. GIRK1/GIRK4C216T, GIRK1C179A/GIRK4C185A) (41, 42). Both of these mutants retained almost the same sensitivity to intracellular acidification as the WT GIRK1/GIRK4 (Fig.
3). These results as well as our
single-channel recordings in the absence of exogenous G-proteins
suggest that the inhibition appears independent of G-proteins.
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The G-protein independence suggests that protons may directly act on the channel protein. If this is true, there should be specific amino acid residues that are titratable, exist in intracellular protein domains, and play a critical role in the pH sensitivity of the channels. We thus performed studies on the molecular mechanism for the channel inhibition. To understand which subunit has such proton sensors, we first studied homomeric GIRK channels. Although some previous studies showed homomeric expression of GIRK1 and GIRK4 (43, 44), we found that neither the WT GIRK1 nor the WT GIRK4 expressed detectable basal currents recorded 2-4 days after cDNA injections. Similarly, we failed to express GIRK4 currents with cRNA injections. The GIRK1F137S and GIRK4S143T mutants have been shown to be expressed as homomeric channels (45) and are widely used in the study of GIRK channel modulations (7, 8, 12, 19, 20). However, we found that intracellular acidification activated both the channels rather than inhibiting them (data not shown). Because these mutations per se affected the channel sensitivity to pHi, they were rejected for further studies. Since mutations of cysteine 185 in GIRK4 and cysteine 179 in GIRK1 enable expression of constitutively active heteromeric GIRK1C179A/GIRK4C185A channels (42), it is possible that these cysteine mutations also allow homomeric expressions. Therefore, we injected these two mutants individually and observed that the GIRK4 but not GIRK1 expressed detectable inward rectifying currents. When the homomeric GIRK4C185A was studied, the mutant channel was inhibited by intracellular acidification to almost the same degree as the WT channels (34.7 ± 3.6%, n = 5, Fig. 3A). Thereafter, we decided to perform further studies using the homomeric GIRK4C185A channel.
To identify amino acid residues involved in proton sensing, we carried
out systematic mutational analysis on all histidine residues in the
intracellular protein domains as histidine is likely to be protonated
in the pH range 7.2-6.6 with its side chain pK of 6.04. Eight histidine residues are located in the cytosolic N and C termini
of GIRK4. When they were individually mutated to non-titratable
glutamine, all mutants except H278Q expressed functional channels. The
channel inhibition by intracellular protons was completely eliminated
in the H64Q and H228Q mutants. Instead, they were activated by
47.0 ± 4.9% (n = 6) and 57.0 ± 7.3%
(n = 5), respectively (Fig.
4). The H352Q mutant showed a significant
decrease in pH sensitivity (9.8 ± 2.6%, n = 6;
Fig. 4C). Mutations of all other histidines did not produce
any significant change in the pH sensitivity (Fig. 4C).
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Since mutation of critical amino acids in either GIRK1 or GIRK4 could
completely eliminate basal channel activity and agonist-induced currents of heteromeric GIRK1/GIRK4 channels (7, 8), it is possible
that mutation of histidines in either GIRK1 or GIRK4 subunit can also
remove the channel inhibition by intracellular protons. To test this
hypothesis, we examined the effect of histidine mutations in either
GIRK4 or GIRK1 on the heteromeric GIRK1/GIRK4 channels. The GIRK4 with
histidine mutations was co-injected with WT GIRK1 into oocytes, and
the cells expressing functional channels were challenged with
intracellular acidification. Consistent with the homomeric studies,
individual mutation of His-64 or His-228 in GIRK4 completely eliminated
the pHi-induced inhibition of the heteromeric channels (their
currents were enhanced by 21.8 ± 4.4%, n = 5, and 27.7 ± 6.8%, n = 12, respectively). The
inhibition of GIRK1/GIRK4H352Q by acidification was significantly
decreased in comparison with the WT GIRK1/GIRK4 (9.6 ± 3.1%,
n = 12; p < 0.05). In contrast, all
the other mutants, including GIRK1/GIRK4H22Q, GIRK1/GIRK4H63Q,
GIRK1/GIRK4H278Q, and GIRK1/GIRK4H 400Q, were still inhibited by
intracellular acidification (Fig. 5).
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To elucidate whether the corresponding histidines in GIRK1 were also
critical for the inhibition, we co-injected cDNAs of WT GIRK4
with GIRK1H57Q, GIRK1H222Q, or GIRK1H346Q, counterparts of His-64,
His-228, and His-352 in GIRK4. They all interrupted the channel
inhibition by intracellular protons (GIRK1H57Q-GIRK4: 0.0 ± 5.4%, n = 7; GIRK1H222Q-GIRK4: 85.6 ± 3.9%,
n = 4; GIRK1H346Q-GIRK4: 5.9 ± 3.2%,
n = 9). Furthermore, we studied heteromeric GIRK1/GIRK4 with multiple mutations of the histidine residues critical for the
pHi sensitivity. Effects similar to single mutations were
observed (Fig. 5). Joint mutations of other histidines had no
additional effect (Fig. 5), further suggesting that they are not
involved. Taken together, these results indicate that His-64, His-228,
and His-352 in GIRK4 and His-57, His-222, and His-346 in GIRK1 are
critical sites, any mutation of which is sufficient to disrupt the
pHi sensitivity of the heteromeric GIRK1/GIRK4 channels.
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DISCUSSION |
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In the present study, we have elucidated the inhibition of GIRK1/GIRK4 channels by intracellular acidification and shown evidence for the biophysical and molecular mechanisms underlying the inhibition. The inhibition of whole-cell GIRK currents is produced by strong suppression of channel open-state probability with modest inhibition of the single-channel conductance. Such an inhibition does not seem to require G-proteins and is likely to be a result of the direct interaction of protons with three histidine residues in the channel protein.
GIRK channels belong to a subfamily of inward rectifying K+ channels. Recent studies have shown that most of these channels are sensitive to intra- and/or extracellular acidification (33-37,46). In the present study, we have shown evidence for the pHi sensitivity of the heteromeric GIRK1/GIRK4 channels. Using 90 mM KHCO3 to replace all KCl in the extracellular solution, we selectively reduced the pHi without changing the pHo, a technique that has been widely used in lowering the pHi of Xenopus oocytes by several groups (33, 37, 47). We measured the pHi in our previous studies and found that the pHi was reduced to 6.6 under such an experimental condition. At this pHi level, whole-cell GIRK1/GIRK4 currents are inhibited by ~35%. We have studied the biophysical mechanism for the inhibition. We have found that acidic pHi strongly inhibits the Po with pK of 6.95, suggesting that these GIRK channels can be regulated under most physiological and pathophysiological conditions. Intracellular acidification also suppresses the single-channel conductance. The latter effect, however, is rather small (2-3% at a pHi of 7.0) in comparison with the pHi effect on Po (~20% at a pHi of 7.0). Therefore, the inhibition of whole-cell GIRK currents seems to attribute mainly to the inhibition of Po.
Intracellular acidification has been previously shown to inhibit cell-endogenous KACh channels in atrial myocardium (48). Consistent with our current observations, intracellular protons act on the Po in the cardiac K+ channels with modest inhibitory effect on single-channel conductance.
The involvement of G-proteins in the pH sensitivity has been examined
in the present study. Our results support the idea that the pH effect
does not require G-proteins for the following reasons. First, both
GIRK1/GIRK4C216T and GIRK1C179A/GIRK4C185A are equally sensitive to
pHi as the WT GIRK1/GIRK4. Whereas the former is known to be
less sensitive to G (41), the latter can neither be activated by
G
nor inhibited by G
(42). Second, in excised patches in the
absence of G-proteins, we observed similar inhibition of single-channel
currents. Third, a previous report has shown that G-proteins themselves
are activated at a low pHi (49). If the channel inhibition were
mediated by G-proteins, such G-protein activation would lead to a
stimulation of the GIRK channels rather than inhibition at acidic
pHi, which is exactly opposite to what we found. Thus, the
intracellular proton-induced inhibition of GIRK channels does not
appear to be a result of the interference of the GIRK channel-G-protein interaction.
Our results support the direct interaction of protons with the channel protein. By systematic scanning of all histidine residues in the channel proteins, three sites have been found to be critical, i.e. His-64, His-228, and His-352 in the GIRK4. Mutation of any of them totally abolished or greatly diminished the pHi sensitivity of the homomeric GIRK4C185A and the heteromeric GIRK1/GIRK4. These histidines are seen in the GIRK1 channel, too, i.e. His-57, His-222, His-346 in corresponding positions. They play an equally important role as mutations of them have the same effect on the pHi sensitivity of the heteromeric channels as the mutations in GIRK4. His-228 and His-352 in GIRK4 are conserved in all GIRK channels, whereas the His-64 is absent in GIRK3. The existence of some of these histidine residues in GIRK2 and GIRK3 channels suggests that they are likely to be pHi-sensitive as well, although seemingly to a less degree.
It is worth noting that the GIRK1F137S and GIRK4S143T mutants are activated by intracellular acidification instead of inhibition. The Phe-137 in GIRK1 and its corresponding site Ser-143 in GIRK4 have been shown to be critical for channel gating (50-52). The GIRK1F137S and GIRK4S143T mutants are widely used for homomeric expressions (7, 8, 12, 19, 20). The opposite effect on these channel gating-defective mutants to the WT channels suggests that the channel gating mechanism may be separate from proton binding in these GIRK channels.
Although the pHi sensitivity does not seem to interfere with G-protein binding in these GIRK channels, protonations of certain intracellular protein domains have been shown to affect other cellular functions. Previous studies have indicated that pH differentially modulates the effects of GTP and ATP on GIRK channels (48). Our recent studies have shown that protonation of a histidine residue in the intracellular C terminus affects the ATP sensitivity of the ATP-sensitive K+ channels through the allosteric mechanism (36, 53). In the Kir1.1 channel, intracellular protons produce direct inhibition, and reciprocally affect the interaction of the channel protein with phospholipids and protein phosphorylation (54, 55). Thereby, the possibility that protonations of histidines affect other functions of the GIRK channels cannot be excluded.
Recently, we have shown that GIRK1/GIRK4 channels are activated by
extracellular acidification, likely through protonation of a histidine
residue in the extracellular domain (20). Why do protons act on these
channels in such an opposite manner? The pHo sensitivity
appears to enhance synaptic transmission because it is known that a
large quantity of protons is released from neurotransmitter-containing
vesicles after vesicle fusion to presynaptic membranes (56). The
pHi sensitivity shown in the present study may be involved in
the regulation of membrane excitability within the postsynaptic cell.
Inhibition of the GIRK channels results in depolarization and an
increase in cellular excitability. In the brain, this may help recruit neurons and raise the arousal state of the central nervous system. In
the heart, the inhibition of the GIRK channels during intracellular acidosis may affect myocardium excitability (57, 58). It is also
possible that these channels detect a transmembrane pH gradient instead
of the absolute pH levels. In light of the inequality of the
pHi and pHo in most cells, and their relatively
independent regulations by cellular versus systemic mechanisms, the detection of pH gradient across the plasma membranes may constitute an important cellular function in neurons, myocardium, and other cells. Thus, the information about the pHi and
pHo sensitivities of the GIRK channels contributes to the
understanding of GIRK channel functions under physiological and
pathophysiological conditions.
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ACKNOWLEDGEMENT |
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Special thanks are due to Dr. Norman Davidson at California Institute of Technology, Pasadena, CA for the GIRK1 and GIRK4 cDNAs.
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FOOTNOTES |
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* This work was supported by the National Institutes of Health (Grant HL58410) and by the American Diabetes Association (Grant 1-01-RA-12).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
A Career Investigator of the American Lung Association. To whom
correspondence should be addressed: Dept. of Biology, Georgia State
University, 24 Peachtree Center Ave., Atlanta, GA 30302-4010. Tel.:
404-651-0913; Fax: 404-651-2509; E-mail: cjiang@gsu.edu.
Published, JBC Papers in Press, December 25, 2002, DOI 10.1074/jbc.M211461200
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ABBREVIATIONS |
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The abbreviations used are: GIRK, G-protein-coupled inward rectification K+; WT, wild type.
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