Institut für Physiologie, Medizinische Fakultät, Otto-von-Guericke Universität, D-39120 Magdeburg, Germany
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ABSTRACT |
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Shah, Mukesch Johannes, Susanne Meis, Thomas Munsch, and Hans-Christian Pape. Modulation by Extracellular pH of Low- and High-Voltage-Activated Calcium Currents of Rat Thalamic Relay Neurons. J. Neurophysiol. 85: 1051-1058, 2001. The effects of changes in the extracellular pH (pHo) on low-voltage- (LVA) and high-voltage- (HVA) activated calcium currents of acutely isolated relay neurons of the ventrobasal thalamic complex (VB) were examined using the whole cell patch-clamp technique. Modest extracellular alkalinization (pH 7.3 to 7.7) reversibly enlarged LVA calcium currents by 18.6 ± 3.2% (mean ± SE, n = 6), whereas extracellular acidification (pH 7.3 to 6.9) decreased the current by 24.8 ± 3.1% (n = 9). Normalized current amplitudes (I/I7.3) fitted as a function of pHo revealed an apparent pKa of 6.9. Both, half-maximal activation voltage and steady-state inactivation were significantly shifted to more negative voltages by 2-4 mV on extracellular alkalinization and to more positive voltages by 2-3 mV on extracellular acidification, respectively. Recovery from inactivation of LVA calcium currents was not significantly affected by changes in pHo. In contrast, HVA calcium currents were less sensitive to changes in pHo. Although extracellular alkalinization increased maximal HVA current by 6.0 ± 2.0% (n = 7) and extracellular acidification decreased it by 11.9 ± 0.02% (n = 11), both activation and steady-state inactivation were only marginally affected by the moderate changes in pHo used in the present study. The results show that calcium currents of thalamic relay neurons exhibit different pHo sensitivity. Therefore activity-related extracellular pH transients might selectively modulate certain aspects of the electrogenic behavior of thalamic relay neurons.
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INTRODUCTION |
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Both normal and pathological
neuronal activity is accompanied by distinct changes in extracellular
and/or intracellular pH (for review see Chesler and Kaila
1992; Kaila and Ransom 1998
). Even small changes
in pHo and/or pHi have been
shown to affect normal synaptic transmission and a variety of
ligand-gated and voltage-gated ion channels, as, for instance,
glutamate and GABA receptors (Pasternack et al. 1992
;
Traynelis 1998
) and voltage-dependent calcium channels
(Church 1999
; Kiss and Korn 1999
;
Mironov and Lux 1991
; Tombaugh and Somjen
1997
, 1998
). In light of the importance of
calcium ions to the pathogenesis of epileptogenic activity, pH-dependent modulation of voltage-dependent calcium conductances may
be a key mechanism leading to the pathological synchronization of
neuronal activity. Calcium currents in acutely dissociated adult rat
CA1 hippocampal neurons, for instance, are sensitive to changes in both
pHo and pHi
(Tombaugh and Somjen 1996
). Moreover, low-voltage- (LVA)
and high-voltage- (HVA) activated calcium currents seem to exhibit
differential sensitivity to changes in pHi
(Tombaugh and Somjen 1997
). Alkaline
pHo has been shown to increase calcium currents,
whereas acidic pHo decreases the current
(Tombaugh and Somjen 1998
). However, it remains unclear
to what extent changes in pHo might
differentially affect either LVA or HVA calcium currents.
Thalamic relay neurons possess both HVA and LVA calcium
currents (Hernandez-Cruz and Pape 1989), which
critically determine their firing pattern. HVA calcium currents in
thalamic relay neurons are thought to regulate tonic firing of action
potentials at depolarized membrane potentials (Budde et al.
2000
; Guyon and Leresche 1995
; Kammermeier and Jones 1997
; Zhou et al.
1997
). The pH sensitivity of HVA calcium currents has been
studied in detail in other types of neurons (Mironov and Lux
1991
; Tombaugh and Somjen 1996
,
1997
; Zhou and Jones 1996
).
The LVA calcium current, by comparison, is considered an
important element in the generation of rhythmic oscillatory electrical activity, in that it underlies the generation of a low-threshold calcium spike (LTS) (Llinas and Jahnsen 1982;
McCormick and Bal 1997
; Steriade et al.
1993
). Oscillatory generation of LTSs is controlled by the
interplay of LVA calcium current and a hyperpolarization-activated cation current (Ih), and synaptic
mechanisms (McCormick and Bal 1997
). Recently we have
shown that voltage dependence of activation of
Ih is influenced by
pHi rather than pHo
(Munsch and Pape 1999
). In comparison, pH-modulation of
LVA calcium currents in neurons is less well understood.
We have therefore addressed the question of the pHo sensitivity of LVA and HVA calcium currents in thalamic relay neurons and the consequences of a modulation of LVA currents by extracellular protons for LTS generation by the use of patch-clamp techniques on thalamic relay neurons either acutely isolated from the rat ventrobasal thalamic complex (VB) or within an in situ slice preparation of VB.
Part of this work was performed in partial fulfillment of an
MD thesis by M. J. Shah. Part of this work has been presented in
abstract form (Munsch et al. 2000).
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METHODS |
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Preparation
Long Evans rats of either sex [postnatal day 11-17 (P11-P17)] were anesthetized with halothane and decapitated. A block of tissue containing the VB was quickly removed from the rest of the brain and placed in chilled oxygenated PIPES-buffered solution containing (in mM) 123 NaCl, 2.4 KCl, 10 MgSO4, 0.5 CaCl2, 20 PIPES, 23 sucrose, and 10 dextrose; pH 7.25. Coronal slices (300-400 µm) containing the VB were made from the thalamus. For in situ whole cell patch-clamp measurements, thalamic slices were immediately transferred to a submersion chamber filled with artificial cerebrospinal fluid (ACSF) containing (in mM) 120 NaCl, 2.5 KCl, 22-26 NaHCO3, 1.25 NaH2PO4, 2 MgSO4, 2 CaCl2, and 10 glucose, pH adjusted to 7.35 with 95% O2-5% CO2. Slices were maintained at room temperature (22-25°C).
For acute isolation of neurons, the VB was carefully dissected from neighboring tissue under stereoscopic observation. Slices were placed in an oxygenated PIPES solution containing (in mM) 20 PIPES, 115 NaCl, 5 KCl, 2.6 MgCl2, 25 dextrose, and 0.5 CaCl2, pH 7.35 and warmed up to 30°C. Slices were then incubated in protease-containing (1 mg/ml) PIPES solution for 25-40 min.
After washing in enzyme-free medium, the neurons were mechanically dissociated by triturating with fire-polished Pasteur pipettes.
Electrophysiological recording
For in situ recordings, whole slices were transferred to an
experimental chamber. Individual cells were approached by visual control with differential interference contrast infrared (DIC-IR) videomicroscopy. Recordings were made with an EPC-9 amplifier operating
Pulse software (HEKA, Lambrecht, Germany). Measurements were performed
at room temperature (22-25°C) in ACSF. The composition of the
pipette solution used for current-clamp experiments was as follows (in
mM): 95 K-gluconate, 20 K-citrate, 10 NaCl, 10 HEPES, 1 MgCl2, 0.5 CaCl2, 1 K-bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid
(BAPTA), 3 Mg-ATP, and 0.5 Na2-GTP, pH adjusted
to 7.4 with KOH. The pH of HCO
Acutely isolated cells were electrophysiologically analyzed with the
patch-clamp technique in whole cell voltage-clamp mode using an EPC-7
amplifier (List Medical System, Darmstadt, Germany). Patch pipettes
were made from borosilicate glass (GC150TF-10, Clark Electromedical
Instruments, Pangbourne, UK). Typical electrode resistance was 2-5
M, with access resistance of 6.75 ± 0.07 M
(mean ± SE, n = 68). Records were low-pass filtered at
2.5 kHz (8-pole Bessel filter). Voltage-clamp experiments were
performed using PClamp software operating via Labmaster interface (Axon Instruments, Foster City, CA) on an IBM computer. Each test pulse at
the various pH values was preceded by a prepulse to
110 mV to regard
possible pH effects on leakage currents. However, there was no
significant difference between leakage currents at the pHo values used. Leakage currents were then
compensated off-line by estimating the electronic, ohmic component of
the membrane current during the prepulses and subtracting each of the
scaled values from the corresponding current traces.
Isolated neurons were continuously superfused (0.1-1 ml per minute) with extracellular solution containing (in mM) 134 NaCl, 10 HEPES, 2 KCl, 15 dextrose, 15 D-mannitol, 2 CaCl2, 3 MgCl2 (pH 7.34 with NaOH). A multibarreled laminar-flow perfusion system (0.1 ml/min) was placed close to the recorded neuron allowing to completely change the solution surrounding the recorded cell within <500 ms. For analyzing the current-voltage (I-V) relationship, kinetics, activation, and inactivation at pH 6.9, 7.3, 7.7, and 8.1 we used a HEPES-buffered solution containing (in mM) 120 NaCl, 1 KCl, 3 CaCl2, 1 MgCl2, 20 dextrose, 10 mannitol, 6 4-aminopyridine, 20 TEA, 0.0015 TTX, and 10 HEPES. pH was adjusted to the desired value with NaOH or HCl. For all acidic solutions below a pH of 6.9, HEPES was replaced with equimolar PIPES.
The internal solution contained 70 mM CsCl, 15 mM CsOH, 10 mM NaCl, 1 mM KCl, 11 mM EGTA, 2 mM MgCl2, 1 mM
CaCl2, 20 mM TEA, 5 mM ATP, 0.5 mM GTP, 15 mM
phosphocreatin, 50 units creatine kinase, and 50 mM HEPES to minimize
changes of internal pH (Irisawa and Sato 1986). The
internal solution was adjusted to pH 7.2 with CsOH. Experiments were
performed at room temperature (21-24°C). After establishing the
whole cell configuration, active currents were monitored by periodic
voltage steps until the current had stabilized. Recordings begun
typically 3-6 min after rupturing the membrane patch.
Isolation of LVA and HVA currents
LVA and HVA calcium components were separated using a
conditioning pulse protocol (Tsakiridou et al. 1995). A
100-ms prepulse to
50 mV between the hyperpolarizing pulse to
110
mV (1,000 ms) and the depolarizing voltage steps was used to inactivate LVA calcium currents. LVA calcium currents were isolated by digital subtraction of records obtained with and without the conditioning prepulse. Pharmacological tools to separate different calcium currents
were renounced, because of possible pH-dependent effects of the drugs
(Galizzi et al. 1984
; Platt et al. 1993
).
Data analysis
Analyses of current records, including curve fitting, were done
with the use of PClamp software. Inactivation and activation curves
were obtained by fitting the data points to a Boltzmann function
{y = A2 + (A1 A2)/[1 + exp(x
x0/dx)], where
x0 is the half-maximal activation voltage, dx is
the slope factor, and A1 and A2 are constants}. Data are presented as
means ± SE and were statistically evaluated using a paired
t-test (Origin 4.1).
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RESULTS |
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pH effects on LVA calcium currents
LVA calcium currents could be evoked by depolarizing voltage
steps from a prepotential of 110 mV to various test potentials in
acutely isolated VB relay neurons (Fig.
1, A and C).
Currents activated at
70 mV and peaked near
40 mV. Modest changes
in the pH of the superfusing solution led to distinct shifts in the peak amplitude of the I-V curve. Extracellular acidification
from pH 7.3 to 6.9 caused a decrease of the peak amplitude of LVA
calcium currents of 24.8 ± 3.1% (n = 9), whereas
extracellular alkalinization reversibly increased peak currents by
18.6 ± 3.2% (n = 6; Fig. 1, B and
D). By fitting normalized current amplitudes
(I/I7.3) as a function of
the pH of the extracellular solution (pHo), a Hill-plot was obtained, from which an apparent
pKa of 6.9 was estimated (slope factor = 14.5; Fig. 1E). Peaks of the I-V curves were not
significantly shifted along the voltage axis by either acidic
pHo (6.9) or alkaline pHo
(7.7; Fig. 1, B and D).
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To determine the effects of pHo on gating
properties of LVA calcium channels, activation curves were obtained
from tail currents elicited by stepping to 80 mV from various
depolarizing test potentials as shown in inset of Fig.
2A. Activation curves were reversibly shifted by changes in pHo. Acidic
pHo (6.9) moved the half-maximal activation
voltage from
54.9 ± 1.9 mV (n = 4) to
51.6 ± 1.3 mV (n = 4; Fig. 2A).
Conversely, alkaline pHo (7.7) significantly
(P
0.05) shifted the half-maximal activation voltage to more negative potentials from
53.8 ± 1.2 mV
(n = 5) to
55.8 ± 1.1 mV (n = 5; Fig. 2B). Time for current activation showed typical
voltage dependence with time constants (
) becoming larger at more
negative voltages (Fig. 2, C and D). Time
constants were not significantly affected by changes in
pHo at potentials more positive than
30 mV but
increased at more negative potentials for acidic
pHo (Fig. 2C) and decreased for
alkaline pHo (Fig. 2D), respectively.
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Steady-state inactivation of LVA calcium current was analyzed with
conditioning pulses of 1,000 ms duration between 110 and
45 mV
followed by a constant test pulse to
40 mV (inset of Fig. 3A). Inactivation curves were
obtained from plots of the normalized current
(I/Imax) versus the
potential of the conditioning prepulse. The inactivation curves were
fitted with a Boltzmann equation. Changing the external pH from 7.3 to
6.9 caused a significant (P
0.05) shift of the
half-maximal voltage (V1/2) of
1.85 ± 0.6 mV (n = 7) to more positive potentials
(Fig. 3A). Conversely, alkaline pHo
(7.7) reversibly shifted V1/2 by
2.6 ± 0.7 mV (n = 7) to more negative potentials
without changing slope parameters (Fig. 3B). Time constants
of inactivation were determined by fitting single exponential equations
to the decay of individual LVA calcium current traces. The rate of
current inactivation became faster at more depolarized potentials (Fig.
3, C and D). Again, time constants of
inactivation were unaffected by pHo at
30 mV or more positive potentials, but differed significantly at more negative potentials. For instance, at
50 mV, extracellular acidification resulted in a slower rate of inactivation and, conversely, alkaline pHo led to a faster inactivation of LVA calcium
currents (Fig. 3, C and D).
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To study the time course of recovery from inactivation, cells were held
at a potential of 50 mV to inactivate LVA calcium currents
completely. Hyperpolarizing command steps to
110 mV were then applied
with increasing intervals followed by a constant test pulse to
40 mV.
Increasing the duration of hyperpolarization was accompanied by an
increase in current amplitudes. The fractional recovery of normalized
currents (I/Imax) showed
the typical time dependence (data not shown). A significant
pHo dependence was not observed.
pH effects on HVA calcium currents
HVA calcium currents were isolated through inactivation of the LVA
currents by a 100 ms prepulse to 50 mV (Kammermeier and Jones
1997
). Sustained inward currents could be activated by step depolarizations above
40 mV (Fig. 4,
A and B), and the I-V curve revealed a
peak near
5 mV (Fig. 4, C and D). At acidic
pHo (6.9), HVA calcium currents became depressed
by 11.9 ± 2.6% (n = 11), while current
amplitudes were increased at alkaline pHo (7.7) by 6.0 ± 2.0% (n = 7; Fig. 4, C and
D). A Hill-fit of normalized peak HVA calcium currents
(I/I7.3) plotted as a
function of pHo revealed an apparent
pKa of 6.5. The moderate changes in
pHo used in the present experiments only
marginally affected the half-maximal activation and the rate of HVA
calcium current activation (data not shown). Also, neither moderate
extracellular acidosis (6.9, n = 5) nor alkalosis (7.7, n = 6) altered the steady-state inactivation (P
0.3). The rate of HVA calcium current
inactivation was only marginally affected by either acidic
pHo (6.9, n = 11) or alkaline pHo (7.7, n = 7), but this effect
was more variable and seldom reached statistical significance (data not
shown). The small effects of pHo changes on HVA
channel gating are likely due to differential pHo
sensitivity of HVA subtypes, but this was not further investigated.
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Effects of bath pH on burst activity
The consequences of pHo effects on calcium
currents for the electrogenic activity of relay neurons were tested in
an in vitro slice preparation of the thalamus containing the VB. We
concentrated on low-threshold burst activity in view of the
well-documented contribution of the LVA current to this type of
activity. By repetitive hyperpolarization through injection of negative
current pulses, rebound LTSs can be evoked, which mimics rhythmic
oscillatory activity as occurs during burst mode, the major activity
pattern of relay neurons at relatively hyperpolarized membrane
potentials (McCormick and Bal 1997). In a brain slice
superfused with a bath solution of pH 7.3 (control, HEPES-buffered)
repetitive injection of hyperpolarizing current (15 × 100 ms
pulses at a frequency of 2.5 Hz) under current-clamp conditions was
adjusted to elicit LTSs at every second current pulse (Fig.
5A, top trace,
n = 6). These LTSs were typically crowned by 2-5 fast
sodium/potassium action potentials (inset of Fig. 5,
A and B). Changing bath pH to 6.9 significantly
reduced the frequency of LTS generation, whereas a more alkaline bath
pH (7.7) caused a significant increase in LTS frequency (Fig.
5A, middle and bottom traces). The
same results were obtained for brain slices bathed in solutions with HCO
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DISCUSSION |
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The present investigation indicates that extracellular H+ may act as a modulator of LVA calcium currents due to proton-induced changes in the activation properties of the underlying channels, thereby relating activity-dependent extracellular pH transients to changes in the activity patterns of thalamic relay neurons.
pHo versus pHi effects on calcium currents
Changes in pHo have been shown to affect
intracellular pH in smooth muscle (Klöckner and Isenberg
1994) and in rat hippocampal neurons (Church et al.
1998
). In the present study, indirect effects of
pHo changes on calcium currents via intracellular
H+ were minimized by modest changes in
pHo within a physiological range and using
strongly buffered pipette solutions (50 mM HEPES) (Irisawa and Sato 1986
; Tombaugh and Somjen
1996
; Tytgat et al. 1990
; Zhou and Jones
1996
). Moreover, the effects of changes in pHo were always fast, which would require either
appreciable proton currents or proton exchange systems for rapidly
changing pHi. In contrast, in an in situ slice
preparation of VB we have previously shown that changes in
pHi due to changes of bath pH were typically slow
and too small to significantly affect other
pHi-dependent conductances, as for instance, the
hyperpolarization-activated cation current
(Ih), present in VB neurons
(Munsch and Pape 1999
). In summary, these findings argue
in favor of an extracellular proton modulatory effect on calcium
currents when changing the pH of the bathing solution, at least in a
physiological range. An extracellular proton sensitivity for calcium
currents has been described in a variety of cell types (Church
1999
; Irisawa and Sato 1986
; Tombaugh and
Somjen 1996
; Tytgat et al. 1990
; Zhou and
Jones 1996
).
Possible mechanisms of pHo sensitivity of LVA and HVA
The effects of physiologically relevant changes of
pHo on LVA and HVA calcium currents, described in
the present paper, could in principle be due to
pHo-induced gating shifts or changes in channel
conductance, or both. The underlying mechanisms are thought to either
involve screening of fixed negative charges facing the surface of the
cell membrane, which will cause a shift in the membrane potential
perceived by the voltage sensor of a particular ion channel, or direct
binding of H+ ions to negative groups within the
channel, which will interfere with current flow through the channel
pore. The effects of changes in pHo on activation
and inactivation kinetics of both LVA and HVA calcium currents of
thalamic relay neurons support the interpretation that the primary
effect is due to surface charge screening by extracellular
H+. For heterologously expressed 1H and
1G
currents, it was recently shown that a proton-induced change in channel
gating accounted for most of the effects of extracellular pH shifts on
current amplitude (Delisle and Satin 2000
; Kozlov
et al. 2000
). Also, the voltage dependence of deactivation of
1H currents was not affected by changes in pHo
(Delisle and Satin 2000
), which is consistent with a
reduction of negative surface potential by protons (Hille et al.
1975
). Evidence for both surface charge screening and channel
block by H+ was found for T-type cardiac calcium
channels (Tytgat et al. 1990
), N-type calcium channel
currents of bullfrog sympathetic neurons (Zhou and Jones
1996
), and L-type Ca2+ channels expressed
in Xenopus oocytes (Chen et al. 1996
).
Differential pHo sensitivity of LVA and HVA currents
In isolated rat hippocampal CA1 neurons, a differential
sensitivity to changes in pHi has been observed
between LVA and HVA calcium currents. LVA calcium currents were more
sensitive to changes in pHo and appeared
relatively insensitive to pHi changes, as
revealed by exposing CA1 neurons to weak acids and bases, thereby affecting pHi, and by exposure to bathing
solutions of different pHo (Tombaugh and
Somjen 1997). The results of our study suggest a similar
preferential pHo sensitivity of LVA calcium
currents in thalamic relay neurons. However, HVA calcium currents in
thalamic relay neurons were much less affected by changes in
pHo than HVA Ca2+ channels
in cortical neurons (Ou-Yang et al. 1994
), hippocampal neurons (Church et al. 1998
; Tombaugh and Somjen
1996
), and retinal photoreceptors (Barnes et al.
1993
), which is supported by a lower pKa
value as compared with hippocampal neurons (pKa = 6.5, this study vs. pKa = 7.1, Tombaugh
and Somjen 1996
; and pKa = 7.2, Church et al.
1998
). For LVA calcium currents of relay neurons, we found a
pKa of 6.9, which is very similar to that found
for the cloned T-type calcium channel subunit
1G expressed in
HEK-293 cells (6.5-7.0) (Kozlov et al. 2000
). Through
this differential pHo sensitivity of LVA and HVA
calcium currents, certain aspects of the electrogenic activity of
thalamic relay neurons might be selectively modulated during
activity-related pHo transients, which have been
shown to accompany normal as well as pathological neuronal activity
(for review see Kaila and Ransom 1998
).
Functional significance for oscillatory activity in the thalamocortical network
HVA calcium currents in thalamic neurons are thought to regulate
tonic firing of fast action potentials, thereby providing Ca2+ influx necessary for calcium-induced release
of Ca2+ from intracelluar stores (CICR) and
subsequent activation of Ca2+-dependent
K+ currents (Budde et al. 2000;
Guyon and Leresche 1995
; Hernandez-Cruz and Pape
1989
; Kammermeier and Jones 1997
). The tonic
firing mode has been associated with functional states of arousal
(McCormick and Bal 1997
; Steriade et al.
1993
). The LVA calcium current, by comparison, is thought to
regulate rhythmic oscillatory patterns of electrical activity, which
appears as burst discharges during slow-wave sleep or generalized
epilepsy (McCormick and Bal 1997
; Sherman and
Guillery 1996
). Recently we have shown that the two major modes of relay cell activity are accompanied by distinct patterns
of intracellular pH changes that potentially contribute to the control
of mode switching of relay neurons (Meyer et al. 2000
).
The differences in the H+ sensitivity of calcium
channels of thalamic relay cells provide a mechanism by which certain
aspects of the electrogenic behavior of relay neurons can be
selectively modulated. For instance, depression of HVA calcium currents
by intracellular acidosis may limit Ca2+ influx
during tonic activity representing a negative feedback to prevent
excessive Ca2+ influx. Such a
pHi sensitivity of Ca2+
dynamics has recently been shown in rat CA1 neurons (Tombaugh 1998
). The situation seems more complicated, however, in view of recent reports on an inhibition of
Ca2+-dependent K+ current
activation by a fall in pHi (Church et al.
1998
; Tombaugh 1998
), possibly resulting in
broadening of action potentials and increase in
Ca2+ influx. The net effect of those interactions
for neuronal integrative behavior in thalamus remains to be elucidated.
By comparison, the preferential pHo sensitivity
of LVA calcium currents might provide a mechanism more relevant in the
control of Ca2+ influx during burst mode. During
this activity mode, relay neurons rhythmically generate LTSs
(Llinas and Jahnsen 1982; Steriade et al.
1993
). Since modulation of LVA calcium currents is considered an important element for rhythmic membrane potential oscillations in
thalamocortical neurons (McCormick and Bal 1997
), it is
obvious that activity-related pHo changes
potentially influence bursting behavior by affecting the generation of
LTSs. Indeed, we found that extracellular acidification by 0.4 pH units
drastically reduced the frequency of LTS generation in relay neurons,
maintained in an in vitro slice preparation, whereas extracellular
alkalinization caused the opposite effect (Fig. 5). By afferent
stimulation (5 s at 10 Hz) of the dorsal lateral geniculate nucleus,
transient alkaline pHo shifts of 0.04 ± 0.02 pH units followed by acid shifts of 0.05 ± 0.03 pH units
could be elicited in rat thalamic slices (Tong and Chesler
1999
). The magnitude of these activity-dependent pHo shifts is smaller than the artificial changes
in pHo used in the present study. However, the
tips of ion-selective microelectrodes cause artificial enlargement of
the extracellular space volume. Therefore pHo
shifts recorded by ion-selective microelectrodes in brain slice
preparations are likely attenuated representations of the actual
pHo shifts occurring in the small extracellular space. Also, during sustained electrical activity and synchronized oscillatory activity, as occurs in the thalamocortical network under
normal and pathological conditions, activity-related
pHo shifts may amount to several tenths of a pH unit.
Recently, in rat CA1 hippocampal pyramidal neurons, the
Ca2+-mediated rebound response following a
current-evoked hyperpolarization was shown to be modulated by changes
in pHo (Church 1999). Therefore it
seems feasible to speculate that activity-related
pHo transients may contribute to the control of
rhythmic oscillatory activity also in the thalamocortical network.
Synchronized oscillations in the thalamocortical network have been
shown to be under control of a hyperpolarization-activated cation
conductance as a key mechanism for the modulation of duration and
frequency of network oscillations (Bal and McCormick
1996
). Previously we have shown that activity-related intracellular pH shifts may contribute to the temporal control of
network oscillations by modulation of the hyperpolarization-activated cation conductance (Munsch and Pape 1999
). The
pHo sensitivity of LVA calcium channels make them
likely candidates for an additional mechanism by which the propensity
of the thalamocortical network to generate synchronized oscillations
can be controlled. However, it remains to be demonstrated whether
activity-related pHo shifts during synchronized
network activity are large enough to affect LTS generation in
thalamocortical neurons. The steep pHo dependence of LVA current amplitude and activation kinetics in particular at
physiologically relevant pH clearly argues in favor of such a mechanism.
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ACKNOWLEDGMENTS |
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Thanks are due to R. Ziegler for excellent technical assistance.
This work was supported by the Deutsche Forschungsgemeinschaft (Pa 336/13-1).
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FOOTNOTES |
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Address for reprint requests: T. Munsch, Institut für Physiologie, Medizinische Fakultät, Otto-von-Guericke Universität, Leipziger Str. 44, D-39120 Magdeburg, Germany (E-mail: thomas.munsch{at}medizin.uni-magdeburg.de).
Received 12 June 2000; accepted in final form 20 November 2000.
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REFERENCES |
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