Department of Physiology and Pharmacology, State University of New York Health Science Center, Brooklyn, New York 11203
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ABSTRACT |
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Chuang, Shih-Chieh,
Riccardo Bianchi, and
Robert K. S. Wong.
Group I mGluR Activation Turns on a Voltage-Gated Inward Current
in Hippocampal Pyramidal Cells.
J. Neurophysiol. 83: 2844-2853, 2000.
A unique property of the group
I metabotropic glutamate receptor (mGluR)-induced depolarization in
hippocampal cells is that the amplitude of the depolarization is larger
when the response is elicited at more depolarized membrane potentials.
Our understanding of the conductance mechanism underlying this
voltage-dependent response is incomplete. Through the use of
current-clamp and single-electrode voltage-clamp recordings in guinea
pig hippocampal slices, we examined the group I mGluR-induced
depolarization in CA3 pyramidal cells. The group I mGluR agonists
(S)-3-hydroxyphenylglycine and (S)-3,5-dihydroxyphenylglycine turned on a voltage-gated
inward current (ImGluR(V)), which was
pharmacologically distinct from the voltage-gated sodium and calcium
currents intrinsic to the cells. ImGluR(V)
was a slowly activating, noninactivating current with a threshold at
about 75 mV. In addition to the activation of
ImGluR(V), group I mGluR stimulation also
produced a voltage-independent decrease in the K+
conductance. Our results suggest that the depolarization induced by
group I mGluR activation is generated by two ionic mechanisms
a heretofore unrecognized voltage-gated inward current
(ImGluR(V)) that is turned on by
depolarization and a voltage-insensitive inward current that results
from a turn-off of the K+ conductance. The low-threshold
and noninactivating properties of ImGluR(V)
allow the current to play a significant role in setting the resting
potential and firing pattern of CA3 pyramidal cells.
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INTRODUCTION |
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Metabotropic glutamate receptors (mGluRs) have
been classified into three groups based on structural homology and
their link to intracellular second-messenger systems (Nakanishi
1994). Physiological studies suggest that the different
subgroups of mGluRs have distinct roles in neuronal signal processing.
Activation of group II and III mGluRs causes presynaptic inhibition
(Baskys and Malenka 1991
; Desai et al.
1994
; Vignes et al. 1995
). In contrast,
activation of group I mGluRs affects postsynaptic function, causing
neuronal depolarization and excitation (Bianchi and Wong
1995
; Miles and Poncer 1993
; Pozzo Miller
et al. 1995
; Whittington et al. 1995
).
In the hippocampus, stimulation of group I mGluRs elicits neuronal
population oscillations at the (30-50 Hz) and
(12-30 Hz)
frequencies.
oscillations are elicited primarily by an
mGluR-induced excitation of inhibitory interneurons (Whittington
et al. 1995
), whereas
activities and the associated
epileptiform afterdischarges are mediated by an excitation of pyramidal
cells (Merlin and Wong 1997
; Taylor et al.
1995
).
oscillations, once induced, are long-lasting and
cannot be reversed by washing out the agonist (Merlin and Wong
1997
; Merlin et al. 1998
). At present, the
mechanisms whereby group I mGluRs excite hippocampal cells and elicit
long-lasting
oscillations remain unclear. We begin to explore this
issue by examining the action of group I mGluRs on individual pyramidal cells in hippocampal slices.
Group I mGluR activation produces depolarization of hippocampal neurons
via a suppression of potassium conductances (Charpak et al.
1990; Guérineau et al. 1994
; Harata
et al. 1996
; Lüthi et al. 1997
;
Stratton et al. 1989
) and an activation of nonspecific cationic conductances (Congar et al. 1997
;
Guérineau et al. 1995
; Pozzo Miller et al.
1995
). It has been noted that the amplitude of the
mGluR-mediated depolarization is enhanced significantly with cell
depolarization (Charpak et al. 1990
;
Guérineau et al. 1994
; Lüthi et al.
1997
). One study suggests that the voltage dependency of the
mGluR response is caused by an increase in potassium channel blockade
as the cell is depolarized (Lüthi et al. 1997
). We
have carried out experiments to describe more fully the conductance mechanism underlying the mGluR-mediated depolarization in CA3 neurons
and to evaluate whether the voltage dependency can be adequately
explained by the mechanisms described in the preceding text.
Portions of this work have been presented in abstract form
(Chuang et al. 1997, 1998
).
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METHODS |
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Slice preparation
Transverse hippocampal slices 400-µm thick were prepared from
adult guinea pigs as described previously (Bianchi and Wong 1995) and placed on nylon mesh in an interface recording
chamber (Fine Science Tools, BC, Canada). The control solution
consisted of (in mM) 157 Na+, 136 Cl
, 5 K+, 1.6 Mg2+, 2 Ca2+, 26 HCO3
, and 11 D-glucose. In some
experiments, Mn2+-containing solutions were used.
These solutions had added Mn2+ (1.5 mM) and
reduced Ca2+ (0.5 mM). Perfusion media were
bubbled with 95% O2-5%
CO2 to maintain the pH near 7.4, and the
temperature was at 34-36°C.
Electrophysiological recordings
Intracellular recordings of CA3 neurons were carried out using
an Axoclamp 2A amplifier (Axon Instruments; Foster City, CA). Electrodes were pulled with thin-wall glass tubings and had resistance of 30-40 M when filled with K-acetate (2 M) solution. Voltage and
current signals were digitized and stored in an Intel 386-based computer using a 12-bit A/D converter controlled by pClamp software (Axon Instruments). Voltage-clamp experiments were performed using the
single-electrode discontinuous clamp mode. The headstage output was
monitored continuously on an oscilloscope, and the switching frequency
(4-6 kHz) and gain (0.5-1.0 nA/mV) were adjusted so that the decay of
voltage transients was complete between switch cycles. Synaptic
stimulation was applied using tungsten bipolar electrodes (100 µs;
0.1-1 mA).
Pharmacological agents
To optimize the conditions for studying the mGluR-mediated
inward current, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 20 µM),
3-((R,S)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic
acid (CPP, 20 µM), and tetrodotoxin (TTX, 0.6 µM) were added to the perfusing solution. The mGluR agonists and antagonists
(S)-3-hydroxyphenylglycine (3HPG),
(S)-3,5-dihydroxyphenylglycine (DHPG),
(+)--methyl-4-carboxyphenylglycine (MCPG), and
(S)-4-carboxyphenylglycine (4CPG) also were added to the
perfusate. About 15-20 min was required for added drugs to equilibrate
in the recording chamber. The peak inward current amplitude induced by
Group I mGluR agonists was usually reached 45-60 min after the
addition of the agonist. This delay is probably caused by the slow
development of the inward current after mGluR stimulation. Glutamate
receptor ligands were purchased from Tocris Cookson (Ballwin, MO); all
the other chemicals were from Sigma (St. Louis, MO).
Estimation of the reversal potential of ImGluR(V)
Maximum outward current (I1, indicated in
Fig. 2C) activated during a hyperpolarizing pulse
(V2, indicated in Fig. 2A) is given
by
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(1) |
Peak inward ImGluR(V)
(I2) observed after the return of the
membrane potential back to the holding level
(V1) after the hyperpolarization is
given by
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(2) |
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(3) |
Student's t-test was applied and differences between data sets were considered to be significant when P < 0.05.
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RESULTS |
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Effects of group I mGluR agonists on the subthreshold properties of CA3 pyramidal cells
In CA3 pyramidal cells with resting potentials of 60 to
65 mV,
bath perfusion of the selective group I mGluR agonists 3HPG (50 µM,
n = 6) or DHPG (50 µM, n = 32)
produced depolarization of 7-18 mV and altered neuronal responses to
hyperpolarizing current injection. Before agonist application,
current-induced hyperpolarizations developed with an exponential time
course (Fig. 1A, Control). In
the presence of an mGluR agonist, responses to hyperpolarizing current
developed in two steps
an initial fast hyperpolarization followed by a
slower second phase (Fig. 1, DHPG). Recovery from hyperpolarization at
the end of the current pulse mirrored that recorded at the onset of the
hyperpolarization, consisting of an initial fast depolarization
followed by a slow phase (Fig. 1, DHPG). The amplitude and duration of
the slow phase during repolarization increased in proportion to the
amplitude of the hyperpolarization.
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Group I mGluR agonist-induced responses examined under voltage clamp
Because the subthreshold responses elicited by group I mGluR agonists exhibit voltage- and time-dependent changes, we examined the agonist effects under voltage clamp. All voltage-clamp experiments were carried out in the presence of TTX (0.6 µM) to suppress Na+ current.
Under the control condition, a hyperpolarizing command pulse from 45
to
80 mV elicited a brief surge of inwardly directed capacitive
current followed by a sustained inward ionic current (Fig.
2A). At the end of the
hyperpolarizing pulse, an outwardly directed surge of capacitive
current was observed before the membrane current returned to the
holding level (Fig. 2A). The time courses of the capacitive
current at the onset and release of the hyperpolarizing pulse were
fitted with single exponential functions with time constants of 24.7 and 29.5 ms, respectively.
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Addition of the group I mGluR agonist DHPG produced an inward shift of the holding current (Fig. 2B). A hyperpolarizing command pulse elicited the capacitive current and an inward ionic current as in the control condition. However, the inward current was not maintained for the duration of the hyperpolarizing pulse. A slow outward current appeared that was not observed in the control condition (Fig. 2B). At the end of the hyperpolarizing pulse, an initial outward surge of capacitive current was followed by a slow inward ionic current I2, returning the current to the holding level. The current responses activated during and after the hyperpolarization could be fitted with biexponential functions. The initial shorter time constants, largely reflecting the capacitive currents, were 24.5 and 32.6 ms for the responses at the onset and release of the hyperpolarization, respectively. These values for the capacitive current are similar to those of the control response. The longer time constants, reflecting the slow ionic current elicited in the presence of the agonist, had values of 543.5 and 894.0 ms for the responses during and after the hyperpolarization, respectively. In the presence of the mGluR antagonists MCPG (0.5-1 mM; n = 3) and 4CPG (0.5 mM; n = 2) in the bath, the effects of DHPG were prevented.
Conductance mechanism underlying the hyperpolarization-activated outward current induced by group I mGluR agonist
The outward ionic current I1 observed during the hyperpolarizing pulse may represent the turn-off of an inward current (involving a conductance decrease) or the turn-on of an outward current (involving a conductance increase). We measured the conductance changes accompanying the outward current to distinguish between the two possibilities.
The amplitude of the instantaneous current elicited by command voltage step is directly proportional to the cell conductance. The instantaneous current elicited at the onset of the hyperpolarization (providing an indication of the cell conductance at rest, prior to the hyperpolarizing step) was obtained by extrapolating the single-exponential fit for the outward current to the start of the hyperpolarizing pulse (Fig. 2C; I1 + I'). The instantaneous current associated with the termination of the hyperpolarizing step was similarly obtained (Fig. 2C; I2 + I'). Assuming that there is no significant shift in the relevant ionic concentrations during the hyperpolarizing pulse, the difference in the instantaneous currents elicited at the onset and at the termination of the hyperpolarizing pulse represents the difference in cell conductances at the onset of the hyperpolarization and at its termination. This difference in instantaneous currents is determined by subtracting (I2 + I') from (I1 + I') or, simply, I2 from I1. If I1 is larger than I2, the cell conductance at the start of the hyperpolarizing pulse is larger than that at the termination of the pulse.
We measured I1 and I2 of cells in response to a series of hyperpolarizations (Fig. 3). The values of the currents were measured from traces obtained after subtraction of the responses recorded in the presence of the agonist from those recorded in the control condition (Fig. 3, C and D). For any given hyperpolarization, I1 was always larger than I2 (Fig. 3, E and F), indicating that the cell conductance was always larger at the beginning of the hyperpolarizing pulse than at the end. The data indicate that the outward current activated during the hyperpolarization was associated with a conductance decrease, suggesting that group I mGluR stimulation activates an inward current (ImGluR(v)) that is turned off by hyperpolarization and turned on by depolarization.
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The values of I1 and
I2 also allowed an estimation of the
reversal potential for ImGluR(V) (see
METHODS). For the cell shown in Fig. 3, corresponding
I1 and
I2 measurements were obtained for
seven levels of hyperpolarizations (from 80 to
110 mV in 5-mV
steps) yielding seven values for
EmGluR(V). The mean
ImGluR(V) reversal potential for this
cell was
12.3 ± 3.0 mV. The mean reversal potential value
obtained in this way for a total of five cells was
9.8 ± 1.1 mV.
In addition to the activation of
ImGluR(V), stimulation of group I
mGluRs also produced a voltage-independent decrease in the baseline
conductance of the cell. The group I mGluR agonist reduced the
instantaneous currents elicited both at the onset (Ion) and release
(
Ioff) of the hyperpolarizing
pulses compared with the control response. This was reflected in
subtracted traces where the instantaneous currents
(
Ion and
Ioff) were always outward (e.g.,
Fig. 3D). Figure 3D also shows that
Ioff is larger than
Ion. This observation is consistent
with the hypothesis that an outward current is turned off by the
hyperpolarizing pulse.
Relationship between ImGluR(V) and ICa
CA3 pyramidal cells have robust voltage-gated Na+ and Ca2+ conductances. Because the preceding experiments were carried out in TTX, the contribution of intrinsic Na+ conductance to ImGluR(V) appears unlikely. We explored a possible relationship between ImGluR(V) and intrinsic voltage-gated Ca2+ currents.
Nimodipine (10 µM), a blocker of L-type calcium channels, was applied to the perfusing solution after group I mGluR stimulation (n = 4; Fig. 4). The agent blocked the generation of Ca2+ spikes (Fig. 4C). The amplitude of ImGluR suppressed by hyperpolarization was not affected by nimodipine (Fig. 4, A and B). In addition, current-clamp experiments carried out in the same cell showed that the slow hyperpolarization responses induced by group I mGluR agonists persisted in the presence of nimodipine (Fig. 4C, right).
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In other experiments, Mn2+ (1.5 mM), a nonspecific Ca2+ conductance blocker, was added to a solution with reduced Ca2+ (0.5 mM) after ImGluR(V) was elicited. Mn2+ blocked Ca2+ spikes in the recorded cell without affecting the amplitude of ImGluR(V). The hyperpolarization-activated outward current induced by DHPG persisted under Mn2+ treatment (n = 3; data not shown).
Effects of group I mGluR agonists on the baseline conductance of CA3 pyramidal cells
On occasion, the two effects of DHPG, namely the reduction in cell conductance and the activation of ImGluR(V), appeared separately during the wash in of the agonist. DHPG first produced a decrease in instantaneous current responses to hyperpolarizing steps (early response; Fig. 5, A(ii) and B). Over time and with continued agonist perfusion, the slowly developing hyperpolarization-activated outward current appeared (delayed response; Fig. 5A(iii)).
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Figure 5B shows a plot of the current-voltage
(I-V) relationship of the cell in response to
hyperpolarizing pulses. The I-V relationship for both the
control () and early (
) test responses were linear. The two lines
cross at about
90 mV, close to the K+
equilibrium potential of the cell. The separation of early and delayed
responses was observed in two other cells. Extrapolated reversal
potentials obtained from these two cells were
123 and
127 mV.
Activation properties of the group I mGluR-induced current
The data suggest that ImGluR(V)
is an inward current activated by depolarization. We examined the
activation threshold of the current (Fig.
6). In the presence of DHPG, cells were
held at 45 mV. A conditioning pulse of
90 mV was applied for 750 ms
to deactivate ImGluR(V) (Fig.
6B). This was followed by a series of 2-s test pulses from
90 to
20 mV in 5-mV increments. Inward current
(ImGluR(V)) first appeared when the
test pulses were at about
75 mV (Fig. 6, D and
E). The amplitude of
ImGluR(V) progressively increased with
increasing levels of test depolarization and peaked between
30 to
20 mV. Once activated, ImGluR(V)
persisted for the duration of the depolarization without showing
inactivation.
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Effects of group I mGluR stimulation on the resting potential and firing pattern of CA3 pyramidal cells
Because ImGluR(V) is a persistent current and it had a threshold close to the resting potential, we observed interesting modulations of the resting potential by group I mGluR agonists. In current-clamp recordings, cells exhibiting stable depolarized resting potentials in the presence of DHPG occasionally shifted their resting potential to a more hyperpolarized level. The hyperpolarized resting potential was brought about by an incomplete recovery of the cell response from a hyperpolarizing pulse (Fig. 7A). Figure 7B shows that an inhibitory postsynaptic potential (IPSP) could reset the resting potential of the cell to a more hyperpolarized level in a manner similar to that elicited by intracellular current injection.
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For cells resting at the more hyperpolarized state in the presence of the group I mGluR agonist, depolarizing currents could reset the resting potential to a more depolarized level (Fig. 7C). In such instances, the fast phase depolarization elicited by the applied current was succeeded by a slow phase depolarization usually causing phasic cell firing. After the phasic firing, the membrane potential often stayed at the depolarized level. In six cells examined, bilevel swings of the resting potential ranged from 8 to 19 mV. This phenomenon of bilevel resting potential was not observed in cells under the control condition.
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DISCUSSION |
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The data indicate that stimulation of group I mGluRs activates a
novel inward current (ImGluR(V)) with
a reversal potential at about 10 mV. A distinct property of
ImGluR(V) is its voltage dependency:
the amplitude of ImGluR(V) increases
monotonically when the cell is depolarized (Fig. 6). The threshold of
activation of ImGluR(V) is between
75 and
70 mV and maximum activation of
ImGluR(V) occurs at about
30 mV. In
addition to the activation of
ImGluR(V), group I mGluR agonists also
elicit a voltage-independent reduction in K+ conductance.
The low activation threshold of
ImGluR(V) (75 to
70 mV)
facilitated our studies of the current.
ImGluR(V) can be examined using
hyperpolarizing pulses from
45 mV. Hyperpolarizing pulses at this
range also should activate Iq
(Perkins and Wong 1995
), the only other time- and
voltage-dependent persistent current operating at this voltage range.
Activation of Iq would produce a slow
inward current opposing the development of
ImGluR(V). Current- (Fig. 1) and
voltage-clamp (e.g., Figs. 2 and 3) recordings show that no noticeable
time-dependent current developed in response to hyperpolarizing pulses
under the control condition. This observation is consistent with
previous findings suggesting that Iq
is not prominently expressed in CA3 cells (Bianchi et al.
1999
) in contrast to its significant presence in CA1 cells
(Perkins and Wong 1995
). To further ensure that the
properties of ImGluR(V) were examined in isolation from other contaminating responses, all the data used for
analysis were obtained by subtracting the control responses from those
recorded in the presence of an agonist.
Group I mGluRs activate a voltage-dependent depolarizing current
There are two major mechanisms by which stimulation of mGluRs has
been shown to depolarize hippocampal cells. These are an increase in a
nonspecific conductance to cations (Congar et al. 1997;
Guérineau et al. 1995
) and a decrease in
conductance to potassium ions (Charpak et al. 1990
;
Guérineau et al. 1994
; Harata et al.
1996
; Lüthi et al. 1997
; Stratton
et al. 1989
). A recent study also shows that group I mGluR
stimulation elicited an inward current in basolateral amygdala neurons
via an activation of the Na+-Ca2+exchange
(Keele et al. 1997
). This current differs from
ImGluR(V) in that it is not
accompanied by conductance change and is insensitive to voltage
(Keele et al. 1997
). The mGluR-activated opening of nonspecific cationic channels involves the direct action of G proteins
(Pozzo Miller et al. 1995
) or intracellular
Ca2+ increase (Congar et al. 1997
)
and does not show voltage-gated properties. In contrast,
voltage-dependent mGluR-induced depolarizations have been noted by
others (Charpak et al. 1990
; Guérineau et al. 1994
; Lüthi et al. 1997
) and have been
attributed to an mGluR-mediated suppression of potassium conductances.
The voltage dependency of the response was explained by assuming that
the agonist-mediated conductance block itself is voltage dependent and
is markedly increased at depolarized potentials (Lüthi et
al. 1997
). However, the voltage-dependent depolarization
induced by group I mGluR activation in our studies is accompanied by a
conductance increase and therefore cannot be attributed to a blockade
of potassium channels. Instead, the present data indicate that the
depolarization is mediated by a novel, voltage-gated inward current
that requires cell depolarization for its activation.
A similar dichotomy exists for the explanation of the muscarinic
receptor-mediated depolarization. Although a decrease in potassium
conductance had been suggested to be a primary action mediated by
muscarinic receptors (Benardo and Prince 1982;
McCormick and Prince 1986
), more detailed examination
revealed that the fundamental mechanism for the muscarinic
depolarization of cortical neurons involves the activation of a
voltage-dependent inward current with a reversal potential near
15 mV
(Haj-Dahmane and Andrade 1996
). The authors suggest that
the voltage-dependent properties of the current led to the apparent
conductance decrease observed under voltage- or current-clamp measurements.
A recent study by Wu and Barish (1999) showed that group
I or group II mGluR agonists suppressed
ID (Wu and Barish 1992
)
by acceleration of its inactivation. The activation threshold of ID is depolarized to
45 mV, thus the
effects of mGluR agonists on ID should
not affect most of our study demonstrating the deactivation of I
mGluR(V) by hyperpolarizing pulses from a
holding potential of
45 mV (Figs. 2-5). The action of mGluR agonists
on ID may introduce overestimation in
our measurements of ImGluR(V) at
depolarizing pulses above
45 mV (Fig. 6). However the peak activation
of ID is depolarized to +30 mV,
whereas ImGluR(V) is maximally
activated at around
30 to
20 mV. This suggests that the inward
current caused by an accelerated inactivation of
ID may not be a major factor affecting
the measurement of ImGluR(V). However,
the detailed effects of the mGluR agonist modulation of
ID on the kinetic properties of
ImGluR(V) can only be derived from
studies using dendrotoxin and 4-aminopyridine.
Because of the complex structure of hippocampal pyramidal cells, the data obtained in this study will be affected by space-clamp errors. However, these errors may not be a major factor in determining the measurements of the time course of ImGluR(V) because different amplitudes of ImGluR(V) appearing at the same membrane potential developed with comparable time constants (Fig. 3), whereas systematic variations in the activation time constant were observed when ImGluR(V) developed at different levels of depolarization (Fig. 6).
Although we emphasize the existence of a voltage-dependent inward current turned on by group I mGluR activation, we also observed the coexistence of a voltage-independent conductance decrease contributing to the production of mGluR-mediated depolarization. The reduction in cell conductance can occur separately over time from the development of ImGluR(V) during agonist wash in (Fig. 5) and is expected to amplify the ImGluR(V)-mediated effects on the membrane potential and firing pattern of pyramidal cells.
ImGluR(V): a novel, voltage-dependent inward current activated by group I mGluRs
Five unique features of ImGluR(V)
distinguish this current from other known voltage-dependent inward
currents, including Na+ and
Ca2+ currents, intrinsic to hippocampal neurons:
1) ImGluR(V) is
pharmacologically distinct from Na+ and
Ca2+ currents because it persists in the presence
of TTX and Ca2+ channel blockers (Fig. 4).
2) ImGluR(V) is
noninactivating. In voltage-clamp experiments, after a hyperpolarizing
test pulse (80 mV) and on the return of the membrane potential to the
holding level (
45 mV; e.g., Fig. 2), the current develops slowly to a maximum in ~1 s. This maximum current shows no sign of decay and is
maintained for
10 s, the longest interval at which cells were held at
the holding potential in our experiments before another test pulse. The
extent to which ImGluR(V) persists
greatly exceeds that demonstrated for intrinsic
Na+ and Ca2+
conductances. 3) The voltage-dependent activation and
deactivation of ImGluR(V) develops
with time constants on the order of several hundred
milliseconds
significantly longer than those of the
Na+ and Ca2+ currents.
4) The threshold of
ImGluR(V) is more hyperpolarized (between
75 to
70 mV) than that of other known noninactivating or
slowly-inactivating voltage-dependent inward currents. The latter
include L-, N-, P-, and Q-type Ca2+ currents, all
of which require depolarization beyond
45 mV for activation
(Bean 1989
; Hess 1990
; Hillman et
al. 1991
; Zhang et al. 1993
). And 5)
ImGluR(V) is not detectable under
normal conditions and appears only when group I mGluRs are stimulated.
Bistable resting potentials associated with the generation of ImGluR(V)
The low-threshold and noninactivation properties
of ImGluR(V) allowed it to
contribute in an unexpected way to the maintenance of the resting
membrane potential. The data suggest that persistent activation of
ImGluR(V) sustained the pyramidal
cells in a depolarized state. As a result, in current-clamp recordings,
hyperpolarizing pulses turned off the persistent
ImGluR(V) and produced the slow phase
hyperpolarization (e.g., Fig. 1, A and B).
Conversely, on release of the hyperpolarization,
ImGluR(V) was turned on. Activation of
ImGluR(V) caused regenerative
depolarization of the cell, eliciting the slow phase depolarization
(Fig. 1, A and B). A stable depolarized membrane
potential then is attained when the inward
ImGluR(V) is balanced by counteracting
outward currents. Intrinsic Na+ and
Ca2+ currents also can have a tonic influence on
the resting potential within the range where the activation and
inactivation curves overlap (e.g., Hughes et al. 1999).
The contribution of these intrinsic currents to the group I
mGluR-mediated changes in the resting potential is not significant
because robust voltage-dependent shifts in membrane potential still are
elicited by hyperpolarizing pulses in pyramidal cells exposed to DHPG
when Na+ and Ca2+ channels
are blocked (Fig. 4C). In this way, group I mGluR-mediated effects on pyramidal cells differ from those shown in other
preparations (dorsal horn cells, Morisset and Nagy 1996
;
turtle motoneurons, Svirskis and Hounsgaard 1998
), where
Ca2+ channel antagonists blocked the responses
mediated by mGluRs. Svirskis and Hounsgaard (1998)
showed that group I mGluR action on the turtle motoneurons consisted
solely of a depolarization associated with a reduction in input
conductance and that this response was sufficient to promote
regenerative events driven by intrinsic Ca2+ currents.
ImGluR(V) also enables the hippocampal cells to rest at two membrane potentials. Sufficient hyperpolarization can totally deactivate ImGluR(V) and reset the membrane potential to a hyperpolarized level. Physiologically, hippocampal cells can be driven to operate at the hyperpolarized state by inhibitory postsynaptic potentials (Fig. 7).
The properties of ImGluR(V) can be
compared with those of the inward current activated by
hyperpolarization (Iq) (Perkins and Wong 1995). Previous studies emphasized the importance of Iq in regulating the pacing activities
of hippocampal neurons (Maccaferri and McBain 1996
;
Perkins and Wong 1995
). The two features of
Iq that allowed it to contribute in
this manner are its activation at the subthreshold range and its
noninactivation properties
properties similarly possessed by
ImGluR(V). The major difference
between these two currents is that Iq
is activated by hyperpolarization, whereas
ImGluR(V) is activated by
depolarization. Accordingly, Iq may
have a more dominant role in shaping the subthreshold properties of the
cell, whereas ImGluR(V) may play a
bigger role in regulating the pacing and firing activities of the cell.
We have shown previously that via intrinsic and synaptic interactions,
group I mGluR agonists induce hippocampal neurons into prolonged
epileptiform afterdischarges (Taylor et al. 1995
).
Additional studies showed that these group I mGluR-mediated effects are
long-lasting (Merlin and Wong 1997
; Merlin et al.
1998
) and may constitute an epileptogenic process. By eliciting
periodic prolonged depolarizations of hippocampal pyramidal cells,
ImGluR(V) may sustain the prolonged
epileptiform afterdischarges. We used group I mGluR agonists to induce
the slow inward current and the epileptiform activity. At present there
is no information on whether synaptically released glutamate can induce
these responses. Experiments involving tetanization of glutamatergic
pathways will be needed to establish the duration and extent of
synaptic stimulation that is required to induce ImGluR(V) and epileptiform afterdischarges.
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ACKNOWLEDGMENTS |
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We thank Dr. Lisa Merlin for providing helpful input to the manuscript.
This work was supported by National Institute of Neurological Disorders and Stroke Grant NS-35481.
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FOOTNOTES |
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Address for reprint requests: R.K.S. Wong, SUNY Health Science Center, 450 Clarkson Ave., Box 29, Brooklyn, NY 11203.
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.
Received 22 October 1999; accepted in final form 7 February 2000.
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