Department of Physiology, Marshall University School of Medicine, Huntington, West Virginia 25755-9340
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ABSTRACT |
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Grover, Lawrence M. and Chen Yan. Blockade of GABAA receptors facilitates induction of NMDA receptor-independent long-term potentiation. An N-methyl-D-aspartate (NMDA)-independent form of long-term potentiation (LTP), which depends on postsynaptic, voltage-dependent calcium channels (VDCCs), has been demonstrated in area CA1 of hippocampus. GABA acting at GABAA receptors limits postsynaptic depolarization during LTP induction. Blockade of GABAA receptors should therefore enhance activation of postsynaptic VDCCs and facilitate the induction of this NMDA receptor-independent, VDCC-dependent LTP. In agreement with this hypothesis, pharmacological blockade of GABAA receptors in the in vitro rat hippocampal slice increased the magnitude of LTP resulting from a normally effective, high-frequency (200 Hz) tetanic stimulation protocol. In addition, GABAA receptor blockade allowed a lower frequency (25 Hz) and normally ineffective tetanic stimulation protocol to induce this form of LTP. Intracellular recordings from CA1 pyramidal cells revealed that blocking GABAA receptors during tetanic stimulation allowed greater postsynaptic depolarization, increased the number of postsynaptic action potentials fired during the tetanization, and also increased the duration of synaptically evoked action potentials. To mimic the increased action potential firing observed when GABAA receptors were blocked, we paired 25-Hz antidromic stimulation with 25-Hz orthodromic stimulation. Paired antidromic + orthodromic 25-Hz stimulation induced NMDA receptor-independent LTP, whereas neither antidromic nor orthodromic stimulation alone induced LTP. Increased action potential firing can therefore at least partially account for the facilitation of NMDA receptor-independent LTP caused by blockade of GABAA receptors. This conclusion is consistent with prior studies demonstrating that action potentials are particularly effective stimuli for the gating of VDCCs in CA1 pyramidal cell dendrites.
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INTRODUCTION |
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High-frequency (200 Hz) tetanization can induce a
slow onset, N-methyl-D-aspartate (NMDA)
receptor-independent long-term potentiation (LTP) in hippocampal area
CA1 (Cavus and Teyler 1996; Grover 1998
; Grover and Teyler 1990
, 1992
, 1994
, 1995
). Previous
studies (Cavus and Teyler 1996
; Grover and Teyler
1990
) indicated that induction of this NMDA
receptor-independent LTP requires Ca2+ influx through
postsynaptic voltage-dependent Ca2+
channels (VDCCs). During tetanization, GABAA
receptor-mediated inhibition limits the degree of postsynaptic
depolarization and postsynaptic action potential firing and thus limits
the magnitude of Ca2+ influx into postsynaptic neurons
(van der Linden et al. 1993
). Relief from
GABAA inhibition should therefore enhance postsynaptic Ca2+ entry and thereby facilitate induction of LTP.
Although it was shown that LTP can be enhanced by blockade of
GABAA receptors (Wigström and Gustafsson
1983
), this enhancement was attributed to an increase in
postsynaptic Ca2+ influx through NMDA receptors
(Wigström and Gustafsson 1985
) because the
blockade of GABAA receptors allows greater postsynaptic depolarization and thus more complete relief from the voltage-dependent block of NMDA receptors by Mg2+. The first objective of
this study was to determine if GABAA receptors regulate the
induction of an NMDA receptor-independent LTP similar to the GABAergic
regulation of NMDA receptor-dependent LTP.
A previous study (Grover 1998) found that firing of
action potentials by the postsynaptic CA1 pyramidal neurons during
tetanization was required for induction of NMDA receptor-independent
LTP. Other studies have shown that action potential waveforms are
effective stimuli for the gating of high-threshold Ca2+
currents (McCobb and Beam 1991
; Scroggs and Fox
1992
). In addition, propagation of action potentials into
dendrites generates VDCC-dependent dendritic Ca2+ signals
(Magee and Johnston 1995
; Spruston et al.
1995
). These observations suggest that postsynaptic action
potential discharge during tetanization is critical for triggering
Ca2+ entry into the postsynaptic neurons through VDCCs. A
second objective of this study then was to determine if blockade of
GABAA receptors would alter postsynaptic action potential
discharge in a manner that would be expected to facilitate
Ca2+ influx through VDCCs.
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METHODS |
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Slice preparation
Hippocampal slices were prepared from 1- to 2-mo-old male and female Sprague-Dawley rats (Hilltop Laboratory Animals). Animals were sedated by inhalation of a CO2-air mixture and decapitated. The skull was opened, and the brain was removed and submerged in chilled, oxygenated (95% O2-5% CO2), low-Ca2+/high-Mg2+ artificial cerebrospinal fluid (ACSF) composed of (in mM) 124 NaCl, 26 NaHCO3, 3 KCl, 0.5 CaCl2, 5.0 MgSO4, and 10 glucose. After cooling in the low-Ca2+/high-Mg2+ ACSF, the brain was trimmed and glued to the stage of a vibratome and immersed in a bath of chilled, oxygenated, low-Ca2+/high-Mg2+ ACSF. Coronal sections, 400 µm thick, were then cut. Sections containing the hippocampus in transverse profile were selected and transferred to a small petri dish, where they were further dissected to free the hippocampus from surrounding tissue. In most slices, the CA3 region was also removed to prevent any spontaneous synaptic activity driven by the bursting of CA3 neurons in the presence of bicuculline. However, identical results were obtained in slices where area CA3 was left intact. After dissection, hippocampal slices were transferred to a holding chamber, where they were stored for later use.
Slices were maintained in the holding chamber at room temperature
(18-22°C) at the ACSF-atmosphere (95% O2-5%
CO2) interface. The holding chamber was filled with
standard ACSF containing (in mM) 124 NaCl, 26 NaHCO3, 3.4 KCl, 1.2 NaH2PO4, 2.0 CaCl2, 2.0 MgSO4, and 10 glucose. Slices were incubated in the holding
chamber for 1 h before use.
Slices were withdrawn from the holding chamber as needed and placed in a low-volume (~200 µl) interface recording chamber, where they were continuously perfused at a rate of 1-2 ml/min with standard ACSF. The recording chamber was kept at a temperature of 35 ± 0.15°C. After transferring slices from the holding to the recording chamber, a minimum 30-min period was allowed for recovery.
Electrophysiology
Extracellular potentials were recorded through low-impedance
(1-2 M) glass micropipettes, filled with ACSF, and placed into the
stratum radiatum of area CA1. Signals were amplified (gain 100-1000)
and filtered (DC-3,000 Hz or 0.1-10,000 Hz). After processing, signals
were digitized at 10-40 kHz and stored on a personal computer. Excitatory postsynaptic potentials (EPSPs) were evoked by delivery of
constant current stimuli through a bipolar stimulating electrode placed
into stratum radiatum. EPSPs were evoked at an interstimulus interval
of 30 s. In some slices a second stimulating electrode was placed
into the alveus, on the subicular side of the slice, to evoke
antidromic population spikes (see Fig. 8, B and
C, for examples). Electrode placements and stimulus
intensities were adjusted to maximize the antidromic spike without
evoking an orthodromic response, which can result if sufficient
stimulus current flows into the adjacent stratum oriens to excite
afferent fibers. Antidromic population spikes were typically 2-3 mV in
amplitude. In some slices, an antidromic population spike could not be
evoked without contamination by orthodromic responses; these slices
were excluded from analysis.
Intracellular recordings were used to observe postsynaptic potentials
evoked by single stimuli and by tetanic stimulation. Individual CA1
pyramidal cells were impaled with glass microelectrodes filled with
2-4 M potassium acetate (60-110 M), connected to the headstage of
an Axoclamp 2B amplifier operating in bridge mode. Cell input
resistances were estimated by injection of small hyperpolarizing and
depolarizing currents. Output from the amplifier was low-pass filtered
at 1-2 kHz, digitized at 2-5 kHz, and stored on a personal computer.
Most extracellular and intracellular records were stored and analyzed with custom routines written in the Axobasic (Axon Instruments) programming environment. These routines allowed the extraction of waveform maxima, minima, slopes, and areas. Some recordings were made with Axotape (Axon Instruments) or WCP (Strathclyde Electrophysiology Software, John Dempster, University of Strathclyde) programs. Supplemental data analysis was performed with Origin (Microcal Software) and Excel (Microsoft).
Slices were tetanized in the presence of 50-100 µM
D,L-2-amino-5-phosphonopentanoic acid (AP5) or
AP5 + MK-801 (dizocilpine, 20 µM) to test for induction of NMDA
receptor-independent LTP (Grover 1998; Grover and
Teyler 1990
, 1992
, 1994
, 1995
) and to observe postsynaptic
responses to tetanic stimulation. Tetanic stimulation consisted of
either a single 25-Hz, 1-s stimulus train or a series of four 200-Hz,
0.5-s stimulus trains presented at an intertrain interval of 5 s.
In some slices, the GABAA antagonist bicuculline (10 µM)
was applied with AP5 or AP5 + MK-801. The stimulus intensity used for
tetanization was set to evoke a population spike, measured with an
extracellular electrode in s. pyramidale, of 1-2 mV. In slices
perfused with bicuculline, stimulus intensities were selected before
application of the drug. Test stimuli, delivered before and after
tetanic stimulation, were evoked with the stimulus intensity set to
one-half that used for tetanization.
Drug application
AP5, MK-801, and bicuculline methiodide (obtained from Tocris and RBI) were first dissolved in water to make concentrated stock solutions (5-100 mM). Stock solutions were diluted 250- to 1,000-fold into ACSF for delivery to slices. Drugs were applied to the tissue by switching between reservoirs containing ACSF and ACSF + drug.
Data analysis
For each slice, EPSP slopes were averaged over a 10-min baseline period preceding tetanization. Each EPSP was then normalized relative to the averaged baseline response. EPSP measurements are reported as percentage change from the baseline.
Postsynaptic responses to tetanic stimulation were quantified in two ways. First, the response area (in mV · ms) of the intracellularly recorded membrane potential during tetanization was determined. For each cell, the response area was calculated by summing the area under each point in the digitized waveform, measured relative to the pretetanus resting membrane potential. This was used as an index of the overall magnitude of postsynaptic depolarization. Second, the number of action potentials occurring during a stimulus train was manually counted. Action potential duration was determined by measurements taken at the base of the action potential (defined as 10 mV positive to threshold).
For statistical analysis, responses were averaged over all the cells or slices in a group. Values are reported as means ± 1 SE. Statistical comparisons were made by use of independent or dependent samples t-tests, as appropriate, with P < 0.05 (two-tailed) considered significant.
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RESULTS |
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Block of GABAA receptors enhances NMDA receptor-independent LTP
In previous studies (Grover 1998; Grover and
Teyler 1990
, 1992
, 1994
, 1995
), NMDA receptor-independent LTP
was induced by a series of four 0.5-s, 200-Hz stimulus trains delivered
in the presence of the competitive NMDA receptor antagonist AP5. As in prior reports, slices tetanized with this protocol displayed an LTP
requiring ~10-20 min to reach maximum amplitude (Fig.
1A). Some degree of
potentiation was seen in all 11 slices examined (range: 15-38%
increase in EPSP). Slices tetanized at 200 Hz in AP5 showed a mean
increase in EPSP slope of 26 ± 2% (n = 11, averaged over a period from 40-45 min post-tetanus). Addition of
bicuculline to the ACSF enhanced the degree of NMDA
receptor-independent LTP obtained (Fig. 1A). For slices
tetanized in AP5 + bicuculline, the mean increase in EPSP was 55 ± 12% (n = 10, range from 10-140% increase, 40-45
min post-tetanus). The magnitude of LTP obtained in AP5 + bicuculline
was significantly greater than in AP5 alone (P < 0.02, Fig. 1C).
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Although our previous studies (Grover 1998;
Grover and Teyler 1990
, 1994
) found no contribution of
NMDA receptors to the LTP induced by the same procedures used in this
experiment, others (Pananceau and Gustafsson 1997
), with
similar procedures, reported that NMDA receptor activation could
contribute to LTP induction, despite the presence of AP5. To determine
whether residual, AP5-resistant, NMDA receptor activation might have
contributed to the facilitation of LTP produced by bicuculline, we
repeated the experiment of Fig. 1A, but this time we
tetanized slices at 200 Hz in the presence of both AP5 (a competitive
NMDA receptor antagonist) and MK-801 (20 µM, a noncompetitive
antagonist). If some fraction of NMDA receptors was unblocked by AP5
alone, then the combination of AP5 + MK-801 should reduce the magnitude
of LTP obtained. Again we found that bicuculline (10 µM) increased
the magnitude of LTP (Fig. 1B). In AP5 + MK-801, EPSPs were
potentiated by 39 ± 9% (n = 13, range from
7-105% increase, measured 40-45 min post-tetanus), whereas the
addition of bicuculline increased the magnitude of LTP to 75 ± 14% (n = 11, range from 22-165% increase, 40-45 min post-tetanus). This difference was significant (P < 0.04, Fig. 1C). As previously noted (Grover
1998
), MK-801 did not affect the magnitude of LTP obtained by
200-Hz tetanization in the presence of AP5; LTP resulting from
tetanization in AP5 + MK-801 was on average slightly greater than LTP
resulting from tetanization in AP5 alone, but this small difference
(26 ± 2% vs. 39 ± 9%) was not significant
(P > 0.17, see Fig. 1C). LTP resulting from tetanization in bicuculline was also unaffected by MK-801 (AP5 + bicuculline vs. AP5 + MK-801 + bicuculline, P > 0.27, see Fig. 1C). These observations are inconsistent with any
contribution of AP5-resistant NMDA receptors to the LTP studied here.
Under conditions where GABAA inhibition is intact,
high-frequency tetanic stimulation is required for the induction of
NMDA receptor-independent LTP. Lower-frequency tetanic stimulation (25 Hz) can induce NMDA receptor-dependent LTP but ordinarily does not
result in NMDA receptor-independent LTP (Grover and Teyler 1990,
1994
). Consistent with these prior reports, a single 25-Hz, 1-s
stimulus train delivered in the presence of AP5 failed to induce LTP
(Figs. 2 and 9). However, when
bicuculline was perfused with AP5, 25-Hz stimulation was sufficient to
induce an NMDA receptor-independent LTP in ~50% of the slices
examined (5/11 slices, Fig. 2). In this subset of slices, the mean
increase in EPSP after 25-Hz tetanization in AP5 + bicuculline (31 ± 3%) was significantly greater (P < 0.01) than the
change in EPSP seen in the slices tetanized at 25 Hz in AP5 only
(
1 ± 7%). In 6 of 11 slices tetanized at 25 Hz in AP5 + bicuculline, the change in EPSP (
8 ± 5%) was not significantly
different (P > 0.40) from slices tetanized at 25 Hz in
AP5 only.
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In some slices, perfusion of AP5 + bicuculline was continued throughout the post-tetanus recording period (Fig. 3A), whereas in other slices AP5 and bicuculline were washed out (Fig. 3B). LTP was observed under both of these conditions, indicating that the maintenance of NMDA receptor-independent LTP does not require a change in the strength of GABAA inhibition.
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Basis for enhancement of NMDA receptor-independent LTP by GABAA receptor blockade
Intracellular recordings were obtained from CA1 pyramidal cells to gain insight into the mechanisms by which GABAA receptors might regulate NMDA receptor-independent LTP. The general procedure in these experiments was to record the postsynaptic membrane potential during tetanization (while perfusing AP5) and then apply bicuculline and repeat the previous tetanic stimulation. Cells were exposed to series of four 200-Hz stimulus trains or to single 25-Hz stimulus trains.
One potential complication of these experiments was that the first tetanization (in AP5) might induce LTP, leading to an enhanced postsynaptic response to the second tetanic stimulation (in AP5 + bicuculline). Any enhancement caused by LTP might then be confused with changes caused by the blockade of GABAA receptors. Two approaches were taken to circumvent this problem.
The first approach took advantage of the fact that NMDA
receptor-independent LTP is expressed with a delayed onset
(Grover 1998; Grover and Teyler 1990
, 1992
, 1994
,
1995
). After an initial enhancement resembling post-tetanic
potentiation, which typically lasts from 1 to 3 min, the response
returns toward the baseline level (returning all the way to the
baseline in ~1/2 of the slices examined previously). This is
followed by the gradual emergence of a sustained potentiation. With the
perfusion apparatus used here, solution exchanges could be completed
within 3 min. It was therefore possible to tetanize a slice, wash
bicuculline onto the slice, and tetanize the slice again at a time
point after the decay of PTP and preceding the development of LTP. An
example of this procedure is illustrated in Fig.
4. Single EPSPs were recorded immediately
before the first series of tetanic stimuli (in AP5) and immediately
before the second series of tetanic stimuli (in AP5 + bicuculline). As
shown in Fig. 4A, the rising slopes of the EPSPs were
identical, demonstrating an identical level of excitatory synaptic
transmission for both series of tetanic stimuli. Figure 4A
also shows that the fast inhibitory postsynaptic potential (IPSP) was
adequately blocked by the GABAA antagonist at the time of
the second set of tetanic stimuli. In addition, the cell input
resistance was measured before tetanization in AP5 and again after
tetanization in AP5 + bicuculline (Fig. 4B). There was no
detectable change in input resistance.
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A second approach was also used to ensure that the only difference between the first and second episodes of tetanic stimulation was the blockade of GABAA inhibition. In this second approach, responses to 25-Hz tetanization were examined. Because 25-Hz tetanization in AP5 alone fails to induce LTP (Figs. 2 and 8A), it was possible to record responses to 25-Hz, 1-s stimulus trains in AP5 and then add bicuculline to the ACSF and record a response to the same stimulus train with confidence that the level of excitatory synaptic transmission was unaltered.
Both approaches yielded similar results (Figs. 4C and 6).
The most salient effect of bicuculline on the response to tetanization was an increase in the number of action potentials fired during the
stimulus train. In the experiments with 200-Hz tetanic stimulation, this effect was especially pronounced on the first train in each series
of four (see Train 1, Fig. 4C; Fig.
5). Although this difference was
maintained to some degree during later trains in the series, the effect
became less prominent. This is most likely a reflection of the
use-dependent depression of inhibition that follows high-frequency stimulation (McCarren and Alger 1985). In other words,
by the third and fourth trains of a series delivered in AP5 alone,
GABAA inhibition is already depressed so that the addition
of the GABAA antagonist has considerably less effect.
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Bicuculline increased the underlying envelope of depolarization during
200-Hz tetanization (Figs. 4C and 5); this effect presumably was responsible for the increased action potential discharge. Two
hundred-hertz tetanization can elicit a depolarizing GABAA response (Grover et al. 1993), which itself can evoke
action potential discharge. Because the depolarizing GABAA
response is blocked by bicuculline, bicuculline potentially might have
decreased the number of action potentials fired during 200-Hz
tetanization. This turned out not to be the case, however, because the
loss of the hyperpolarizing GABAA response component (Fig.
4C, Train 1, single arrows) had a greater overall impact on
the postsynaptic membrane potential during tetanization than did the
loss of the depolarizing GABAA response (Fig.
4C, Train 1, double arrows).
Bicuculline similarly affected the postsynaptic responses to 25-Hz tetanization and 200-Hz tetanization. The number of action potentials fired during 25-Hz stimulation was increased by bicuculline (Fig. 6, A-C). With the lower frequency of tetanization, individual EPSPs could be easily discerned. Bicuculline facilitated action potential firing by increasing the peak amplitude of the EPSPs (Fig. 6D), thereby allowing a greater proportion of the responses to reach spike threshold. Bicuculline did not affect the rising slope of the EPSPs (Figs. 4A and 6D).
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In addition to changing the number of action potentials evoked during tetanic stimulation, blocking GABAA receptors also changed the shape of individual, synaptically evoked action potentials (Fig. 7). In the presence of bicuculline, action potential duration, measured at the base of the action potential, was increased from 1.25 ± 0.07 ms to 1.54 ± 0.09 ms (P < 0.02). This effect was reversed by washing bicuculline off the slice (Fig. 7).
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Bicuculline increased both the number and duration of action
potentials, and either of these changes could facilitate induction of
NMDA receptor-independent LTP. We conducted a final experiment to test
the importance of the increase in number of action potentials fired.
Three groups of slices were examined in this experiment. In all three
groups, tetanic stimulation was delivered in AP5, and GABAA
inhibition was intact (bicuculline was not administered). One group was
tetanized with a single 25-Hz, 1-s stimulus train delivered through a
stimulating electrode placed in s. radiatum (orthodromic tetanization,
identical to the tetanization protocol used in Fig. 2). A second group
was simultaneously tetanized with 25-Hz, 1-s stimulation of the
Schaffer collaterals and 25-Hz, 1-s stimulation of the alveus
(orthodromic + antidromic tetanization) to mimic the increased
postsynaptic action potential firing observed when GABAA
inhibition was blocked. A third group received only the 25-Hz, 1-s
antidromic (alvear) stimulation. The results of this final experiment
are shown in Fig. 8. As in the previous experiment (Fig. 2), 25-Hz tetanization in the presence of AP5 failed
to induce LTP (Fig. 8A, mean EPSP change measured 30 min post-tetanus was 2 ± 5%). Orthodromic tetanization paired with antidromic tetanization, however, was effective for inducing LTP (Fig.
8B, mean EPSP change = 17 ± 3%,
P < 0.02 compared with results with orthodromic
tetanization alone). Antidromic tetanization by itself (Fig.
8B) failed to induce LTP and instead resulted in a
persistent decrease in EPSPs (mean change = 12 ± 7%) as previously reported (Christofi et al. 1993
;
Pockett et al. 1990
). These results are consistent with
the previous finding (Grover 1998
) that postsynaptic
action potential firing is required for induction of NMDA
receptor-independent LTP. These results also indicate an important role
for GABAA inhibition in regulating NMDA
receptor-independent LTP by controlling the number of postsynaptic action potentials fired during tetanization but do not rule out a
further contribution through control of action potential duration.
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DISCUSSION |
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The data presented here demonstrate that blockade of GABAA receptor-mediated inhibition increases the magnitude of NMDA receptor-independent LTP obtained with a normally effective stimulation protocol (200-Hz tetanization). Furthermore, a normally ineffective stimulation protocol (a single 25-Hz tetanus) can become effective if GABAA-mediated inhibition is blocked. Observations of postsynaptic responses to tetanic stimulation under control conditions (AP5 alone) and with GABAA receptors blocked (AP5 + bicuculline) revealed an increase in the number of action potentials fired during tetanization and an increase in the duration of individual action potentials.
We conducted an experiment to test whether an increase in the number of
action potentials fired during tetanic stimulation was sufficient to
change the normally ineffective 25-Hz tetanization protocol to an
effective protocol for inducing NMDA receptor-independent LTP. In this
experiment, we paired simultaneous 25-Hz, 1-s orthodromic stimulation
of the Schaffer collateral/commissural fibers with 25-Hz, 1-s
antidromic stimulation of alvear fibers. The orthodromic + antidromic
pairing protocol was effective for LTP induction, confirming the
critical role of postsynaptic action potential firing in NMDA
receptor-independent LTP (Grover 1998). Although the
orthodromic + antidromic tetanization protocol mimicked the increased
number of postsynaptic action potentials fired in bicuculline, this
protocol did not mimic the increased action potential duration observed
in bicuculline. For this reason, the importance of action potential
duration for the induction of NMDA receptor-independent LTP will need
to be assessed by further experimentation. Nonetheless, this experiment
does support the conclusion GABAA receptors can regulate
the induction of NMDA receptor-independent LTP by controlling action
potential discharge in postsynaptic neurons.
The conclusions reached above are consistent with previous studies
where Ca2+ influx into CA1 pyramidal neurons was directly
examined. Action potentials propagate into the dendritic tree
(Spruston et al. 1995) where they open VDCCs
(Christie et al. 1995
; Magee and Johnston 1995
; Miyakawa et al. 1992
; Spruston et
al. 1995
). The entry of Ca2+ into the dendrites of
CA1 pyramidal cells during orthodromic stimulation is largely dependent
on propagation of action potentials into dendritic processes
(Christie et al. 1995
; Miyakawa et al. 1992
). The influx of Ca2+ through VDCCs
depends not only on the number of action potentials fired but also on
the shape of the action potential, with longer action potentials being
more effective for gating VDCCs, especially high-threshold VDCCs
(McCobb and Beam 1991
; Scroggs and Fox
1992
). Because bicuculline increased both the duration and the
number of action potentials during tetanization, it is almost certain that the block of GABAA receptors led to a greater
VDCC-mediated Ca2+ influx into the postsynaptic neurons.
This postulated increase in Ca2+ influx provides a
reasonable explanation for the facilitation of NMDA
receptor-independent LTP produced by treating slices with bicuculline.
This explanation is consistent with the observations of Tsubokawa and
Ross (1996)
, who found that GABAA receptors limit action
potential propagation and the associated increase in intracellular Ca2+ concentration in dendrites of CA1 pyramidal neurons.
The increased duration of synaptically evoked action potentials during
block of fast, GABAA IPSPs may have additional consequences for VDCC-dependent forms of synaptic plasticity. As noted previously, increasing the duration of action potentials promotes the opening of
high-threshold VDCCs. Because VDCCs differ in activation and inactivation kinetics and voltage dependency (Bertolino and
Llinás 1992; Perez-Reyes and Schneider
1994
; Tsien et al. 1991
), an increase in the
duration of action potentials might result in the opening of additional
types of VDCCs. This could allow other types of VDCCs, in addition to
the dihydropyridine sensitive (L-type) VDCCs previously implicated
(Cavus and Teyler 1996
; Grover and Teyler 1990
), to participate in the induction of NMDA
receptor-independent LTP. In agreement with this suggestion, it was
shown (Ito et al. 1995
) that T- and P-type VDCCs
participate in the induction of LTP during theta burst stimulation, a
type of patterned tetanic stimulation that is particularly effective in
promoting disinhibition (Pacelli et al. 1989
) and that
might therefore increase the duration of postsynaptic action potentials
similarly to pharmacological disinhibition.
The findings reported here may shed some light on the control over
induction of NMDA receptor-independent LTP in situ. The stimulation
protocols used in previous in vitro studies (Cavus and Teyler
1996; Grover 1998
; Grover and Teyler
1990
, 1992
, 1994
, 1995
) to induce NMDA receptor-independent LTP
relied on high-frequency (200 Hz) tetanic stimulation. However, as
shown in this study, reduction of fast, GABAA
receptor-mediated inhibition can allow lower stimulation frequencies to
induce NMDA receptor-independent LTP. The hippocampus is innervated by
noradrenergic, serotonergic, cholinergic, and GABAergic systems
(Amaral and Kurz 1985
; Eckenstein et al.
1988
; Frotscher and Léránth 1985
;
Houser et al. 1985
; Köhler et al.
1984
; Lysakowski et al. 1989
; Molliver
1987
; Oleskevich et al. 1989
; Steinbusch
1981
; Wainer et al. 1985
; Woolf et al. 1984
), and GABAergic interneurons are targeted by at least some of these inputs (Freund and Antal 1988
; Freund et
al. 1990
; Milner and Bacon 1989
). Because evoked
inhibition in the hippocampus is modulated by these transmitters
(Marty and Llano 1995
; Thompson 1994
),
the regulation of hippocampal interneurons by extrinsic neuromodulatory
inputs might serve a gating function, either facilitating or
suppressing the induction of NMDA receptor-independent LTP.
Although the results of this and previous (Grover 1998;
Grover and Teyler 1990
) investigations have highlighted
the role of postsynaptic VDCCs in the induction of NMDA
receptor-independent LTP, at least one additional process is also
required, the activation of metabotropic glutamate receptors (mGluRs)
(Grover 1998
; Little et al. 1995
).
Although the specific role of mGluRs is not yet known, we have
suggested that mGluR activation, and not postsynaptic VDCC activation,
is responsible for the synapse specificity of NMDA receptor-independent
LTP (Grover 1998
; Grover and Teyler 1992
). It is possible that blockade of GABAA
receptors could, under some conditions, enhance mGluR activation and
thereby facilitate NMDA receptor-independent LTP. For example, blocking
GABAA receptors enhances the excitatory interactions
between CA3 pyramidal neurons (Colom and Saggau
1994
; Miles and Wong 1987
); this could lead to
greater recruitment of presynaptic neurons during tetanic stimulation, greater release of glutamate during tetanization, and enhanced activation of mGluRs. Although this scenario is possible, it most likely did not occur during our experiments because we removed the CA3
area from most slices exposed to the GABAA antagonist, thereby preventing this possibility. Furthermore, there were no differences in LTP between slices with and without an intact CA3 region, suggesting that the enhanced activation of mGluRs is not a
major mechanism allowing facilitation of NMDA receptor-independent LTP
by GABAA receptor antagonism.
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ACKNOWLEDGMENTS |
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We thank Drs. Idil Cavus and Pascal DiScenna for comments and suggestions.
This work was supported by National Institute of Neurological Disorders and Stroke Grant NS-34650.
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FOOTNOTES |
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Address for reprint requests: L. M. Grover, Dept. of Physiology, Marshall University School of Medicine, 1542 Spring Valley Dr., Huntington, WV 25755-9340.
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 24 June 1998; accepted in final form 1 March 1999.
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