1Department of Neurophysiology, Institute of Physiology, University Hospital Charité, Humboldt University Berlin, 10117 Berlin, Germany; and 2Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California 94141-0450
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ABSTRACT |
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Behr, Joachim,
Tengis Gloveli,
Dietmar Schmitz, and
Uwe Heinemann.
Dopamine Depresses Excitatory Synaptic Transmission Onto Rat
Subicular Neurons Via Presynaptic D1-Like Dopamine Receptors.
J. Neurophysiol. 84: 112-119, 2000.
Schizophrenia is considered to be associated with an abnormal
functioning of the hippocampal output. The high clinical potency of
antipsychotics that act as antagonists at dopamine (DA) receptors indicate a hyperfunction of the dopaminergic system. The subiculum obtains information from area CA1 and the entorhinal cortex and represents the major output region of the hippocampal complex. To
clarify whether an enhanced dopaminergic activity alters the hippocampal output, the effect of DA on alveus- and perforant path-evoked excitatory postsynaptic currents (EPSCs) in subicular neurons was examined using conventional intracellular and whole cell
voltage-clamp recordings. Dopamine (100 µM) depressed
alveus-elicited (S)--amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor-mediated EPSCs to 56 ± 8% of control while perforant path-evoked EPSCs were attenuated to only 76 ± 7% of control. Dopamine had no effect on the EPSC kinetics. Dopamine reduced the
frequency of spontaneous miniature EPSCs without affecting their
amplitudes. The sensitivity of subicular neurons to the glutamate
receptor agonist (S)-
-amino-3-hydoxy-5-methyl-4-isoxazolepropionic acid was unchanged by DA pretreatment, excluding a postsynaptic mechanism for the observed reduction of excitatory synaptic
transmission. The effect of DA on evoked EPSCs was mimicked by the D1
receptor agonist SFK 38393 and partially antagonized by the D1 receptor antagonist SCH 23390. While the D2 receptor agonist quinelorane failed
to reduce the EPSCs, the D2 receptor antagonist sulpiride did not block
the action of DA. The results indicate that DA strongly depresses the
hippocampal and the entorhinal excitatory input onto subicular neurons
by decreasing the glutamate release following activation of presynaptic
D1-like DA receptors.
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INTRODUCTION |
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Although the hippocampal complex is not classified
as a part of the dopaminergic neuronal system, previous studies have
shown that dopamine (DA) acts as a neurotransmitter in the hippocampus (Hsu 1996; Otmakhova and Lisman 1999
).
This idea was given support by anatomical and biochemical studies
suggesting that the hippocampal area CA1 and the subiculum receive a
dense mesencephalic dopaminergic projection from the ventral tegmental
area (VTA) (Descarries et al. 1987
; Gasbarri et
al. 1994
; Verney et al. 1985
) and express high
levels of D1- and D2-like DA receptors (Bruinink and Bischoff 1993
; Fremeau et al. 1991
; Martres et al.
1985
; Meador-Woodruff et al. 1994
).
The subiculum controls most of the
entorhinal-hippocampal output. It receives strong input from area CA1
(Amaral et al. 1991; Finch and Babb 1981
;
Tamamaki and Nojyo 1990
) and layer III of the entorhinal
cortex (Steward and Scoville 1976
; Witter
1993
) and relays information to other regions of the subicular
complex, to the deep and superficial layers of the entorhinal cortex,
as well as to a variety of distant cortical and subcortical structures (Witter 1993
; Witter and Groenewegen
1990
; Witter et al. 1990
). The subicular
projection to the nucleus accumbens has received increasing attention
as alterations of its activity seem to be involved in schizophrenia
(Gray 1998
; Gray et al. 1995a
;
Greene 1999
; Joyce 1993
; Mogenson
et al. 1993
). The subicular output onto the nucleus accumbens
appears to functionally interact with a substantial dopaminergic
projection from the VTA, thereby balancing the activity of the nucleus
accumbens (Blaha et al. 1997
; Brudzynski and
Gibson 1997
; Harvey and Lacey 1996
;
Nicola et al. 1996
; Totterdell and Smith
1989
; Wu and Brudzynski 1995
). As DA
hyperfunction has been implicated in schizophrenia (Gray et al.
1995b
; Joyce 1993
; Joyce and
Meador-Woodruff 1997
) the present study examines how DA
modulates the synaptic excitability of subicular neurons. Using intracellular- and whole cell voltage-clamp recordings, the results obtained indicate that DA strongly suppresses glutamatergic hippocampal and entorhinal neurotransmission onto subicular neurons by activation of presynaptic D1-like DA receptors.
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METHODS |
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Slice preparation
The experiments were performed on horizontal slices containing the entorhinal cortex, the subiculum, and the hippocampal formation obtained from adult 180- to 230-g female Wistar rats. The rats were decapitated under deep ether anesthesia, the brains were quickly removed, and 400-µm-thick slices were prepared with a Campden vibroslicer (Loughborough, UK). The slices were transferred to an interface recording chamber continuously perfused with an aerated (95% O2-5% CO2), prewarmed (34°C) artificial cerebrospinal fluid (ACSF) containing (in mM) 124 NaCl, 1.25 Na2PO4, 26 NaHCO3, 3 KCl, 1.6 CaCl2, 1.8 MgSO4, and 10 glucose at pH 7.4.
Electrophysiological recordings
Recordings from subicular neurons were made exclusively in the
cell band in extension of the pyramidal cell layer in area CA1 within
close proximity to the perforating fibers of the perforant path.
Discontinuous single-electrode voltage-clamp recordings were performed
with conventional sharp microelectrodes by using a SEVC amplifier
(SEC10L, NPI Electronic, Tamm, Germany). Only cells with resting
potentials more negative than 60 mV and with overshooting action
potentials (prior QX-314 diffusion) were accepted. Electrodes were
pulled from borosilicate glass (1.2 mm OD, resistance 50-80 M
) and
filled with 2.5 M K-acetate. In all experiments we additionally
included 50 mM QX-314 in the recording electrode to minimize effects on
intrinsic K+ currents following application of
DA. After clamping the cell close to the resting membrane potential, we
optimized the gain, capacitance compensation, and switching frequency
(25-35 kHz). Stimulation of afferent fibers in the alveus or the
perforant path was performed with glass-insulated bipolar platinum wire electrodes (tip diameter 50 µm, tip separation 100-200 µm).
Synaptic potentials were evoked following electrical stimulation every 10 s (intensity: 1-5 V; duration: 0.05 ms). Local application of
glutamate agonist onto subicular dendrites was conducted with electrodes pulled from borosilicate glass (2.0 mm OD, 1.0 mm ID, 3 µm
tip diam) using high pressure (1.5 bar) application of 20 ms duration.
Miniature excitatory postsynaptic currents (mEPSCs) were recorded in
whole cell voltage-clamp mode using a continuous feedback patch-clamp
amplifier (EPC-7, HEKA, Lambrecht, Germany). Electrodes were pulled
from borosilicate glass (2.0 mm OD, 1.0 mm ID, 3-8 M resistance).
Intrapipette solutions contained (in mM) 135 Cs-gluconate, 6 CsCl2, 2 MgCl2, 10 HEPES,
and 1 QX-314; CsOH was used to adjust pH to 7.2. Whole cell recordings
were obtained by lowering patch electrodes into the subicular cell
layer while monitoring current responses to 10-mV voltage pulses and
applying suction to form >G
seals. Access resistance was monitored
throughout each experiment, and only recordings with access resistance
of <15 M
were considered acceptable for analysis. Access resistance
(not compensated) was repeatedly checked during the experiments, and
recordings showing an increase of >20% were rejected. Neurons were
voltage-clamped at
70 mV for recordings of mEPSCs.
Signals were filtered at 3 kHz, digitized at 9-12 kHz by a TIDA interface card (Batelle, Frankfurt, Germany) or an ITC-16 interface (Instrutech, Great Neck, NY), and subsequently stored on an IBM-compatible computer. Peak amplitudes of evoked EPSCs were measured from an average of 8-10 sweeps. All data were analyzed off-line using TIDA software (HEKA, Lambrecht/Pfalz, Germany). Detection of individual mEPSCs was done off-line using ISO2/ANA3 (MFK, Niedernhausen, Germany). Statistical evaluation was performed on the means ± SE by applying a Student's t-test (Origin 4.1, Microcal). Significance level was set to P < 0.05.
Compounds
The following drugs were bath-applied: bicuculline methiodide
(BCM) (SIGMA, Deisenhofen, Germany), 5 µM; 2-amino-5-phosphonovaleric acid (APV) (Research Biochemicals, Natick, MA), 60 µM;
6-nitro-7-sulfamoylbenzo(f)quinoxaline-2,3-dione (NBQX; a gift from
NOVO Nordisk), 10 µM;
3-N-[1-(s)-(3,4-dichlorophenyl)ethyl]amino-2-(s)-hydroxypropyl-P-benzyl-phosphinic acid (CGP55845A; a gift from CIBA-GEIGY, Basel), 2 µM; dopamine-HCl, 100 µM; (±)-sulpiride, 20 µM;
R(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1 h-3-benzazepine hydrochloride [(+)-SCH-23390], 20 µM (all from Research Biochemicals, Natick, MA);
(±)-1-phenyl-2,3,4,5-tetrahydro-(1H)-3-benzazepine-7,8-diol (SFK 38393 hydrobromide), 20 µM;
(4aR-trans)-4,4a,5,6,7,8,8a,9-octahydro-5-propyl-1H-pyrazolo[3,4-g]quinoline [()-quinpirole hydrochloride], 20-400 µM (both from Tocris,
Bristol, UK). As DA oxidates rapidly in warm ACSF, we generally applied 20 µM
Na2S2O5
(Merck, Darmstadt, Germany) to prevent oxidation (Sutor and ten
Bruggencate 1990
). For mEPSCs recordings, we applied 1 µM TTX
to avoid activation of Na+ currents. High
pressure application was conducted with the glutamate receptor agonist
(S)-
-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA),
100-1,000 µM (Tocris, Bristol, UK).
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RESULTS |
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Voltage-clamp recordings were used to examine the effect of DA (100 µM) on stimulus-evoked glutamate receptor-mediated EPSCs in subicular neurons. GABAA receptor-mediated inhibitory postsynaptic currents (IPSCs) were blocked by bicuculline (5 µM), and GABAB receptor-mediated IPSCs were eliminated by CGP 55845A (2 µM) and by QX-314-containing intracellular solutions. AMPA/kainate receptor (in the following indicated as AMPAR)-mediated responses were isolated by application of APV (60 µM) while N-methyl-D-aspartate (NMDA) receptor (NMDAR)-mediated EPSCs were recorded in the presence of NBQX (10 µM).
Dopamine reduces alveus and perforant path-evoked EPSCs
Stimulation of alvear fibers evokes a glutamatergic postsynaptic
response in subicular neurons consisting of a NBQX-sensitive AMPAR-mediated response and a strong NMDAR-mediated component that is
predominantly APV sensitive (Behr et al. 1998). In
addition, subicular cells express a small APV-insensitive
NMDAR-mediated component (Hetka et al. 1998
) that shows
an attenuated Mg2+ blockade reminiscent to
recently described neocortical NMDAR-mediated EPSCs (Fleidervish
et al. 1998
). Bath application of DA for 10 min reversibly
depressed AMPAR- and NMDAR-mediated EPSCs in subicular neurons evoked
by stimulation of alvear fibers, which represent the dominant
hippocampal input from area CA1. Both glutamatergic components were
decreased by the same extent: While the AMPAR-mediated EPSCs declined
to 56 ± 8% (mean ± SE, n = 9) of
control, the NMDAR-mediated response attenuated to 54 ± 11%
(n = 4) of control (Fig.
1, A and B). The
effect of DA could not be prevented by intracellular perfusion of the
cell in patch-clamp recording technique (n = 4).
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In addition to the alvear input from area CA1, the subiculum receives
an excitatory monosynaptic input from the entorhinal cortex via the
perforant path projection (Behr et al. 1998;
Steward and Scoville 1976
; Witter 1993
)
(Fig. 2A). As both pathways
excite subicular neurons and determine the activity of the hippocampal output, we extended our investigation examining the effect of DA on the
excitatory entorhinal input. As previously described, stimulation of
perforant path fibers originating in layer III of the entorhinal cortex
evokes a glutamatergic postsynaptic response in subicular neurons
consisting of an AMPAR- and NMDAR-mediated component (Behr et
al. 1998
). Using drop application of glutamate onto layer III
of the entorhinal cortex to elicit orthodromic activation of subicular
cells, we had ascertained that perforant path-evoked responses in
subicular cells are not due to the antidromic activation of
subiculo-entorhinal axons. Comparing the kinetics of perforant
path-evoked AMPAR- and NMDAR-mediated components with those elicited
by alveus stimulation recorded in the same cell revealed no significant
difference (Fig. 2B). The decay time constant of
AMPAR-mediated currents was 8.6 ± 0.74 ms following perforant
path stimulation and 7.7 ± 0.85 ms following alveus stimulation
(n = 7) when monoexponentially fitted. The
corresponding values for the NMDAR-mediated currents were 41.9 ± 4.3 ms and 43.2 ± 6.4 ms, respectively (n = 8).
Interestingly, the hippocampal and entorhinal inputs onto the subiculum
seem to be differentially affected by DA (100 µM). While
alveus-evoked AMPAR-mediated EPSCs were depressed to 56 ± 8%
(n = 9) of control, the perforant path-evoked EPSCs
recorded in the same cell were decreased to only 76 ± 7% (n = 9, P = 0.013; Fig. 2, C
and D).
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Depression of EPSCs is mediated by a D1-like DA receptor
Dopamine receptors can be subdivided into two pharmacologically
and biochemically distinct classes (Civelli et al. 1993;
Sibley 1995
), the D1- and the D2-like DA receptors. To
elucidate the DA receptor subtype responsible for the depression of
excitation, first we examined the effects of D1 and D2 receptor
agonists and antagonists on alveus-evoked AMPAR-mediated EPSCs (Fig.
3). Like DA, SFK 38393 (20 µM), an
agonist of D1-like DA receptors, decreased the EPSC peak amplitude to
73 ± 8% of control (n = 6). Although DA
attenuated EPSCs to 56 ± 8% of control (n = 9),
the effects were not significantly different (P = 0.16). In contrast, the D2 receptor agonist quinelorane failed to
suppress EPSCs even when applied at higher concentrations (up to 400 µM; n = 6). Consistent with this result, the D2
receptor antagonist sulpiride (20 µM) did not prevent the DA-induced
depression to 57 ± 8% of control (n = 9), which
is not different from the observed decline in the absence of any
antagonist (56 ± 8%, n = 9). However, the D1
receptor antagonist SCH 23390 (20 µM) did not completely block the
depressant action of DA (82 ± 7% of control, n = 8), despite the fact that this antagonist was used at concentrations
exceeding the ones applied by other groups (Harvey and Lacey
1996
; Nicola et al. 1996
). Finally, we examined
the DA receptor involved in the attenuation of perforant path-evoked
EPSCs. As shown for alveus-evoked EPSCs, application of the D1
agonist SFK 38393 (20 µM) mimicked the effect of DA and attenuated
AMPAR-mediated EPSCs to 80 ± 8% of control (n = 5, data not shown).
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Depression of EPSCs is mediated by a presynaptic mechanism
The mechanism by which DA exerts its effect on excitatory synaptic transmission may involve a decrease in presynaptic glutamate release and/or a decreased postsynaptic sensitivity to glutamate. To clarify the site of action, we first examined the kinetics of the isolated alveus-evoked AMPAR- and NMDAR-mediated EPSCs before and after application of DA. There was no DA-induced difference in the decay time, making a modulation of glutamate receptors by competitive interaction with binding sites or a shunting of the EPSCs in the dendrites unlikely. The decay time constants of AMPAR- and NMDAR-mediated EPSCs before and after application of DA were 7.8 ± 0.63 ms versus 8.2 ± 0.65 ms (n = 5) and 43.5 ± 10.7 ms versus 39.8 ± 7.0 ms (n = 4), respectively, when monoexponentially fitted (Fig. 1, C and D). In addition, the sensitivity of subicular neurons to pressure application of AMPA (100-1,000 µM) was unchanged by DA (100 µM) pretreatment (151 ± 24 pA vs. 137 ± 9 pA, n = 7, paired t-test analysis, P = 0.58), suggesting a presynaptic mechanism for the observed effect (Fig. 1E).
To further determine whether a pre- or postsynaptic mechanism is involved, we recorded miniature EPSCs (mEPSCs) in the presence of the Na+-channel blocker tetrodotoxin (TTX; 1 µM) in whole cell voltage-clamp recordings to study the effect of DA receptor activation on spontaneous glutamate release (Fig. 4). Miniature EPSCs are believed to represent the random release of single neurotransmitter packets. Analysis of the mEPSC frequency before and after application of DA provides information about possible changes in the presynaptic release process, while changes in the amplitudes of the miniature currents reflect postsynaptic alterations in receptor properties including their number at the synapse. DA (100 µM) prolonged the mean inter-event interval from 198 ± 57 ms to 1.149 ± 0.249 s (n = 4, paired t-test analysis, P < 0.05). The amplitude distribution of mEPSCs remained unaltered by DA (9.61 ± 0.64 pA vs. 9.12 ± 0.86 pA), suggesting that DA reduces glutamate release onto subicular neurons. A purely postsynaptic origin for the more than 80% decrease of mEPSC frequency in the presence of DA can be excluded because, as calculated for the cell shown in Fig. 2, a 56% decrease of mEPSC amplitude (percentage reduction of evoked AMPAR-mediated EPSCs) would increase the mean inter-event interval by only 27%, from 409 ± 18 ms to 562 ± 36 ms.
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Although the reduction in mEPSC frequency strongly suggests a
presynaptic action of DA, the excitatory synapses that contribute to
the mEPSC distribution remain unknown. To exclude distinct pathway-dependent mechanisms for the DA-induced depression, we tested
the effect of DA (100 µM) on alveus and perforant path-evoked paired-pulse facilitation (PPF), which is considered to depend on a
presynaptic mechanism (Zucker 1989) (Fig.
5). Surprisingly, DA inconsistently
altered the paired-pulse ratio (PPR; 50-ms interevent interval, 2nd
response relative to the 1st one) of alveus and perforant path-evoked
AMPAR-mediated EPSCs. In the majority of cells (12 of 16), the PPR
declined significantly during application of DA. The PPR of
alveus-evoked responses decreased from 1.54 ± 0.07 to 1.30 ± 0.07 (n = 12); in five of these cells the PPR following perforant path stimulation declined from 1.53 ± 0.11 to
1.27 ± 0.06. In these five cells there was no pathway-dependent difference in the change of the PPRs. In four cells, however, we
observed either an increase of both alveus- and perforant path-evoked PPF (n = 1) or coinciding increases and decreases of
the PPR (n = 3).
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DISCUSSION |
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An increasing number of studies suggest that the subiculum, the
major output region of the hippocampal formation, seems to be relevant
to schizophrenia (Gray 1998; Gray et al.
1995a
; Greene 1999
; Joyce 1993
;
Mogenson et al. 1993
). Morphological studies showed
neuropathological alterations of the subiculum in schizophrenic patients (Arnold et al. 1991
, 1995
;
Eastwood et al. 1995
; Gray et al. 1991
).
The subicular involvement may lie in the presumed functional alteration
of hippocampal output and in the convergence of subicular and
dopaminergic inputs onto the nucleus accumbens (Totterdell and
Smith 1989
), a structure that is considered a principal
component of the mesolimbic DA system (Paxinos and Watson 1986
; Pennartz et al. 1994
) and a site of action
of psychostimulants as well as antipsychotic drugs (Goldstein
and Deutch 1992
; Gray et al. 1991
; Nicola
et al. 1996
). In an effort to clarify how the subiculum might
be affected by dopaminergic hyperfunction, the present study
investigated the effect of DA on excitatory synaptic transmission in
this region. Our study demonstrates that DA strongly depresses the
hippocampal and the entorhinal excitatory input onto subicular cells by
decreasing the glutamate release following activation of presynaptic
D1-like DA receptors. While DA depressed alveus-elicited AMPA
receptor-mediated EPSCs in subicular neurons to 56 ± 8% of
control, perforant path-evoked EPSCs, however, were attenuated to only
76 ± 7% of control. The presynaptic mechanism for the observed
reduction of excitatory synaptic transmission was confirmed by
employing paired-pulse protocols, recordings of spontaneous miniature
EPSCs, wash out of postsynaptic G-proteins, the use of AMPA-pressure
application, and the analysis of EPSC kinetics before and after
application of DA.
The subiculum receives strong glutamatergic input from area CA1
(Behr et al. 1998; Taube 1993
) and is
thus part of the polysynaptic loop from the entorhinal cortex layer II
through the dentate gyrus, area CA3, area CA1 and the subiculum back to
the entorhinal cortex. In addition, the subiculum receives a
monosynaptic excitatory perforant path projection from layer III of the
entorhinal cortex, which bypasses the classic trisynaptic hippocampal
loop and terminates exclusively in area CA1 and the subiculum
(Steward and Scoville 1976
; Witter 1993
).
The functional interaction of both inputs balances the activity of the
subiculum and hence controls the hippocampal output. Considering the
recently described pronounced and selective DA-induced inhibition of
the excitatory perforant path input onto CA1 pyramidal cells
(Otmakhova and Lisman 1999
), we examined the effect of
DA on the entorhinal input and compared it with that originating in
area CA1. As previously described, both synaptic inputs consist of
AMPAR- and NMDAR-mediated components (Behr et al. 1998
).
Neither the AMPAR- nor the NMDAR-mediated components of each input
differed in their kinetics, suggesting similar pathway-independent
postsynaptic receptor sites. Interestingly, application of DA affected
both pathways selectively: while the CA1 input was attenuated to 56%
of control, the entorhinal input was diminished to only 76% of control.
Our data indicate that the D1-like DA receptor most likely mediates the
decrease in transmitter release onto subicular cells. The D1 receptor
agonist SFK 38393, but not the D2 receptor agonist quinelorane,
mimicked the action of DA. Consistent with these results, the D2
antagonist sulpiride failed to inhibit the depressive effect of DA.
However, preapplication of the D1 receptor antagonist SCH 23390 could
not completely block the DA effect. Considering previous studies that
had also difficulties to mimic or antagonize DA effects (Nicola
et al. 1996; O'Donnell and Grace 1994
;
Otmakhova and Lisman 1999
; Pralong and Jones
1993
), the present result appears likely to result from either
a decreased sensitivity of subicular D1-like DA receptors to SCH 23390 or a subset of atypical DA receptors that are not blocked by this
antagonist. As the D1 receptor agonist SFK 38393 was less effective
than DA in depressing evoked EPSCs, we favor the latter explanation.
Alternatively, DA exerts its action by activating other monoamine
receptors (Aguayo and Grossie 1994
; Malenka and
Nicoll 1986
); however, the concentrations to be needed were
reported to be at least 10-fold higher than those we employed
(Haas and Konnerth 1983
). Several studies have
demonstrated that D1-like DA receptors control glutamate release in
different regions of the brain including the nucleus accumbens
(Nicola et al. 1996
; Pennartz et al.
1992
), the entorhinal cortex (Pralong and Jones
1993
), the prefrontal cortex (Law-Tho et al.
1994
; Williams and Goldman-Rakic 1999
), and the
basal forebrain (Momiyama et al. 1996
). As each of these
structures receive dopaminergic projections from the VTA, one may
conclude that the dopaminergic system controls excitatory
neurotransmission by D1-like DA receptors. These receptors regulate the
activity of cyclic AMP (cAMP)-dependent protein kinase (PKA)
(Kebabian and Calne 1979
) and were shown to reduce
N-type Ca2+ currents (Surmeier et al.
1995
), an effect that might account for the reduction of evoked
EPSCs. However, it is noteworthy that recent studies in the hippocampal
area CA1 (Hsu 1996
) as well as in layer III of the
entorhinal cortex (Stenkamp et al. 1998
) indicated a
D2-like DA receptor-mediated presynaptic inhibition of glutamate release.
As the decreases in the AMPAR- and NMDAR-mediated alveus-evoked EPSC components were similar and DA had no effect on the kinetics of AMPAR- and NMDAR-mediated EPSCs, we assumed a presynaptic mechanism for the DA-induced depression. It is conceivable that postsynaptic ion channels are modulated indirectly via the activation of G-proteins and second-messenger pathways. However, altering the cellular metabolism by employing intracellular perfusions with a patch pipette could not prevent the DA-induced depression of EPSCs. Therefore a postsynaptic G-protein-mediated cellular pathway appears unlikely to underlie the observed depression. In contrast, DA had no effect on the membrane inward currents induced by pressure application of AMPA indicating that the depression of EPSC by DA is not mediated by a postsynaptic action on glutamate receptors. To support the hypothesis of a presynaptic mechanism, we examined the effect of DA on mEPSCs. Dopamine significantly reduced the frequency of mEPSCs without affecting their amplitude, thereby excluding an interaction of DA with postsynaptic glutamate receptors and giving strong evidence for a suppression of glutamate release from synaptic terminals. As the DA-induced suppression was recorded in the presence of the GABAB receptor antagonist CGP 55845A, we can exclude an interaction of DA with presynaptic GABAB receptors.
To elucidate pathway-specific mechanisms for the depression of EPSCs,
we finally investigated the effect of DA on alveus and perforant
path-evoked PPF, a phenomenon sensitive to changes in release
probability (Zucker 1989). While PPF usually increases under conditions where transmitter release is decreased in our study,
the majority of cells showed a decreased PPF in the presence of DA
independent of the input stimulated. Presently, we do not have a
conclusive explanation for the discrepancy. Even more intriguing was
the observation that in some cells increases and decreases of PPF
coincided within one cell, suggesting that DA does not alter the
release machinery only by reducing Ca2+ influx.
The moderate pathway-dependent difference in DA-induced EPSC depression
stands in sharp contrast to the findings in area CA1 where perforant
path-elicited field EPSPs were strongly depressed by DA while the
Schaffer collateral-mediated input was rather unaffected
(Otmakhova and Lisman 1999). Interestingly, the
DA-induced depression of the excitatory perforant path input in area
CA1 seems to be mediated by a presynaptic mechanism as well and shows a
similar dose effect like in the subiculum. Hence, it appears conceivable that the perforant path projection from layer III of the
entorhinal cortex to area CA1 and the subiculum is similarly modulated
by DA receptors located on perforant path terminals. While in the
present study the hippocampal input to the subiculum was strongly
attenuated by DA, the Schaffer collateral mediated synaptic
transmission to area CA1 was reported to be unaffected by DA
(Marciani et al. 1984
; Otmakhova and Lisman
1999
; but see Hsu 1996
). This different
pathway-dependent depression of excitation is consistent with the
distinct distribution of DA receptors in area CA1 and the subiculum:
while area CA1 contains a pathway-dependent laminar distribution of DA
receptors (Goldsmith and Joyce 1994
; Swanson et
al. 1987
), in the subiculum D1-like DA receptors are rather
homogeneously (Köhler et al. 1991
) distributed
throughout its molecular layer, the target site for both, the perforant
path and the alvear projection, respectively. Nonetheless, with respect to the stronger DA effect on alveus-evoked responses, alvear terminals seem to have a slightly higher density of D1-like DA receptors or a
distinct DA receptor subunit expression.
The concentration of DA used in this study must be related to the in
vitro conditions of the experiment. Full equilibrium of applied drugs
within slices takes at least 1 h within the interface type of
recording chamber (Müller et al. 1988). The
demonstrated effects, however, were recorded after 10 min of DA
perfusion. In addition, rapid oxidation (Sutor and ten
Bruggencate 1990
) as well as uptake of DA will reduce the final
concentration of DA. Hence, the nominal applied concentrations of DA
were considerably higher than those that actually induced the observed
effects. This is in agreement with studies in the hippocampus
(Gribkoff and Ashe 1984
), the entorhinal cortex
(Pralong and Jones 1993
; Stenkamp et al.
1998
), the nucleus accumbens (Nicola et al.
1996
), and the basal forebrain (Momiyama et al.
1996
), which reported that similar concentrations were
necessary to show effects of DA on synaptic responses under in vitro conditions.
The mesolimbic dopaminergic system is known to be implicated in
impairments of memory and cognitive functions that are closely connected with the hippocampus. Alterations of the dopaminergic innervation as well as of the density of DA-sensitive receptors within
the hippocampus may influence the hippocampal output and contribute to
dementing and neuropsychiatric diseases. Schizophrenia is considered to
be associated with an attenuated flow of hippocampal output
(Gray 1998; Gray et al. 1991
;
Joyce 1993
; Mogenson et al. 1993
).
Functional neuroimaging studies of schizophrenics suggest an abnormal
information flow between the hippocampal formation and the prefrontal
cortex (Weinberger et al. 1992
). As schizophrenia is
associated with a hyperfunction of the dopaminergic system (Gray
et al. 1995b
; Joyce 1993
; Joyce and
Meador-Woodruff 1997
), an enhanced DA-induced depression of the
excitatory drive onto subicular cells may cause abnormalities in the
hippocampal output. Considering that antagonists at DA receptors have a
high antipsychotic potency, the subiculum may represent a potential
site of action. A reduction of the DA-induced inhibition of glutamate
release by D1 antagonists may partially restore the diminished
hippocampal information transfer.
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ACKNOWLEDGMENTS |
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We are indebted to A. Piechotta and A. Duerkop for excellent technical assistance and editorial help.
This work was supported by a grant from the Bundesministerium für Bildung und Forschung and Deutsche Forschungsgemeinschaft Grant 2022/2-1, 2-2 to J. Behr.
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FOOTNOTES |
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Address for reprint requests: J. Behr, Dept. of Neurophysiology, Institute of Physiology, University Hospital Charité, Humboldt University Berlin, Tucholskystr. 2, 10117 Berlin, Germany (E-mail: joachim.behr{at}charite.de).
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 10 November 1999; accepted in final form 22 March 2000.
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REFERENCES |
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