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Increased Hippocampal Excitability in the 3xTgAD Mouse Model for Alzheimer's Disease In Vivo


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Mouse Alzheimer's disease (AD) models develop age- and region-specific pathology throughout the hippocampal formation. One recently established pathological correlate is an increase in hippocampal excitability in vivo. Hippocampal pathology also produces episodic memory decline in human AD and we have shown a similar episodic deficit in 3xTg AD model mice aged 3-6 months. Here, we tested whether hippocampal synaptic dysfunction accompanies this cognitive deficit by probing dorsal CA1 and DG synaptic responses in anaesthetized, 4-6 month-old 3xTgAD mice. As our previous reports highlighted a decline in episodic performance in aged control mice, we included aged cohorts for comparison. CA1 and DG responses to low-frequency perforant path stimulation were comparable between 3xTgAD and controls at both age ranges. As expected, DG recordings in controls showed paired-pulse depression; however, paired-pulse facilitation was observed in DG and CA1 of young and old 3xTgAD mice. During stimulus trains both short-latency (presumably monosynaptic: 'direct') and long-latency (presumably polysynaptic: 're-entrant') responses were observed. Facilitation of direct responses was modest in 3xTgAD animals. However, re-entrant responses in DG and CA1 of young 3xTgAD mice developed earlier in the stimulus train and with larger amplitude when compared to controls. Old mice showed less DG paired-pulse depression and no evidence for re-entrance. In summary, DG and CA1 responses to low-frequency stimulation in all groups were comparable, suggesting no loss of synaptic connectivity in 3xTgAD mice. However, higher-frequency activation revealed complex change in synaptic excitability in DG and CA1 of 3xTgAD mice. In particular, short-term plasticity in DG and CA1 was facilitated in 3xTgAD mice, most evidently in younger animals. In addition, re-entrance was facilitated in young 3xTgAD mice. Overall, these data suggest that the episodic-like memory deficit in 3xTgAD mice could be due to the development of an abnormal hyper-excitable state in the hippocampal formation.
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Increased Hippocampal Excitability in the 3xTgAD Mouse
Model for Alzheimer’s Disease
In Vivo
Katherine E. Davis, Sarah Fox, John Gigg*
Faculty of Life Sciences, University of Manchester, Manchester, United Kingdom
Mouse Alzheimer’s disease (AD) models develop age- and region-specific pathology throughout the hippocampal
formation. One recently established pathological correlate is an increase in hippocampal excitability in vivo. Hippocampal
pathology also produces episodic memory decline in human AD and we have shown a similar episodic deficit in 3xTg AD
model mice aged 3–6 months. Here, we tested whether hippocampal synaptic dysfunction accompanies this cognitive
deficit by probing dorsal CA1 and DG synaptic responses in anaesthetized, 4–6 month-old 3xTgAD mice. As our previous
reports highlighted a decline in episodic performance in aged control mice, we included aged cohorts for comparison. CA1
and DG responses to low-frequency perforant path stimulation were comparable between 3xTgAD and controls at both age
ranges. As expected, DG recordings in controls showed paired-pulse depression; however, paired-pulse facilitation was
observed in DG and CA1 of young and old 3xTgAD mice. During stimulus trains both short-latency (presumably
monosynaptic: ‘direct’) and long-latency (presumably polysynaptic: ‘re-entrant’) responses were observed. Facilitation of
direct responses was modest in 3xTgAD animals. However, re-entrant responses in DG and CA1 of young 3xTgAD mice
developed earlier in the stimulus train and with larger amplitude when compared to controls. Old mice showed less DG
paired-pulse depression and no evidence for re-entrance. In summary, DG and CA1 responses to low-frequency stimulation
in all groups were comparable, suggesting no loss of synaptic connectivity in 3xTgAD mice. However, higher-frequency
activation revealed complex change in synaptic excitability in DG and CA1 of 3xTgAD mice. In particular, short-term
plasticity in DG and CA1 was facilitated in 3xTgAD mice, most evidently in younger animals. In addition, re-entrance was
facilitated in young 3xTgAD mice. Overall, these data suggest that the episodic-like memory deficit in 3xTgAD mice could be
due to the development of an abnormal hyper-excitable state in the hippocampal formation.
Citation: Davis KE, Fox S, Gigg J (2014) Increased Hippocampal Excitability in the 3xTgAD Mouse Model for Alzheimer’s Disease In Vivo. PLoS ONE 9(3): e91203.
Editor: Christian Holscher, University of Lancaster, United Kingdom
Received July 5, 2013; Accepted February 11, 2014; Published March 12, 2014
Copyright: ß2014 Davis et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: The authors are grateful for RCUK funding to KED (BBSRC PhD studentship), SF (BBSRC CASE PhD studentship) and JG (BBSRC grant BB/D011159/1;
Royal Society grant RSRG 24519). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have the following interests. JG is an in vivo electrophysiology consultant for GlaxoSmithKline Neurodegeneration, China.
This does not alter the authors’ adherence to PLOS ONE policies on sharing data and materials.
* E-mail:
Alzheimer’s disease (AD) is characterised phenotypically by
profound declarative memory deficits. The hippocampus, vital for
both the formation and retrieval of declarative memory, is one of
the first areas affected by AD pathological hallmarks of amyloid-
beta (Ab), extracellular plaques and tau neurofibrillary tangles
[1,2]. Interestingly, however, overt plaque burden correlates
poorly with cognitive decline in AD patients [3]. A more recent
hypothesis is that intracellular oligomeric amyloid species, present
before the accumulation of extracellular plaque pathology, may
instead play a pivotal role in disease progression [4,5]. However,
despite extensive research examining the causes of AD pathology
and characterising behavioural phenotypes, there is still little
knowledge about the physiological basis for memory loss in
pathological states such as AD [6].
The hippocampal formation is comprised of the dentate gyrus
(DG), hippocampus proper (CA fields), subiculum (SUB), para-
subiculum, presubiculum and entorhinal cortex (EC; medial and
lateral divisions; MEC and LEC). CA1 and subiculum are the
principal output structures of the hippocampus [7] and EC
provides the interface between incoming and outgoing information
from surrounding cortex [8]. Hippocampal output can return to
surrounding neocortex via deep layers of EC and/or re-enter the
hippocampal formation via projections from deep to superficial
EC [9,10]. The latter reverberation (or ‘re-entrance’) is considered
to be pivotal in memory formation, possibly acting as a
comparator mechanism to allow processed input to be evaluated
alongside new information and/or as a memory consolidation
mechanism during sleep [7,11–13]. Selective lesions to all
hippocampal formation structures have profound effects on
memory performance [14–18]. Functional abnormalities have
also been detected in the hippocampus during memory encoding
in human AD patients [19]. It is of major interest, therefore, to
determine the pathophysiological profile of the hippocampal
formation in AD models and to elucidate, in particular, whether
reverberation (re-entrance) occurs in control and AD model mice.
The triple transgenic mouse model (3xTgAD) carries familial
AD transgenes for Amyloid Precursor Protein (APP
), Pre-
senilin-1(M146V) and an additional tauopathy mutation
). This model develops Aband tau pathology targeted
to the hippocampus and other medial temporal lobe structures in a
manner temporally and spatially similar to human AD [20,21].
The model develops cognitive deficits at a young age [21,22] and,
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as we have demonstrated recently, a specific decline in episodic-
like memory from 3 months that becomes a complete deficit at 6
months of age [23,24]. 3xTgAD mice also show abnormalities in
basal synaptic transmission and deficits in LTP in CA1 hippo-
campus in vitro [21]. However, to date, there have been no
electrophysiological recordings in vivo in the 3xTgAD mouse and,
to our knowledge, there is only one in vivo account in another AD
model examining evoked responses [25]. Thus, the hippocampal
system in AD mouse models has largely been examined only in the
reduced brain slice preparation [21,26], that is, without the
substantial bi-directional cortical connectivity that also contributes
to memory processes [27]. In the 3xTgAD mouse, only CA3-CA1
connectivity has been examined directly in vitro [21], therefore,
there is a need to examine basic synaptic transmission more widely
in hippocampal formation and also hippocampal network function
through examining response reverberation.
One question that arises is the extent to which cognitive deficits
seen in early AD stem from abnormalities in the fibre pathways
projecting to the hippocampus and/or within hippocampal
subfields. Recent research has indicated the presence of early
myelination abnormalities in the 3xTgAD mouse Schaffer
collateral pathway, prior to the occurrence of Aband tau
pathology [28]. In human AD, tau pathology is restricted to the
stellate cells in EC layer II, and leads to degenerative changes in
the perforant path [29]. In addition, afferents from CA1 and SUB
to EC are major sites for Abpathology in human AD [16]; thus,
there is a strong possibility that information flow through the
hippocampal formation could become impaired in AD. However,
synaptic deficits first arise in AD models in vitro at an age prior to
overt extracellular plaque and tangle pathology, suggesting that
mechanisms such as intracellular Abaccumulation or abnormal-
ities in calcium homeostasis are major contributing factors to early
cognitive symptoms [21,26].
Here, we examined the functional state of the hippocampal
formation in the 3xTgAD mouse through the recording of
extracellular field potentials in vivo. Whereas most similar studies
focus on one region of hippocampus in isolation (e.g., CA1 or DG),
in this study we measured synaptic integrity and short-term
plasticity at three sites simultaneously, specifically, the granule cell
layer of DG, CA1 stratum radiatum (CA1sr) and CA1 stratum
lacunosum-moleculare (CA1slm), using a multi-site electrode
recording approach [9]. These sites were chosen as regions that
receive monosynaptic input from the perforant path and, for
CA1sr, polysynaptic feed forward input from DG via CA3. We
measured extracellular field excitatory post-synaptic potentials
(fEPSPs) in these target regions and performed current-source-
density (CSD) analyses on CA1-DG axis responses. Through this
we aimed to map synaptic current flow, indicative of synaptic-
activity throughout the laminar structure, to test the integrity of
synaptic connections within the EC-hippocampal tri-synaptic
Electrophysiological recordings conducted in AD mouse lines in
vitro support a long-term decrease in synaptic function [21,30–32].
However, in vivo recordings support the presence of hyper-
excitability and epileptiform activity, at least in early pathological
stages, which are thought to contribute to cognitive decline
[33,34]. In addition, recent findings indicate the presence of
spontaneous seizure activity in the 3xTgAD model [35]. Thus, in
the current study, we hoped to further examine these contradic-
tory reports of decreased/increased synaptic function and test
whether our observed decline in episodic-like memory in the
3xTgAD model [23,24] correlates with excitability changes within
hippocampal circuits in vivo.
In contrast to in vitro reports, we found little evidence for
decreased functional connectivity in either young or old 3xTgAD
mice. Indeed, in measures of short-term plasticity we saw evidence
for increased DG and CA1sr excitability in 3xTgAD mice
compared to controls, particularly in young mice. This was
coupled, however, with relatively less facilitated fEPSP amplitudes
to a stimulus train alongside an enhanced re-entrant response into
the hippocampal circuit in 3xTgAD mice. Thus, our data support
increased hippocampal excitability at two loci in the 3xTgAD
mouse: firstly, EC layer II input to DG and CA1sr (the latter
presumably via propagation of DG output via CA3) in young and
aged mice; and secondly, facilitation of hippocampal re-entrance
in young 3xTgAD animals, presumably through potentiated deep-
to-superficial EC connectivity.
Materials and Methods
Triple-transgenic mice (3xTgAD) carrying APP
, PS1
and Tau
transgenes and matched controls were bred at the
University of Manchester from a colony donated by the La Ferla
group [21]. Mice were housed in same-sex and genotype groups of
5–6 individuals on a 12:12 light/dark cycle with access to food and
water ad libitum. The 3xTgAD colony was maintained through the
pairing of homozygous individuals and the presence of transgenes
was confirmed by genotyping a subset of 3xTgAD mice in each
cage. All procedures conformed to UK Home Office licensing
(project license PPL 40/3231) and were approved by the
University of Manchester Ethical Review Panel. Electrophysio-
logical data were collected from a ‘young’ sample of mice aged 4–6
months comprising 12 control (6 males: 6 female) and 11 3xTgAD
(5 males: 6 female) mice. The age of the young group was selected
to coincide with the early presence of hippocampal intracellular
Abpathology (plaques and tangles accumulate from approximate-
ly 9 months of age in 3xTgAD mice) and hippocampal-dependent
memory deficits [21–24]. A second ‘old’ sample of mice at 17–18
months of age comprised 5 control and 4 3xTgAD mice (all
female); at this age there is extracellular plaque and tau pathology
within hippocampus and subiculum. Thus, these groups allowed
us to explore the additive effect of both pathology and general
ageing on DG and CA1 responsiveness [20–22]. All female mice
had previously experienced a battery of behavioural testing
(spontaneous recognition tasks) which ceased at least a month
before electrophysiological recordings were conducted. Male mice
used for electrophysiology were experimentally naive. In the
present experiments data from young male and female mice were
combined as no sex differences were seen in either their basal
input/output response, or in their CSD profiles to a 5 Hz train
(data not shown).
Surgery and equipment
Mice were anaesthetised (urethane 30% w/v in dH
O, 1.6 g/
kg, i.p) and monitored until areflexia was achieved. Where reflexes
remained, a small additional dose of urethane (10% w/v in dH
20–30 ml, i.p.) was given. The mouse was held in a stereotaxic
frame (Kopf, 1430, USA) with lamda and bregma in the same
horizontal plane to match the mouse atlas of Paxinos and Franklin
[36]. A homoeothermic heating blanket (Harvard Apparatus, UK)
and rectal probe maintained core body temperature at 37uC. A
midline scalp incision was made and the skull exposed. Craniot-
omies were drilled above CA1 (B–2 mm, ML 1.5 mm) for the
recording electrode and dorsal subiculum (B-3.85 mm, ML
1.5 mm) for the stimulating electrode. Recordings were made
using a linear multi-electrode recording array containing either 16
Hippocampal Hyperexcitability in 3xTgAD Mouse
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contacts spaced100 mm, or 32 contacts spaced 50 mm apart (both
413 mm
contact area; NeuroNexus Tech, USA; there were no
clear differences in response profiles between the different contact
arrangements so data from different arrays were pooled.). The
array was lowered vertically into the brain until the tip was
approximately 2.5 mm below brain surface; at this depth
recording contacts spanned the CA1-DG axis fully ([9]; see
Figure 1A). A bipolar stimulating electrode (twisted 125 mm
diameter Teflon-insulated stainless-steel wires; Advent RM, UK)
was inserted 1.5 mm from brain surface at 30urelative to vertical
into the molecular layer of the mid antero-posterior part of the
subiculum (Figure 1A). At this position, stimulus pulses would
activate the hippocampus both directly (via activation of perforant
path (PP) fibres traversing subiculum) and indirectly (via re-
entrance through subicular output to deep layers of entorhinal
cortex which then activate superficial entorhinal PP output [9,10].
It was expected that upon stimulation, a characteristic pattern of
current sources and sinks would be identifiable across the CA1-
DG axis similar to those seen in the rat, reflecting the termination
pattern of PP inputs to DG and CA1 ([9]; see Figure 1B for typical
evoked laminar response and C for hippocampal circuit diagram).
Stimulation Protocols
Stimulus pulses were timed using a National Instruments card
(PCI-6071E, NI, UK) controlled using LabVIEW software (8.2,
NI, UK). These triggered a constant-current source (DS3,
Digitimer, UK) connected to the stimulating electrode. Stimulus
duration was set to 0.2 ms throughout the experiments. Stimulus
protocols consisted of single pulses (0.33 Hz), paired pulses
(PPulse) and trains of 20 pulses.
We first confirmed that the recording electrode spanned CA1sr,
CA1slm and the superior and inferior blades of DG. This was
achieved by monitoring the presence of stable evoked local field
potentials (LFPs) in these regions to single-pulse stimulation at
200 mA [9,10]. In all experiments, a characteristic DG molecular
layer positive-going LFP component was recorded with, in most
cases, a granule cell layer population spike (PS; latter visible with
high intensity PPulse or train stimulation; Figure 1B point C). The
CA1slm response (presumably from activation of direct EC layer
III input) was not seen in all experiments, likely due to variations
in the placement of recording/stimulating electrodes (Figure 1B
point B). CA1sr was characterised by a longer-latency negative-
going LFP (Figure 1B point A). Due to the fixed distance between
contacts in the recording array, upon identification of a typical
response in one layer, those in other layers could reliably be
confirmed due to the distance between contacts and LFP profile
(CA1sr 100 microns dorsal to CA1slm, CA1slm approximately
100 microns dorsal to the largest DG molecular layer response; see
Figure 1B for illustration of spacing).
Once a characteristic laminar LFP profile was achieved, a
current-voltage (input-output) response curve was recorded for
each LFP component by applying pairs of pulses at 50 ms intervals
over a range of current intensities (50–600 mA). The first pulse of
each pair was analysed and used to plot the curve. Thereafter, for
PPulse and train protocols, the stimulus current was set to the
value required to evoke a half-maximal response (i.e., 50% of the
value required to elicit the maximum fEPSP in CA1sr); this was
typically in the range 100–200 mA.
To examine short-term plasticity, PPulses were delivered at
inter-pulse-intervals (PPI) of 25, 50, 100, 200, 500 and 1000 ms.
All pairs were separated by 3 seconds and repeated 20 times. To
examine the effects of repetitive stimulation and re-entrance into
the hippocampal circuit, a single train of 20 pulses was applied at a
frequency of 5 Hz. This frequency was chosen to provide a
comparison with PPulse intervals and we predicted that 5 Hz
trains would not produce any long-term plastic changes [9,10].
Upon completion of all stimulation protocols, lesions were
created to mark electrode placements displaying particular
response components [37]. Mice were perfused transcardially
with 0.2 M sodium phosphate buffer and 4% paraformaldehyde
and brains removed for fixation. Electrode placements were
confirmed from 30 mm thick, Nissl-stained sagittal brain sections.
Data Acquisition and Analysis
Signals were amplified at source through an AC-coupled
headstage (x20 gain) and further amplified for a total gain of
500x (Recorder64 system, Plexon, USA). Signals were filtered
(0.1 Hz–6 kHz) and sampled at 10 kHz per channel (12-bit
Raw fEPSP response amplitudes to single-pulse stimulation
were plotted to describe the current-response relationship (input-
output; I/O) in control and 3xTgAD mice. In addition, PPulse
and train data were normalised to the first pulse and results shown
as a percentage change for both amplitude and latency to peak/
trough to demonstrate any facilitation or depression of responses.
Finally, train data were subjected to 1D CSD analysis [38] using
custom Matlab software (see below).
All numerical data were analysed using Prism (v5; Graphpad).
Mixed ANOVA with Bonferroni post hoc comparisons were
applied for identification of pair-wise genotype differences for each
of the three measures (I/O, PPulse and trains) for each response
(CA1sr, CA1slm and DG). Due to sex differences in the number of
animals present in each sample, young and old data sets were
analysed separately to control for the small female-only older
mouse group; however, young male and female data were then
pooled as no sex difference was seen for any measure (data not
shown). If recordings became unstable during an experiment,
stimulation protocol data were excluded for that animal. Final
sample sizes for each hippocampal layer in each stimulation
protocol are shown in Table 1.
Amplitude, Slope and Latency measurements
Amplitude and the latency of evoked responses were extracted
through custom Matlab software (Mathworks, version 7.0). In
brief, each response was first visually inspected to ensure it had the
expected shape with no noise or other artefacts. Five cursor points
were then marked on different components of a mean response
(see below) averaged over the 20 stimulus-response repeats.
Thereafter, the programme used this five-point template to
calculate the amplitude and latency of each original response.
For train data, the waveform of each pulse was measured ‘by
hand’ to account for the rapid changes in response profile
associated with repetitive stimulation. In both cases, five points on
each response were marked as follows: point 1 as the sample point
before the start of the stimulus artefact; points 2 and 3 surrounded
the response onset and points 5 and 6 surrounded the response
peak (or trough).
For response amplitude (mV), the mean value between points
2–3 was subtracted from the corresponding maximal (response
peak) or minimal (response trough) value between points 4 and 5.
For latency to response peak/trough (ms), the centre timestamp
between points 4 and 5 was subtracted from that for point 1. We
did not attempt to measure latency to response onset, as there was
seldom a defined level period in the recording between the
stimulus artefact and response. All data presented in this paper are
the average of 20 pulses. For DG population spikes (PS), the
probability of evoking a PS was calculated out of 20 sweeps for
Hippocampal Hyperexcitability in 3xTgAD Mouse
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Figure 1. The hippocampal tri-synaptic circuit. A: Hippocampal formation with approximate electrode placements marked. Input and output
pathways with principal direction of synaptic flow (arrows) and termination of fibre pathways from Schaffer collaterals [A], PP layer III [B] and PP layer
II [C] are marked. Evoked activity at these points corresponds to CA1sr [A], CA1slm [B] and DG responses [C]. Stimulating electrode placement was in
the deep dendritic layer of subiculum (S). B: Typical laminar field response to single-pulse stimulation at half-maximal current intensity. A, B and C
correspond to responses recorded within regions A–F in Figure 3. C: Block diagram of the connectivity displayed in [A] for reader’s clarity. Dashed
lines represent the presence of sparse regional connectivity. Calibration applies to both profiles in B.
Table 1. Summary of data included in statistical analyses.
Young (4–6 months)
Input/Output Paired Pulse Trains
Control 3xTgAD Control 3xTgAD Control 3xTgAD
CA1sr n = 12 n = 11 n = 12 n = 10 n = 10 n = 9
CA1slm n = 12 n = 11 n = 12 n = 10 n = 7 n = 8
DG n=12 n=11 n=12 n=10 n=11 n=9
Old (17–18 months)
Input/Output Paired Pulse Trains
Control 3xTgAD Control 3xTgAD Control 3xTgAD
CA1sr n=5 n=4 n=5 n=4 n=5 n=3
CA1slm n = 5 n = 4 n = 5 n = 4 n = 5 n = 3
DG n=5 n=4 n=5 n=4 n=5 n=3
Hippocampal Hyperexcitability in 3xTgAD Mouse
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each PPI in animals that demonstrated PS at half-maximum
Current Source Density Analysis
Current Source Density (CSD) analysis when applied to
extracellular recordings taken from a laminar structure provides
a highly detailed, anatomically aligned, spatiotemporal map of
current sinks and sources produced through synaptic activity.
Single responses from train data were analysed by one-
dimensional CSD analysis in a custom built Matlab programme,
estimating the second-order spatial derivative using the formula of
Freeman and Nicholson [38]. For this the distance between
electrodes was either 100 or 50 mm and the degree of spatial
smoothing applied was 2 as we analysed neighbouring contacts. It
was assumed that (a) the major extracellular currents ran in
parallel to the recording electrode (along the CA1-DG axis) and (b)
tissue conductivity was spatially homogenous across the recording
array (that is, any residual currents would represent local synaptic
sinks and sources). CSD values at locations between those
calculated for each electrode position were estimated by linear
interpolation (20 steps) to produce a smooth and continuous
mapped depiction of synaptic currents. Mapped CSD values are
presented here in arbitrary units: current sources were mapped as
reds, oranges and yellows; current neutral regions in light green;
and regions showing current sinks as dark green to dark blue. The
colour map for each CSD plot was normalised to itself (max/min)
to represent the current gradient running from its maximum
positive current (hot colours), to the maximum negative current
(cool colours) with green equivalent to resting state or neutral.
Current-response relationship
To examine the integrity of CA1 sr/slm-DG hippocampal
circuitry, amplitude and latency of responses to activation of PP
fibres were determined over a range of current intensities. There
were no significant differences in amplitude measurements
between control and 3xTgAD animals for CA1sr, CA1slm or
DG responses in either young or old mice (Figure 2A–F; genotype
(2) by PPI (5) mixed ANOVA). However, there was a significant
effect of current intensity on all response amplitudes in all animals
(F (4,28) = 5.74–23.06 P,0.0001), with higher currents eliciting
larger responses as would be expected by recruitment of PP fibres
with increasing current. In addition, DG population spikes
developed from 200 mA stimulus current onwards in 5/12 control
and 8/11 3xTgAD young mice (data not shown). In old mice, two
of the four 3xTgAD animals developed DG PS at high current
intensities; however, PS were not seen in old control mice (data not
shown). While these observations in DG PS frequency are merely
qualitative for the purpose responses to single stimuli, their
occurrence was strain-dependent in 3xTgADs and controls during
PPulse stimulation, as described below.
To investigate differences in response latency between 3xTgAd
and control mice we compared responses from each hippocampal
layer at a stimulus intensity of 200 mA (typical half-maximal
stimulation current; see Table 2). The order of latencies (to
response peak or trough) was consistent with our expectations for
the associated synaptic delays in vivo of CA1sr, CA1slm and DG in
response to PP stimulation as used here (i.e., fastest for DG and
CA1slm; refer to Figure 1C) and no genotype differences were
found (data not shown). Thus, from the similar I/O amplitude and
relative response latency patterns seen with this stimulus protocol,
we concluded that there were no genotype differences in
functional connectivity during low-frequency activation in the
PP fibre pathways from EC layers II and III traversing subiculum
to the hippocampal circuit in both young and old animals.
Short-term synaptic plasticity
To assess short-term synaptic plasticity, we examined the effect
of varying PPI on the amplitude and latency of responses to half-
maximal stimulus current (range for all mice 100–200 mA).
Measurements from the second pulse were normalised to those
from the first and are presented as percentage change, such that
positive values represent paired-pulse facilitation (or a decreased
latency) and negative values represent paired-pulse depression (or
an increased latency). CA1sr, CA1slm and DG responses were
analysed in separate genotype (2) by PPI (6) mixed ANOVA with
Bonferroni post-hoc comparisons.
For young animals, there was a significant effect of PPI on the
amplitude of CA1sr and CA1slm responses (F(5,100) = 7.32, P,
0.0001 and F(5,100) = 4.93, P,0.0005, respectively) with maximal
facilitation seen at 25 and 50 ms intervals. While there were no
significant genotype differences for CA1slm responses (data not
shown), for CA1sr there was an interaction (F(5,100) = 2.54, P,
0.05) and a pair-wise genotype difference at 50 ms (t(20) = 3.27,
P,0.01; see Figure 3A). This was evident as 3xTgAD responses
facilitating significantly more than that for controls, in stark
contrast to the depression of CA1sr responses frequently seen in
AD models in vitro during LTP protocols [21,31,32]. For the DG
response, there was again a significant genotype difference (F
(1,100) = 11.80, P,0.005) and an interaction (F(5,100) = 4.88,
P,0.001). Furthermore, there were pair-wise genotype differences
at 25 ms (t(20) = 5.12, P,0.001) and 50 ms (t(20) = 3.54, P,0.01),
where response amplitudes facilitated in 3xTgAD mice and
depressed in controls (Figure 3B), suggestive of DG hyper-
excitability in this AD model. As expected, population spikes to
the second pulse (P2) were seen in the DG granule cell layer in 4/
10 3xTgAD and 6/12 controls during half-maximal amplitude PP
stimulation. However, 3xTgAD mice had a higher tendency to
exhibit PS at shorter PPI (25 ms–100 ms), whereas for control
animals this relationship was reversed (most PS in 200 ms–
1000 ms PP interval range; examples of paired-pulse responses at
50 ms and 200 ms PPI left panel Figure 3C, probability
distribution across PPI for PS to P2 displayed in right panel
Figure 3C).
To determine whether genotype differences in the velocity of
synaptic transmission could have contributed to observed short-
term amplitude differences, response latencies were examined for
each hippocampal layer in response to pairs of pulses (latency of
second pulse to peak/trough relative to the first). There was a
significant effect of PPI on latency for CA1sr (F(5,100) = 2.63, P,
0.05) with PPI 25 ms eliciting a slightly faster response for both
genotypes (3xTgAD 14.99%63.17 SEM faster and control
5.82%61.7 SEM faster than the first pulse). For DG, a significant
effect of PPI was also seen (F(5,100) = 7.41,P,0.001), with PPI
100 ms and 200 ms eliciting a slightly slower response in both
genotypes (data not shown); however, no effects of latency were
observed in CA1slm. Thus, despite a significant effect of PPI on
the time to peak/trough of the second pulse, there were no obvious
effects of AD pathology. To summarise the data from young
animals, we found an indication of increased short-term excitabil-
ity in 3xTgAD mice in the form of increased LFP amplitude in
CA1sr, and with DG facilitation to short interval delays; however,
there was no apparent change in overt hippocampal synaptic
connectivity as reflected in the absolute latencies of CA1sr and DG
molecular layer LFPs.
For old animals there was a significant effect of PPI on response
amplitude for CA1sr (F(5,35) = 2.54, P,0.05) and a significant
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interaction (F(5,35) = 5.94, P,0.0005). This manifested as a
significant pair-wise genotype difference at PPI 50 ms (t(7)
= 5.65,P,0.0001; Figure 4A) where 3xTgAD animals again
facilitated more than controls. There were no significant effects
for CA1slm; however, there was an interaction for DG (F(5,35)
= 3.46,P,0.05) and a pair-wise genotype difference at PPI 25 ms
(t(7) = 3.08), P,0.05; Figure 4B), where again 3xTgAD responses
facilitated in contrast to control responses which depressed. Of
note, 3xTgAD DG responses tended to depress more strongly than
controls at PPI of 200 ms or longer, however, these differences did
not reach significance. Also of interest, control DG appeared to
show little if any modulatory changes across the range of PPIs,
neither depressing nor facilitating (compare Figure 3B with
Figure 4B). We assume these were age-related changes in DG
function, as they present a different profile to that of aged 3xTgAD
animals. The implication of this reduction in paired-pulse
depression for DG with normal ageing is that it may be causative
in the known age-related impairment in pattern separation that
may, in turn, contribute to our observed age-related deficit in
episodic-like memory in older control mice ([23]; see Discussion).
For latency to peak/trough, there was a significant effect of PPI
for CA1sr (F(5,35) = 2.69, P,0.05), an interaction (F(5,35) = 4.23,
P,0.005) and a pair-wise genotype difference at PPI 50 ms
(t(7) = 3.99, P,0.05). In this case, 3xTgAD mice had a signifi-
cantly longer latency relative to P1, whereas control mice had a
slightly shorter latency (3xTgAD change 59.42% 627.83 SEM
slower, control 8.08%63.49 SEM faster). This difference can be
explained by the larger CA1sr amplitudes in 3xTgAD mice
compared to controls (3xTgAD responses have a longer period
from stimulus to response trough). For CA1slm, there was a
significant interaction (F(5,35) = 4.35, P,0.01) and a pair-wise
genotype difference at PPI 50 ms (t(7) = 3.01,P,0.05) with
3xTgAD mice showing a faster P2 response than control mice
(3xTgAD 10.79%64.28 SEM faster, control 1.47%65.26 SEM
slower). For DG, there was an effect of PPI only (F(5,35) = 3.02,
P,0.05). In these old animals, only one 3xTgAD mouse showed
evidence of a DG PS and, as per the young animals, this was more
prevalent at shorter PPI (PP25ms and PP50ms; data not shown).
Thus, as per the young data, it appeared that old 3xTgAD mice
demonstrate increased short-term plasticity and excitability in
CA1sr and DG, albeit to a smaller degree when compared to
younger mice.
To summarise the findings of the PPulse protocol, both young
and old animals showed response facilitation to PPulse stimulation
at short intervals in CA1sr; however, excitability in terms of
increases in DG PS to short PPI was most pronounced at a young
age. 3xTgAD mice had a significantly larger CA1sr response than
controls and this was seen in both young and old age groups,
suggesting a specific change at the CA1 level could alter
excitability in this region. In addition, 3xTgAD DG responses at
a short PPI were facilitated relative to controls (which depressed in
young controls), indicating changes in the DG synaptic network.
The DG paired-pulse depression in young controls appeared to be
absent in old controls, suggesting an ageing-related shift in DG
Figure 2. Input-output relationships in young and old 3xTgAD mice are similar to controls. A to F: Stimulus-response (I/O) curves in
young (4–6 month) and old (17–18 months) animals in relation to increasing input current intensity at CA1sr (A and D), CA1slm (B and E) and DG (C
and F). Error bars are (6SEM).
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function; this attenuation may have mechanistic implications for
age-related deficits in the pattern separation role carried out by
DG and episodic-like memory capacity in older mice.
Train stimulation
To examine neuronal reverberation within the hippocampal
formation, mice experienced low-frequency stimulation in the
form of a 20-pulse train, delivered at 5 Hz. Train data were
analysed in genotype (2) by pulse number (20) Mixed ANOVA as
follows: raw amplitudes and peak/trough latencies were examined
for each response (results labelled as ‘raw’) and data were also
converted to a percentage change from P1, as a measure of the
summated effect of train stimulation (labelled as ‘% change’). Data
from young and old mice were also subjected to CSD analyses to
identify the spatiotemporal relationships between current sources
and sinks along the CA1-DG axis and to shed light on the origins
and relative strength of the synaptic inputs to different laminae.
5 Hz Train analysis: Young animals at 4–6 months of age
For young animals, there was a significant effect of pulse
number on the amplitude for all responses; however, for latency,
there was a significant effect of pulse number only for CA1slm and
DG responses and a genotype difference for CA1sr.
CA1sr (n = 10 control, n = 9 3xTgAD) showed a significant
effect of pulse number on amplitude (F(19, 323) = 5.94, P,0.0001
raw; F(19,323) = 6.18, P,0.0001%change; Figure 5A), that is,
subsequent pulses facilitated with respect to pulse 1. However,
despite the tendency for 3xTgAD mice to show less facilitation
throughout the train this genotype difference did not reach
significance. There was also no effect of pulse number on response
latency, however, there was a significant genotype difference
(F(1,323) = 11.01, P,0.005%change) and an interaction
(F(19,323) = 1.63, P,0.05), with Bonferroni pair-wise differences
at pulses 5–8 and 13 (t(17) = 3.071–3.725, P,0.05/P,0.01;
Figure 5B). In this case, 3xTgAD response latencies remained
the same or were slightly shorter compared to that of the first
pulse, whereas control latencies increased.
CA1slm response amplitudes (n = 7 control, n = 8 3xTgAD)
showed a significant effect of pulse number only (F(19, 247) = 5.96,
P,0.0001 raw; F(19, 247) = 6.07, P,0.0001, %change) as did
CA1slm latencies (F(19,247) = 3.37, P,0.0001 raw; F(19,
247) = 1.84, P,0.05%change) with progressive pulses evoking a
larger and slightly longer latency response (data not shown). There
were no genotype differences.
DG response amplitudes (n = 11 control, n = 9 3xTgAD)
showed a significant effect of pulse number (F(19,342) = 3.5, P,
0.0001 raw; F(19,343) = 10.36, P,0.0001%change) as did re-
sponse latencies (F(19, 342) = 3.69, P,0.0001 raw; F(19, 342) = 8,
P,0.0001), with subsequent responses depressing slightly and
latencies increasing compared to the first response. There were no
genotype differences for either response amplitude or latency (data
not shown).
In summary, there was a robust effect of pulse number on both
the amplitude and latencies of responses. We found no significant
evidence for genotype differences in the absolute amplitude of
CA1 and DG responses; however, CA1sr clearly demonstrated a
trend for poorer facilitation in 3xTgAD animals with an
accompanying genotype difference in CA1sr latency in 3xTgAD
Re-entrance into CA1 and DG: young mice
Half of the young mice from each genotype were classified as
‘excitable’ (5/11 control and 5/9 3xTgAD), that is, they showed
increasing levels of DG population spiking for subsequent pulses in
Table 2. Response latencies in hippocampal formation.
Control 4–6 mo 3xTgAD 4–6 mo Control 17–18 mo 3xTgAD 17–18 mo
Latency SEM n Latency SEM n Latency SEM n Latency SEM n
CA1sr 7.84 0.56 12 7.99 0.54 11 9.79 1.21 5 9.25 1.64 4
CA1slm 6.01 0.49 12 5.26 0.42 11 7.30 1.21 5 6.73 0.82 4
DG 8.03 0.65 12 7.46 0.36 11 6.69 2.24 5 7.81 1.26 4
Figures show peak response latencies (ms) in response to subiculum stimulation at 200 mA.
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the train and, in some cases, re-entrance into the hippocampal
circuit in both CA1 and DG (Figure 5 panels C and D; see arrows).
This is better illustrated using 1-dimensional CSD analyses, where
the evoked synaptic currents in each layer of the hippocampus can
be computed and visualised over time (see next section). All
animals that showed re-entrance did so by pulse 20: within the 5
excitable 3xTgAD animals, 3 showed clear re-entrance into CA1sr
(at pulses 10, 17 and 20; 15.7160.33 ms; mean latency 6SEM), 4
showed re-entrance into CA1slm (at pluses 15, 18 and twice by 20;
14.7160.12 ms) and 3 exhibited re-entrance into DG (at pulses 7,
15 and 20 pulses; 20.6961.98; Figure 5D). Re-entrance latencies
were similar in the 5 excitable control mice; CA1sr (n = 4;
17.561.5 ms) and CA1slm (n = 3; 15.4962.08 ms), whereas DG
had a shorter latency re-entrance wave (n = 2; 16.3262.22 ms).
However, in contrast to 3xTgAD mice, all re-entrance was seen at
pulse 20 of the train and not earlier in the sequence. It appears,
therefore, that young control and 3xTgAD mice (of 4–6 months of
age) can show self-sustained reverberation through the hippocam-
pal formation following repetitive low-frequency stimulation; in
addition, these responses tend to occur in 3xTgAD mice at an
earlier stage in the pulse sequence.
Current Source Density analysis: young mice
Stimulation of the PP with a 5 Hz frequency train elicited a
profile of current sources and sinks consistent with an electrode
traversing the CA1-DG axis, with positioning parallel to the
dendrites of pyramidal cells in CA1 and spanning DG [9]. Typical
CSD responses in and their layers of origin are outlined in
Figure 6A (control mouse response to the 5
The CSD responses for control and 3xTgAD animals can be
viewed for pulses 1, 5, 10 and 20 in Figure 6 C and D. To
orientate the reader to the appearance of each synaptic
component, the CSD responses to pulse 1 are referred to using
the example in Figure 6A. In response to pulse 1, both 3xTgAD
and control animals show an early latency current sink in CA1slm
(Figure 6A: component 1) consistent with an excitatory synaptic
response to activation of perforant path (EC layer III tempero-
ammonic) terminals on the apical dendrites of CA1 pyramidal
cells. Simultaneous with this, a sink in the molecular layer of the
inferior blade of DG is seen (Figure 6A: component 2) and an
accompanying current source in the granule cell layer (Figure 6A:
component 3). For this control mouse, a further small current
source is seen in the CA1 pyramidal cell layer, perhaps
representing the passive source of a CA1slm sink-source pair
(Figure 6A: component 4). In both genotypes a weak sink is visible
in CA1 stratum oriens, consistent with synaptic delay from the DG
mossy fibres to CA3 and output of CA3 Schaffer collaterals onto
basal CA1 dendrites (Figure 6A: component 5). These CSD
patterns were typical of first pulse responses for all animals,
including those classified as ‘excitable’ (DG PS and/or re-
entrance) and in those that did not show any evidence of PS in
DG or CA1.
Figure 3. Increased short-term plasticity in young 3xTgAD mice. Panels A and B show fEPSP amplitude of pulse 2 normalised to pulse 1
(6SEM) for CA1sr [A], and DG [B]. No differences were observed for CA1slm (data not shown). Inserts show representative traces from single
experiments with pulse 1 (black line) and pulse 2 (grey line) taken from PP50ms in all examples. Pair-wise genotype differences are evident (**P,0.01;
***P,0.0001). Calibration 12.5 ms/2 mV. C: Left panel shows examples of population spikes (PS) evoked at 50 or 200 ms in control and 3xTgAD mice
(average of 20 sweeps each; calibration 10 ms/3 mV). Right panel shows the probability of evoking a PS to the second pulse of a stimulus pair versus
paired-pulse interval. Note that the PS probability is highest at short intervals for 3xTgAD and longer intervals for controls. Error bars are (6SEM).
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In response to pulse 5, a clear PS could be seen in the DG of the
example 3xTgAD animal and this DG population spike also
begins to emerge in the control animal (Figure 6, pulse 5 upward
arrow). Visible in the control mouse, there was also the
appearance of a slightly longer component extending the existing
CA1slm sink, perhaps within the region of CA1 where CA3
Schaffer collaterals synapse onto CA1sr (also visible in Figure 6C
pulses 10 and 20). It is likely that this response was also present in
the selected 3xTgAD mouse; however, the sink was likely obscured
by the large DG PS.
In response to pulse 10, long-latency re-entrance (presumably
transferring through EC) in the 3xTgAD animal is seen in the
form of a sink-source pair in CA1 cell layer at around 15 ms
latency (Fig. 6D: component 6). There is still a clear early PS in
DG, and there is a long-latency DG source, perhaps resulting from
the inferior blade cell layer.
By pulse 20, the example control animal is also showing a clear
long-latency CA1 current source; the CA1 response being
accompanied by a population spike in the 3xTgAD example
(Figure 6D upper black arrow). In the 3xTgAD profile the early
DG cell layer response is curtailed, by a presumed inhibitory after-
potential (Figure 6D: IAP); however, there is a clear re-entrant
population spike in the inferior blade of DG (Figure 6D lower
arrow). Figure 6B shows pulse 20 for the 3xTgAD animal in more
detail, where the re-entrance of the CA1 and DG response is
clearly reflected in the overlaid LFPs. Thus, the 3xTgAD animal
appears to demonstrate increased excitability in DG, as indicated
by a larger early response to activation of the PP.
To summarize our CSD findings in young animals, we saw a
pattern of sources and sinks revealed by CSD analyses that were
consistent across controls and 3xTgAD mice with the expected
connectivity of the layer II and III EC inputs to the hippocampal
formation. In addition, comparative excitability was increased in
DG and CA1 of 3xTgADs to both the direct effect of PP activation
and indirect, delayed re-entrant input, presumably also arriving
via PP afferents.
5 Hz train analysis: Old animals of 17–18 months of age
For old animals, there were no significant effects on the
amplitude or latency for any response (when analysed as
percentage change, i.e., normalized values); however, it is likely
that some effects could have been masked due to large variances in
the control sample (n = 5 control, n = 3 AD). Nevertheless,
analysing ‘raw’ response amplitudes did show genotype differenc-
es. In detail;
CA1sr response amplitudes showed a significant effect of
genotype for raw amplitude (F(1,114) = 6.96, P,0.05), with
3xTgAD animals showing little fEPSP facilitation following the
first stimulus pulse compared to controls (Figure 7A). There was a
trend for controls to show facilitation over the course of the train
but this effect did not reach significance. For pulse 20, the
3xTgAD fEPSP (early PP-evoked response) peaked at
9.8761.64 ms (n = 3) and for control animals at 8.6361.45 ms
(n =5); however, no long latency, re-entrant responses were seen in
either group.
CA1slm responses showed no significant genotype differences
for either fEPSP amplitude or latency (data not shown), however,
there was again a trend for CA1slm amplitude to remain steady
over the train in 3xTgAD mice (Figure 7B). Again, responses in
controls appeared to facilitate over the course of the train but this
group difference could have been masked by the large variance of
the control data set. The CA1slm response to pulse 20 peaked at
6.6962.75 ms (n = 4) in 3xTgAD and at 7.7161.26 ms (n = 5) in
control mice; however, no long-latency re-entrant responses were
DG responses showed a significant effect of pulse number on
response amplitude (F(19, 114) = 4.20, P,0.0001 raw; F(19, 114)
= 4.67, P,0.0001% change) as pulses depressed over each
subsequent trial; however, in contrast to young animals, there
was little evidence of DG PS activity (except for one 3xTgAD
animal: see Figure 7D). 3xTgAD early responses peaked faster
than those for control animals (5.7661.17 ms (n = 3) versus
7.4761.95 (n = 5) respectively); however, again there was a large
within group variance, perhaps due to an effect of age and sample
size. Also, as there were no long-latency DG responses this again
supports a lack of re-entrance in all old mice.
Laminar response profiles during pulse trains are shown in
Figure 7 panels C (control) and D (3xTgAD). In contrast to the
traces seen in young animals (Figure 5 panels C and D), very little
facilitation was seen in any cell layer. Thus, in summary, the
latencies to the initial response peak/trough in old animals were
comparable, if slightly longer (c.1ms) than those in young animals;
however, no long-latency response components were seen in CA1
and DG of old mice at delays that would be associated with
Figure 4. Increased short-term plasticity in old 3xTgAD mice.
Panels A and B as per Fig 3 for CA1sr [A] and DG [B]. Pairwise genotype
differences are *P,0.05; ***P,0.0001. Note lack of modulatory effect of
PPI in controls for DG.
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hippocampal re-entrance in younger animals. The lack of an effect
of pulse number in CA1 responses, and of PS activity in animals of
both genotypes, likely reflects age-related changes in excitability.
Current source density analysis: old animals
Mice aged 17–18 months demonstrated a pattern of synaptic
current sources and sinks consistent with activation of the EC PP
from layers II and III. This profile was similar to that seen in
young animals. However, whereas young 3xTgAD and control
mice demonstrated increasing excitability over a pulse train, the
CSD profiles of old animals remained largely unchanged (Figure 7
panels E and F). Both genotypes showed what appeared to be an
inhibitory after-potential in the DG granule cell layer following the
source seen in the main DG response (Figure 7 panels E and F).
Control mice showed a large sink in CA1 stratum oriens (smaller
in 3xTgAD) at a slightly longer latency than the CA1slm response,
consistent with a synaptic delay from the CA3 Schaffer collaterals.
In the example shown (Figure 7F), the 3xTgAD animal showed a
current source in CA1sr/stratum pyramidale.
To summarize the results from aged animals, CSD analyses
revealed that the expected synaptic components reflecting PP
inputs to hippocampus were present in animals of both genotypes;
however, progressive changes in excitability to 5 Hz stimulation
were not observed. There was a significant effect of genotype on
the absolute response amplitude evoked in CA1sr and a clear
trend for a similar lack of facilitation in CA1slm (although, not
significant). Thus, we found evidence for decreased short-term
potentiation in the form of train responses in old 3xTgAD mice,
above the general decline of amplitude seen in control mice in
normal aging.
Alzheimer’s disease (AD) is characterised phenotypically by a
loss of episodic memory and pathologically by the stereotypical
accumulation of Aband tau protein. We have shown previously
that the 3xTg murine model for AD exhibits a selective deficit in
hippocampal-dependent, episodic-like memory at 6 months of age
with intact but declining performance at 3 months [23,24]. At six
months of age, 3xTgAD mice show pronounced intracellular
accumulation of Ab[20,21], strongly suggesting a causal role for
this AD pathology in our observed episodic deficits. In the present
study, we sought to examine whether this episodic-like memory
deficit is accompanied by changes in hippocampal synaptic
function through in vivo electrophysiological recordings in 3xTgAD
and control mice. A positive outcome would provide strong
evidence for a neural correlate of memory loss in early AD or
MCI. To this end, we examined synaptic responsiveness to single
pulses of increasing current intensity and short-term synaptic
plasticity through PPF and circuit reverberation (re-entrance) in
Figure 5. Changes in response latency and occurrence of hippocampal re-entrance during 5 Hz stimulus trains in young 3xTgAD
animals. A & B: fEPSP amplitude [A] and latency [B] for CA1sr response, expressed as a percentage change from the response to pulse 1. Pair-wise
genotype differences at *P,0.05 and **P,0.01. C & D: Example to show differential fEPSP responses to selected stimuli of a 5 Hz train in control [C]
and 3xTgAD [D] mice classified as ‘excitable’. Responses in ‘non-excitable’ animals during 5 Hz trains remained as per the response to pulse 1. Re-
entrance (longer latency responses, likely propagating through EC) are shown with upward pointing black arrows for pulse 20 in both genotypes and,
in this instance, by pulse 10 in [D]. Calibration bar [C,D] 2 mV by 10 ms.
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response to train stimulation in 3 regions of the hippocampal
circuit: CA1 stratum radiatum, CA1 stratum lacunosum-molecu-
lare and DG.
Synaptic connectivity and the ‘tri-synaptic’ pathway
We found no significant genotype difference in the amplitude of
evoked responses to increasing input currents in 3xTgAD and age-
matched control mice at 4–6 or 17–18 months of age. This is in
contrast to previous in vitro reports in the same model and the
mouse suggesting a depression in the input/output curve
upon the accumulation of intracellular Ab[21,31] but agrees with
other findings, showing no basal response change [30]. The
discrepancies between reports could be due to methodological
differences (in vitro slice recordings versus in vivo recordings in the
present study) and/or stimulation protocol. More specifically, in
the research cited above, single-pulse responses were examined
through direct stimulation of a monosynaptic pathway (Schaffer
collateral input to CA1 or PP input to DG [21,30,31]). In the
current study, responses were generated through stimulation
within the molecular layer of the subiculum, creating several
discrete responses: (1) an early mono-synaptic (‘direct’) response in
DG and CA1slm through activation of en passage PP fibres (the EC
layer II and III inputs to DG and CA1slm, respectively); (2) di-/tri-
synaptic responses in CA1sr and CA1so (presumably via
propagation through CA3); and (3) a longer-latency polysynaptic
(‘indirect’) response in DG and CA1. The latter represents ‘re-
entrance’ of synaptic activity, shown in the rat to depend on
activation of subicular/CA1 output that targets deep layers of EC,
from where recruitment of deep-to-superficial connectivity within
EC then allows ‘re-entrance’ of evoked activity into the DG and
hippocampus via the PP [9,10]. This re-entrant response is most
apparent during repetitive stimulation, suggesting that it requires
substantial temporal synaptic facilitation within EC. Thus, in our
in vivo 3xTgAD model, the combination of activating direct and
Figure 6. Laminar CSD profiles for responses to 5 Hz train stimulation in young mice. A: CSD from pulse 10 of control mouse with
approximate layer boundaries indicated (SO-stratum oriens, SP-stratum pyramidale, SR-stratum radiatum, SLM-stratum lacunosum-moleculare and
DG-Dentate gyrus and Alveus). Left side of all CSD panels represents point of stimulus onset. Current sources are yellow/red, current sinks light/dark
blues and neutral regions are in green. The overlaid numbers 1–6 seen in [A] and in [C–D] correspond to synaptic events as follows: slm sink (1), DG
molecular sink (2), DG granule cell source (3), CA1 source (4), SO sink (5) and CA1 long latency source (6). B: 3xTgAD CSD laminar profile with voltage
overlay of evoked response corresponding to pulse 20 in [D]. Long-latency re-entrance and PS can be seen in CA1. CSD figures from control [C] and
3xTgAD [D] mice correspond to the field response profiles for the pulses seen in Figure 5C and D. CSD analysis of pulses 1, 5, 10, and 20 of a 20-pulse
train shows progressive increases in excitability in CA1 and DG cell layers in control [C] and 3xTgAD [D] mouse. Black arrows point to short-early DG
PS (pulse 5), inferior blade DG PS (pulse 20 3xTgAD) and long-latency CA1 PS that appears to propagate away from CA1 through the alveus (pulse 20
3xTgAD). Presumed inhibitory-afterpotentials labelled as IAP. Scale bar is 20 ms. CSD figures are scaled to the same parameters for each pulse and
between genotypes.
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indirect pathways could, perhaps, have offset abnormalities
present in the monosynaptic CA3-CA1 pathway of the 3xTgAD
as found by Oddo et al. [21]. Due to a lack of any abnormality in
the basic synaptic connectivity of the 3xTgAD model shown here
in vivo, we propose that any differences found in the current study
for PPF or trains between the control and 3xTgAD mouse cannot
be attributed simply to a lower capacity or conductance of
damaged fibre pathways [28] as the current required to elicit a
half-maximum fEPSP was not different between genotypes. Thus,
in the following sections we discuss evidence for AD-specific
changes in short-term synaptic plasticity that are separate from the
basal synaptic abnormalities reported previously in the 3xTgAD
mouse [21].
Increased synaptic facilitation in the 3xTgAD model
supports the early development of hyper-excitability
To examine short-term synaptic plasticity, mice were subjected
to paired-pulse stimulation over a variety of intervals [39]. In DG,
the granule cell response to such short interval PPulse stimulation
is modulated through a combination of: (a) feed forward, GABAa-
dependent inhibition driven via PP input to local interneurons; (b)
pre-synaptic metabotropic glutamate receptor activity
(mGluR;[40,41]); and (c) feedback inhibition elicited from
recurrent interneuron collaterals in the vicinity of the activated
granule cell [41,42]. The net effect of these mechanisms is that DG
responses normally show paired-pulse depression to PP activation.
In the present study, control mice, as expected [42], exhibited
paired-pulse depression to short PPIs (25–50 ms). This depression
is presumably produced through polysynaptic GABAergic feed-
back, driven by the first pulse, causing hyperpolarisation of GCs
(mediated by GABA
and GABA
receptors[43,44]). This
inhibitory modulation not only results in depression of the field
response to PPulse stimulation, but also depresses the GC PS to
the second pulse of the pair; this effect wanes as the hyperpolar-
ising current decays during longer PPulse stimulus intervals (from
100 ms onwards; [45]). In agreement with the above mechanism,
we found control mice were most likely to display a PS to the
second pulse of a pair at PPIs of 100 ms or more, that is, at longer
intervals where presumably inhibition has subsided. In contrast,
we found the PPulse response profile of 3xTgAD mice in DG was
quite different from that of controls. Young and old 3xTgAD mice
showed pronounced synaptic facilitation at short PPIs (25 and
Figure 7. Train stimulation at 5 Hz in old animals. A. There is a significant genotype difference in raw fEPSP amplitude in old animals upon 5 Hz
frequency stimulation in CA1sr (P,0.05). B. CA1 responses did not facilitate following pulse 1 in old 3xTgAD animals (data shown for CA1slm). Control
[C] and 3xTgAD [D] representative responses for old animals. Note a lack of increase in response magnitude as the train progresses. 3xTgAD animal in
[D] was the only old animal to show DG PS. Calibration 2 mV/10 ms. CSD analysis of pulses 10 and 20 in representative old control [E] and 3xTgAD [F]
animals. CSD components are comparable to those seen in young mice at pulse 1 (Fig 6C & D; current sinks in blue, sources in red). A prolonged,
facilitating afterpotential is clearly visible from 20 ms in the DG of both control and 3xTgAD animals (marked IAP).
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50 ms), often accompanied by a PS to the second pulse. In
particular, 3xTgAD mice were more likely to display a DG PS to
the second pulse at short inter-stimulus intervals with an apparent
increasing suppression of PS at longer paired-pulse intervals (that
in controls were more likely to elicit PS). We suggest that these
results demonstrate a long-term shift to hyper-excitability in the
layer II input to DG in these AD mice for high-frequency inputs.
In contrast, we saw a shift to increased depression for low-
frequency inputs at longer PPI and in the train stimuli. Overall,
these data suggest that DG in 3xTgADs may also show a more
pronounced ‘late’ inhibitory component compared to controls,
perhaps representing a homoeostatic compensatory mechanism to
ameliorate the impact of reduced ‘early’ inhibition. Such an
increase in late inhibition may also underlie the poor facilitation
shown by 3xTgADs to low-frequency train stimulation (see below).
When examining PPulse responses in CA1 sr and slm, we found
no overall genotype differences in PPF, and CA1 responses
facilitated relative to the first pulse at short intervals in accordance
with previous reports in young AD mice [30,46]. However, we
found 3xTgAD animals demonstrated significantly more facilita-
tion in CA1sr relative to controls at a PPI of 50 ms, in accordance
with the findings of Gengler et al at the same age [46]. In old
animals, the CA1sr response of both genotypes was shown to
facilitate at short intervals but, again, there was a significantly
increased CA1sr response in old 3xTgAD animals at PPI 50 ms.
Thus, it appears from examining the responses of DG (fEPSP and
PS data), that there is a subtle reduction in the short-term,
recurrent inhibitory feedback system in the 3xTgAD mouse at
short PPIs. It is also possible that an excess of excitability in DG
and subsequent polysynaptic propagation of ‘overly excitatory’
responses through CA3 to CA1sr could account for the facilitation
that we witnessed in CA1sr. Such a mechanism could contribute
to aberrant excitation and seizure-like activity, phenomena
witnessed in vivo in other AD mouse models [33–35].
Mechanisms for increased short-term excitability
Due to the unexpected differences between control and
3xTgAD mice (i.e., the facilitation seen in 3xTgAD PPulse
responses), it is worthwhile discussing mechanisms though which
the AD phenotype could exert a hyper-excitable effect. There is
little evidence for intra or extracellular Abpathology in the DG of
the 3xTgAD model and no tau pathology localised to the DG in
older animals [20,21]. This would suggest that any change in
inhibition is separate from these signature AD pathologies. Indeed,
in human AD, the DG is one of the hippocampal regions least
affected by pathology [1]. Previous research in other AD mouse
models (APP/PS1, APOE e4 and hAPP J20) has demonstrated a
loss of interneurons within CA1 and DG. Specifically, those
expressing calcium binding proteins parvalbumin, calbindin and
calretinin, resulting in alterations in GABAergic interneuron
populations and paralleling interneuron loss in human AD patients
[47–49]. Recently, degeneration of GABAergic interneurons in
DG expressing Neuropeptide Y was seen in a novel triple-
transgenic AD model (TauPS2APP mouse; [50]). Interestingly,
enhanced LTP to PP stimulation was seen in this triple mutation
mouse in DG (in vitro), therefore, it is possible that loss of
GABAergic tone in this and other models could contribute to
hyper-excitability originating in DG [33–35,50]. Although this
could account for the excessive facilitation seen in DG and CA1sr,
we know from immunohistological studies that the 3xTgAD
mouse model does not experience overt cell loss [20,21]. Thus,
there may be similar mechanisms present in the 3xTgAD mouse
detectable through the application of antibodies sensitive to GABA
interneurons expressing calcium-binding proteins. Alternative/
additional factors include the abnormal calcium homeostasis
linked to PS1mutations [26,51,52], a reduction in nicotinic a7
acetylcholine receptors (a7nAchRs [53]) and abnormalities in the
mGluR I-III families [54,55].
3xTgAD field responses to low-frequency pulse trains are
depressed; however laminar synaptic connectivity
profiles appear normal and response re-entrance into the
hippocampal circuit is enhanced
Alongside responses to single and PPulse stimulation, we
examined the effects of low-frequency trains on presumed
monosynaptic and polysynaptic/re-entrant CA1 and DG respons-
es [7,11–13]. Young mice displayed synaptic excitability (increased
DG population cell spiking) and reverberation within the
hippocampal formation, in a subset of both 3xTgAD and control
animals. This confirms for the first time the presence of
hippocampal output and re-entrant pathways in the mouse, as
seen previously in rat ([9,10]. As an effect of genotype, small
changes in fEPSPs could be seen in CA1sr (and to a lesser extent in
CA1slm) in young 3xTgAD mice. Over a train of 20 pulses, the
fEPSP amplitude of 3xTgAD mice tended not to facilitate as much
as seen in controls (although there were no significant genotype
differences), further, they displayed relatively constant response
latencies over the train where response latencies in control mice
generally slowed. In the 3xTgAD mouse, this effect was
independent of response amplitude, thus, represented a faster rise
time to fEPSP peak.
CSD profiles in both 3xTgAD and control mice to train
stimulation showed a pattern of current sinks and sources
consistent with excitatory synapses onto DG molecular layer
(GC dendrites) and apical dendrites in CA1 [9,56]. If the
connectivity between subiculum, EC and hippocampus was intact
in the early stages of AD we expected to see: 1) a current sink in
DG, reflecting layer II EC PP inputs to the dendrites of granule
cells within the molecular layer and a positive-going fEPSP
(presumably a reversal of the population cell fEPSP); 2) a
concomitant current sink (reflected by a fast, negative-going local
field potential; LFP) within CA1slm, marking the excitatory input
from layer III PP (stimulated directly by the traversing of fibres
through subiculum); 3) a delayed CA1sr negative going LFP, of a
latency reflecting synaptic delay through the tri-synaptic connec-
tivity to EC, DG, CA3 and ending with the Schaffer collateral
input to CA1sr [9,10].
The presence of a clear CA1slm sink was variable in both
genotypes, likely due to the relative success of electrode placement
within the narrow CA1slm layer, rather than an abnormality in
the PP input. In all animals, a large DG current source could be
seen upon population spiking, thus, synaptic transmission through
PP layer II appeared intact in the 3xTgAD mouse. CSD sinks also
indicated that the (presumed) Schaffer collateral input from CA3
terminated in both stratum oriens and stratum radiatum of CA1,
suggesting that the connectivity of hippocampal fibre pathways are
functionally intact in the 3xTgAD mouse (but see [28]). Note that
we cannot discount a contribution from CA2 input to CA1 so/sr
[57]. However, the CA1sr latency would appear rather too long
for a di-synaptic response (EC layer III input to CA2 which then
projects to CA1 so/sr). Thus, the CA1sr response is much more
likely to reflect feed-forward excitation from DG via CA3 [9,10].
Animals that demonstrated response re-entrance also showed
progressive DG (and CA1 in 3xTgAD) population cell spiking and,
within a train of 20 pulses, there was evidence for hippocampal
excitability within each layer of the CA1-DG axis. For 3xTgAD in
particular, the re-entrant input to CA1 elicited PS, which could be
Hippocampal Hyperexcitability in 3xTgAD Mouse
PLOS ONE | 13 March 2014 | Volume 9 | Issue 3 | e91203
seen within as few as ten pulses of a train. However, 3xTgAD
excitatory responses were also frequently coupled with what were
presumably prolonged inhibitory after potentials (also seen in non-
excitable 3xTgAD mice), perhaps indicating abnormalities in the
balance of inhibitory feedback within hippocampus [33]. The
presence of putative inhibitory currents during a 5 Hz train is
supported by the lack of a DG second PS during seen PPulse
stimulation, where a 100 ms or more ISI suppressed DG PS
activity in 3xTgADs.
The highly specific topographic connectivity of the hippocam-
pal-entorhinal system [8–10,58–60] may also have meant that
subtle differences in recording and stimulating placements
produced a range of response profiles within each genotype;
however, placement of electrodes was consistent across all animals
at the targeted coordinates. Thus, the present response profiles
strongly suggest that there are subtle differences in the laminar
profile of hippocampal responses in 3xTgAD and control mice, in
response to low-frequency stimuli.
By 17–18 months of age, we found significant differences in the
size of CA1sr fEPSP amplitude generated in the 3xTgAD mouse;
further, their CA1 responses did not facilitate relative to the first
pulse of the train. This effect appeared to be independent of the
normal ageing process, as control mice retained some degree of
facilitation; therefore, we conclude that this change in the
3xTgAD mouse is due to AD-like pathological progression. The
latter likely includes the appearance of extracellular plaques and
hyperphosphorylated tau [20,21] in addition to a range of other
pathological markers, including increase in de-myelination [28],
inflammatory processes [20] and/or loss of synaptic density [32].
There was, however, a general decrease in excitability with age in
both genotypes, which could be due to an age-related increase in
inhibitory after-potentials, such as those after-hyperpolarisation
currents seen when recording intracellularly from hippocampal
slices derived from old animals [61]. The latter conclusion is
supported by the CSD profiles of aged animals which appear to
show: (a) a sustained sink current in DG; (b) the lack of
hippocampal re-entrance; and (c) a decrease in PS probability
during both PPulse and train stimulation. Due to the small
numbers of animals in the old group, conclusions must be drawn
with caution; nevertheless, it appears in the aged 3xTgAD mouse
there is augmentation of the usual decline in synaptic facilitation
associated with ageing. Of note, there is also an age-related
decrease in DG paired-pulse depression in control mice, a change
that may underlie the known ageing-related deficit in pattern
separation (see [62] for review) and, perhaps in turn, the age-
related deficit in episodic-like memory in older mice [23].
What do the present data contribute to our understanding of
early hippocampal dysfunction in AD? We found previously that
the 3xTgAD model exhibits an early deficit in episodic-like
memory that develops fully by 6 months of age [23,24]. We show
here that this behavioural deficit coincides with subtle abnormal-
ities in neuronal excitability and paucity of fEPSP facilitation seen
in the DG and CA1 regions, suggesting an imbalance of inhibitory
feedback in the model. However, our single-pulse responses show
no obvious evidence of degradation in hippocampal fibre
pathways [28]. Therefore, we suggest that cellular/synaptic
abnormalities (be it intracellular Abaccumulation, abnormal
calcium homeostasis or dysfunction of recurrent inhibition) are
mediating the effects witnessed here, at least in young mice. These
findings do not necessarily contradict deficits seen in LTP (which
are likely interlinked with intracellular mechanisms) seen in this
and other AD mouse models [21,25,32,46]. However, a major
conclusion from the present experiments is that hippocampal
synaptic integrity in the 3xTgAD model of familial human AD is
largely intact (as measured through single and paired-pulse low
frequency stimulation in vivo) but that responses to brief high-
frequency activation display hyper-excitability. If the 3xTgAD
model does indeed mirror the pathological progression of familial
human AD (bearing in mind that no familial patient would
experience multiple mutations in APP or PS1 together as in this
AD model), these results provide some hope that in the early
stages, the hippocampus is functionally complete. Thus, treat-
ments that can alleviate intracellular abnormalities that underlie
AD-related synaptic changes (such as chelation of excess calcium;
[26] or clearance of intracellular Ab[22]) may improve cognitive
function and quality of life in individuals at the onset of AD by
correcting the hippocampal hyper-excitability shown here by us
and others in AD models.
Author Contributions
Conceived and designed the experiments: KED JG. Performed the
experiments: KED. Analyzed the data: KED SF JG. Contributed
reagents/materials/analysis tools: SF. Wrote the paper: KED SF JG.
1. Braak H, Braak E (1991) Neuropathological stageing of Alzheimer-related
changes. Acta Neuropathol 82: 239–59.
2. Braak H, Braak E (1995) Staging of Alzheimer’s disease-related neurofibrillary
changes. Neurobiol Aging 16: 271–8.
3. Berg L, McKeel DW Jr, Miller JP, Storandt M, Rubin EH, et al. (1998)
Clinicopathologic studies in cognitively healthy aging and Alzheimer’s disease:
relation of histologic markers to dementia severity, age, sex, and apolipoprotein
E genotype. Arch Neurol 55: 326–35.
4. LaFerla FM, Green KN, Oddo S (2007) Intracellular amyloid-beta in
Alzheimer’s disease. Nat Rev Neurosci 8: 499–509.
5. Selkoe DJ (2008) Soluble oligomers of the amyloid beta-protein impair synaptic
plasticity and behavior. Behav Brain Res 192: 106–13.
6. Hwang DY, Golby AJ (2006) The brain basis for episodic memory: insights from
functional MRI, intracranial EEG, and patients with epilepsy. Epilepsy Behav 8:
7. Naber PA, Witter MP, and Lopes da Silva, FH (2000) Networks of the
hippocampal memory system of the rat. The pivotal role of the subiculum.
Ann N Y Acad Sci 911: 392–403.
8. Van Groen T, Miettinen P, Kadish I (2003) The entorhinal cortex of the mouse:
organization of the projection to the hippocampal formation. Hippocampus 13:
9. Kloosterm an F, Van Haeften T, Lopes da Silva FH (2004) Two reentrant
pathways in the hippocampal-entorhinal system. Hippocampus 14: 1026–39.
10. Kloosterman F, Van Haeften T, Witter MP, Lopes da Silva FH (2003)
Electrophysiological characterization of interlaminar entorhinal connections: an
essential link for re-entrance in the hippocampal-entorhinal system.
Eur J Neurosci 18: 3037–52.
11. Egorov AV, Hamam BN, Franse´n E, Hasselmo ME, Alonso AA (2002) Graded
persistent activity in entorhinal cortex neurons. Nature 420: 173–8.
12. Gerrard JL, Burke SN, McNaug hton BL, Barnes CA (20 08) Sequence
reactivation in the hippocampus is impaired in aged rats. J Neurosci 28:
13. Ribeiro S, Gervasoni D, Soares ES, Zhou Y, Lin SC, et al. (2004) Long-lasting
novelty-induced neuronal reverberation during slow-wave sleep in multiple
forebrain areas. PLoS Biol 2: e24. doi:10.1371/journal.pbio.0020024
14. Backman L, Small BJ, Fratiglioni L (2001) Stability of the preclinical episodic
memory deficit in Alzheimer’s disease. Brain 124: 96–102.
15. Buckmaster CA, Eichenbaum H, Amaral DG, Suzuki WA, Rapp PR (2004)
Entorhinal cortex lesions disrupt the relational organization of memory in
monkeys. J Neurosci 24: 9811–25.
16. Gouras GK, Tsai J, Naslund J, Vincent B, Edgar M, et al. (2000) Intraneuronal
Abeta42 accumulation in human brain. Am J Pathol 156: 15–20.
17. Reed JM, Squire LR (1997) Impaired recognition memory in patients with
lesions limited to the hippocampal formation. Behav Neurosci 111: 667–75.
18. Zola-Morgan S, Squire LR, Amaral DG (1986) Human amnesia and the medial
temporal region: enduring memory impairment following a bilateral lesion
limited to field CA1 of the hippocampus. J Neurosci 6: 2950–67.
Hippocampal Hyperexcitability in 3xTgAD Mouse
PLOS ONE | 14 March 2014 | Volume 9 | Issue 3 | e91203
19. Golby A, Silverberg G, Race E, Gabrieli S, O’Shea J, et al. (2005) Memory
encoding in Alzheimer’s disease: an fMRI study of explicit and implicit memory.
Brain 128: 773–87.
20. Mastrangelo MA, Bowers WJ (2008) Detailed immunohistochemical character-
ization of temporal and spatial progression of Alzheimer’s disease-related
pathologies in male triple-transgenic mice. BMC Neurosci 9: 81.
21. Oddo S, Caccamo A, Shepherd JD, Murphy MP, Golde TE, et al. (2003) Triple-
transgenic model of Alzheimer’s disease with plaques and tangles: intracellular
Abeta and synaptic dysfunction. Neuron 39: 409–21.
22. Billings LM, Oddo S, Green KN, McGaugh JL, LaFerla FM (2005)
Intraneuronal Abeta causes the onset of early Alzheimer’s disease-related
cognitive deficits in transgenic mice. Neuron 45: 675–88.
23. Davis KE, Eacott MJ, Easton A, Gigg J (2013) Episodic-like memory is sensitive
to both Alzheimer’s-like patholgical accumulation and normal ageing processes
in mice. Behav Brain Res 254: 73–82.
24. Davis KE, Easton A, Eacott MJ, Gigg J (2013) Episodic-Like Memory for What-
Where-Which Occasion is Selectively Impaired in the 3xTgAD Mouse Model of
Alzheimer’s Disease. J Alzheimers Dis 33: 681–698.
25. Gureviciene I, Ikonen S, Gurevicius K, Sarkaki A, Van Groen T, et al. (2004)
Normal induction but accelerated decay of LTP in APP +PS1 transgenic mice.
Neurobiol Dis 15: 188–95.
26. Wang Y, Greig NH, Yu QS, Mattson MP (2009) Presenilin-1 mutation impairs
cholinergic modulation of synaptic plasticity and suppresses NMDA currents in
hippocampus slices. Neurobiol Aging 30: 1061–8.
27. Witter MP (2003) Organization of cortico-hippocampal networks in rats related
to learning and memory. International Congress Series 1250: 131–145.
28. Desai MK, Sudol KL, Janelsins MC, Mastrangelo MA, et al. (2009) Triple-
transgenic Alzheimer’s disease mice exhibit region-specific abnormalities in
brain myelination patterns prior to appearance of amyloid and tau pathology.
Glia 57: 54–65.
29. Van Hoesen GW, Hyman BT (1990) Hippocampal formation: anatomy and the
patterns of pathology in Alzheimer’s disease. Prog Brain Res 83: 445–57.
30. Chapman PF, White GL, Jones MW, Cooper-Blacketer D, Marshall VJ, et al.
(1999) Impaired synaptic plasticity and learning in aged amyloid precursor
protein transgenic mice. Nat Neurosci 2: 271–6.
31. Fitzjohn SM, Morton RA, Kuenzi F, Rosahl TW, Shearman M, et al. (2001)
Age-related impairment of synaptic transmission but normal long-term
potentiation in transgenic mice that overexpress the human APP695SWE
mutant form of amyloid precursor protein. J Neurosci 21: 4691–8.
32. Jacobsen JS, Wu CC, Redwine JM, Comery TA, Arias R, et al. (2006) Early-
onset behavioral and synaptic deficits in a mouse model of Alzheimer’s disease.
Proc Natl Acad Sci U S A 103: 5161–6.
33. Palop JJ, Chin J, Roberson ED, Wang J, Thwin MT, et al. (2007) Aberrant
excitatory neuronal activity and compensatory remodeling of inhibitory
hippocampal circuits in mouse models of Alzheimer’s disease. Neuron 55:
34. Ziyatdinova S, Gurevicius K, Kutchiashvili N, Bolkvadze T, Nissinen J, et al.
(2011) Spontaneous epileptiform discharges in a mouse model of Alzheimer’s
disease are suppressed by antiepileptic drugs that block sodium channels.
Epilepsy Res 94: 75–85.
35. Yan XX, Cai Y, Shelton J, Deng SH, Luo XG, et al. (2012) Chronic Temporal
Lobe Epilepsy Is Associated with Enhanced Alzheimer-Like Neuropathology in
3xTg-AD Mice. PLoS One 7: e48782.doi:10.1371/journal.pone.0048782
36. Paxinos G, Franklin KJB (2001) The mouse brain instereotaxic coordinates. San
Diego: Academic Press.
37. Townsend G, Peloquin P, Kloosterman F, Hetke JF, Leung LS (2002) Recording
and marking with silicon multichannel electrodes. Brain Res Brain Res Protoc 9:
38. Freeman JA, Nicholson C. (1975) Experimental optimization of current source-
density technique for anuran cerebellum. J Neurophysiol 38: 369–82.
39. Zucker RS (1989) Short-term synaptic plasticity. Annu Rev Neurosci 12: 13–31.
40. Brown RE, Reymann KG (1995) Metabotropic glutamate receptor agonists
reduce paired-pulse depression in the dentate gyrus of the rat in vitro. Neurosci
Lett 196: 17–20.
41. Buzsaki G (1984) Feed-forward inhibition in the hippocampal formation. Prog
Neurobiol 22: 131–53.
42. Blaise JH, Bronzino JD (2000) Modulation of paired-pulse responses in the
dentate gyrus: effects of normal maturation and vigilance state. Ann Biomed Eng
28: 128–34.
43. Lambert NA, Wilson WA (1994) Temporally distinct mechanisms of use-
dependent depression at inhibitory synapses in the rat hippocampus in vitro.
J Neurophysiol 72: 121–30.
44. Sloviter RS (1991) Feedforward and feedback inhibition of hippocampal
principal cell activity evoked by perforant path stimulation: GABA-mediated
mechanisms that regulate excitability in vivo. Hippocampus 1: 31–40.
45. Bampton ET, Gray RA, Large CH (1999) Electrophysiological characterisation
of the dentate gyrus in five inbred strains of mouse. Brain Res 841: 123–34.
46. Gengler S, Hamilton A, Holscher C (2010) Synaptic plasticity in the
hippocampus of a APP/PS1 mouse model of Alzheimer’s disease is impaired
in old but not young mice. PLoS One 5: e9764.doi: 10.1371/journ al.-
47. Andrews-Zwilling Y, Bien-Ly N, Xu Q, Li G, Bernardo A, et al. (2010)
Apolipoprotein E4 causes age- and Tau-dependent impairment of GABAergic
interneurons, leading to learning and memory deficits in mice. J Neurosci 30:
48. Baglietto-Vargas D, Moreno-Gonzalez I, Sanchez-Varo R, Jimenez S, Trujillo-
Estrada L, et al. (2010) Calretinin interneurons are early targets of extracellular
amyloid-beta pathology in PS1/AbetaPP Alzheimer mice hippocampus.
J Alzheimers Dis 21: 119–32.
49. Takahashi H, Brasnjevic I, Rutten BP, Van Der Kolk N, Perl DP, et al. (2010)
Hippocampal interneuron loss in an APP/PS1 double mutant mouse and in
Alzheimer’s disease. Brain Struct Funct 214: 145–60.
50. Loreth D, Ozmen L, Revel FG, Knoflach F, Wetzel P, et al. (2012) Selective
degeneration of septal and hippocampal GABAergic neurons in a mouse model
of amyloidosis and tauopathy. Neurobiol Dis 47: 1–12.
51. Guo Q, Sebastian L, Sopher BL, Miller MW, Ware CB, et al. (1999) Increased
vulnerability of hippocampal neurons from presenilin-1 mutant knock-in mice to
amyloid beta-peptide toxicity: central roles of superoxide production and caspase
activation. J Neurochem 72: 1019–29.
52. Smith IF, Hitt B, Green KN, Oddo S, LaFerla FM (2005) Enhanced caffeine-
induced Ca2+release in the 3xTg-AD mouse model of Alzheimer’s disease.
J Neurochem 94: 1711–8.
53. Oddo S, Caccamo A, Green KN, Liang K, Tran L, et al. (2005) Chronic
nicotine administration exacerbates tau pathology in a transgenic model of
Alzheimer’s disease. Proc Natl Acad Sci U S A 102: 3046–51.
54. Lee HG, Zhu X, O’Neill MJ, Webber K, Casadesus G, et al. (2004) The role of
metabotropic glutamate receptors in Alzheimer’s disease. Acta Neurobiol Exp
(Wars) 64: 89–98.
55. Phillips T, Rees S, Augood S, Waldvogel H, Faull R, et al. (2000) Localization of
metabotropic glutamate receptor type 2 in the human brain. Neuroscience 95:
56. Leung LS, Roth L, Canning KJ (1995) Entorhinal inputs to hippocampal CA1
and dentate gyrus in the rat: a current-source-density study. J Neurophysiol 73:
57. Chevaleyre V, Siegelbaum SA (2010) Strong CA2 pyramidal neuron synapses
define a powerful disynaptic cortico-hippocampal loop. Neuron 66: 560–72.
58. McNaughton BL, Barnes CA (1977) Physiological identification and analysis of
dentate granule cell responses to stimulation of the medial and lateral perforant
pathways in the rat. J Comp Neurol 175: 439–54.
59. Steward O, Scoville SA (1976) Cells of origin of entorhinal cortical afferents to
the hippocampus and fascia dentata of the rat. J Comp Neurol 169: 347–70.
60. Tamamaki N (1997) Organization of the entorhinal projection to the rat dentate
gyrus revealed by Dil anterograde labeling. Exp Brain Res 116: 250–8.
61. Disterhoft JF, Oh MM (2007) Alterations in intrinsic neuronal excitability during
normal aging. Aging Cell 6: 327–36.
62. Yassa MA, Stark CE (2011) Pattern separation in the hippocampus. Trends
Neurosci 34: 515–25.
Hippocampal Hyperexcitability in 3xTgAD Mouse
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... 5XFAD mice first begin to exhibit subclinical epileptiform electroencephalographic activity and increased susceptibility to PTZ at a prodromal stage (around 4 months of age) (Abe et al. 2020;Gourmaud et al. 2022), at which time mild behavioral impairment and AD pathology are present (Kimura and Ohno 2009;Oakley et al. 2006). To further understand the mechanisms and progression of hyperexcitability in 5XFAD mice and to extend the evidence of E:I imbalance reported in other AD models at early Kazim et al. 2017;Davis, Fox, and Gigg 2014) and more advanced pathological stages (Palop, Mucke, and Roberson 2011;Palop et al. 2007;Verret et al. 2012;Hazra et al. 2016;Krantic et al. 2012) we assessed the hippocampi and cortices from prodromal 5XFAD and WT mice for E:I markers by western blot ( Figure S3). We found a significant decrease in GABAARa1/GABAARa2 ratio in the hippocampus (p<0.05, ...
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... Hippocampal-dependent impairment of learning and memory has been linked to aberrant calcium signaling (Chakroborty et al., 2009) and cellular excitability in AD mice (Davis et al., 2014). The elevated firing rate of subiculum neurons is involved in spatial navigation novelty. ...
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Alzheimer's disease (AD) is becoming increasingly prevalent worldwide. It represents one of the greatest medical challenges as no pharmacologic treatments are available to prevent disease progression. Astrocytes play crucial functions within neuronal circuits by providing metabolic and functional support, regulating interstitial solute composition, and modulating synaptic transmission. In addition to these physiological functions, growing evidence points to an essential role of astrocytes in neurodegenerative diseases like AD. Early-stage AD is associated with hypometabolism and oxidative stress. Contrary to neurons that are vulnerable to oxidative stress, astrocytes are particularly resistant to mitochondrial dysfunction and are therefore more resilient cells. In our study, we leveraged astrocytic mitochondrial uncoupling and examined neuronal function in the 3xTg AD mouse model. We overexpressed the mitochondrial uncoupling protein 4 (UCP4), which has been shown to improve neuronal survival in vitro. We found that this treatment efficiently prevented alterations of hippocampal metabolite levels observed in AD mice, along with hippocampal atrophy and reduction of basal dendrite arborization of subicular neurons. This approach also averted aberrant neuronal excitability observed in AD subicular neurons and preserved episodic-like memory in AD mice assessed in a spatial recognition task. These findings show that targeting astrocytes and their mitochondria is an effective strategy to prevent the decline of neurons facing AD-related stress at the early stages of the disease.
... Elle pourrait être la conséquence tardive de l'hyperactivation provoquée par l'amyloïdogénèse, mais pourrait également être la réponse des effets toxiques provenant de la synergie entre la pathologie amyloïde et celle des DNF (Figure 8). Malgré l'amélioration significative des techniques d'investigation dans le domaine de la recherche biomédicale, l'analyse des mécanismes cellulaires (Busche et al., 2012 ;Davis et al., 2014 ;Korzhova et al., 2021 ;Vitale et al., 2021). Dès lors, les études ont tenté de déterminer de quelle façon la plasticité synaptique au sein des réseaux hippocampiques était impactée par l'Aβ. ...
Les échecs cliniques de ces dernières années concernant le traitement de la maladie d’Alzheimer (MA) poussent aujourd’hui les chercheurs à mieux en comprendre la physiopathologie. L’accumulation toxique du peptide amyloïde-β (Aβ) est l’une des caractéristiques cardinales de la MA. L’amyloïdogénèse est impliquée dans la perturbation de l’homéostasie des réseaux neuronaux et contribue à terme aux déficits cognitifs retrouvés chez les patients. L’acide aminé D-sérine est important pour cette homéostasie en étant le co-agoniste préférentiel du récepteur N-methyl-D-aspartate (NMDA-R) du glutamate, un acteur indispensable au contrôle de la plasticité fonctionnelle des réseaux neuronaux de l’hippocampe. Plusieurs études ont directement lié la D-sérine à la MA en rapportant une modification de ses taux ou ceux de son enzyme de conversion – la sérine racémase (SR) – dans le sang, le LCR ou bien le tissu cérébral de patients. Cependant, la direction des modifications et l’implication réelle de la D-sérine dans la MA restent encore débattues. Ce travail de thèse à préciser le rôle de cet acide aminé à l’aide d’un modèle animal innovant associant une amyloïdogénèse à une délétion de la SR (souris 5xFAD/SR-KO). Des études in vivo, ex vivo et in vitro ont été menées de manière à avoir une vision la plus intégrée possible. Dans un premier temps, les travaux ont montré l’implication de la D-sérine dans de nombreux déficits cognitifs retrouvés chez les souris 5xFAD âgées présentant une amyloïdogénèse accentuée. Ces déficits étaient associés à une altération de la plasticité fonctionnelle au niveau des synapses CA3-CA1 de l’hippocampe. Ces atteintes comportementales et fonctionnelles n’étaient pas retrouvées chez la souris 5xFAD/SR-KO indiquant une contribution majeure de la D-sérine dans la physiopathologie amyloïde. Un des résultats majeurs de ce travail est la mise en évidence d’une augmentation précoce et transitoire des taux hippocampiques de D-sérine apparaissant conjointement au début de l’accumulation amyloïde chez les souris 5xFAD et qui se corrèle à une diminution progressive du recrutement des NMDA-R synaptiques. L’ensemble de ces résultats permet une meilleure compréhension de la physiopathologie associée à l'augmentation précoce des taux d’Aβ, en apportant la preuve in vivo d'une implication précoce de la D-sérine dans les désordres fonctionnels des réseaux hippocampiques induits dans un contexte d’amyloïdogénèse accentuée et dans les déficiences cognitives qui en résultent.
... It is known that oligomeric Aβ may induce neuronal hyper-excitability even in the early phases of AD patients [39][40][41][42][43] and mouse models of AD [44][45][46][47]. Therefore, we observed c-Fos expression, a marker for neuronal excitation, in glutamatergic neurons of the frontal cortex. ...
Non-cognitive behavioral and psychological symptoms often occur in Alzheimer's disease (AD) patients and mouse models, although the exact neuropathological mechanism remains elusive. Here, we report hyperactivity with significant inter-individual variability in 4-month-old APP/PS1 mice. Pathological analysis revealed that intraneuronal accumulation of amyloid-β (Aβ), c-Fos expression in glutamatergic neurons and activation of astrocytes were more evident in the frontal motor cortex of hyperactive APP/PS1 mice, compared to those with normal activity. Moreover, the hyperactive phenotype was associated with mislocalization of perivascular aquaporin 4 (AQP4) and glymphatic transport impairment. Deletion of the AQP4 gene increased hyperactivity, intraneuronal Aβ load and glutamatergic neuron activation, but did not influence working memory or anxiety-like behaviors of 4-month-old APP/PS1 mice. Together, these results demonstrate that AQP4 mislocalization or deficiency leads to increased intraneuronal Aβ load and neuronal hyperactivity in the motor cortex, which in turn causes locomotor over-activity during the early pathophysiology of APP/PS1 mice. Therefore, improving AQP4 mediated glymphatic clearance may offer a new strategy for early intervention of hyperactivity in the prodromal phase of AD.
... However, recent work suggests that early cognitive dysfunction is uncoupled from these aggregates (Arroyo-García et al., 2021;Nuriel et al., 2017;Shimojo et al., 2020). Several alternative models for early cognitive decline are under consideration (De Strooper and Karran, 2016;Frere and Slutsky, 2018), including abnormal circuit activity (Busche and Konnerth, 2015;Busche et al., 2008;Cirrito et al., 2005;Davis et al., 2014;Pooler et al., 2013;Wu et al., 2016). Circuit hyperexcitability is evident in several mouse models of familial (FAD) and sporadic AD (Lamoureux et al., 2021;Minkeviciene et al., 2009;Nuriel et al., 2017), including at prodromal stages (Bai et al., 2017;Busche and Konnerth, 2015). ...
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In Alzheimer's disease (AD), a multitude of genetic risk factors and early biomarkers are known. Nevertheless, the causal factors responsible for initiating cognitive decline in AD remain controversial. Toxic plaques and tangles correlate with progressive neuropathology, yet disruptions in circuit activity emerge before their deposition in AD models and patients. Parvalbumin (PV) interneurons are potential candidates for dysregulating cortical excitability, as they display altered AP firing before neighboring excitatory neurons in prodromal AD. Here we report a novel mechanism responsible for PV hypoexcitability in young adult familial AD mice. We found that biophysical modulation of K v 3 channels, but not changes in their mRNA or protein expression, were responsible for dampened excitability in young 5xFAD mice. These K ⁺ conductances could efficiently regulate near-threshold AP firing, resulting in gamma-frequency specific network hyperexcitability. Thus biophysical ion channel alterations alone may reshape cortical network activity prior to changes in their expression levels. Our findings demonstrate an opportunity to design a novel class of targeted therapies to ameliorate cortical circuit hyperexcitability in early AD.
... 29 Brain slices from 3xTgAD and control animals aged 3, 6, and 10 months were examined for immunoreactivity to 6e10 (targeting the N-terminus of beta-amyloid, Ab, Figure 2(a) to (f)) and AT8 (targeting neurofibrillary tangle-specific phospho-tau (Ser202, Thr205) Figure 2(g) to (l)). Consistent with previous work, 23,44 no extracellular immunoreactivity was observed to 6e10 at 3 and 6 months of age (Figure 2(a) to (d)), while AT8 binding was revealed in the amygdala at 3 months (Figure 2(h)) and also observed in the hippocampus at 6 and 10 months (Figure 2(i) to (l)). However, as no immunoreactivity was detected in the ENTl area at this stage, we conclude that 3xTgAD mice aged 3-6 months represent a pre-plaque and pre-tangle stage of AD-like pathology, partially overlapping to Braak stage $II. ...
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Functional network activity alterations are one of the earliest hallmarks of Alzheimer's disease (AD), detected prior to amyloidosis and tauopathy. Better understanding the neuronal underpinnings of such network alterations could offer mechanistic insight into AD progression. Here, we examined a mouse model (3xTgAD mice) recapitulating this early AD stage. We found resting functional connectivity loss within ventral networks, including the entorhinal cortex, aligning with the spatial distribution of tauopathy reported in humans. Unexpectedly, in contrast to decreased connectivity at rest, 3xTgAD mice show enhanced fMRI signal within several projection areas following optogenetic activation of the entorhinal cortex. We corroborate this finding by demonstrating neuronal facilitation within ventral networks and synaptic hyperexcitability in projection targets. 3xTgAD mice, thus, reveal a dichotomic hypo-connected:resting versus hyper-responsive:active phenotype. This strong homotopy between the areas affected supports the translatability of this pathophysiological model to tau-related, early-AD deficits in humans.
... 16 Despite the low half-maximal inhibitory concentration (IC50) (24.6 nM), a higher concentration of TC-2153 is required to increase Tyr phosphorylation of STEP 61 substrates in primary cortical neuronal culture (1-10 μM) and the cortex in vivo (10 mg/kg) and to reverse cognitive deficits in a 3xTg-AD mouse model of Alzheimer's disease (AD), 16 in which these mice display hippocampal hyperactivity and spontaneous seizures. 17,18 With a low level of acute toxicity (LD 50 [Lethal Dose 50] >1000 mg/kg ), 19 TC-2153 can alleviate audiogenic seizures in the FXS mouse model 15 and block pentylenetetrazol-induced convulsions, 19 although the sex dependence of the antiseizure effects of TC-2153 and the underlying mechanism were not described. ...
Objective: STriatal-Enriched protein tyrosine Phosphatase (STEP) is a brain-specific tyrosine phosphatase. Membrane-bound STEP61 is the only isoform expressed in hippocampus and cortex. Genetic deletion of STEP enhances excitatory synaptic currents and long-term potentiation in the hippocampus. However, whether STEP61 affects seizure susceptibility is unclear. Here we investigated the effects of STEP inhibitor TC-2153 on seizure propensity in a murine model displaying kainic acid (KA)-induced status epilepticus and its effect on hippocampal excitability. Methods: Adult male and female C57BL/6J mice received intraperitoneal injection of either vehicle (2.8% dimethylsulfoxide [DMSO] in saline) or TC-2153 (10 mg/kg) and then either saline or KA (30 mg/kg) 3 h later before being monitored for behavioral seizures. A subset of female mice was ovariectomized (OVX). Acute hippocampal slices from Thy1-GCaMP6s mice were treated with either DMSO or TC-2153 (10 μM) for 1 h, and then incubated in artificial cerebrospinal fluid (ACSF) and potassium chloride (15 mM) for 2 min prior to live calcium imaging. Pyramidal neurons in dissociated rat hippocampal culture (DIV 8-10) were pre-treated with DMSO or TC-2153 (10 µM) for 1 h before whole-cell patch-clamp recording. Results: TC-2153 treatment significantly reduced KA-induced seizure severity, with greater trend seen in female mice. OVX abolished this TC-2153-induced decrease in seizure severity in female mice. TC-2153 application significantly decreased overall excitability of acute hippocampal slices from both sexes. Surprisingly, TC-2153 treatment hyperpolarized resting membrane potential and decreased firing rate, sag voltage, and hyperpolarization-induced current (Ih ) of cultured hippocampal pyramidal neurons. Significance: This study is the first to demonstrate that pharmacological inhibition of STEP with TC-2153 decreases seizure severity and hippocampal activity in both sexes, and dampens hippocampal neuronal excitability and Ih . We propose that the antiseizure effects of TC-2153 are mediated by its unexpected action on suppressing neuronal intrinsic excitability.
... In vivo anesthetized recordings in older 4-6 and 17-18 months 3xTg mice comparing evoked responses in CA1 and DG of hippocampus following performant path stimulation revealed that short-term plasticity in response to brief high frequency stimulation is facilitated in both young and old 3xTg mice. Though responses to single or low frequency paired-pulses indicate that synapse integrity is nominal (Davis et al., 2014). This hippocampal hyperexcitability has been associated with pilocarpine-evoked (Yan et al., 2012), audiogenic (Kazim et al., 2017), and corneal kindled seizures (Vyver et al., 2020). ...
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Continually emerging data indicate that sub-clinical, non-convulsive epileptiform activity is not only prevalent in Alzheimer’s disease (AD) but is detectable early in the course of the disease and predicts cognitive decline in both humans and animal models. Epileptiform activity and other electroencephalographic (EEG) measures may hold powerful, untapped potential to improve the translational validity of AD-related biomarkers in model animals ranging from mice, to rats, and non-human primates. In this review, we will focus on studies of epileptiform activity, EEG slowing, and theta-gamma coupling in preclinical models, with particular focus on its role in cognitive decline and relevance to AD. Here, each biomarker is described in the context of the contemporary literature and recent findings in AD relevant animal models are discussed.
... Hippocampal-dependent impairment of learning and memory has been linked to aberrant calcium signaling (Chakroborty et al., 2009) and cellular excitability in AD mice (Davis et al., 2014). The elevated firing rate of subiculum neurons is involved in spatial navigation (Kitanishi et al., 2021), a cognitive function jeopardized in AD patients. ...
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Alzheimer’s disease (AD) is becoming increasingly prevalent worldwide. It represents one of the greatest medical challenge as no pharmacologic treatments are available to prevent disease progression. Astrocytes play crucial functions within neuronal circuits by providing metabolic and functional support, regulating interstitial solute composition, and modulating synaptic transmission. In addition to these physiological functions, growing evidence points to an essential role of astrocytes in neurodegenerative diseases like AD. Early stage AD is associated with hypometabolism and oxidative stress. Contrary to neurons that are vulnerable to oxydative stress, astrocytes are particularly resistant to mitochondrial dysfunction and are therefore more resilient cells. In our study, we leveraged astrocytic mitochondrial uncoupling and examined neuronal function in the 3xTg AD mouse model. We overexpressed the mitochondrial uncoupling protein 4 (UCP4), which has been shown to improve neuronal survival in vitro . We found that this treatment efficiently prevented alterations of hippocampal metabolite levels observed in AD mice, along with hippocampal atrophy and reduction of basal dendrite arborization of subicular neurons. This approach also averted aberrant neuronal excitability observed in AD subicular neurons and preserved episodic-like memory in AD mice assessed in a spatial recognition task. These findings show that targeting astrocytes and their mitochondria is an effective strategy to prevent the decline of neurons facing AD-related stress at the early stages of the disease.
Alzheimer's disease (AD) is the commonest neurodegenerative disease with slow progression. Pieces of evidence suggest that the GABAergic system is impaired in the early stage of AD, leading to hippocampal neuron over-activity and further leading to memory and cognitive impairment in patients with AD. However, the precise impairment mechanism of the GABAergic system on the pathogenesis of AD is still unclear. The impairment of neural networks associated with the GABAergic system is tightly associated with AD. Therefore, we describe the roles played by hippocampus-related GABAergic circuits and their impairments in AD neuropathology. In addition, we give our understand on the process from GABAergic circuit impairment to cognitive and memory impairment, since recent studies on astrocyte in AD plays an important role behind cognition dysfunction caused by GABAergic circuit impairment, which helps better understand the GABAergic system and could open up innovative AD therapy.
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The neuropathological correlates of Alzheimer's disease (AD) include amyloid-β (Aβ) plaques and neurofibrillary tangles. To study the interaction between Aβ and tau and their effect on synaptic function, we derived a triple-transgenic model (3×Tg-AD) harboring PS1M146V, APPSwe, and tauP301L transgenes. Rather than crossing independent lines, we microinjected two transgenes into single-cell embryos from homozygous PS1M146V knockin mice, generating mice with the same genetic background. 3×Tg-AD mice progressively develop plaques and tangles. Synaptic dysfunction, including LTP deficits, manifests in an age-related manner, but before plaque and tangle pathology. Deficits in long-term synaptic plasticity correlate with the accumulation of intraneuronal Aβ. These studies suggest a novel pathogenic role for intraneuronal Aβ with regards to synaptic plasticity. The recapitulation of salient features of AD in these mice clarifies the relationships between Aβ, synaptic dysfunction, and tangles and provides a valuable model for evaluating potential AD therapeutics as the impact on both lesions can be assessed.
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Episodic memory depends on the hippocampus and is sensitive to both Alzheimer's disease (AD) pathology and normal ageing. We showed previously that 3xTgAD mice express a specific, episodic-memory deficit at 6 months of age in the What-Where-Which occasion (WWWhich) task (Davis, Easton, Eacott and Gigg, 2013). This task requires the integration of object-location and contextual cues to form an episodic-like memory. Here, we explore the cumulative effect of AD pathology on WWWhich memory by testing very young and middle-aged mice (3 and 12 months old, respectively). For comparison, we included an alternative episodic-like task (What-Where-When; WWWhen) and an object temporal order (Recency) task to explore claims that WWWhen types of memory are open to non-episodic solutions. We found that in contrast to their performance at 6 months, 3-month-old 3xTgAD mice formed WWWhich episodic-like memories; however, their performance at this age was poorer than in matched controls. In contrast, 3xTgAD and control animals aged 12 months were both impaired on the WWWhich task. Finally, 3xTgAD mice with a WWWhich deficit were unimpaired in both Recency and WWWhen tasks. These results support conclusions that: (1) young 3xTgAD mice express episodic-like memory, albeit depressed relative to controls; (2) age-related changes result in a deficit on the hippocampal-dependent WWWhich episodic memory task; and (3) control and 3xTgAD mice can use recency (trace strength) rather than episodic-like memory for tasks that contain a temporal 'When' component. These results, in combination with our previous findings, support an age-related decline in WWWhich episodic-like memory in mice. Furthermore, this decline is accelerated in the 3xTgAD model.
During the past 100 years clinical studies of amnesia have linked memory impairment to damage of the hippocampus. Yet the damage in these cases has not usually been confined to the hippocampus, and the status of memory functions has often been based on incomplete neuropsychological information. Thus, the human cases have until now left some uncertainty as to whether lesions limited to the hippocampus are sufficient to cause amnesia. Here we report a case of amnesia in a patient (R.B.) who developed memory impairment following an ischemic episode. During the 5 years until his death, R.B. exhibited marked anterograde amnesia, little if any retrograde amnesia, and showed no signs of cognitive impairment other than memory. Thorough histological examination revealed a circumscribed bilateral lesion involving the entire CA1 field of the hippocampus. Minor pathology was found elsewhere in the brain (e.g., left globus pallidus, right postcentral gyrus, left internal capsule), but the only damage that could be reasonably associated with the memory defect was the lesion in the hippocampus. To our knowledge, this is the first reported case of amnesia following a lesion limited to the hippocampus in which extensive neuropsychological and neuropathological analyses have been carried out.
Transgenic mouse models have been created that mimic many of the neuropathologic and behavioral phenotypes of Alzheimer's disease (AD). Using mutations found in familial AD, the mouse models exhibit some of the cardinal features of the human disease. Wong et al. [1] and Higgins and Jacobsen [2] have written reviews of this topic. The current review extends a previous one [3] and will describe the similarities in the neuropathology of AD and the mouse models of the disease, specifically regarding neurodegeneration, and also describe treatments being developed using the mouse models.
Eighty-three brains obtained at autopsy from nondemented and demented individuals were examined for extracellular amyloid deposits and intraneuronal neurofibrillary changes. The distribution pattern and packing density of amyloid deposits turned out to be of limited significance for differentiation of neuropathological stages. Neurofibrillary changes occurred in the form of neuritic plaques, neurofibrillary tangles and neuropil threads. The distribution of neuritic plaques varied widely not only within architectonic units but also from one individual to another. Neurofibrillary tangles and neuropil threads, in contrast, exhibited a characteristic distribution pattern permitting the differentiation of six stages. The first two stages were characterized by an either mild or severe alteration of the transentorhinal layer Pre-alpha (transentorhinal stages I-II). The two forms of limbic stages (stages III-IV) were marked by a conspicuous affection of layer Pre-alpha in both transentorhinal region and proper entorhinal cortex. In addition, there was mild involvement of the first Ammon's horn sector. The hallmark of the two isocortical stages (stages V-VI) was the destruction of virtually all isocortical association areas. The investigation showed that recognition of the six stages required qualitative evaluation of only a few key preparations.
: The hippocampal system, consisting of the hippocampus, subiculum, and adjacent parahippocampal region, is known to play an important role in learning and memory processes. It is also known that the originally proposed trisynaptic circuit is a simplified representation of the organization of this system. In this paper, we present evidence, both anatomically and electrophysiologically, for the existence of direct and indirect parallel pathways through the hippocampal memory system arising from the perirhinal and postrhinal cortex. These pathways form nested loops. The subiculum occupies a central position within these loops. In the subiculum, both “raw” and highly processed information will converge. Therefore, we propose that the subiculum occupies a pivotal position in the hippocampal memory system, both as recipient and comparator of signals and as a distributor of processed information.
Progressive memory loss and cognitive dysfunction are the hallmark clinical features of Alzheimer's disease (AD). Identifying the molecular triggers for the onset of AD-related cognitive decline presently requires the use of suitable animal models, such as the 3xTg-AD mice, which develop both amyloid and tangle pathology. Here, we characterize the onset of learning and memory deficits in this model. We report that 2-month-old, prepathologic mice are cognitively unimpaired. The earliest cognitive impairment manifests at 4 months as a deficit in long-term retention and correlates with the accumulation of intraneuronal Abeta in the hippocampus and amygdala. Plaque or tangle pathology is not apparent at this age, suggesting that they contribute to cognitive dysfunction at later time points. Clearance of the intraneuronal Abeta pathology by immunotherapy rescues the early cognitive deficits on a hippocampal-dependent task. Reemergence of the Abeta pathology again leads to cognitive deficits. This study strongly implicates intraneuronal Abeta in the onset of cognitive dysfunction.