Hippocampal Neuronal Atrophy and Cognitive Function in Delayed Poststroke and Aging-Related Dementias

Article (PDF Available)inStroke 43(3):808-14 · December 2011with37 Reads
DOI: 10.1161/STROKEAHA.111.636498 · Source: PubMed
We have previously shown delayed poststroke dementia in elderly (≥75 years old) stroke survivors is associated with medial temporal lobe atrophy; however, the basis of the structural and functional changes is unknown. Using 3-dimensional stereological methods, we quantified hippocampal pyramidal neuronal volumes and densities in a total of 95 postmortem samples from demented and nondemented poststroke survivors within our prospective Cognitive Function after Stroke study and subjects pathologically diagnosed with vascular dementia, Alzheimer disease, and mixed Alzheimer disease and vascular dementia syndrome. Hippocampal CA1 but not CA2 subfield neuron density was affected in poststroke, Alzheimer disease, vascular dementia, and mixed dementia groups relative to control subjects (P<0.05). Neuronal volume was reduced in the poststroke dementia relative to poststroke nondemented group in both CA1 and CA2, although there were no apparent differences in neuronal density. Poststroke nondemented neuronal volumes were similar to control subjects but greater than in all dementias (P<0.05). Neuronal volumes positively correlated with global cognitive function and memory function in both CA1 and CA2 in poststroke subjects (P<0.01). Degrees of neuronal atrophy and loss were similar in the poststroke dementia and vascular dementia groups. However, in the entorhinal cortex layer V, neuronal volumes were only impaired in the mixed and Alzheimer disease groups (P<0.05). Our results suggest hippocampal neuronal atrophy is an important substrate for dementia in both cerebrovascular and neurodegenerative disease.


Hippocampal Neuronal Atrophy and Cognitive Function in
Delayed Poststroke and Aging-Related Dementias
Elizabeth Gemmell, MRes; Helen Bosomworth, MRes; Louise Allan, PhD; Roslyn Hall, BSc;
Ahmad Khundakar, PhD; Arthur E. Oakley, BSc; Vincent Deramecourt, MD;
Tuomo M. Polvikoski, MD; John T. O’Brien, DM; Raj N. Kalaria, FRCPath
Background and Purpose—We have previously shown delayed poststroke dementia in elderly (75 years old) stroke
survivors is associated with medial temporal lobe atrophy; however, the basis of the structural and functional changes
is unknown.
Methods—Using 3-dimensional stereological methods, we quantified hippocampal pyramidal neuronal volumes and
densities in a total of 95 postmortem samples from demented and nondemented poststroke survivors within our
prospective Cognitive Function after Stroke study and subjects pathologically diagnosed with vascular dementia,
Alzheimer disease, and mixed Alzheimer disease and vascular dementia syndrome.
Results—Hippocampal CA1 but not CA2 subfield neuron density was affected in poststroke, Alzheimer disease, vascular
dementia, and mixed dementia groups relative to control subjects (P0.05). Neuronal volume was reduced in the
poststroke dementia relative to poststroke nondemented group in both CA1 and CA2, although there were no apparent
differences in neuronal density. Poststroke nondemented neuronal volumes were similar to control subjects but greater
than in all dementias (P0.05). Neuronal volumes positively correlated with global cognitive function and memory
function in both CA1 and CA2 in poststroke subjects (P0.01). Degrees of neuronal atrophy and loss were similar in
the poststroke dementia and vascular dementia groups. However, in the entorhinal cortex layer V, neuronal volumes
were only impaired in the mixed and Alzheimer disease groups (P0.05).
Conclusions—Our results suggest hippocampal neuronal atrophy is an important substrate for dementia in both
cerebrovascular and neurodegenerative disease. (Stroke. 2012;43:808-814.)
Key Words: Alzheimer disease hippocampus poststroke dementia stroke vascular dementia
Patients with stroke who do not develop dementia as a
direct result of a stroke have a 9-fold increased risk of
developing delayed poststroke dementia (PSD),
which can
affect up to 50% of all stroke survivors and is associated with
poor long-term survival.
However, the pathological pro-
cesses that increase vulnerability of nondemented stroke
survivors to cognitive decline are unknown. We have previ-
ously shown that medial temporal lobe atrophy (MTA) on
MRI was a predictor of delayed PSD in elderly patients with
stroke who were not cognitively impaired 3 months post-
Given that hippocampal atrophy is considered a
sensitive marker of Alzheimer disease (AD),
we postulated
that MTA reflects the presence of asymptomatic Alzheimer
pathology uncovered or exacerbated by ischemic injury.
However, there is growing evidence for MTA and hippocam-
pal degeneration associated with cerebrovascular disease in
ischemic vascular dementia (VaD)
and dementia caused
by sporadic and familial small vessel disease,
postmortem verification of neurodegenerative pathology was
not available in most of these studies.
The basis of MTA and associated functional impairment in
delayed PSD is unclear. In AD, the cause is generally thought
to be hippocampal neurodegeneration, particularly in the CA1
This may also be true in PSD because CA1
neurons are vulnerable to ischemia and hypoperfusion,
and human and experimental studies suggest there is signif-
icant CA1 neuron loss after ischemic stroke.
Greater neu-
rodegeneration in stroke survivors who developed delayed
PSD may therefore account for more severe cognitive impair-
ment compared with those who maintained normal cognitive
function. However, loss of neurons is not the only factor that
contributes to brain atrophy and functional decline. This has
been demonstrated in the hippocampus, where although
atrophy is strongly associated with memory impairment in
Received August 23, 2011; final revision received October 12, 2011; accepted October 26, 2011.
From the Centre for Brain Ageing and Vitality, Institute for Ageing and Health, Newcastle University, Campus for Ageing & Vitality, Newcastle upon
Tyne, UK.
The online-only Data Supplement is available with this article at http://stroke.ahajournals.org/lookup/suppl/doi:10.1161/
Correspondence to Raj N. Kalaria, FRCP, Institute for Ageing and Health, Campus for Ageing & Vitality, Newcastle upon Tyne, NE4 5PL, UK. E-mail
© 2011 American Heart Association, Inc.
Stroke is available at http://stroke.ahajournals.org DOI: 10.1161/STROKEAHA.111.636498
AD and normal aging,
there is limited neuron loss with
and neurodegeneration in AD makes only a weak
contribution to tissue atrophy.
Furthermore, conflicting
reports of CA1 neuron loss in VaD suggest other mechanisms
are involved.
Volumetric or atrophic changes in neuro-
nal morphometry have also been suggested as a major
contributory factor to gross structural and functional changes
in dementia.
This study therefore investigated the impact
of cellular pathology on cognitive dysfunction in cerebrovas-
cular and neurodegenerative diseases associated with MTA
and dementia. Hippocampal pyramidal neuron density and
soma volume were examined in relation to cognitive function
in prospectively assessed stroke survivors who had developed
delayed PSD or remained nondemented.
Subjects and Methods
Subject Selection and Clinical Diagnosis
The demographic details of the different subjects and pathological
findings are presented in Table 1. Hippocampal tissue was analyzed
from 36 poststroke subjects from the prospective Cognitive Function
After Stroke (CogFAST) study as described previously.
patients 75 years old were selected if they were not demented 3
months poststroke and did not exhibit disabilities that would prevent
them from completing cognitive tests. They received annual clinical
assessments and a neuropsychological test battery from baseline
including the Cognitive Drug Research battery, the Mini-Mental
State Examination, and the Cambridge Assessment of Mental Dis-
orders in the Elderly, which generated subscores for various cogni-
tive domains, including memory and executive function.
Subjects were diagnosed as demented if they met Diagnostic and
Statistical Manual of Mental Disorders, Third Edition Revised
criteria for dementia. Control subjects 75 years old were only
selected if they demonstrated no evidence of cognitive impairment;
however, they were not psychologically tested. Ethical approval was
granted by local research ethics committees for this study (Newcastle
on Tyne Hospitals Trust, UK) and permission for postmortem
research using brain tissue was granted for this project.
Neuropathological Examination
Final diagnoses of demented subjects were assigned based on
established neuropathological diagnostic criteria.
Briefly, hematox-
ylin and eosin staining was used for assessment of structural integrity
and infarcts, Nissl and Luxol fast blue staining for cellular pattern
and myelin loss, Bielschowsky silver impregnation for Consortium
to Establish a Registry for Alzheimer’s Disease (CERAD) rating of
neuritic plaques, and tau immunohistochemistry for Braak staging of
neurofibrillary tangles. A diagnosis of VaD was made when there
were multiple or cystic infarcts, lacunae, microinfarcts, and small
vessel disease and Braak stage III.
A diagnosis of AD was made
when there was evidence of significant Alzheimer-type pathology
(Braak Stage V–VI and moderate to severe CERAD score) and
absence of significant vascular pathology. Subjects were diagnosed
as “mixed” when there was evidence of concomitant VaD with AD,
Lewy body pathology, or tauopathy. We found that 70% of delayed
PSD cases met pathological criteria for VaD at autopsy with
negligible neurodegenerative pathology.
Global vascular pathology
was assessed (V.D., R.K.) and calculated as the sum of ratings of
vascular pathology in the hippocampus, frontal lobe, temporal lobe,
and basal ganglia to generate a score /20 (Deramecourt et al,
unpublished data).
Tissue Acquisition
To investigate the effects of different disease processes, we also
analyzed neurons in elderly control subjects and VaD, AD, and
dementia with mixed AD and VaD pathology cases. Brain tissues
were retrieved from the Newcastle Brain Tissue Resource (New-
castle, UK) except 4 control cases, which were obtained from the
Medical Research Council London Brain Bank for Neurodegenera-
tive Diseases (Institute of Psychiatry, London, UK). Brain tissue was
cut from predefined, paraffin-embedded blocks of the hippocampus
according to the Newcastle Brain Map.
The coronal samples were
taken between the level of the pregeniculate nucleus and the pulvinar
at which the emergence of the ventricle is visible.
Table 1. Demographic Details of Groups Analyzed*
Control PSND PSD VaD Mixed AD
Maximum no. of subjects
analyzed (total95)
14 22 16 13 16 14
Age, y
Mean (range) 81 (72–92) 84 (78 –94) 88 (80 –98) 86 (71–97) 84 (72–94) 84 (70 –91)
Braak stage
Mean (range) NPD 2.6 (1–5) 2.6 (0– 4) 2.1 (1–4) 5.1 (4– 6) 5.2 (4– 6)
Mean (range) NPD 1.5 (0 –2) 1 (0 –3) 1.3 (0 –2) 2.6 (2–3) 3 (3)
Vascular pathology
Mean (range) NPD 12.6 (7–16) 11.9 (8 –17) 13 (12–14) 3 (6–14) N/A
PMD, h
Mean (2 SE) 25 (7) 46 (12) 48 (13) 43 (10) 36 (11) 56 (18)
Fixation duration, wk
Mean (2 SE) 15 (4) 11 (3) 8 (2) 12 (6) 17 (5) 10 (3)
Section thickness,
Mean (2 SE) 24.1 (1.6) 26.3 (0.3) 27 (0.3) 27.1 (0.6) 26.2 (0.7) 26.2 (0.9)
PSND indicates nondemented poststroke subjects; PSD, delayed poststroke dementia; VaD, vascular dementia, “mixed” Alzheimer,
and vascular dementia; AD, Alzheimer disease; CERAD, Consortium to Establish a Registry for Alzheimer’s Disease; PMD, postmortem
delay; NPD, no pathological diagnoses; ECV, entorhinal cortex Layer V.
*Maximum number in control group ECV n8, PSND CA2 n20, and ECV n20; PSD CA2 n21 and ECV n11; VaD CA2 n12
and ECV n11; mixed CA2 n12 and ECV n11; AD CA2 n13 and ECV n12. Total no. in CA1 n95, in CA2 n82 and ECV
Gemmell et al Neuronal Atrophy and Poststroke Dementia 809
neuroimaging did not detect differences in the degree of MTA in the
left versus right hemisphere; therefore, sections were taken from
either the left or right hippocampus in each case provided there were
no apparent gross lesions. At least 3 30-
m sections per case were
cut using a Shandon FinessErotary microtome. Sections were
stained using the Nissl method and checked for quality, staining
consistency, and penetration. Slides were coded so the investigators
(E.G., H.B.) were blind to disease group. To minimize differential
tissue effects from processing and staining, all cases were collected,
treated, and analyzed in a standardized manner, allowing accurate
and valid comparisons to be made.
Stereological Analysis
Pyramidal neurons were analyzed in hippocampal areas CA1, CA2,
and entorhinal cortex Layer V (ECV), except 13 cases in which CA2
and 22 cases where ECV was not present or suitable for analysis
(Table 1). Sections were imaged using a Zeiss Axioplan Photomi-
croscope with a Pixelink PL-B623CF color digital camera linked to
a computer. The reference area was defined by the investigator with
a2.5 objective lens using stereological analysis software (Visiop-
harm Integration System, Hørsholm, Denmark). At this magnifica-
tion, CA1 and CA2 were easily distinguishable because CA1 was
broader with lower neuron density than CA2. The entorhinal cortex
was defined according to Insausti et al.
Layer V was identified as
a band of darkly stained large- to medium-sized pyramidal neurons
superficially bordered by cell-sparse Layer IV as described by Canto
et al.
A motorized stage (Prior ProScan II; Prior Scientific Instruments
Ltd, Cambridge, UK) with an accuracy of 1
m was used to take up
a uniform random sampling procedure within the reference area to
select approximately 33 frames from each of the 3 sections per case.
Frames were viewed at 100 magnification using an oil immersion
objective with a numeric aperture of 1.25. The optical disector
method was used for estimation of neuron density.
Based on x–y
axis length, each disector frame had an area of 2548.66
Heidenhain z-axis microcator (Heidenhain GB Ltd, London, UK),
accurate to 0.5
m, was used to precisely measure disector depth and
tissue section thickness, which was recorded every 10 frames (mean,
m). Each disector probe had a z-axis depth of 18
excluding a guard volume 4
m. Pyramidal neurons were identi-
fied using established criteria, that is, characteristic triangular soma,
Nissl-stained cytoplasm, and darkly stained single nucleolus.
pilot study was performed to ensure that the neurons were sampled
in sufficient numbers to provide precise estimates based on the mean
coefficient of error. Approximately 150 neurons were analyzed per
subfield per case, ensuring that the average sampling error reached a
satisfactory level.
The soma volume of each sampled neuron was
measured using an independent uniform random orientated nucleator
probe (Figure 1).
Statistical Analyses
Statistical analyses were carried out using SPSS Version 17.0.
Significance was set at P0.05. The Shapiro-Wilk test was used to
check for normal distribution and data nonnormally distributed were
analyzed using nonparametric tests. Group means were compared
using the Kruskal-Wallis test. Pairwise analyses were performed
using the Mann-Whitney UTest. The Wilcoxon signed rank test
was used for comparison of related means. Correlations were assessed
using Spearman rank correlation. Pearson
test or Fisher exact tests
were used to determine associations between categorical data.
There were no differences in the mean age or the distribution
of males and females between groups. Poststroke cases were
divided into 2 groups based on cognitive status at their final
assessment (mean 7.6 months before death) to allow compar-
ison between nondemented stroke survivors (PSND) and
those who developed delayed PSD (Tables 1 and 2). We
found no apparent differences in the Braak stage, CERAD
score, vascular pathology scores, or time from stroke to death
between PSND and PSD groups (Table 1). There was no
association between Braak stage 2 and delayed PSD.
Sixty-three percent (n10) of delayed PSD cases met path-
ological criteria for a final diagnosis of VaD; the remainder
were mixed AD and VaD. There were no associations
between lesion location and delayed PSD (Fisher exact test
P0.743). The coefficient of error values for neuronal
densities were CA1 P0.06, CA2 P0.002, ECV P0.02,
and for neuronal volumes were CA1 P0.04, CA2 P0.02,
ECV P0.05 demonstrating a high level of precision. Period
of postmortem delay or length of fixation (up to 40 weeks)
was not found to influence neuronal density or volume results
among the various groups (P0.05).
Neuronal Densities
As shown in Figure 2, neuronal density was different between
all regions CA2ECVCA1 (P0.001) and positively cor-
related between CA1 and CA2 and CA1 and ECV (r0.311,
Figure 1. The nucleator method for neuronal volume. Six ran-
domly oriented rays originating from the nucleolus were marked
where they crossed the border of the neuronal soma, and soma
volume was calculated using the nucleator.
P indicates pyram-
idal cell; N, nonpyramidal cell; G, glial cell. Red and green lines
delineate the disector frame. Bar10
Table 2. Clinical Findings in Poststroke Subjects
Time from baseline- death (mo)
Mean (2 SE) 63.5 (22) 64.4 (14)
Total CAMCOG score (/100)
Mean (range) 88.6 (76–99) 64 (24–80)
Memory subscore (/27)
Mean (2 SE) 17 (1.6) 10 (2)
Executive function subscore (/28)
Mean (2 SE) 22 (0.8) 16 (2.3)
Hemisphere with visible lesion on CT
(right, left, both, none) (5, 2, 5, 6) (3, 6, 2, 3)
PSND indicates nondemented poststroke subjects; PSD, delayed poststroke
dementia; CAMCOG, Cambridge Assessment of Mental Disorders in the Elderly.
810 Stroke March 2012
P0.005 and r0.247, P0.037, respectively). CA1 neuro-
nal density was different between the groups (P0.001).
Compared with control subjects, CA1 neuronal density was
reduced in the PS group (P0.025), PSND (P0.027), VaD
(P0.012), mixed (P0.001), and AD groups (P0.001).
In CA1, the mixed dementia and AD groups had lower
neuronal density than the PSND group (P0.001 and
P0.015, respectively). Surprisingly, there was no difference
in CA1 neuronal density between the PSD and PSND groups
(P0.643). CA2 and ECV neuronal densities were not
different between groups (P0.562 and P0.303, respec-
tively); however, there were trends for lower ECV neuronal
density in the mixed group compared with control subjects
and PSND groups (P0.083 and P0.08, respectively).
Neuronal Volumes
We found neuronal volume was different across all areas
(CA2CA1ECV; (P0.001; Figure 3). Providing consis-
tency in our results, neuronal volume was correlated in all 3
areas (CA1 versus CA2 r0.406, P0.001; CA1 versus ECV
r0.231, P0.05; CA2 versus ECV r0.311 P0.012).
CA1 neuronal volume was different among all groups
(P0.015). Compared with control subjects, CA1 neuronal
volume was reduced in VaD (P0.047), mixed dementia
(P0.039), and AD (P0.037). Compared with the PSND
group, CA1 neuronal volume was reduced in all dementia
groups (PSD P0.028, VaD P0.026, mixed P0.009, and
AD P0.01). However, there was no difference in neuronal
volume between the PSND group and control subjects
CA2 neuronal volume was different among all groups
(P0.002). Compared with control subjects, CA2 neuronal
volume was reduced in all dementia groups (PSD P0.014,
VaD P0.021, mixed P0.003, and AD P0.016). Com-
pared with the PSND group, CA2 neuron volume was
reduced in all dementia groups (PSD P0.009, mixed
P0.003, and AD P0.019), and there was a trend to
significance with the VaD group (P0.08). We did not find
any differences between the PSND and control subjects
(P0.959). A negative correlation was found between CA2
neuronal volume and age (r⫽⫺0.246, P0.031).
ECV neuronal volumes were different among all groups
(P0.001). ECV neuron volumes were reduced compared
with control subjects and PSND in mixed dementia (P0.001
and P0.001, respectively) and AD (P0.008 and P0.019,
respectively). There were no differences found among the PS,
VaD, and control groups. We did not find differences in
neuronal volume or density between the PSD cases that were
classified as “VaD” or “mixed” (data not shown). There were
no differences in neuronal volume or density between male
and female subjects.
Poststroke Neuronal Volume and
Cognitive Function
Final total Cambridge Assessment of Mental Disorders in the
Elderly scores were positively correlated with neuronal vol-
ume in CA1 (r0.399, P0.01) and CA2 (r0.445,
P0.007; Table 2; Figure 4). CA2 neuronal volume was
positively correlated with total memory (r0.481, P0.006),
recent memory (r0.692, P0.001), remote memory
(r0.428, P0.016) and attention Cambridge Assessment of
Figure 2. Neuronal densities in hippocampal subregions CA1, CA2, and entorhinal cortex Layer V (ECV). All PS indicates all poststroke
subjects; PSND, nondemented poststroke subjects; PSD, delayed poststroke dementia; VaD, vascular dementia, “mixed” mixed Alzhei-
mer, and vascular dementia; AD, Alzheimer disease. Means2 SE. Asterisks indicate significantly different to controls (black) or PSND
(gray; P0.05). Dots indicate trend to significance (P0.01).
Figure 3. Neuronal volumes in hippocampal
subregions CA1 , CA2, and entorhinal cortex
Layer V (ECV). All PS indicates all poststroke
subjects; PSND, nondemented poststroke
subjects; PSD, delayed poststroke dementia;
VaD, vascular dementia, “mixed” mixed Alz-
heimer, and vascular dementia; AD, Alzheimer
disease. Means2 SE. Asterisks indicate sig-
nificantly different to controls (black) or PSND
(gray; P0.05). Dots indicate trend to signifi-
cance (P0.01).
Gemmell et al Neuronal Atrophy and Poststroke Dementia 811
Mental Disorders in the Elderly subscores (r0.379,
P0.035). There were trends to significant correlations
between CA2 neuronal volume and learning (r0.352,
P0.052). ECV neuronal volume was correlated with remote
memory subscores (r0.372, P0.043). We did not find
correlations between neuronal density and cognitive function.
Neurodegenerative Pathology
CA1 neuron density was negatively correlated with Braak
stage and CERAD score across all groups (r⫽⫺0.379,
P0.002 and r⫽⫺0.0392, P0.001) even where Braak
staging was not diagnostic for AD.
CA2 and ECV neuronal
volume were negatively correlated with Braak stage
(r⫽⫺0.392, P0.003 and r⫽⫺0.395, P0.004) and
CERAD score (r⫽⫺0.261, P0.059 and r⫽⫺0.419,
P0.002). There were no correlations between vascular
pathological burden and neuronal volumes or between neu-
rodegenerative or vascular pathology and neuronal density.
Our results provide novel evidence that reduced neuronal
volume or “neuronal atrophy” is associated with cognitive
impairment in delayed PSD and other aging-related demen-
tias with both vascular etiology and AD. Pyramidal neuronal
volumes in the hippocampal subfields CA1 and CA2 were
10% to 20% smaller in delayed PSD, VaD, mixed, and AD
groups compared with cognitively normal elderly subjects (online-
only Supplementary Table, http://stroke.ahajournals.org). Interest-
ingly, the PSND group had neuronal volumes similar to
control subjects, whereas neuronal volumes in the delayed
PSD group were 20% smaller than PSND. Because the PS
groups had comparable vascular and negligible Alzheimer-
type lesion burden, neuronal atrophy appears as a selective
pathological feature discriminating nondemented and de-
mented elderly stroke survivors. This suggests that reduced
neuronal volume reflects mechanistic change occurring in
some PS survivors accelerating cognitive decline. This was
supported by the positive correlation between neuronal vol-
umes and cognitive test scores in stroke survivors. Further-
more, because neuronal volumes were reduced in VaD, AD,
and mixed dementia, our results suggest that mechanisms
causing neuronal atrophy are associated with cognitive de-
cline in both cerebrovascular disease and neurodegenerative
In agreement with the well-documented differential vul-
nerability of the entorhinal cortex to AD pathology,
nal volumes in Layer V of the entorhinal cortex were
negatively correlated with Braak stage and were reduced in
AD and mixed dementia but not affected in PSD and VaD.
The finding that neuronal volume and density changes in the
delayed PSD group were similar to those in VaD supported
clinicopathological findings that delayed PSD is associated
with vascular-type dementia driven by vascular rather than
Alzheimer or other neurodegenerative pathology.
We found that CA2 neuronal volumes were related to
various memory Cambridge Assessment of Mental Disorders
in the Elderly subscores, which supports a recent study
implicating CA2 neurons in hippocampal function and mem-
ory formation.
Our observations were also consistent with
an MRI study, which found that CA2 atrophy was more
strongly associated with mild cognitive impairment and
progression to dementia than CA1 atrophy in AD.
atrophy appears to be a potential mechanism relating CA2
atrophy to cognitive impairment in the absence of extensive
CA2 neuron loss.
Soma size is thought to reflect the amount of cellular
machinery needed to support the axodendritic arbor.
may therefore conclude that a reduction in neuronal soma
volume in patients with dementia could be related to a loss of
or reduced complexity of dendrites and/or axons.
mechanisms that cause volumetric changes in neuronal mor-
phometry could contribute to disruption of hippocampal
circuitry and cognitive impairment in neurodegenerative and
cerebrovascular disease.
Our results extend previous studies that suggested neuronal
volume is decreased in AD.
However, our findings differ
from a previous study by Zarow et al
that found no
significant change in hippocampal neuronal volume in AD
and ischemic VaD compared with control subjects. Because
they used a 3-dimensional method similar to ours, we
reasoned that the small numbers of subjects analyzed per
group may have limited their detection of statistically signif-
icant differences.
Appropriate human material is universally difficult to
obtain. However, we were fortunate to have acquired suffi-
cient material of suitable quality to allow accurate analyses
and reliable statistical results. During this study, tissue
sections were taken from predefined paraffin-embedded
blocks with a fixed starting point. This limitation therefore
precluded the strict sampling procedure necessary to conduct
Cavalieri calculations on brain region volume as well as the
Figure 4. Poststroke cognitive function
correlated with CA2 neuronal volumes. A,
CA2 neuronal volumes versus total Cam-
bridge Assessment of Mental Disorders in
the Elderly (CAMCOG) scores (r0.445,
P0.007). B, CA2 neuronal volumes ver-
sus CAMCOG memory subscores
(r0.481, P0.006). x indicates post-
stroke demented; o, poststroke
812 Stroke March 2012
subsequent estimation of total cell number within the struc-
ture using the desirable “fractionator” method.
density measures are based on the relationship between the
numerator (the cell counts) and denominator (extracellular
matrix), the effect of tissue processing cannot be ruled out.
However, all sections were processed and handled in a
standardized manner, allowing valid comparisons to be made
as any introduced artifact would be common to all sections.
Our results were consistent with the literature describing
significant loss of CA1 neurons in VaD and AD.
We were
surprised to find significantly decreased neuronal density in
the CA1 of the PSND group compared with control subjects
and no significant difference in CA1 neuron density between
PS groups because we expected greater neuron loss to explain
progression to PSD. This suggests an important role for
mechanisms causing neuronal atrophy in cognition even after
neuron loss. However, because we have shown that delayed
PSD patients have greater MTA than nondemented stroke
survivors on MRI,
the PSD neuronal density measurements
may be affected by tissue atrophy bringing surviving neurons
closer together. Because the neurodegenerative dementias we
studied are associated with MTA, the true level of neuron loss
is likely to be greater than our results suggested and closer to
values reported in previous studies.
The greatest reduction
in CA1 neuron density was found in the mixed AD and VaD
groups, which supports clinicopathological studies demon-
strating that coexistence of vascular and Alzheimer-type
pathology has an additive effect on cognition.
Our results provide novel evidence of hippocampal neuronal
atrophy in delayed PSD, VaD, mixed AD and VaD, and AD.
ECV neuronal volumes were only reduced in the mixed and
AD groups, which was consistent with the known vulnera-
bility of the ECV to AD pathology. Hippocampal CA1 and
CA2 neuronal volumes were significantly reduced in delayed
PSD subjects compared with nondemented stroke survivors
and were related to global cognitive function as well as
learning and memory function. However, there were no
differences in hippocampal neuronal density between nonde-
mented and demented stroke survivors. Our findings suggest
that even after significant neurodegeneration, therapeutic
strategies to maintain or restore functional morphology in
surviving neurons could prevent further cognitive decline in
PS and aging-related dementias.
We are grateful to the patients, families, and clinical house staff for
their cooperation in the investigation of this study. We thank
Michelle Widdrington, Carein Todd, Jean Scott, Deborah Lett, and
Anne Nicholson for assistance in managing and screening the cohort.
Sources of Funding
Our work is supported by grants from the Newcastle Centre for Brain
Ageing and Vitality (Lifelong Health and Wellbeing programme
funded by Research Councils United Kingdom), Medical Research
Council (MRC) of the United Kingdom (G0500247), and Alzhei-
mer’s Research United Kingdom (ARUK). Tissue for this study was
collected by the Newcastle Brain Tissue Resource, which is funded
in part by a grant from the MRC (G0400074), by the Newcastle
National Institute of Health Research Biomedical Research Centre in
Ageing and Age Related Diseases award to the Newcastle upon Tyne
Hospitals National Health Service Foundation Trust, and by a grant
from the Alzheimer’s Society and ARUK as part of the Brains for
Dementia Research Project.
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814 Stroke March 2012
    • "Post-stroke cerebral hypoperfusion may also provide an explanation for the loss of hippocampal CA1 area neurons observed in post-stroke demented patients, in whom neuronal volumes positively correlated with global cognitive function and memory function. Post-stroke non-demented hippocampal volumes were similar to control subjects (Gemmell et al., 2012). These data provide evidence that ET-1 mediated vascular and Ab-mediated pathology play a role in the etiology of poststroke dementia. "
    [Show abstract] [Hide abstract] ABSTRACT: In the normal central nervous system, endothelin-1 (ET-1) is found in some types of neurons, epithelial cells of the choroid plexus, and endothelial cells of microvessels, but it is usually not detectable in glial cells. However, in different pathological conditions, astrocytes adapting a reactive phenotype express high levels of ET-1 and its receptors, mainly the ETB receptor. ET-1 released by reactive astrocytes appears mainly to have neurodeleterious effects by mechanisms that include constriction of cerebral arterioles leading to impairment of the cerebral microcirculation, increase of blood brain barrier permeability, inflammation, excitotoxicity, impairment of fast axonal transport, and astrogliosis. A few studies in rodents found that ET-1 increased the astrocytic expression of brain-derived neurotrophic factor, glial cell-line derived neurotrophic factor and neurotropin-3, and the production of endocannabinoids. However, whether this occurs in physiological or pathological conditions is unclear. This review summarizes current knowledge about the role of the astrocytic ET-1 system in acute and chronic neurological conditions, including multiple sclerosis, ischemic stroke and hypoxic/ischemic brain injury, traumatic brain injury, subarachnoid hemorrhage, Alzheimer's disease, Binswanger's disease and post-stroke dementia, amyotrophic lateral sclerosis, and CNS infections. Counteracting the harmful effects of astrocytic ET-1 may represent a promising therapeutic target for mitigating secondary brain damage in a variety of neurological diseases. We also briefly address the role of astrocytic ET-1 in astrocytic tumors and pain.
    Full-text · Article · Apr 2016
    • "However , WM changes in the frontal and parieto-occipital regions correlate with hippocampal atrophy, suggesting that there is a tangible link between vascular pathology and hippocampal atrophy [115]. That such a link exists, there is selective hippocampal neuronal shrinkage which does not only appear to be an important substrate for AD but also delayed dementia after stroke in the absence of any neurodegenerative pathology [116]. This is consistent with the findings in animal models that long-term hypoperfusion does not require co-existing neurodegenerative changes to induce hippocampal atrophy [117] and demonstrates the vascular basis for hippocampal neurodegeneration and dementia. "
    [Show abstract] [Hide abstract] ABSTRACT: The global burden of ischaemic strokes is almost 4-fold greater than haemorrhagic strokes. Current evidence suggests that 25–30% of ischaemic stroke survivors develop immediate or delayed vascular cognitive impairment (VCI) or vascular dementia (VaD). Dementia after stroke injury may encompass all types of cognitive disorders. States of cognitive dysfunction before the index stroke are described under the umbrella of pre-stroke dementia, which may entail vascular changes as well as insidious neurodegenerative processes. Risk factors for cognitive impairment and dementia after stroke are multifactorial including older age, family history, genetic variants, low educational status, vascular comorbidities, prior transient ischaemic attack or recurrent stroke and depressive illness. Neuroimaging determinants of dementia after stroke comprise silent brain infarcts, white matter changes, lacunar infarcts and medial temporal lobe atrophy. Until recently, the neuropathology of dementia after stroke was poorly defined. Most of post-stroke dementia is consistent with VaD involving multiple substrates. Microinfarction, microvascular changes related to blood–brain barrier damage, focal neuronal atrophy and low burden of co-existing neurodegenerative pathology appear key substrates of dementia after stroke injury. The elucidation of mechanisms of dementia after stroke injury will enable establishment of effective strategy for symptomatic relief and prevention. Controlling vascular disease risk factors is essential to reduce the burden of cognitive dysfunction after stroke.
    Full-text · Article · Jan 2016
    • "Astrocyte cultures from the hippocampus (Hulse et al., 2001), after exposure to acidic Ringer's solution and mitochondrial inhibition, showed that the greatest degree of clasmatodendrosis occurred in the pyramidal cell body layers and closely paralleled that seen in vivo (Friede and van Houten, 1961). Given these observations, we cannot rule out that low level clasmatodendrosis occurs in the pyramidal cell layers of the cortex above the white matter in our cases and together with white matter astrocyte pathology could directly or indirectly causes neuronal damage, resulting in cognitive dysfunction (Gemmell et al., 2012; Foster et al., 2014). We have provided clinical, pathological and experimental evidence to support the interpretation of our findings on how cognitive function in ageing subjects with greater volumes of white matter hyperintensities may decline. "
    [Show abstract] [Hide abstract] ABSTRACT: White matter hyperintensities as seen on brain T2-weighted magnetic resonance imaging are associated with varying degrees of cognitive dysfunction in stroke, cerebral small vessel disease and dementia. The pathophysiological mechanisms within the white matter accounting for cognitive dysfunction remain unclear. With the hypothesis that gliovascular interactions are impaired in subjects with high burdens of white matter hyperintensities, we performed clinicopathological studies in post-stroke survivors, who had exhibited greater frontal white matter hyperintensities volumes that predicted shorter time to dementia onset. Histopathological methods were used to identify substrates in the white matter that would distinguish post-stroke demented from post-stroke non-demented subjects. We focused on the reactive cell marker glial fibrillary acidic protein (GFAP) to study the incidence and location of clasmatodendrosis, a morphological attribute of irreversibly injured astrocytes. In contrast to normal appearing GFAP+ astrocytes, clasmatodendrocytes were swollen and had vacuolated cell bodies. Other markers such as aldehyde dehydrogenase 1 family, member L1 (ALDH1L1) showed cytoplasmic disintegration of the astrocytes. Total GFAP+ cells in both the frontal and temporal white matter were not greater in post-stroke demented versus post-stroke non-demented subjects. However, the percentage of clasmatodendrocytes was increased by >2-fold in subjects with post-stroke demented compared to post-stroke non-demented subjects (P = 0.026) and by 11-fold in older controls versus young controls (P < 0.023) in the frontal white matter. High ratios of clasmotodendrocytes to total astrocytes in the frontal white matter were consistent with lower Mini-Mental State Examination and the revised Cambridge Cognition Examination scores in post-stroke demented subjects. Double immunofluorescent staining showed aberrant co-localization of aquaporin 4 (AQP4) in retracted GFAP+ astrocytes with disrupted end-feet juxtaposed to microvessels. To explore whether this was associated with the disrupted gliovascular interactions or blood-brain barrier damage, we assessed the co-localization of GFAP and AQP4 immunoreactivities in post-mortem brains from adult baboons with cerebral hypoperfusive injury, induced by occlusion of three major vessels supplying blood to the brain. Analysis of the frontal white matter in perfused brains from the animals surviving 1-28 days after occlusion revealed that the highest intensity of fibrinogen immunoreactivity was at 14 days. At this survival time point, we also noted strikingly similar redistribution of AQP4 and GFAP+ astrocytes transformed into clasmatodendrocytes. Our findings suggest novel associations between irreversible astrocyte injury and disruption of gliovascular interactions at the blood-brain barrier in the frontal white matter and cognitive impairment in elderly post-stroke survivors. We propose that clasmatodendrosis is another pathological substrate, linked to white matter hyperintensities and frontal white matter changes, which may contribute to post-stroke or small vessel disease dementia.
    Full-text · Article · Dec 2015
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