Billard, J-M

Freret, T

Gorisse, L.

Ploux, E

Radzishevsky, I

Wolosker, H

English

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HAL Id: hal-02331616
https://hal-normandie-univ.archives-ouvertes.fr/hal-02331616
Submitted on 24 Oct 2019
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D- -SERINE IS INVOLVED IN THE
- -AMYLOID- -RELATED PATHOPHYSIOLOGY IN
ALZHEIMER’S DISEASE
E Ploux, L. Gorisse, I Radzishevsky, H Wolosker, T Freret, J-M Billard
To cite this version:
E Ploux, L. Gorisse, I Radzishevsky, H Wolosker, T Freret, et al.. D- -SERINE IS INVOLVED IN THE
- -AMYLOID- -RELATED PATHOPHYSIOLOGY IN ALZHEIMER’S DISEASE. 14eme Colloque de
la Société des Neurosciences, May 2019, Marseille, France. ￿hal-02331616￿
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1 
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300 400 500 
WT	  	  (n=15)	  
5xFAD	  (n=17)	  
5xFAD/SR-­‐/-­‐	  (n10)	  
fE
PS
P/
PF
V  
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+o

S+mulus  intensity  (μA)
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D-­‐serine	  level	  12	  month-­‐old	  
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D-­‐serine	  level	  3-­‐4	  month-­‐old	  
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WT (n=11) 
5xFAD (n=17) 
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WT	  (n=11)	  
5xFAD	  (n=13)	  
5xFAD/SR-­‐/-­‐	  (n=11)	  
10-­‐12	  month-­‐old	  
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Time  (min)
0 30 6015 45
#
	  
Extracellular	   recording	   in	   CA1	   stratum	   radiatum	   hippocampal	   slices	   from	   either	  
3-­‐4	  or	  10-­‐12	  month-­‐old	  mice	  (aEer	  electrical	  sFmulaFon	  of	  Schaﬀer	  collaterals)	  
	  	  	  	  	  
•  High	  frequency	  (HFS)-­‐induced	  long-­‐term	  potenLaLon	  (LTP):	  
	  	  	  
-­‐  TesFng	  sFmulaFon	  (average	  of	  3	  sFmulaFons	  at	  0,1	  Hz)	  for	  15min	  baseline	  and	  	  
then	  resumed	  for	  60	  min	  aEer	  condiFoning	  
-­‐  CondiFoning	  :	  1	  electrical	  train	  of	  sFmulaFon	  at	  100	  Hz	  for	  1	  sec	  	  
	  	  	  	  	  	  	  
•  Isolated	  NMDAr-­‐mediated	  fEPSPs	  (ﬁeld	  Excitatory	  PostSynapLc	  PotenLals):	  
	  	  	  
-­‐  PresynapFc	   Fiber	   Volley	   (PFV)	   and	   ﬁeld	   Excitatory	   PostsynapFc	   PotenFals	  
(fEPSPs)	   were	   recorded	   and	   fEPSP/PFV	   raFo	   were	   calculated	   for	   Input/Output	  
curves	  	  
-­‐  Recording	   were	   performed	   (low	   magnesium	   supplemented	   medium	   with	   the	  
non-­‐NMDAr	  antagonist	  NBQX,	  10µM)	  either	  before	  and	  15	  min	  aVer	  addiLon	  of	  
D-­‐serine	  (100	  µM)	  
D-­‐SERINE  IS  INVOLVED  IN  THE  β-­‐AMYLOID-­‐RELATED  PATHOPHYSIOLOGY  
IN  ALZHEIMER’S  DISEASE
1Ploux	  E.,	  Gorisse-­‐Hussonnois	  L.,	  2Radzishevsky	  I.,	  2Wolosker	  H.,	  1Freret	  T.,	  1Billard	  J-­‐M.	  	  
1Normandie	  Univ,	  UNICAEN,	  INSERM,	  COMETE,	  GIP	  CYCERON,	  14000	  Caen,	  France	  
2Department	  of	  biochemistry,	  Technion,	  Israel	  InsItute	  of	  Technology,	  Haifa,	  Israel	  	  
	   	   	   	   	   AcFvaFon	   of	   N-­‐methyl-­‐D-­‐aspartate	   subtype	   of	  
glutamate	   receptors	   (NMDAr),	   key	   regulators	   of	   funcFonal	  
brain	  plasFcity	  and	  memory	  process,	  requires	  the	  binding	  of	  
the	  co-­‐agonist	  D-­‐serine.	  The	  homeostasis	  of	  these	  receptors	  
are	   aﬀected	   by	   soluble	   oligomers	   of	   the	   beta-­‐amyloid	  
pepLde	   (Aßo)	   in	  Alzheimer´s	  disease	   (AD).	  Aßo	  toxic	  eﬀects	  
possibly	   pass	   through	   mechanisms	   involving	   D-­‐serine	   since	  
Aßo	   sFmulate	   in	   vitro	   the	   producFon	   of	   the	   co-­‐agonist.	  
However,	   the	   actual	   in	   vivo	   contribuFon	   of	   D-­‐serine	   in	   the	  
funcFonal	  NMDAr-­‐related	  deregulaFons	  mediated	  by	  Aßo	   is	  
sFll	  unknown.	  	  
Spontaneous	  alternaFon	  test	  
	  
	  
-­‐  12	  month-­‐old	  mice	  were	  tested	  for	  
working	  memory	  performances	  	  
	  
-­‐  Mice	   were	   let	   free	   to	   explore	   Y-­‐
maze	   for	   8-­‐min	   and	   successive	  
entry	  of	  the	  3	  arms	  of	  the	  maze	  was	  
considered	  as	  an	  alternaFon	  
	  
-­‐  The	  percentage	  of	  alternaLon	  was	  
calculated	   as	   follows:	   NB	   of	  
alternaFons	   /	   (total	   NB	   of	   arms	  
visited	  –	  2)	  x	  100	  
Hippocampal  phenotype  of  the  experimental  groups
5xFAD	  and	  5xFAD/SR-­‐/-­‐	  display	  similar	  hippocampal	  level	  of	  Aßo	  	  
(only	  traces	  were	  observed	  in	  WT	  mice,	  used	  as	  negaIve	  control)	  
5xFAD/SR-­‐/-­‐	  mice	  at	  both	  ages	  show	  markedly	  reduced	  hippocampal	  level	  of	  D-­‐serine	  	  
(either	  compared	  to	  WT	  mice	  or	  5xFAD)	  
Func+onal  plas+city  (HFS-­‐induced  LTP)
SR	  deleFon	  (5xFAD/SR-­‐/-­‐)	  reverses	  LTP	  deﬁcits	  displayed	  in	  5xFAD	  mice,	  
both	  in	  3-­‐4	  and	  10-­‐12	  month-­‐old	  animals	  
NMDAr  ac+va+on  (isolated  synap+c  poten+als)
No	  genotype	  diﬀerence	  of	  NMDAr	  acFvaFon	  is	  observed	  in	  basal	  condiFons	  (A)	  
Only	  WT	  and	  5xFAD/SR-­‐/-­‐	  mice	  show	  signiﬁcant	  D-­‐serine-­‐induced	  increase	  in	  NMDAr	  acFvaFon	  (B)	  
Our	   results	   show	   that	   deleLon	   of	   serine	   racemase	  
prevents,	  at	  least	  parLally,	  memory-­‐related	  behavioral	  
deﬁcits	   observed	   in	   mice	   with	   prominent	   features	   of	  
amyloïdogenesis.	   Besides,	   impairment	   of	   NMDAr-­‐
dependent	  funcLonal	  plasLcity,	   indicated	  a	  signiﬁcant	  
contribuLon	   of	   D-­‐serine	   in	   NMDAr-­‐dependent	   β-­‐
amyloid	  related	  pathophysiology	  of	  AD.	  	  
1	  
1	  
2	  
Working  
memory
SR	  deleLon	  (5xFAD/SR-­‐/-­‐)	  
	  reverses	  working	  memory	  deﬁcits	  
displayed	  by	  5xFAD	  
SR	  deleFon	  (5xFAD/SR-­‐/-­‐)	  parFally	  reverses	  reference	  memory	  
deﬁcits	  displayed	  by	  5xFAD	  mice	  aEer	  a	  second	  spaFal	  learning	  
Spa+al  learning,  Flexibility,  Reference  memory
Contact	  :	  
eva.ploux@unicaen.fr	  
5xFAD	  and	  5xFAD/SR-­‐/-­‐	  mice	  display	  reduced	  spaFal	  
learning	  performances	  (compared	  to	  WT	  mice)	  
5xFAD	  mice	  display	  ﬂexibility	  deﬁcits	  (compared	  to	  WT	  
mice),	  fully	  reversed	  in	  5xFAD/SR-­‐/-­‐	  mice	  
Learning Relearning
Probe-­‐test1 Probe-­‐test2
Unaltered	  reference	  memory	  in	  5xFAD	  and	  5xFAD/SR-­‐/-­‐	  mice	  
Morris	  Water	  Maze	  
	  	  
	  	  	  	  
-­‐  12	   month-­‐old	   mice	   were	   trained	   to	   learn	   locaFon	   of	   hidden	  
plaporm	  (through	  disFnct	  visual	  cues)	  
-­‐  Learning	   (spaIal	   learning	   performance)	   and	   relearning	  
(ﬂexibility)	   session:	   4	   trials/day	   during	   5	   days	   (60	   sec	   inter-­‐trial	  
interval)	  
	  
	  
-­‐  Probe	  tests	  (spaFal	  reference	  memory	  performance):	  performed	  
48	   aEer	   last	   trial	   of	   (respecFvely,	   learning	   and	   relearning	  
session).	   Plaporm	  was	   removed	   and	  mice	  were	   free	   to	   explore	  
the	  maze	  during	  60	  sec	  	  
Phenotype	  	  
characterizaLon	  
	  
•  Hippocampal	   Aβ42	   	   levels	  	  
3-­‐4	  months	  old	  mice	   (n=4	  
to	  6	  per	  group)	  
	  
•  Hippocampal	   D-­‐serine	  
levels	   3-­‐4	   and	   12	   month	  
old	   mice	   (n=6	   for	   all	  
groups)	  
Behavioral	  characterizaLon	  
5xFAD/SR-­‐/-­‐	  
?	  
♂	  
3	  
In	   this	   study,	   we	  wonder	   if	   D-­‐serine	   contributes	   in	   vivo	   to	  
NMDAr	  deregulaFons	  mediated	  by	  Aßo.	  Behavioral	  analysis	  
combined	  to	  extracellular	  electrophysiological	  recordings	  at	  
hippocampal	  synapses	  is	  conducted	  in	  the	  5xFAD	  transgenic	  
mice	  model	  of	  amyloïdogenesis	  displaying	  marked	  increase	  
in	  Aßo	  rates.	  They	  are	  compared	  to	  5xFAD	  animals	  in	  which	  
the	   homozygous	   gene	   of	   the	   serine	   racemase	   (SR)	   that	  
synthesizes	  D-­‐serine,	  has	  been	  jointly	  invalidated.	   B
fE
PS
P/
PF
V  
ra
+o

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WT 5xFAD 5xFAD/SR-­‐/-­‐
+  
D
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+  
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Day	  1	   Day	  2	   Day	  3	   Day	  4	   Day	  5	  
Sw
im
m
in
g	  
di
st
an
ce
	  (c
m
)	  
WT	  
5xFAD	  
5xFAD/SR-­‐/-­‐	  
N=19	  
*	  
***	  
***	  
#	  #	  #	  
#	  #	  #	  
N=22	  
N=19	  
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Day	  7	   Day	  8	   Day	  9	   Day	  10	   Day	  11	  
Sw
im
m
in
g	  
di
st
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	  (c
m
)	  
WT	  
5xFAD	  
5xFAD/SR-­‐/-­‐	  
#	  #	  #	  
*	  
***	  
***	  
N=19	  
N=18	  
N=22	  
WT
5x
FA
D -/-
5x
FA
D/S
R
40
45
50
55
60
65
%
 A
lte
rn
at
io
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$ $
N=18	   N=18	   N=21	  
Electrophysiology	  
Mean	  +	  sem	  ;	  ANOVA	  repeated	  measures	  *	  p<	  0.05	  ;	  ***	  p<	  0.001	  ;	  ###	  p<	  0.001	  
Mean	  +	  sem	  ;	  Univariate	  t-­‐test	  :	  $	  p<0.05	  vs	  50%	  
Mean	  +	  sem	  
ANOVA	  one-­‐way	  No	  group	  eﬀect	  
Univariate	  t-­‐test	  :	  $	  p<0.05	  vs	  25%	  
KO	  Serine	  
Racemase	  (SR-­‐/-­‐)	  
D-­‐Serine	  
L-­‐Serine	  
2	   3	  
#	  #	  #	  #	  
N=19	   N=19	   N=22	   N=19	   N=19	   N=22	   N=19	   N=18	   N=22	   N=19	   N=18	   N=22	  
#
Mean	  +	  sem	  	  
ANOVA	  one-­‐way	  	  #	  p<0.05	  
Mean	  +	  sem	  	  
ANOVA	  one-­‐way	  No	  group	  eﬀect	  
Mean	  +	  sem	  	  
ANOVA	  repeated	  measures	  #	  #	  p<0.01	  
Mean	  +	  sem	  
ANOVA	  one-­‐way	  *	  p<0.05	  
Mean	  +	  sem	  
ANOVA	  one-­‐way	  **	  p<0,01	  
Univariate	  t-­‐test	  :	  $	  p<0.05	  vs	  25%	  
Mean	  +	  sem	  
ANOVA	  one-­‐way	  *	  p<0.05	  
Mean	  +	  sem	  ;	  ANOVA	  repeated	  measures	  *	  p<	  0.05	  ;	  ***	  p<	  0.001	  ;	  ###	  p<	  0.001	  
3-­‐4	  month-­‐old	   3-­‐4	  month-­‐old	  
***	   ***	   ***	  
Mean	  +	  sem	  	  
ANOVA	  one-­‐way	  	  ***	  p<0.001	  vs	  WT	  
WT	  	  
(C57Bl/6)	   5xFAD	  
Ca²+	  
LTP	  
Memory	  process	  
D-­‐serine	  Glutamate	  
+	  
Morphology	  
NMDA	  
Receptor	  
ää	  D-­‐serine	  ?	  
Ca²+	  
LTP	  
Memory	  process	  
Impaired	  
 ää	  
Glutamate	  
	  Neurodegenerescence	  
NMDAR	  
deregulaLon	  	  
Aβ	  oligomers	  
♂	   ♂	  


D-­‐SERINE IS INVOLVED IN THE β-­‐AMYLOID-­‐RELATED PATHOPHYSIOLOGYIN ALZHEIMER’S DISEASE

HAL - Normandie Université

D-serine is involved in the β-amyloid-related pathophysiology in Alzheimer’s disease

https://hal-normandie-univ.archives-ouvertes.fr/hal-02331616/document

D-‐SERINE IS INVOLVED IN THE β-‐AMYLOID-‐RELATED PATHOPHYSIOLOGYIN ALZHEIMER’S DISEASE

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Archive Ouverte en Sciences de l'Information et de la Communication

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D-­‐SERINE IS INVOLVED IN THE β-­‐AMYLOID-­‐RELATED PATHOPHYSIOLOGYIN ALZHEIMER’S DISEASE

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Archive Ouverte en Sciences de l'Information et de la Communication

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D-‐SERINE IS INVOLVED IN THE β-‐AMYLOID-‐RELATED PATHOPHYSIOLOGYIN ALZHEIMER’S DISEASE