Maternal Insult and Embryonic Neurodevelopmental Damage

Hava Golan, Mahmoud Huleihel, Ira Kashtuzki-Philip, Maya Gerstein, Revital Mane, Miriam Stilman, Tamar Levav, Vered Kaplan

Dept of Developmental Genetics & Zlotowski center for Neuroscience

Energy failure

Transcriptional and translational regulation

Fetal hypoxia

RegenerationCell death

Immediate response Delayed response

Animals: Pregnant, Jackson Black C-57 mice at gestation day 17 were assigned to one of four groups:

1. Saline injections (i.p) in room air exposure (S+A), 2. Saline injections (i.p) with hypoxia chamber exposure (S+H), 3. Mg injections (i.p) in room air exposure (Mg+A),4. Mg injections (i.p) with hypoxia chamber exposure (Mg+H).

Study Design

Mg administration: 270 mg/kg followed by 27 mg/kg every 20 minutes for 4 hours. A second loading dose of 270 mg/kg was given at the end of the 4-hour period. Control mice were injected with saline following the same volume and schedule.

P 180P 14P 1

E17 E20E18 P1 P7 P14 P21 P120-P180

4h pretreatmentSaline or Mg2h hypoxia or air

9% O23% CO2

IS MAGNESIUM-SULFATE LOAD A PROPHLACTIC TREATMENT FOR PRENATAL HYPOXIA?

S+H Mg+HBehaviorMotor impaired partial protectionLearning impaired short and long term Sort-term normal long-term not

Brain morphologyCerebellum low PC density PC density normal

thinner granular layer not preventedCortex low cell density L 2-3,5-6 L5-6 protectedHippocampus no effect thinner CA1 layer

GABAergic neuronsCortex PV and CB decreased L2-3, 5-6 L2-3 partial protected Hippocampus PV in CA3 decreased not prevented Inhibitory synaptogenesisCortex not affected increased in all layersHippocapus VGAT in CA3 & hilus decreased not prevented

Glutamatergic synaptogenesisCerebellum reduced NR2AB density in PC at P1 normal NR2AB densityCortex no effect no effectHippocampus reduced NR2AB in all regions at P1 normal NR2AB density

First minute Fifth minute

S+

AS

+H

Mg+

HBehavior - Exploration in open field

Vertical Pole

0

20

40

60

80

100

vertical pole -12 cm vertical pole -18 cm

% o

f m

ice

fai

led

Wire Clinging

0

5

10

15

20

25 Mg+H

S+H

S+A

% o

f m

ice

faile

d

Str

ide

dist

ance

(m

m)

0

10

20

30

40

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80

1

Footprint Pattern

0

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50

S + A

% o

f M

ice

Sto

ppe

d

*

Motor Function

N=11-15 animals in each group Golan et al., 2004

S+A Mg+HS+H

Cerebellum Clb 3

0.0

1.0

2.0

3.0

4.0

5.0

P u r k in je c e lls

S+A

S+H

Mg+H

0

50

100

150

200

250

G r a n u la r la y e r

Gra

nula

r L

ayer

Wid

th (m

)

Pur

kinj

e ce

lls

dens

ity

(cel

l/10

0m

)Morphology - Cerebellum

**

**

N=3-4 animals in each group, 3-4 section per animal Golan et al., 2004

2h

24h

P-value < 0.05

1912 1108 594

SH - Control MgH - Control

P-value < 0.01

634 291 196

SH - Control MgH - Control

P-value < 0.05

57 48 250

SH - Control MgH - Control

P-value < 0.01

9 12 66

SH - Control MgH - Control

-800

-700

-600

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-200

-100

0

0 5 10 15 20 25 30

Cont - SH

Cont - MgH

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0 5 10 15 20 25 30

Cont - SH

Cont - MgH

Differential expression of genes

Time after hypoxia (hours)

Nu

mb

er o

f g

enes

Nu

mb

er o

f g

enes

folds change > 1.3, P<0.05

Spearman Rank CorrelationWith centered data, Pairwise centroid linkage

Pattern 1

Pattern 2

Pattern 3

Pattern 1

Pattern 2

Pattern 3