Research trends in understanding how maternal

stress and health interacts with prenatal brain development

Hanna Stevens, M.D., Ph.D. Department of Psychiatry

Iowa Neuroscience Institute Carver College of Medicine

University of Iowa

Disclosures

• Advisory board – Klingenstein Third Generation Foundation

• Research funding (past year): – Roy J Carver Trust – Nellie Ball Trust – Chromadex – Environmental Health Sciences Research Center

(NIEHS) – Hypertension Research Center

Important Questions

• What happens during prenatal brain development?

• Does maternal prenatal stress and health matter for brain development that occurs in utero?

• How can we think about buffering brain development in utero? – What molecular mechanisms are involved?

Prenatal Brain Development

Andersen (Neurosci and Biobehav Rev; 2003)

General Principles of Development

Patterning the Nervous System: regions defined by unique combinations of factors

Patterning and Regional Differences • Different regions

become defined by these early patterning signals to be the place where complex sensory, motor, emotional, executive and integrated processing occurs.

Development of the Nervous System Sanes, Reh and Harris, Copyright 2006.

Expanding Cell number

Vaccarino et al. J. Autism Dev. Dis., Vol 39, 2009

Human Brain Expansion

Dorus et al. Cell, Vol 119, 2004

Processes for Cortex Formation • Neuronal Migration:

– newer cells migrate along processes of older, parent cell

Development of the Nervous System Sanes, Reh and Harris, Copyright 2006.

Marin et al (CSH Perspectives; 2010)

Processes for Cortex Formation

• The caudate is >95% inhibitory GABAergic neurons of multiple types

– Decreased caudate GABAergic interneurons in Tourette Syndrome patients

– Overgrowth of GABAergic progenitors in macrocephalic Autism

Childhood psychiatric disorders and GABAergic neurons of the Caudate nucleus

García-Montes et al 2012; Health

García-Montes et al 2012; Health

Andersen (Neurosci and Biobehav Rev; 2003)

Maternal Health and Stress Factors • Anxiety and mood disorders are

common and can disrupt adaptive physiology & health behaviors

• The demands of pregnancy are high and can be difficult to meet

• Psychological and biological problems that arise or continue in pregnancy are highly interactive

• Medication treatment during pregnancy should be personalized, but access to care impedes this process

Christian 2012

Modeling Maternal Health and Stress Factors

Bale 2015

Bergman et al., 2007

Prenatal Stress and Later Brain Function

Prenatal stress and stress-related states are associated with:

•Neonatal outcomes: –Preterm birth and low birth weight

•Childhood outcomes: –Cognitive developmental delay –Increased behavioral & stress reactivity –Altered cortical development

•Childhood diagnosis: –Autism Spectrum Disorder risk –Tourette Syndrome symptomatology –Mood, anxiety, and ADHD symptoms

Buss et al., 2009

Model System of Prenatal Stress

Repetitive Restraint Stress 45 min of physical restraint Three times daily Days E12-E19

Embyonic Day 0 (E0)

E12

Postnatal Day 0

Elevated Plus: increased anxiety

Forced Swim: increased immobility

HPA axis dysregulation

Electrophysiological Studies showing reduced hippocampal LTP

Serotonin receptor and ligand changes in prefrontal cortex

REM sleep alterations

Increased sensitivity to drugs of abuse

Decreased dopamine transporter in ventral telencephalon

Pregnant Dam

Changes in Caudate GABAergic Neuron populations

Neurobiological differences • Increased production and populations of GABAergic neurons in

caudate: Only in Males

Lussier et al in preparation

Parker Abbott

Stephanie Lussier

Caudate GABAergic neurons

Childhood Psychiatric Disorders and the Caudate Nucleus

• Autism, ADHD, and Tourette syndrome affect the same brain functions: – Repetitive behavior and procedural learning

– Disruptions of motor regulation (hyperactivity, impulsivity)

• These brain functions depend on the caudate nucleus, a subcortical region in the forebrain that contributes to cognitive and motor behaviors

• MRI changes in caudate – Activation during motor tasks, habits, and procedural learning

– Volume changes in patients with schizophrenia, autism, Tourette syndrome, and ADHD

Anatomography

Persistence in Caudate GABAergic Neuron populations Adult GABAergic differences • Increased Caudate

GABAergic numbers: Only in Males

Behavioral differences • Learning impairments and

motor hypoactivity: Only in Males

Lussier et al in preparation

Stephanie Lussier

Rotarod: Motor learning

* *

P24 P150 P24 P150

NS PS NS PS NS PS NS PS

Fcon|Fps|Mcon|Mps

Moderators of Prenatal Stress: Sex Differences Interaction of Sex with Prenatal Stress in effects on Gene Expression in GABAergic Progenitors at Embryonic Day 13

Upregulated in Males • Cellular Proliferation/Stem Cell Genes

Sox2; Ankrd17; PCNA; Smc3; Cdc6; Cdk2; Cenph; Kif18a; Kif4; Mtbp; Nusap1; Rad21

• Mitochondrial Genes Aifm1; Atp5a1; Cox7b; Hadhb; Idh2; Ndufa5; Ndufb4; Ndufb5; Ndufs4; Pmpcb; Sdh

Upregulated in Females • Neural Differentiation Genes

Adora1; Alk; Diap1; Epb4.1l1; Hcn3; Hnmt; Kcnc4; Klhl14; Map3k12; Mapt; Nrxn1; Rac3; Rgs7; Sort1; Stx1a; Stxbp1

Jake Michaelson Lab Collaboration

00.10.20.30.40.50.6

NSF

NSM

PSF

PSM

*

*

Sox2

Ankrd17

00.00020.00040.00060.0008

0.0010.00120.0014

05

10152025 Pcna

p=0.1

Male susceptibility to Stress Factors in vitro Neurosphere preparation from embryonic mouse brain • Neuronal proliferation of male and female neural stem cells • The inflammatory stress mediator, interleukin-6 (IL6), increases

neurosphere growth only in males, not females Interleukin-6 blockade during stress blocks proliferation increase

012345

Control IL6

Rel

ativ

e 48

Hr

Neu

rosp

here

gro

wth

Control IL6

Edenia Menezes; Jessica Armer, Stevens Lab 2018

00.5

11.5

22.5

33.5

NS PS NS PS

Males Females

E13 Placenta IGF-1 Expression

Dimasuay et al 2016

Moderators of Prenatal Stress: Sex Differences

*

00.050.1

0.150.2

0.25

NS PS NS PS

Females Males

E13 Placenta Endogenous Antioxidant Glutathione

Peroxidase

• Stress differential impacts on males and females may diverge in the placenta and its support for growth

*

Prenatal Stress and Mechanisms

Mechanisms & Developmental Stage =

potential targets for treatment and prevention

What mechanisms at the molecular

and cellular level may be responsible for:

Changes in the brain during Prenatal

Stress?

What could define maternal stress that is important for these effects?

Consequences for mature brain?

GeneDisRegulationGene DisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulatio

nGeneDisRegulationGene

Abnormal Childhood, Adolescent and Adult

Brain Structure and Function

GeneDisRegulationGene DisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulatio

nGeneDisRegulationGene

Embryonic Brain Development

Cortical Inhibitory Neuron Development

• GABAergic interneurons are born in the ganglionic eminence (at embryonic day 10 to 11 in mice- at about 4 weeks gestation in humans)

• Cortical interneurons migrate tangentially into the cortical plate, then migrate/integrate radially into the cortical layers

Stevens et al 2013; PNEC

Altered distribution of inhibitory cells at Embryonic Day 14

Non-stressed

Prenatally Stressed

GAD67GFP

Embryonic Day 14: GABA Cell Tangential Migration

Rostral Caudal

Stevens et al 2013; PNEC

Non stressed Prenatally Stressed

Postnatal Trajectories of GABA Populations and Inhibited Behavior

Lussier and Stevens 2016

Maternal Stress Factors and the Embryonic Brain

Repetitive • Corticosterone • IL1beta • IL6 injection compared to oil or PBS injection Three times daily Days E12-E13

Embyonic Day 0 (E0)

E12

E14 Collection for Migration Assay

Pregnant Dam

????? No delay

Results: Migration Delay with cytokines but not glucocoticoids

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

NS PS NS/Saline NS/IL-6 NS/anti IL-6 PS/anti IL-6

GPM

as a

% o

f tot

al c

ortic

al p

late

di

stan

ce

* p=.030 * p<.001

* p=.004

IL-6 but no other mediator alone can reproduce the effect of prenatal stress on inhibitory neuron migration

Blocking IL-6 during prenatal stress does not rescue migration delay in embryonic brain

NS/IL-1beta NS/Cort

Jada Bittle and Banu Gumusoglu

Mechanisms of Prenatal Stress

Barry et al 2006

Oxidative stress and the mitochondrial function that regulates it may be critical for effects on migrating cells Ant1 mutants have disrupted mitochondrial ADP/ATP flux and abnormal build up of pro-oxidants. Inhibitory neuron migration is also disrupted

Lin-Hendel et al 2016

• Collaboration with Mike Dailey’s lab to do live imaging of neuronal migration in mouse embryonic cortical slices to directly test mediators

Jada Bittle

Oxidative Stress and Neuronal migration

Oxidative state: Disrupted migration

n = 4 n = 4

01020304050607080

Control H2O2

μm p

er h

our

Velocity

n = 4

n = 4

0

20

40

60

80

100

Control H2O2

Aver

age

devi

atio

n in

deg

rees

Angle Deviation

Oxidative Stress and Neuronal Migration

• There was a significant effect of the antioxidant, N-acetyl cysteine (NAC), on Inhibitory Neuron migration

Jada Bittle

0

20

40

60

80

100

NS PS NS NAC PS NAC

Perc

ent M

igra

tion

into

Co

rtex

Inhibitory Neuron Migration at E13 *

• Insecticides are commonly used to reduce the risk of mosquito born diseases and agricultural pests – This is becoming more

common during pregnancy to protect against the neurodevelopmental risks of mosquito born diseases

– The CDC specifically recommends two insecticides for pregnant women: DEET and permethrin

– Evidence is lacking on neurodevelopmental impacts of permethrin/cypermethrin

Prenatal toxic exposure and GABAergic migration

Prenatal toxic exposure and GABAergic migration %

Mig

ratio

n

Prenatal Stress and Mechanisms

Mechanisms & Developmental Stage =

potential targets for treatment and prevention

What mechanisms at the molecular

and cellular level may be responsible for:

Changes in the brain during Prenatal

Stress?

What could define maternal stress that is important for these effects?

Consequences for mature brain?

GeneDisRegulationGene DisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulatio

nGeneDisRegulationGene

Abnormal Childhood, Adolescent and Adult

Brain Structure and Function

GeneDisRegulationGene DisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulationGeneDisRegulatio

nGeneDisRegulationGene

Embryonic Brain Development

Preventive Measures

Brumbach et al 2017

1980’s Dietary Folate

Supplements and Neural

Tube Defects

1940’s-1960’s Folic Acid and nucleic acid

synthesis

Late 1980’s and 1990’s Folic Acid

RCTs ↓ NTDs by 70-100%

Early 1990’s: CDC recommends

Supplementation for women Mandatory

Fortification by 2000

Crider et al 2011

Acknowledgments Pattern Trust

• Nellie Ball Trust • Nat’l Center for Advancing Translational

Science (Yale Center for Clinical Investigation) • APIRE/Wyeth Pharmaceuticals • NARSAD YI Award: Dr. Mortimer D. Sackler

Developmental Psychobiology Research Program

Glial Development • Begins embryonically

following the production of neurons

• Different types (astrocytes or oligodendrocytes) develop from different regions

• Continues throughout life, particularly in response to damage and injury Development of the Nervous System

Sanes, Reh and Harris, Copyright 2006.

Programmed Cell Death In worms, all neurons have a name and their eventual function/fate is known Death of some cells appears necessary for appropriate fate specification of other cells

www.wormatlas.org

Neuronal Signaling Even as neurons and their branches are being produced, they begin sending nonspecific signals These internally-generated, initial signals play a role in neuronal development