Drug and Chemical Exposures in Animal Models Related to ASD Theodore Slotkin, Ph.D. Department of...
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Transcript of Drug and Chemical Exposures in Animal Models Related to ASD Theodore Slotkin, Ph.D. Department of...
Drug and Chemical Exposures in Animal Models Related to ASD
Theodore Slotkin, Ph.D.Department of Pharmacology & Cancer Biology
Integrated Toxicology & Environmental Health ProgramDuke University
Support: NIH ES10356
Main Points
• Why an increase in neurodevelopmental disorders including ASD?
• Why do neuroactive agents produce permanent alterations with developmental exposures?
• Why is there a critical period for these effects?
• Why do apparently unrelated agents produce similar outcomes?
• Example from environmental chemicals: organophosphate pesticides
• Example from prenatal drug exposure: terbutaline
Developmental Neurotoxicity from Environmental Chemical Exposures
5000 new chemicals/year
EPA estimate: 25% neurotoxic
67% of High Production Chemicals Not Tested for Neurotoxicity
High vulnerability of the developing brain
Increases in ADHD, learning/cognitive problems?• 17% of US schoolchildren suffer from neurobehavioral disabilities
• Annual cost: $80-170 billion
• 250% increase in ADHD diagnosis between 1990-1998
• 190% increase in children in special ed for learning disabilities between 1977-1994
• Increase in autistic spectrum disorders from 4/10,000 (1980s) to 30-60 (1990s)
Developmental Neurotoxicants - The “Silent Pandemic”
LDDI Initiative, 2007 Grandjean & Landrigan, Lancet 2006
Why Neuroactive Agents Disrupt Brain Development —Neurotransmitter Signals Control Cell Fate
NerveTerminal
Receptors
SignalingCascades Nucleus
Gene Transcription
Replicate Differentiate Grow Die Learn
The same neurotransmitter may be used for multiple decisions
Why there is a Critical PeriodWhy there is a Critical Period
Change in Cell DifferentiationChange in Cell Differentiation
Permanent Change in the Permanent Change in the Response to StimulationResponse to Stimulation
Input Input AfterAfter Critical Period Critical Period
Short-Term Response ElicitedShort-Term Response Elicited
Short-Term, Reversible, Short-Term, Reversible, Compensatory AdjustmentsCompensatory Adjustments
Input Input DuringDuring Critical Period Critical Period
Apparently Unrelated Agents Can Produce Similar Outcomes —[maybe we shouldn’t focus on common mechanisms?]
Correct Connection Damage or Loss of Input Damage or Loss of Target
Miswired Connection Mismatched Phenotypes
Corollary - exposure to multiple agents can produce additive or synergistic effects - worsened outcome
Organophosphate Pesticides — Chlorpyrifos
Developmental neurotoxicity unrelated to mechanisms in adults
Effects are subtle but widespread
Originally modeled in animals, neurodevelopmental deficits now confirmed in children (inner-city, agricultural populations)
Developmental exposure increases autism risk
• Widely used - ubiquitous exposure
- OPs = 50% of all insecticide use• Not an endocrine disruptor• Replaced organochlorines• Superfund Site Disposal Problem• OPs: nerve gases in warfare/terrorism
NerveTerminal
Receptors
SignalingCascades Nucleus
Gene Transcription
Replicate Differentiate Grow Die LearnAChE
Inhibition:CPF Oxon
Direct Actions on Cholinergic Receptors
Interaction withSignaling Intermediates
Transcription Factor
Expression, Function
Chlorpyrifos - Multiple Mechanisms Disrupt Neurodevelopment
Critical period in rats: late gestation to early neonatal stage[equivalent - 2nd trimester in human fetus]
Chlorpyrifos - Impact on Serotonin Systems = Miswiring
-20
-10
0
10
20
30
40
50
5HT1A 5HT2 5HTT 5HT1A 5HT2 5HTT 5HT1A 5HT2 5HTT 5HT1A 5HT2 5HTT 5HT1A 5HT2 5HTT
Chlorpyrifos Treatment on PN1-4 — 1 mg/kg
malefemale
cerebralcortex
hippocampus brainstem
ANOVA: Rx, p < 0.0001; Rx x sex, p < 0.0002; Rx x region, p < 0.0001;Rx x measure, p < 0.0003; Rx x region x measure, p < 0.0007
percent change from control
midbrainstriatum
Rx, p < 0.002Rx x sex,p < 0.0006
male: p < 0.0004female: NS
Rx, p < 0.0001Rx x measure,
p < 0.006
Rx x sex, p < 0.004Rx x measure,
p < 0.005
Rx x sex, p < 0.1Rx x measure,
p < 0.001
Rx x measure,p < 0.09
*
**
**
*
*
Male Female
Enhanced neuronalimpulse activity
(serotonin turnover)
Increases in serotonin receptors and transporter
BUT….
…Impaired Serotonergic Function
0
5
10
15
20
25
30
Male Female
Plus Maze: CPF (1 mg/kg)Decreases Anxiety in Males
ControlCPF
*
0
1
2
3
4
5
6
7
Male Female
Chocolate Milk Preference:CPF (1 mg/kg) Causes Anhedonia
ControlCPF
* *
aka: increased risk-taking, impulsive behavior
Chlorpyrifos - Miswiring of Acetylcholine Systems -Serotonin Replaces Acetylcholine for Hippocampal
Circuits and Behaviors
0
2
4
6
8
10
12
Control Chlorpyrifos
PN 1-4 Chlorpyrifos5HT2 Antagonist Drug Challenge0 mg/kg ketanserin0.5 mg/kg ketanserin1.0 mg/kg ketanserin2.0 mg/kg ketanserin
*
*
*
p < 0.0001
Terbutaline Use in Preterm Labor
• Stimulates BARs to inhibit uterine contraction
• Crosses the placenta to stimulate fetal BARs
• Effective for 48 hr max - NOT for maintenance use
• Animal studies from our lab, 1980s-1990s
altered neural cell differentiation
receptor and signaling shifts
permanent changes in responsiveness
• Hadders-Algra 1986 - impaired school performance
• Pitzer 2001 - psychiatric, learning disorders
Control
Terbutaline - 44% decrease in Purkinje cells
CerebellumThinning of cerebellar lobulesThinning of hippocampal CA3Reactive gliosisSomatosensory cortex - loss of
pyramidal cells
Critical Period Newborn Rat - PN2-5 =human 2nd trimester
• Neuroinflammation in cerebral cortex and cerebellum - microglial activation• Morphological changes almost identical to those in postmortem autism samples• Critical period PN2-5• Hyperreactive to novelty, aversive stimuli, sensory input
Decompensation of CVS responses similar to those
in autism
(compare to Ming 2005)
• Continuous terbutaline exposure for 2 weeks: RR=2.0• Male twins with no other affected siblings: RR=4.4
Further increase: BAR polymorphisms (16G, 27E) that prevent desensitization and therefore would enhance terbutaline effects
Terbutaline - Impact on Serotonin Systems = Miswiring ≈ Chlorpyrifos
Enhanced neuronalimpulse activity
(serotonin turnover)
Increases in serotonin receptors and transporter
CONCLUSIONS• Developmental neurotoxicants likely to play an important role in the increased incidence of childhood behavioral disorders including ASD
• Disparate mechanisms and effects converge on common final pathways
— different agents may produce similar outcomes
— different agents may produce additive/synergistic outcomes
• Lasting effects only when exposure occurs in critical periods
• Specific examples with relevance to ASD:
— organophosphate pesticides (ubiquitous exposure)
— terbutaline (use in preterm labor ≈10% US pregnancies)
Neurodevelopmental disorders - CAUSES, not a single ‘cause’
Origins of autism and ASD may not be so distinct from other neurodevelopmental disorders