Introduction: Investigation of Age-Dependent 

Pharmacodynamic Sensitivity to Pyrethroids

Larry Sheets, Bayer CropScience

on behalf of

The Council for the Advancement of Pyrethroid 

Human Risk Assessment (CAPHRA) 
Hypothesis 1: Age-related Sensitivity to 

Pyrethroids in Rats is Limited to High Doses 

• In vivo PD: Assess acute neurotoxic sensitivity in young & 

adult rats at defined internal dose levels

• Acoustic startle response

 Neurologic (reflex-based)

 ↑ Type I and ↓ Type II

 Suited to test young & adult rats

 Sensitive: Low doses PYR in adults

 Less known in young rats

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Deltamethrin effect on ASR in PND 21 and 

Adult LE Rats (1 mL/kg corn oil)

• Similar dose-related decrease in 

ASR at 2 & 4 mg/kg

• PND 21: Acute neurotoxic signs 

at 4 mg/kg; 6 mg/kg was too 

toxic

• Differential sensitivity led us to 

examine toxicity at higher doses

Sheets et al., 1994

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Deltamethrin Lethality Assessment in 

PND 11, 21 and Adult LE Rats (1 mL/kg)*

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*Sheets et al., 1994

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Deltamethrin: Comparable Brain Concentrations in 

PND 21 and Adult LE Rats at Given Effect Level*

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*Sheets et al., 1994

Same effect at similar 

brain conc. at 7x lower 

dose

~2x higher brain conc. 

at same dose & effect

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Model Type I and II Pyrethroids: 

Acoustic Startle Response*

Pyrethroid Doses (mg/kg) Adult PD21 Comment

Cismethrin 0, 3, 6, 12 +300% -24% (ns)

Effect at top dose 

(unless otherwise 

stated)

Permethrin 0, 30, 60, 120 +300% +65% (ns)

(mid-dose)

Deltamethrin 0, 1, 2, 4 -50%* -59%*

Cypermethrin 0, 9, 19, 38 -50%* -55%*

LE hooded rats; 2-hr after gavage dose (corn oil; 1 mL/kg)

Type I: Uncertain effect in PND 21 vs. adult (ns = not statistically significant);

• Did not determine this was the optimal time to test pups; 

• Different strain, dose volume and lab / test equipment in CAPHRA studies

Type II: Sensitivity (↓ASR) comparable in adult & PD21;

• Clinical signs in pups at much lower dose than adults

* Sheets (2000): NeuroToxicology 21 (1-2): 57-64 (Research at EPA ORD).

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Model Type I and II Pyrethroids: 

Acute Oral LD50

Sheets et al., 1994 Adult PND 21 PND 11

I: Cismethrin 55 32 119

I: Permethrin 406 201 (2x) 254 (<2x)

II: Deltamethrin 81 11 (8x) 5 (16x)

II: Cypermethrin 439 73 (6x) 18 (24x)

Cantalamessa, 1993 Adult PND 21 PND 8

I: Permethrin 1500 471 (3x) 340 (4x)

II: Cypermethrin 250 49 (5x) 15 (17x)

LD50 (mg/kg), based on 6 or 7 male rats per dose group at each age; Magnitude of 

the difference from adult shown in parentheses.

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Background: Sensitivity of Juvenile Rats

• Sensitivity of young rats to Type II pyrethroids raised concern 

for sensitivity of children

– Only evident with Type II PYR, at high dose levels & dose rates (e.g., 

not at 1-2 mg/kg DLM)

– Decreased rapidly with age: PND 11 21 Adult rats

– Deltamethrin: Evidence sensitivity associated with limited metabolic 

capacity of young rats (kinetics) vs. tissue sensitivity (dynamics)

• However: Limited data are available to evaluate

– ASR: Deltamethrin, Cypermethrin, Permethrin & Cismethrin

– Target-tissue levels (lethality & ASR) limited to Deltamethrin

– Some PYR with residential uses (potential exposures) have not been 

evaluated

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Pharmacodynamics – Targeted Testing 

Based on the Adverse Outcome Pathway

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Why PND 15 to Model Sensitivity of 1-3 Year 

Old Children?

• No ideal age: Depends on system, region and “process” of 

interest

• Neurobehavioral measures: Limited selection suited for rats 

<PND 12-15 (sensory and motor function)

– ASR: Respond to acoustic stimulus by PND 12 but motor 

function (coordination) is limited until ~PND 15;

– Motor activity: PND 13 low level of activity / highly variable; 

• Brain morphology: General agreement “brain” development 

too immature at PND 10 and maybe too advanced at PND 21

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Presentations

In vivo measures to Assess Age-Related Sensitivity: The Acoustic 

Startle Response & Detailed Clinical Observations

– Charles Vorhees; Cincinnati Children’s Research Foundation, 

University of Cincinnati

Evaluation of Age-Related Pharmacodynamics using Isolated 

Human (cloned) VGSC

– Steve Symington; Salve Regina University

Targeted Evaluation of Age-Related Pharmacodynamics using 

Mammalian CNS Neurolemma Preparations

– John Clark; University of Massachusetts

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In Vivo Rat: Acoustic Startle Response and 

Detailed Clinical Observations

AOP : In vivo relevance but least specific

– Organism: Integrated response

– Brain, spinal cord + PNS; PK + PD; various ion channels

Working hypothesis

– If there is no age-related PD difference in sensitivity in rats, we would 

expect to see comparable neurologic effects at the same internal dose

(PYR concentration in the brain).

Results: Support hypothesis & previous results

– Deltamethrin: NOAEL ~1 mg/kg in PND 15 and adult; Higher brain tissue 

levels on PND 15 vs. adult at a given effect level;

– Permethrin: PND 15 only modest effect at high doses (produces tremor in 

PND 15 and adult rats) – NO difference in dose associated with tremor

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Isolated Human VGSCs

• Model: Mammalian cells transfected with TTX-sensitive human VGSC’s coexpressed with β1 and β 2 subunits (Nav 1.1, 1.2, 1.3 and 1.6)

• Working Hypothesis: If there is no age-dependent PD sensitivity, VGSCs 

expressed in children will not be more sensitive than VGSCs in adults.

• Results (7 core actives)

– Confirm reports that hVGSCs are less sensitive than rat VGSCs & the response 

of Nav 1.1 vs. 1.3 indicates children would be less sensitive than adults

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Cheah et al., 2013 (human brain)

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Rat Neurolemma

Model 

• Effect of PYR on TTX-sensitive VGSCs in brain tissue from PND 15 & 

adult rats incorporated into the plasma membrane of Xenopus oocytes

Working Hypothesis

• If there is no age-dependent PD sensitivity, VGSCs from PND 15 rat 

neurolemma will not express greater sensitivity than from adults

Results (deltamethrin & permethrin)

• The VGSCs expressed in PND 15 rats are not more sensitive to 

deltamethrin or permethrin than those expressed in adult rats

Assessment

• This system lends itself well to evaluate age-dependent sensitivity of 

VGSCs at defined stages of development

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Pharmacodynamics - Conclusions

• In Vivo Rat (DLM and PRM)

– At the common NOAEL for neurotoxicity (1 mg/kg) the predicted brain 

Cmax levels of deltamethrin is ~4x higher in PND 15 than in adults. 

– The results indicate PD sensitivity is less in PND 15 rats than in adults. 

• Isolated human ion channel (7 core actives)

– The channel most prevalent in juveniles aged 1-2 years (and most likely, 

adults) is less sensitive than the isoform more prevalent at birth.

– The evidence argues against any PD difference likely to favor increased 

sensitivity of the young (older than ~6 months). 

– These results also confirm reports that human VGSS’s are less sensitive 

than rat channels to pyrethroids.

• Rat neurolemma studies (DLM and PRM)

– No age-related difference in sensitivity evident at a wide range of 

concentrations. 

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