Question 1.1

Please comment on the extent to which the report adequately summarizes the current state of knowledge. Does the scientific evidence support the conclusion that perturbation of the cholinergic nervous system during development by inhibiting AChE can potentially lead to deficits in the structure and function of the central and peripheral nervous systems?

Age-dependent sensitivity (i.e., young animals can exhibit higher levels of cholinesterase (ChE) inhibition at the same dose or inhibition at lower doses compared to adults) has been observed in several laboratory studies following treatment (acute and/or repeated oral gavage doses) of neonatal, juvenile, and adult rats with organophosphorus (OP) pesticides. The exact mechanisms of this age-dependent sensitivity are not known, but several studies have demonstrated that toxicokinetic factors may be responsible. Most notably, the more limited ability of the young to detoxify OP pesticides by A-esterases and carboxylesterases appears to be an important factor underlying the increased sensitivity of the immature rat to ChE inhibition. There appears to be more rapid recovery of inhibited AChE (synthesis of new ChE enzyme) in postnatal (and fetal) rat tissues, but information on comparative recovery in children and human adults is lacking.

Question 2.1

Please comment on the extent to which the report adequately discussed and summarized the current understanding of age-dependent sensitivity to ChE inhibition, the prevailing views in the scientific community concerning the biological factors involved, and the role esterases may play as a major factor accounting for potential increased sensitivity of the immature rat.

Question 2.2

Please comment on the timing of administration (i.e., the developmental stage treated) and the differential found between adults and the young animal.Question 2.3 Please comment on the extent to which comparative ChE data on six OP pesticides (chlorpyrifos, diazinon, dimethoate, methamidophos, malathion, methyl parathion) may represent a reasonable subset of different structural and pharmacokinetic characteristics of the cumulative group of OP pesticides to define an upper bound on the differential sensitivity that may be expected at different life stages of the immature animal. As an example, there are no chemical-specific comparative cholinesterase data on azinphos-methyl (AZM), an important contributor of risk for the food pathway. Pesticide-specific comparative cholinesterase data on the other six pesticides from the OP class (including data on malathion, a member of the same chemical subgroup as AZM) show a limited range of differential sensitivities -- from one-fold (no increased sensitivity) up to three-fold -- between the young and adults. EPA regards these data on other OPs as providing sufficient evidence to assess the potential for AZM to show age-dependent sensitivity, and to reasonably predict the degree of potential difference in sensitivity between the young and adults. Given the results of the other OPs, EPA concludes that it is unlikely that AZM would exceed a magnitude of difference greater than approximately 3-fold following treatment of PND 11 through 21 pups versus adult animals.

Age dependent sensitivity to cholinesterase inhibition has been associated with the limited ability of the immature rat to detoxify OP pesticides by esterases. In rats, A-esterase activity increases from birth to reach adult levels around postnatal day 21. Fetal rats possess very little carboxylesterase activity with increasing activity as the postnatal rat matures, reaching adult values after puberty (50 days of age). Data showing increased sensitivity of the young animal to cholinesterase inhibition compared to adults has generally been derived from acute dosing of PND 7 or PND 11 pups, or repeated dosing of PND 11 to PND 21 pups. The available data also show as the young rat rapidly matures in its ability to detoxify by esterases, the differential in cholinesterase inhibition becomes smaller. Thus, the relative sensitivities of immature rats found in the studies of dosing pups through PND 11 to 21 are smaller compared to studies of dosing a PND 11 pup. The dosing studies of PND 11 through 21 pups are considered to better approximate the maturation profile of the A-esterases of the highly exposed children's age group in the OP cumulative risk assessment, the one and two year olds, compared to a study of a PND 11 pup which is similar to a newborn. Thus, the repeated rat dosing studies more closely mimic the maturation or developmental profile of A-esterase appearance in children around the one and two year olds where children are reaching adult levels of A-esterase activity. The use of dosing studies of PND 11 through 21 is consistent with the exposure patterns of children. Humans generally do not begin to consume fresh (uncooked) fruits and vegetables until after six months of age or more. Furthermore, repeated dosing studies were used to determine relative sensitivity because people are exposed every day to an OP pesticide through food, and thus an animal study using repeat exposures is considered appropriate. Finally, following exposure to an OP, regeneration of cholinesterase to pre-exposure levels does not occur for days or weeks, making the exposed individual potentially more vulnerable to subsequent exposures during that period.

Question 3.1

Please comment on the maturation profile of A-esterase and the uncertainties surrounding these data in young children. Because no human data are available on the maturation profile of carboxylesterases, please comment on what should be assumed in humans ,especially children age 1 to 2 years, given the animal data and what science understands in general about detoxification maturation profiles.

Question 3.2

Please comment on the extent to which the biological understanding of observed age-dependent sensitivity to cholinesterase inhibition in laboratory animal studies informs our understanding about the likelihood of similar effects occurring in children; in particular, what can be inferred from animal and human information regarding the potential for different age groups to show increased sensitivity if exposed to cholinesterase-inhibiting pesticides. Does the scientific evidence support the conclusion that infants and children are potentially more sensitive to organophosphorus cholinesterase inhibitors?

Question 3.3

Please comment on the conclusions regarding the faster recovery in the young animal of AChE activity. Because there is no human information on the recovery of AChE in children compared to adults, please comment on the extent to which recovery of AChE in children should be factored into conclusions regarding potential risk to children.