TERATOLOGY 60:177–178 (1999)
Letter to the Editor†
More on Chlorpyrifos: Response to the Letter of
Hanley et al. ([1999] Teratology 59:323–324),
Concerning the Article by Roy et al. ([1998]
Teratology 58:62–68)‡
To the Editor:
We are extremely disappointed that you would pub-
lish the letter from Hanley et al. (’99), criticizing our
paper (Roy et al., ’98), without offering us a chance to
respond and to have our response published along with
the letter. In this particular case, Hanley et al. (’99)
chose to ignore explicit statements in our paper that
dealt with the specific question of dose or concentration,
and the applicability of in vitro models. It is worthwhile
to quote that paragraph from our paper:
An inherent limitation of any in vitro approach to
developmental neurotoxicity is the difficulty of ex-
trapolating the concentrations used to relevant doses
in intact animals. Although scant information is
available concerning the actual levels of chlorpyrifos
achieved in fetal brain, we have already demon-
strated that doses that cause only 20% cholinester-
ase inhibition nevertheless depress mitosis in neona-
tal rat brain in vivo (Whitney et al., ’95; Song et al.,
’97; Dam et al., ’98), leading to deficiencies in cell
number (Campbell et al., ’97). A preliminary report
in pregnant rats (Hunter et al., ’98) found that a
comparable degree of cholinesterase inhibition, which
is well below the threshold for any observable signs
of cholinergic hyperstimulation, produces peak fetal
brain concentrations of the major metabolite of
chlorpyrifos of approximately 0.25 µg/g, which on a
molar basis, corresponds to the lowest concentration
of chlorpyrifos used here. On a body weight basis, the
doses of chlorpyrifos needed for adult or developmen-
tal toxicity in rats range up to tens to hundreds of
mg/kg (Pope et al., ’91; Pope and Chakraborti, ’92;
Bushnell et al., ’93; Chakraborti et al., ’93) and
certainly no lower than 2 mg/kg (Whitney et al., ’95).
Mitotic arrest in vivo occurs with brain concentra-
tions of 2 µg/g (Whitney et al., ’95), well within the
concentration range used here (0.5–50 µg/ml). The
likely acute exposure level for infants after home
application of chlorpyrifos is also above this range:
350 µg/kg/day for a 2-week period, for a total of 5
mg/kg (Gurunathan et al., ’98). Although there are
Published 1999 WILEY-LISS, INC. ‡ This article is a
US Government work and, as such, is in the public domain in
the United States of America.
clear limitations of extrapolation across species and
between cultures and intact systems, in vitro evalua-
tions nevertheless can point the way to likely mecha-
nisms and adverse outcomes, and may even be
within the range of specific exposure levels in vivo.
Thus, we clearly indicate that the purpose of in vitro
models is to identify heretofore unsuspected mecha-
nisms by which chlorpyrifos may affect brain develop-
ment. In claiming, incorrectly, that a compound like
chlorpyrifos cannot affect brain development because it
is not dysmorphogenic, Hanley et al. (’99) discount the
entire field of behavioral teratology and ignore the
specific, nonenzymatic role of cholinesterase in develop-
mental events that are specific to the nervous system
(Koenigsberger et al., ’98; Brimijoin and Koenigsberger,
’99). In disparaging ‘‘high-dose’’ exposures, they neglect
the fact that other investigators estimate infant/toddler
exposure levels at much higher than those cited in their
letter (Fenske et al., ’90; Gurunathan et al., ’98). In the
real world, pesticide exposures do not always occur
under properly controlled circumstances; indeed, re-
cent studies in Mexico indicate that, within the same
village, children exposed to supposedly nontoxic levels
of pesticides, displaying neither growth impairment
nor any overt manifestations of systemic toxicity, never-
theless show behavioral and learning disabilities at a
much higher rate than do children of identical socioeco-
*Correspondence to: Theodore A. Slotkin, Department of Pharmacol-
ogy and Cancer Biology, Duke University Medical Center, Durham,
NC 27710.
Received 13 May 1999; Accepted 18 May 1999
†Editor’s Note: The journal has instituted a policy whereby authors
will be given an opportunity to respond to letters written about their
articles. We will publish the letter and the author(s) response in the
same issue. We will give titles to letters in order to facilitate
subsequent literature reviews. LBH
178 SLOTKIN AND ANDREWS
nomic status from homes without contamination (Guil-
lette et al., ’98). By discounting studies conducted with
doses higher than their models, Hanley et al. (’99) foster
an approach that, if adopted, would lead to the abandon-
ment of safety factors for uncertainty, interspecies
extrapolation, and developmental susceptibility.
Our basic findings (Roy et al., ’98) indicate that
chlorpyrifos has the ability to elicit adverse effects on
neural cells undergoing active replication and differen-
tiation, events that are not occurring in the mature
nervous system and that thus escape detection by
examination of adults. The in vitro findings are echoed
by similar results with otherwise nontoxic exposures to
chlorpyrifos in vivo (Whitney et al., ’95; Campbell et al.,
’97; Song et al., ’97; Dam et al., ’98; Johnson et al., ’98).
The main issue here is that the usual biomarker of
organophosphate toxicity, cholinesterase inhibition, may
be inadequate to the task of predicting this type of
developmental vulnerability. The separate issue of risk
extrapolation to humans, which cannot be addressed
simply with an in vitro model, depends first upon the
elaboration of the relevant endpoints of developmental
toxicity, which was the main point of our paper.
Theodore A. Slotkin*
Department of Pharmacology and Cancer Biology
Duke University Medical Center
Durham, North Carolina
James E. Andrews
Developmental Toxicology Division
U.S. Environmental Protection Agency
Research Triangle Park, North Carolina
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