Influence of litter size on the postnatal growth of rat pups: is there a rationale for litter-size standardization in toxicity studies?

The standardization of rodent litter sizes (or culling) in reproductive toxicity studies has become a common practice because it is believed that litter size affects pup postnatal growth and development. A claim has also been made that culling on postnatal day 4 or earlier reduces litter-size-induced variability in the pup growth thereby making statistical detection of toxic effects more sensitive. Although these statements remain controversial, culling has been either recommended or considered optional in current proposals for new OECD guidelines for reproductive/developmental toxicity testing. This study was undertaken to provide data useful for a discussion on the need for culling in reproductive toxicity testing. Along this line, we investigated the effects of rat litter size on maternal body weight gain during pregnancy and lactation as well as on pup growth, somatic maturation and survival to weaning (PND 21). Two-hundred-forty-one untreated pregnant rats (Wistar, Bor:spf, TOR) delivered litters with 1-13 pups, being 10 the most frequent (24%) litter size. Pup mortality was 5.4%, 3.0% and 0.4% in the first, second and third postnatal weeks, respectively. Maternal and pup body weight gains, and the day of appearance of milestones of somatic maturation (fur development, incisor eruption and eye opening) were examined in a subset of litters/mothers (N=180) in which all pups survived to PND 21. An inverse relationship between litter size and pup body weight was found on day of birth (PND 1) and thereafter until PND 21. Delays in the attainment of maturational milestones as litter size increases were also found. These observations indicated that the rate of growth and development of rat pups during suckling period is dependent on the litter size. The consequences of litter-size standardization in toxicity studies are discussed in the light of these findings. Alternative procedures to attenuating litter-size-induced variability in pup growth or its impact on toxicity data evaluation are presented as well.