A previous study from this laboratory showed that sprouting of serotoninergic axons in the hamster's superior colliculus (SC) induced by a single subcutaneous injection of 5,7-dihydroxytryptamine (5,7-DHT) at birth (postnatal day 0; P-0) resulted in an abnormal terminal distribution of the uncrossed retinotectal projection. The present study provided further evidence to support the role of increased 5-HT levels within the SC in this phenomenon. Slow-release polymer (ELVAX) chips impregnated with serotonin (5-HT) were placed over the SC on either P-1 or P-3, and retinotectal projections were assessed via anterograde transport of horseradish peroxidase when animals reached P ΟΎ 18. Analysis of ELVAX chips indicated that they released 5-HT in amounts of Υ1 pmole/hour for at least 12 days. Assessment of the SC of treated hamsters indicated significantly elevated 5-HT concentrations as late as P-12, but not on P-16. Implantation of 5-HT chips, but not control chips, resulted in abnormalities in the uncrossed retinotectal projection similar to those observed in the 5,7-DHT-treated animals. The patches that normally develop in the rostral part of the stratum opticum were not present, and uncrossed axons were distributed densely in this layer and in the lower portion of the stratum griseum superficiale throughout the rostrocaudal and mediolateral extents of the SC. Quantitative analysis of these changes indicated significant differences between the organization of the uncrossed retinotectal projections of 5-HT-treated animals vs. either blank-implant treated or completely untreated animals but not between 5-HT-treated hamsters and animals that received neonatal 5,7-DHT injections. All of these results support the conclusion that increased SC concentrations of 5-HT altered retinotectal development.