The avian frightening agent, 4-aminopyridine (4AP), was tested on five avian and two fish species to assess the effects of long-term 4AP exposure to nontarget vertebrates. Adult bobwhites (Colinus virginianus), mourning doves (Zenaida macroura), and ring-necked pheasants (Phasianus colchicus) were fed diets containing 3 percent 4AP-treated cracked corn diluted with various ratios of untreated corn for 7 to 35 days. Minimum dilution ratios that produced more than 50 percent mortality were 1:29 for bobwhites and 1:99 for doves. Mortality caused by 4AP could not be determined for pheasants because of their inability to subsist only on corn during the test periods. LCso's were determined for doves (316 ppm) and coturnix quail (Coturnix coturnix) (479 ppm) fed exclusively on uniformly treated laboratory rations for 28 to 40 days. Starlings (Sturnus vulgaris) treated by gavage daily for 25 days with 1.78 mg/kg 4AP showed no intoxication symptoms. Mortality in all avian species appeared to be the result of acute poisoning, and no evidence of cumulative toxicity was noted in species fed uniformly-treated rations. In 96-hour static tests, channel catfish (Ictalurus punctatus) and bluegill (Lepomis macrochirus) showed increasing sensitivity to 4AP with time in test waters ranging from very soft to very hard and from 12 to 22 C. The 96-hour LC₅₀'s for both species ranged from 2.43 to 7.56 mg/liter (ppm). Toxicity appeared to decrease with increasing water temperatures but was generally unaffected by water hardness. On the basis of the data presented, and the currently registered application rate for 4AP in agricultural crops (0.34 g/ha), long-term hazards to nontarget wildlife were judged to be minimal.