The Panamanian golden toad's skin contains a potent water-soluble toxin called zetekitoxin, which affects nerve cells and protects the toad from most predators. However, if minor precautions are taken, these toads can be safely handled.

Aspects of its natural history are not fully investigated. Females are larger than males, measuring about 53 mm (2 inches) to the males' 45 mm (1.8 inches). The toad is terrestrial and diurnal (active during the day). Males maintain small territories along forest streams. When males encounter one another, they sometimes engage in waving of their forefeet as a signal of aggression or defense. Males also communicate acoustically, emitting a very soft and trilled mating call. Eggs are presumably laid in strings attached to rocks at the bottom of fast-flowing streams, where the tadpoles develop and grow. The tadpoles have a ventral sucker that may be used for remaining attached to rocks or other substrata in the streams. Panamanian golden toads and other species of the genus Atelopus are members of the family Bufonidae, as are most toads. Some authorities, however, classify the genera Atelopus and Bradycephalus as a separate family called the stub-footed toads (Atelopodidae).