A method to biotinylate and histochemically visualize ibotenic acid for pharmacological inactivation studies

Ibotenic acid (IA) and kainic acid (KA) are commonly used tools to selectively inactivate neuronal perikarya, eventually leading to their degeneration, without affecting fibers of passage. Reversible inactivations and experimental paradigms that do not allow for long survival times, however, do not permit for histological verification of the site and extent of the lesion by identifying the area showing gliosis. We describe here a method in which IA and KA were conjugated with biotin and thus could be easily visualized histochemically. We pressure-injected biotinylated IA and KA into various hindbrain areas of the electrosensory system in electric fish while monitoring neuronal responses at the injection site and assessing effects on the behavior. Whereas the effects of biotinylated IA did not differ from those of the unbiotinylated form, biotinylated KA lost its physiological activity. Thus, only biotinylated IA could be used successfully. The size of the gliosis seen after a survival time of seven days was similar to the extent of biotin label observed after injection of comparable volumes of biotinylated IA. Moreover, this method resulted in labeling of individual neurons presumably affected by IA and yielded information about their projection patterns which was comparable to labeling seen after intracellular injections of neurobiotin or biocytin.