title: Imaging neuronal activity during zebrafish behavior with a genetically encoded calcium indicator.
abstract: Genetically encoded calcium indicators, such as cameleon, have offered the promise of noninvasively monitoring activity of neurons, but no one has demonstrated whether these indicators can report calcium transients in neurons of behaving vertebrates. We show that cameleon can be expressed at high levels in sensory and spinal cord neurons in zebrafish by using neural-specific promoters in both transient expression experiments and in a stable transgenic line. Using standard confocal microscopy, calcium transients in identified motoneurons and spinal interneurons could be detected during escape behaviors produced by a touch on the head of the fish. Small movements of the restrained fish during the behavior did not represent a major problem for analyzing the calcium responses because of the ratiometric nature of cameleon. We conclude that cameleon can be used to noninvasively study the activity of neurons in an intact, behaving vertebrate. The ability to introduce an indicator genetically allows for studies of the functional roles of local interneurons that cannot easily be monitored with other approaches. Transgenic lines such as the one we generated can also be crossed into mutant lines of fish to study both structural and functional consequences of the mutations.
authors.affiliation: Department of Neurobiology and Behavior, State University of New York, Stony Brook, New York 11794-5230, USA.
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Higashijima
authors.forename: Mark A
authors.initials: MA
authors.lastname: Masino
authors.forename: Gail
authors.initials: G
authors.lastname: Mandel
authors.forename: Joseph R
authors.initials: JR
authors.lastname: Fetcho
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001522:Behavior, Animal / Q000502:physiology*; D020013:Calcium Signaling / Q000502:physiology*; D002135:Calcium-Binding Proteins / Q000096:biosynthesis; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D029721:Drosophila Proteins / Q000096:biosynthesis; D004558:Electric Stimulation; D004594:Electrophysiology; D004625:Embryo, Nonmammalian; D004924:Escape Reaction / Q000502:physiology; D007395:Interneurons / Q000502:physiology; D018613:Microscopy, Confocal; D009046:Motor Neurons / Q000502:physiology; D009474:Neurons / Q000502:physiology*; D018408:Patch-Clamp Techniques; D010957:Plasmids / Q000235:genetics; D015027:Zebrafish / Q000502:physiology*
chemical_list: D002135:Calcium-Binding Proteins; D018076:DNA, Complementary; D029721:Drosophila Proteins; C473067:cameleon 2.1 protein, Drosophila