1:名古屋大生物機能開発利用セ / Bioscience and Biotechnology Center, Nagoya University, Nagoya, Japan 2:名古屋大院理生命理学 / Grad. Sch. Science, Nagoya Univ. Nagoya, Japan 3:広島大院生物圏科学 / Graduate School of Biosphere Science, Hiroshima University, Higashihiroshima, Japan 4:国立遺伝研 / Natl Inst Genetics, Mishima, Japan
The cerebellum functions as a neuronal learning machine to control various motor behaviors. Recently, the cerebellum was also shown to be implicated in emotional learning, particularly in fear-related processes in both mammals and teleosts. However little is known about cerebellar contribution to the fear-related processes in teleosts. To address this issue, we adopted a delayed conditioning paradigm with the light extinguishment as the conditioned stimulus (CS) and an electric shock as the unconditioned stimulus (US) in zebrafish. Heart bearts were measured by a photocardiography technique. The US induced cardiac deceleration (bradycardia). After paired-associate conditioning with the CS and US, zebrafish around 20 days post-fertilization could acquire the conditioned bradycardia response. To investigate the involvement of the cerebellum in the classical fear conditioning in zebrafish, we performed the inhibition of granule cell transmission. To this end, we expressed the botulinum toxin using the gSA2AzGFF152B, which expresses a Gal4 in the corpus cerebelli. There was no apparent difference in swimming behavior between the granule-cells-silenced larvae and their siblings. In the control larvae, the CS-dependent bradycardia immediately recovered after the conditioning. The granule-cells-silenced larvae were capable to acquire conditioned bradycardia, but displayed a prolonged bradycardia response. We examined neuronal activities during the classical fear conditioning by expressing a Ca2+ indicator GCaMP7a using a pan-neuronal Gal4-driver line Tg(elavl3:GAL4-VP16). We observed CS-dependent upregulation of the neuronal activities in the corpus cerebelli: some of these neurons quickly responded to the CS, whereas other neurons were activated after the CS presentation period. Our findings suggest that the cerebellum is involved in the fear conditioning and the granule cells control the recovery from conditioned fear responses in zebrafish.