Integrated Behavioral, Genetic and Brain Circuit Visualization Methods to Unravel Functional Anatomy of Zebrafish Amygdala

Abstract

The mammalian amygdala is a complex forebrain structure consisting of a heterogeneous group of nuclei derived from the pallial and subpallial telencephalon. It plays a critical role in a broad range of behaviors such as emotion, cognition, and social behavior; within the amygdala each nucleus has a distinct role in these behavioral processes. Topological, hodological, molecular, and functional studies suggest the presence of an amygdala-like structure in the zebrafish brain. It has been suggested that the pallial amygdala homolog corresponds to the medial zone of the dorsal telencephalon (Dm) and the subpallial amygdala homolog corresponds to the nuclei in the ventral telencephalon located close to and topographically basal to Dm. However, these brain regions are broad and understanding the functional anatomy of the zebrafish amygdala requires investigating the role of specific populations of neurons in brain function and behavior. In zebrafish, the highly efficient Tol2 transposon-mediated transgenesis method together with the targeted gene expression by the Gal4-UAS system has been a powerful tool in labeling, visualizing, and manipulating the function of specific cell types in the brain. The transgenic resource combined with neuronal activity imaging, optogenetics, pharmacology, and quantitative behavioral analyses enables functional analyses of neuronal circuits. Here, we review earlier studies focused on teleost amygdala anatomy and function and discuss how the transgenic resource and tools can help unravel the functional anatomy of the zebrafish amygdala.

Keywords: Gal4-UAS system; amygdala; dorsomedial telencephalon; emotion; pallium; subpallium; transgenic; zebrafish.

Figure 1

Teleost amygdala. (A) The development of telencephalon from neural tube (top) comparing evagination (in mammals) vs. eversion (in teleosts). In mammals, the lateral pallium (LP), putative amygdala region, lies laterally, whereas in teleosts, in a simple eversion, the corresponding structure lies medially. (B) The main neural connectivity of the teleost amygdala complex. Teleost Dm receives sensory information (somatosensory, visual, auditory, lateral line, gustatory, and odor) through thalamus and preglomerular complex (coronal view/top panel). Dm has extensive projections (sagittal view/bottom panel) to the dorsal nucleus (Vd), suppracommissural nucelus (Vs), and postcommissural nucelus (Vp) of the ventral telencephalon. These nuclei project efferents to several hypothalamic nuclei and the periaqueductal gray (PAG). Olfactory projections to Vd, Vs, and Vi are not showed. The dorsal most nucleus (Vdd) is dorsal most part of the ventral telencephalon as defined by Porter and Mueller (2020). ATN, anterior tuberal nucleus; Dl, dorsolateral telencephalon; EN, entopeduncular nucleus; Hd, dorsal zone of periventricular hypothalamus; LH, lateral hypothalamus; OB, olfactory bulb; Ppa, preoptic area; Vi, the intermediate nucleus of the ventral telencephalon (adapted from Nieuwenhuys and Meek, ; Folgueira et al., ; Lal et al., , and Northcutt, 2006).

Figure 2

Genetic dissection of the zebrafish amygdala. (A) Schematic of dorsal and coronal view of the zebrafish brain and telencephalon, respectively. The red line in the dorsal view shows the position of coronal section. (B) Examples of isolated brains of gene trap and enhancer trap transgenic zebrafish lines that show Gal4FF (UAS:GFP) expression in the Dm region. (B1–B8) Dorsal view of the isolated brain. Scale bar, 500 μm. (B1'–B8') Coronal section of the corresponding transgenic fish in (B1–B8). The position and region of the coronal section are showed in A (red line and red box). Scale bar, 200 μm. (C) The Gal4FF (UAS:GFP) expression pattern in the brain of transgenic fish SAGFF120A. SAGFF120A is an emx3-enhancer trap line and labels a population of neurons in the Dm, named 120A-Dm-neurons. (C1) Dorsal view. Scale bar, 500 μm. (C1') Coronal view as above. Scale bar, 200 μm. (D) Gal4FF drivers lines are crossed with botulinum toxin effector fish (UAS:zBoTxBLC:GFP). The double transgenic fish is analyzed in an active avoidance fear conditioning assay. (E) 120A-Dm-neurons have efferent projections to Vd, Vs, EN, Ppa, and hypothalamus. CCe, corpus cerebelli; Dl, dorsolateral telencephalon; EN, entopeduncular nucleus; OB, olfactory bulb; Ppa, preoptic area; TeO, optic tectum; Vd, the dorsal zone of the ventral telencephalon; Vv, the ventral zone of the ventral telencephalon [figures reused and adapted from Lal et al. (2018), https://creativecommons.org/licenses/by/4.0/].