High-throughput sequencing of macaque basolateral amygdala projections reveals dissociable connectional motifs with frontal cortex

Abstract

The basolateral amygdala (BLA) projects widely across the macaque frontal cortex, and amygdalo-frontal projections are critical for optimal emotional responding and decision-making. Yet, little is known about the single-neuron architecture of these projections: namely, whether single BLA neurons project to multiple parts of the frontal cortex. Here, we use MAPseq to determine the projection patterns of over 3000 macaque BLA neurons. We found that one-third of BLA neurons have two or more distinct targets in parts of frontal cortex and of subcortical structures. Further, we reveal non-random structure within these branching patterns such that neurons with four targets are more frequently observed than those with two or three, indicative of widespread networks. Consequently, these multi-target single neurons form distinct networks within medial and ventral frontal cortex consistent with their known functions in regulating mood and decision-making. Additionally, we show that branching patterns of single neurons shape functional networks in the brain as assessed by fMRI-based functional connectivity. These results provide a neuroanatomical basis for the role of the BLA in coordinating brain-wide responses to valent stimuli and highlight the importance of high- resolution neuroanatomical data for understanding functional networks in the brain.

Brainwide mesoscale functional networks revealed by focal infrared neural stimulation of the amygdala

“…That is, within each node are related neurons which share common targets. For example, viral barcode analysis of amygdala-prefrontal connectivity has shown that single amygdala BLA neurons can connect with one to several neurons in different parts of prefrontal cortex (Zeisler et al, 2023). This raises a potential scenario in which dynamics of mesoscale nodal selection is further coupled with intra-node single neuron selection to achieve a broad range of distinct and specific functional circuits.…”

“…Previous studies have shown that the connectivity of these nuclei is quite heterogeneous, appropriate for supporting a broad spectrum of behavioral axes (Klein-Flügge et al, 2022; Scangos et al, 2021). Not only does each nucleus have distinctive cell types and cell densities (Yu et al, 2023), even single neurons within a subdivision of the amygdala exhibit distinct targeting patterns, projecting to either 1, 2, 3, or 4 cortical sites (Zeisler et al, 2023). Electrical stimulation of nuclear-specific targets in the human amygdala combined with neuroimaging revealed the existence of multiple distinct networks and a reproducible temporal pattern of activation (Sawada et al, 2022).…”

Massive Multiplexing of Spatially Resolved Single Neuron Projections with Axonal BARseq

“…However, in previous work (Huang et al, 2020) we have barcoded more than 100,000 neurons in a single brain, and there is no technical barrier to labeling comparable numbers of neurons for axonal BARseq in future studies. Furthermore, Sindbis virus can infect diverse species including primates (Zeisler et al, 2023), so axonal BARseq could potentially be modified to map projections in many model systems, especially those in which conventional tracing-based approaches are impractical. Axonal BARseq thus has the potential to emerge as a powerful tool for massively multiplexed mapping of single neuron projections in diverse model systems.…”

“…However, because spatial resolution in MAPseq is limited by the dissection of brain areas prior to bulk sequencing, axonal projection patterns in this MAPseq/BARseq combined approach can only be crudely resolved. MAPseq and BARseq have been repeatedly validated using multiple methods in a wide range of brain areas (Zeisler et al 2023; Y. Chen et al 2022; Hausmann et al 2022; Tsoi et al 2022; Webb et al 2022; Klingler et al 2021; Mathis et al 2021; Muñoz-Castañeda et al 2021; Sun et al 2021; Gergues et al 2020; Huang et al 2020; X.…”

“…However, because spatial resolution in MAPseq is limited by the dissection of brain areas prior to bulk sequencing, axonal projection patterns in this MAPseq/BARseq combined approach can only be crudely resolved. MAPseq and BARseq have been repeatedly validated using multiple methods in a wide range of brain areas (Kebschull et al, 2016a; Han et al, 2018; Chen et al, 2019, 2022; Gergues et al, 2020; Huang et al, 2020; Klingler et al, 2021; Mathis et al, 2021; Muñoz-Castañeda et al, 2021; Sun et al, 2021; Hausmann et al, 2022; Tsoi et al, 2022; Webb et al, 2022; Zeisler et al, 2023). In particular, we demonstrated that barcode transport is uniform over long distances (>10 mm; Kebschull et al, 2016a; Han et al, 2018; Chen et al, 2019, 2022; Gergues et al, 2020; Huang et al, 2020; Klingler et al, 2021; Mathis et al, 2021; Muñoz-Castañeda et al, 2021; Sun et al, 2021; Hausmann et al, 2022; Tsoi et al, 2022; Webb et al, 2022; Zeisler et al, 2023).…”