Distinct Neuronal Populations in the Zebrafish Nucleus Incertus Exhibit Divergent Connectivity and Functional Properties

Core Concepts

The nucleus incertus (NI) in the zebrafish hindbrain contains genetically distinct neuronal populations, gsc2 and rln3a, that exhibit divergent efferent and afferent connectivity, spontaneous activity, and functional roles in regulating locomotor behavior.

Abstract

The study investigates the properties of two genetically defined neuronal populations, gsc2 and rln3a, within the zebrafish nucleus incertus (NI), which is the proposed analogue of the mammalian NI.
Key findings:

The gsc2 and rln3a neurons are distinct populations located in the NI, with gsc2 neurons anterior to the rln3a neurons.
The gsc2 and rln3a neurons differ in their efferent connectivity patterns. gsc2 neurons project widely to the cerebellum, interpeduncular nucleus (IPN), raphe, diencephalon, and hypothalamus, while rln3a neurons primarily innervate the IPN, medulla, diencephalon, and optic tectum.
The gsc2 neurons, but not the rln3a neurons, show increased calcium activity in response to optogenetic activation of the dorsal habenula-IPN pathway, indicating differential regulation by this circuit.
The gsc2 and rln3a neurons exhibit distinct patterns of spontaneous activity, with gsc2 neurons showing a robust increase in calcium signaling in response to an aversive electric shock, while rln3a neurons display more spontaneous fluctuations in activity.
Selective ablation of rln3a neurons in the NI, but not gsc2 neurons, increases baseline locomotor activity in larval zebrafish, suggesting a role for rln3a neurons in regulating locomotion.

Overall, the study demonstrates that genetically defined neuronal subpopulations within the NI possess divergent connectivity and functional properties, providing new insights into the diversity and roles of NI neurons.

Stats

The gsc2 neuronal population consists of approximately 48.33 ± 2.33 neurons.
The rln3a neuronal population consists of approximately 10.67 ± 1.33 neurons.
An average of 82.43 ± 3.52% of gsc2 neurons and 80.57 ± 5.57% of rln3a neurons are GABAergic.

Quotes

"gsc2 neurons are a distinct population, located anterior to the rln3a neurons."
"Efferents from gsc2 neurons were far more extensive than those of rln3a NI neurons, and were observed in regions not innervated by any rln3a neurons (e.g., cerebellum and caudal hypothalamus)."
"ReaChR activation in the dHb increased calcium transients in gsc2 NI neurons, but similar levels of calcium signaling were detected in the rln3a NI neurons of ReaChR-expressing larvae and negative controls."

Key Insights Distilled From

Genetically defined nucleus incertus neurons differ in connectivity and function

by Spikol,E. D.... at www.biorxiv.org 04-10-2022

https://www.biorxiv.org/content/10.1101/2022.04.07.487414v3

Deeper Inquiries

How do the distinct projection patterns and functional properties of gsc2 and rln3a neurons contribute to the overall role of the nucleus incertus in regulating stress responses, arousal, and memory

The distinct projection patterns and functional properties of gsc2 and rln3a neurons play crucial roles in the overall function of the nucleus incertus (NI) in regulating stress responses, arousal, and memory. The gsc2 neurons, with their extensive projections to regions such as the cerebellum, IPN, raphe, diencephalon, and hypothalamus, are likely involved in coordinating locomotor activity and responses to aversive stimuli. Their activation by electric shock and subsequent increase in calcium signaling suggest a role in immediate behavioral responses to stressors. On the other hand, the rln3a neurons, with projections to the medulla, IPN, diencephalon, lateral hypothalamus, and optic tectum, may be more involved in modulating arousal and memory processes. The spontaneous fluctuations in calcium signaling in rln3a neurons and their response to shock, characterized by shorter duration compared to gsc2 neurons, indicate a role in ongoing neural activity and possibly in encoding and processing aversive stimuli. Together, the heterogeneous properties of gsc2 and rln3a neurons likely contribute to the multifaceted functions of the NI in integrating stress responses, arousal, and memory processes.

What are the potential developmental or genetic mechanisms that give rise to the heterogeneity of neuronal subtypes within the nucleus incertus

The heterogeneity of neuronal subtypes within the nucleus incertus (NI) may arise from a combination of developmental and genetic mechanisms. During embryonic development, differential expression of transcription factors and signaling molecules could lead to the specification of distinct neuronal subpopulations within the NI. For example, the expression of genes like goosecoid homeobox 2 (gsc2) and relaxin-3a (rln3a) may define specific neuronal clusters with unique properties. Additionally, epigenetic modifications and interactions with neighboring brain regions could further refine the identity and connectivity of these subtypes. Genetic factors, including the expression of specific neuropeptides and neurotransmitters, may also contribute to the diversity of NI neurons. The interplay of these developmental and genetic mechanisms likely shapes the heterogeneous nature of neuronal subtypes within the NI.

Could the differential regulation of gsc2 and rln3a neurons by the dorsal habenula-interpeduncular nucleus pathway be linked to their proposed roles in modulating anxiety-like behaviors and responses to aversive stimuli

The differential regulation of gsc2 and rln3a neurons by the dorsal habenula-interpeduncular nucleus (dHb-IPN) pathway could be closely linked to their proposed roles in modulating anxiety-like behaviors and responses to aversive stimuli. The activation of gsc2 neurons by optogenetic stimulation of dHb neurons, leading to increased calcium signaling, suggests that these neurons are part of a circuit involved in processing aversive cues and coordinating stress responses. In contrast, the lack of significant response in rln3a neurons to dHb activation indicates a more specialized role, possibly in modulating arousal or memory processes rather than immediate stress responses. This differential regulation by the dHb-IPN pathway highlights the specificity of neural circuits in modulating different aspects of behavior and suggests a complex interplay between the NI subpopulations and upstream brain regions in regulating anxiety-related behaviors and responses to aversive stimuli.