Agouti-related protein (AgRP) is a hypothalamic regulator of food consumption in mammals. However, AgRP has also been detected in circulation, but a possible endocrine role has not been examined. Zebrafish possess two agrp genes: hypothalamically expressed agrp1, considered functionally equivalent to the single mammalian agrp, and agrp2, which is expressed in pre-optic neurons and uncharacterized pineal gland cells and whose function is not well understood. By ablation of AgRP1-expressing neurons and knockout of the agrp1 gene, we show that AgRP1 stimulates food consumption in the zebrafish larvae. Single-cell sequencing of pineal agrp2-expressing cells revealed molecular resemblance to retinal-pigment epithelium cells, and anatomic analysis shows that these cells secrete peptides, possibly into the cerebrospinal fluid. Additionally, based on AgRP2 peptide localization and gene knockout analysis, we demonstrate that pre-optic AgRP2 is a neuroendocrine regulator of the stress axis that reduces cortisol secretion. We therefore suggest that the ancestral role of AgRP was functionally partitioned in zebrafish by the two AgRPs, with AgRP1 centrally regulating food consumption and AgRP2 acting as a neuroendocrine factor regulating the stress axis.
The master circadian clock in fish has been considered to reside in the pineal gland. This dogma is challenged, however, by the finding that most zebrafish tissues contain molecular clocks that are directly reset by light. To further examine the role of the pineal gland oscillator in the zebrafish circadian system, we generated a transgenic line in which the molecular clock is selectively blocked in the melatonin-producing cells of the pineal gland by a dominant-negative strategy. As a result, clock-controlled rhythms of melatonin production in the adult pineal gland were disrupted. Moreover, transcriptome analysis revealed that the circadian expression pattern of the majority of clock-controlled genes in the adult pineal gland is abolished. Importantly, circadian rhythms of behavior in zebrafish larvae were affected: rhythms of place preference under constant darkness were eliminated, and rhythms of locomotor activity under constant dark and constant dim light conditions were markedly attenuated. On the other hand, global peripheral molecular oscillators, as measured in whole larvae, were unaffected in this model. In conclusion, characterization of this novel transgenic model provides evidence that the molecular clock in the melatonin-producing cells of the pineal gland plays a key role, possibly as part of a multiple pacemaker system, in modulating circadian rhythms of behavior.
The neuropeptide agouti-related protein (AgRP) is expressed in the arcuate nucleus of the mammalian hypothalamus and plays a key role in regulating food consumption and energy homeostasis. Fish express two agrp genes in the brain: agrp1, considered functionally homologous with the mammalian AgRP, and agrp2. The role of agrp2 and its relationship to agrp1 are not fully understood. Utilizing BAC transgenesis, we generated transgenic zebrafish in which agrp1- and agrp2-expressing cells can be visualized and manipulated. By characterizing these transgenic lines, we showed that agrp1-expressing neurons are located in the ventral periventricular hypothalamus (the equivalent of the mammalian arcuate nucleus), projecting throughout the hypothalamus and towards the preoptic area. The agrp2 gene was expressed in the pineal gland in a previously uncharacterized subgroup of cells. Additionally, agrp2 was expressed in a small group of neurons in the preoptic area that project directly towards the pituitary and form an interface with the pituitary vasculature, suggesting that preoptic AgRP2 neurons are hypophysiotropic. We showed that direct synaptic connection can exist between AgRP1 and AgRP2 neurons in the hypothalamus, suggesting communication and coordination between AgRP1 and AgRP2 neurons and, therefore, probably also between the processes they regulate.
Social affiliation emerges from individual-level behavioural rules that are driven by conspecific signals1–5. Long-distance attraction and short-distance repulsion, for example, are rules that jointly set a preferred interanimal distance in swarms6–8. However, little is known about their perceptual mechanisms and executive neural circuits3. Here we trace the neuronal response to self-like biological motion9,10, a visual trigger for affiliation in developing zebrafish2,11. Unbiased activity mapping and targeted volumetric two-photon calcium imaging revealed 21 activity hotspots distributed throughout the brain as well as clustered biological-motion-tuned neurons in a multimodal, socially activated nucleus of the dorsal thalamus. Individual dorsal thalamus neurons encode local acceleration of visual stimuli mimicking typical fish kinetics but are insensitive to global or continuous motion. Electron microscopic reconstruction of dorsal thalamus neurons revealed synaptic input from the optic tectum and projections into hypothalamic areas with conserved social function12–14. Ablation of the optic tectum or dorsal thalamus selectively disrupted social attraction without affecting short-distance repulsion. This tectothalamic pathway thus serves visual recognition of conspecifics, and dissociates neuronal control of attraction from repulsion during social affiliation, revealing a circuit underpinning collective behaviour.
Background: Single-cell RNA sequencing (scRNA-seq) has quickly become one of the most dominant techniques in modern transcriptome assessment. In particular, 10X Genomics Chromium system, with its high throughput approach, turn key and thorough user guide made this cutting-edge technique accessible to many laboratories using diverse animal models. However, standard downstream processing, including the alignment and cell filtering pipelines might not be ideal for every organism or tissue. Here we applied an alternative strategy, based on the pseudoaligner kallisto, on twenty-two publicly available single cell sequencing datasets from a wide range of tissues of eight organisms and compared the results with the standard 10X Genomics Cell Ranger pipeline. Results: In most of the tested samples, kallisto outperformed Cell Ranger in sequencing read alignment rates and total gene detection rates. Although datasets processed with Cell Ranger had higher cell counts, outside of human and mouse datasets, these additional cells were routinely of low quality, containing low gene detection rates. Thorough downstream analysis of one kallisto processed dataset, obtained from the zebrafish pineal gland, revealed clearer clustering, allowing the identification of an additional photoreceptor cell type that previously went undetected. The finding of the new cluster suggests that the photoreceptive pineal gland is essentially a bi-chromatic tissue containing both green and red cone-like photoreceptors and implies that the alignment and processing pipeline can affect the discovery of biologically-relevant cell types. Conclusion: While Cell Ranger favors higher cell numbers, using kallisto results in datasets with higher median gene detection per cell. We could demonstrate that cell type identification was not hampered by the lower cell count, but in fact improved as a result of the high gene detection rate and the more stringent filtering. It is thus beneficial to favor high quality cells and accept a lower cell count, leading to an improved classification of cell types.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.