The calcium-sensing receptor (CaSR) regulates zebrafish sensorimotor decision making via a genetically defined cluster of hindbrain neurons

Shoenhard, Hannah; Jain, Roshan A.; Granato, Michael

doi:10.1016/j.celrep.2022.111790

Cited by 4 publications

(11 citation statements)

References 91 publications

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“…CaSR p190/+ phenotypes were not accurately predicted by any of the three models. Data in panels A and B also appears in Shoenhard, Jain, and Granato [ 19 ].…”

Section: Expected Resultsmentioning

confidence: 88%

“…We collected larvae in rescued (above light blue line) and non-rescued (below pink line) groups. Some data from Panel B also appears in Shoenhard, Jain, and Granato [ 19 ].…”

Section: Expected Resultsmentioning

confidence: 95%

“…CaSR mutants, while capable of escapes, always bias towards the reorientation ( Fig 2A ). Variegated CaSR-EGFP expression in neurons, driven using the Gal4 x UAS system, is sufficient to rescue the reorientation bias of some, but not all, CaSR mutants ( Fig 2B ) [ 19 ]. Specifically, we generated larvae by crossing the αtubulin : Gal4 stable transgenic line [ 20 ] to the 14xUAS : CaSR-EGFP stable transgenic line [ 19 ].…”

Section: Expected Resultsmentioning

confidence: 99%

“…Variegated CaSR-EGFP expression in neurons, driven using the Gal4 x UAS system, is sufficient to rescue the reorientation bias of some, but not all, CaSR mutants ( Fig 2B ) [ 19 ]. Specifically, we generated larvae by crossing the αtubulin : Gal4 stable transgenic line [ 20 ] to the 14xUAS : CaSR-EGFP stable transgenic line [ 19 ]. For the loss-of-function strategy, we divided these CaSR mutant larvae into two groups: those that responded to high intensity stimuli with strong bias towards reorientations (loss-of-function phenotype), and those that responded to the same stimuli with strong bias towards escapes (rescue of loss-of-function phenotype) ( Fig 2B ).…”

Section: Expected Resultsmentioning

confidence: 99%

“…Scale bar = 100 um. C) CaSR overexpression escape-shifted phenotype is exhibited by GFP+ y234; UAS : CaSR-EGFP ; CaSR sibling ( CaSR +/+ and CaSR p190/+ ) larvae responding to a low-intensity acoustic stimulus [ 19 ]. D) Partial rescue of CaSR loss-of-function phenotype in sorted GFP+ y234; UAS : CaSR-EGFP ; CaSR p190/p190 larvae responding to a strong acoustic stimulus.…”

Section: Expected Resultsmentioning

confidence: 99%

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Multivariate analysis of variegated expression in Neurons: A strategy for unbiased localization of gene function to candidate brain regions in larval zebrafish

Shoenhard

Granato

2023

PLoS ONE

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Behavioral screens in model organisms have greatly facilitated the identification of genes and genetic pathways that regulate defined behaviors. Identifying the neural circuitry via which specific genes function to modify behavior remains a significant challenge in the field. Tissue- and cell type-specific knockout, knockdown, and rescue experiments serve this purpose, yet in zebrafish screening through dozens of candidate cell-type-specific and brain-region specific driver lines for their ability to rescue a mutant phenotype remains a bottleneck. Here we report on an alternative strategy that takes advantage of the variegation often present in Gal4-driven UAS lines to express a rescue construct in a neuronal tissue-specific and variegated manner. We developed and validated a computational pipeline that identifies specific brain regions where expression levels of the variegated rescue construct correlate with rescue of a mutant phenotype, indicating that gene expression levels in these regions may causally influence behavior. We termed this unbiased correlative approach Multivariate Analysis of Variegated Expression in Neurons (MAVEN). The MAVEN strategy advances the user’s capacity to quickly identify candidate brain regions where gene function may be relevant to a behavioral phenotype. This allows the user to skip or greatly reduce screening for rescue and proceed to experimental validation of candidate brain regions via genetically targeted approaches. MAVEN thus facilitates identification of brain regions in which specific genes function to regulate larval zebrafish behavior.

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“…CaSR p190/+ phenotypes were not accurately predicted by any of the three models. Data in panels A and B also appears in Shoenhard, Jain, and Granato [ 19 ].…”

Section: Expected Resultsmentioning

confidence: 88%

“…We collected larvae in rescued (above light blue line) and non-rescued (below pink line) groups. Some data from Panel B also appears in Shoenhard, Jain, and Granato [ 19 ].…”

Section: Expected Resultsmentioning

confidence: 95%

Section: Expected Resultsmentioning

confidence: 99%

Section: Expected Resultsmentioning

confidence: 99%

Section: Expected Resultsmentioning

confidence: 99%

See 3 more Smart Citations

Multivariate analysis of variegated expression in Neurons: A strategy for unbiased localization of gene function to candidate brain regions in larval zebrafish

Shoenhard

Granato

2023

PLoS ONE

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The Calmodulin-interacting peptide Pcp4a regulates feeding state-dependent behavioral choice in zebrafish

Zaupa,

Nagaraj,

Sylenko

et al. 2024

Neuron

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Silicon-Rhodamine Functionalized Evocalcet Probes Potently and Selectively Label Calcium Sensing Receptors In Vitro, In Vivo, and Ex Vivo

Bátora,

Fischer,

Kaderli

et al. 2024

ACS Pharmacol. Transl. Sci.

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The calcium sensing receptor (CaSR) is a ubiquitously expressed G-protein coupled receptor (GPCR) that regulates extracellular calcium signals via the parathyroid glands. CaSR has recently also been implicated in noncalcitropic pathophysiologies like asthma, gut inflammation, and cancer. To date, molecular tools that enable the bioimaging of CaSR in tissues are lacking. Based on in silico analyses of available structure− activity relationship data on CaSR ligands, we designed and prepared silicon-rhodamine (SiR) conjugates of the clinically approved drug evocalcet. The new probes EvoSiR4 and EvoSiR6, with differing linker lengths at the evocalcet carboxyl end, both showed a 6-fold and 3-fold increase in potency toward CaSR (EC 50 < 45 nM) compared to evocalcet and the evocalcet-linker conjugate, respectively, in an FLIPR-based cellular functional assay. The specificity of the EvoSiR probes toward CaSR binding and the impact of albumin was evaluated in live cell experiments. Both probes showed strong albumin binding, which facilitated the clearance of nonspecific binding interactions. Accordingly, in zebrafish embryos, EvoSiR4 specifically labeled the high CaSR expressing neuromasts of the lateral line in vivo. EvoSiR4 was also assessed in human parathyroid tissues ex vivo, showing a specific absolute CaSR-associated fluorescence compared to that of parathyroid autofluorescence. In summary, functionalization of evocalcet by SiR led to the preparation of potent and specific fluorescent CaSR probes. EvoSiR4 is a versatile small-molecular probe that can be employed in CaSR-related biomedical analyses where antibodies are not applicable.

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The calcium-sensing receptor (CaSR) regulates zebrafish sensorimotor decision making via a genetically defined cluster of hindbrain neurons

Cited by 4 publications

References 91 publications

Multivariate analysis of variegated expression in Neurons: A strategy for unbiased localization of gene function to candidate brain regions in larval zebrafish

Multivariate analysis of variegated expression in Neurons: A strategy for unbiased localization of gene function to candidate brain regions in larval zebrafish

The Calmodulin-interacting peptide Pcp4a regulates feeding state-dependent behavioral choice in zebrafish

Silicon-Rhodamine Functionalized Evocalcet Probes Potently and Selectively Label Calcium Sensing Receptors In Vitro, In Vivo, and Ex Vivo

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