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Patch-to-Seq and Transcriptomic Analyses Yield Molecular Markers of Functionally Distinct Brainstem Serotonin

Gary C Mouradian1,2, Pengyuan Liu1, Pablo Nakagawa1,3

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Frontiers in Synaptic Neuroscience
|July 18, 2022
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Summary

Researchers linked CO2 sensitivity in brainstem serotonin neurons to distinct gene expression profiles. This study identifies new molecular markers for CO2-sensitive neurons, advancing our understanding of respiratory chemoreception.

Keywords:
5-HT neuronsCO2 chemoreceptionPH sensitivitybrainstem 5-HTmedullary raphepatch-seqraphe magnustranscriptomic analysis

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Area of Science:

  • Neuroscience
  • Physiology
  • Molecular Biology

Background:

  • Acute regulation of CO2 and pH homeostasis relies on respiratory chemoreceptors.
  • Brainstem serotonin (5-HT) neurons exhibit CO2 sensitivity or insensitivity, potentially due to distinct embryonic origins and transcriptional profiles.

Purpose of the Study:

  • To correlate CO2 responses in brainstem 5-HT neurons with unique transcriptional profiles, molecular markers, and pathways.
  • To identify upstream regulators and candidate gene markers for CO2-sensitive 5-HT neurons.

Main Methods:

  • Patch-to-Seq technique applied to fluorescently labeled 5-HT neurons in rat brainstem slices.
  • Measurement of firing rate changes during hypercapnic acidosis.
  • Single-cell RNA sequencing (scRNAseq) and bioinformatic analyses to compare transcriptomes.
  • In situ mRNA hybridization to validate candidate gene expression.

Main Results:

  • CO2-sensitive 5-HT neurons showed lower baseline firing rates.
  • 166 differentially expressed genes were identified between CO2-sensitive and insensitive 5-HT neurons.
  • Predicted upstream regulators include Egr2 and Leptin; CD46 and Iba57 were identified as candidate markers.

Conclusions:

  • Novel insights into the transcriptional control of cellular chemoreception were provided.
  • Unbiased candidate gene markers for CO2-sensitive 5-HT neurons were identified.
  • The Patch-to-Seq technique is valuable for linking gene expression to cellular function.