Single cell transcriptome analysis of mouse carotid body glomus cells

Abstract

Key pointsr Carotid body (CB) glomus cells mediate acute oxygen sensing and the initiation of the hypoxic ventilatory response, yet the gene expression profile of these cells is not available.r We demonstrate that the single cell RNA-Seq method is a powerful tool for identifying highly expressed genes in CB glomus cells.r Our single cell RNA-Seq results characterized novel CB glomus cell genes, including members of the G protein-coupled receptor signalling pathway, ion channels and atypical mitochondrial electron transport chain subunits.r A heterologous cell-based screening identified acetate (which is known to affect CB glomus cell activity) as an agonist for the most highly abundant G protein-coupled receptor (Olfr78) in CB glomus cells.r These data established the first transcriptome profile of CB glomus cells, highlighting genes with potential implications in CB chemosensory function. AbstractThe carotid body (CB) is a major arterial chemoreceptor containing glomus cells whose activities are regulated by changes in arterial blood content, including oxygen. Despite significant advancements in the characterization of their physiological properties, our understanding of the underlying molecular machinery and signalling pathway in CB glomus cells is still limited. To overcome this, we employed the single cell RNA-Seq method by performing next-generation sequencing on single glomus cell-derived cDNAs to eliminate contamination of genes derived from other cell types present in the CB. Using this method, we identified a set of genes abundantly expressed in glomus cells, which contained novel glomus cell-specific genes. Transcriptome and subsequent in situ hybridization and immunohistochemistry analyses identified abundant G protein-coupled receptor signalling pathway components and various types of ion channels, as well as members of the hypoxia-inducible factors pathway. A short-chain fatty acid olfactory receptor Olfr78, recently implicated in CB function, was the most abundant G protein-coupled receptor. Two atypical mitochondrial electron transport chain subunits (Ndufa4l2 and Cox4i2) were among the most specifically expressed genes in CB glomus cells, highlighting their potential roles in mitochondria-mediated oxygen sensing. The wealth of information provided by the present study offers a valuable foundation for identifying molecules functioning in the CB.

Microbial Short-Chain Fatty Acids and Blood Pressure Regulation

“…However, a study in 2016 did show evidence that β ionone is a functional ligand: this study demonstrated that cells natively expressing OR51E2 exhibited an increase in intracellular calcium in response to β ionone which was negated by transfection of siRNA for OR51E2 (27). In late 2015, it was reported that Olfr78 responded to lactate in addition to acetate and propionate(28), and this was confirmed by two other groups in 2016 (29, 30). Notably, microbial metabolism also contributes to circulating lactate levels.…”

Voltage- and receptor-mediated activation of a non-selective cation channel in rat carotid body glomus cells

“…To determine the role of NSC in glomus cell excitability, we need to identify the gene that encodes NSC to delete or silence it. A recent single glomus cell transcriptome analysis indicated that TRPC3, TRPC5 and TRPM7 were most highly expressed TRP ion channels (Zhou et al, 2016). However, the single channel and pharmacological properties of these TRP channels do not match those of NSC, as most TRP ion channel have high single channel conductance (>40-pS) and the modulation by pharmacological agents such as 2-APB are clearly different (Kang et al, 2014).…”

The emerging role of AMPK in the regulation of breathing and oxygen supply

“…normal) cytochrome a 3 , whereas the cytochrome a 3 incorporated in mitochondria of oxygen-sensing cells may have a low affinity for oxygen. Consistent with this hypothesis, recent investigations have demonstrated that NDUFA4L2 [52] and COX4I2 [53,54], two nuclear-encoded atypical subunits of the mitochondrial electron transport chain, are constitutively expressed in carotid body type I cells under normoxia [55]. This contrasts with a number of other cell types where NDUFA4L2 and COX4I2 expression is ordinarily low, but is increased during prolonged hypoxia [52–54].…”

“…The effects on type I cells of hydrogen sulfide may not, therefore, be inconsistent with the conclusion drawn above. This perspective has recently received support from single-cell transcriptome analysis of mouse type I cells which identified few to no reads of the enzymes responsible for generating either carbon monoxide or hydrogen sulfide [55], respectively, haem oxygenase-2, or cystathionine-γ-lyase and cystathionine-β-synthase.As mentioned previously, conditional deletion in tyrosine hydroxylase-positive cells of Ndufs2 , a mitochondrial complex I gene which encodes a protein that participates in ubiquinone binding, has also been shown to selectively block carotid body activation during hypoxia (but not hypercapnia or hypoglycaemia) and thus the hypoxic ventilatory response [61]. The authors concluded that this probably results from loss, during hypoxia, of the capacity for signalling via increased generation of mitochondrial ROS.…”