Landscape of neuronal transcriptional states in the pig RAGP
(A) Expression of 174 genes, each assayed in 321 single neurons (n = 3 animals) through HT-qPCR, yielding six transcriptional states using a combination of clustering and template matching analysis. A majority of the states consisted of both SAN-projecting and non SAN-projecting neurons. Sample annotations at the top of the heatmap indicate whether the single neurons were SAN-projecting or non SAN-projecting and indicate the distribution across animals. A complete heatmap with 405 neurons from all 4 RAGP is shown in Figure S3.
(B) Landscape of neuronal transcriptional states visualized as a tSNE plot. Colors correspond to the states shown in panel (A).
(C) Visualization of the neuronal states within the 3D anatomical framework for a representative RAGP. The bounding box on the lower panel shows 18.8 mm, 19.4 mm, and 16 mm on the x, y, and z axis, respectively.
(D–F) Expression distribution of select genes with enrichment in specific transcriptional states: (D) state (A)Map2, Chat, Th, Dbh; (E) states B and CAdrab2b, Adrb3, Kcnab1, Kcnc1; and (F) states D and FNpff and Kcnip1. Enrichment assessed by a one-way ANOVA and post hoc Tukey Honest Significant Difference test pvalue < 0.01.