Selection of receptors highly expressed and enriched in CSF-cNs from a previous transcriptome (data extracted from Prendergast et al., 2023).

Read counts in Fragments Per Kilobase per Million mapped fragments (FPKM) per transcripts were obtained from 5 replicates for both GFP+ and GFP-cells in pooled Tg(pkd2l1:GAL4; UAS:GFP) transgenic larvae.

The somatostatin receptor sstr2a is predominantly expressed in ventral CSF-cNs and unidentified dorsal spinal cells.

(A) Midline sagittal optical section of the spinal cord of a 3 dpf larval zebrafish showing DAPI staining (A1) outlining dorsal and ventral boundaries (solid lines), the central canal (horizontal dashed lines), and somite boundaries (oblique dashed lines). Orientation: dorsal (D), ventral (V), rostral (R), caudal (C). pkd2l1 labeling (A2, magenta) marks dorsolateral (triangle) and ventral (arrowhead) CSF-contacting neurons (CSF-cNs). sstr2a expression (A3, green) is detected almost exclusively in ventral pkd2l1+ CSF-cNs and in a few unidentified dorsal spinal cells (asterisk). Scale bar, 20 µm. Insets (A2–A4i) show magnified ventral CSF-cNs. Scale bar, 10 µm. (B) Lateral sagittal section showing sstr2a expression (B3, green) in ventral pkd2l1+ CSF-cNs (B2, magenta, filled arrowhead) and in unidentified dorsal pkd2l1− cells (asterisk). Dorsolateral pkd2l1+ CSF-cNs (triangle) lack sstr2a expression. Scale bar, 20 µm. Insets (B2–B4i) show a magnified dorsolateral pkd2l1+/sstr2a− CSF-cN and a sstr2a+/pkd2l1− dorsal cell (asterisk). Scale bar, 10 µm. (C) 3D schematic of a transverse spinal cord section summarizing sstr2a expression (green) primarily in ventral pkd2l1+ CSF-cNs (magenta) and in additional dorsal cells (asterisk). (D) Quantification of sstr2a+ CSF-cNs per larva (n = 3), normalized per 100 µm. In dorsolateral position to the central canal (CC): 1 ± 0.1 sstr2a+/pkd2l1+ cells per 100 µm (grey); in ventral position to the central canal (CC): 12 ± 1.2 sstr2a+/pkd2l1+ cells per 100 µm. Total pkd2l1+ cells (magenta): 18 ± 1.2 dorsolateral CSF-cNs per 100 µm and 18 ± 2.0 ventral CSF-cNs per 100 µm. Mean values are given ± s.e.m.

The metabotropic glutamate receptor 2 grm2a is expressed in ventral and dorsolateral CSF-cNs as well as in unknown dorsal cells in the spinal cord of larval zebrafish.

(A) Sagittal optical section of the spinal cord of a 2 dpf larval zebrafish showing DAPI staining (A1) outlining dorsal and ventral boundaries (solid lines), the central canal (horizontal dashed lines), and somite boundaries (oblique dashed lines). Orientation: dorsal (D), ventral (V), rostral (R), caudal (C). pkd2l1 labeling (A2, magenta) marks dorsolateral (triangle) and ventral (arrowhead) CSF-contacting neurons (CSF-cNs). grm2a expression (A3, green) overlaps with pkd2l1 in both populations and is also detected in unidentified dorsal cells (asterisk). Scale bar, 20 µm. Insets (A2–A4i) show magnified ventral CSF-cNs. Scale bar, 10 µm. (B) Lateral sagittal section showing grm2a expression (B3, green) in dorsolateral pkd2l1+ CSF-cNs (B2, magenta, triangle) and in pkd2l1− cells near the central canal and in unidentified dorsal cells (asterisk). Scale bar, 20 µm. Insets (B2–B4i) show a magnified dorsolateral CSF-cN (pkd2l1+, triangle) and a grm2a+/pkd2l1− cell (asterisk). Scale bar, 10 µm. (C) 3D schematic of a transverse spinal cord section summarizing grm2a expression (green) in ventral and dorsolateral pkd2l1+ CSF-cNs (magenta), in additional dorsal cells and cells near the central canal (asterisk). (D) Quantification of grm2a+ CSF-cNs per larva (n = 3), normalized per 100 µm. In dorsolateral position to the central canal (CC): 10 ± 2.8 per 100 µm grm2a+/pkd2l1+ cells (grey); in ventral position to the central canal (CC): 15 ± 1.7 grm2a+/pkd2l1+ cells per 100 µm (grey). Total pkd2l1+ cells (magenta): 14 ± 2.5 dorsolateral CSF-cNs per 100 µm and 19 ± 1.3 ventral CSF-cNs per 100 µm. Mean values are given ± s.e.m.

The receptor ptprna is expressed in ventral and dorsolateral spinal CSF-Ns.

(A) Lateral sagittal optical section of the spinal cord of a 3 dpf larval zebrafish showing DAPI staining (A1) outlining dorsal and ventral boundaries (solid lines), the central canal (horizontal dashed lines), and somite boundaries (oblique dashed lines). Orientation: dorsal (D), ventral (V), rostral (R), caudal (C). pkd2l1 labeling (A2, magenta) marks dorsolateral (triangle) and ventral (arrowhead) CSF-cNs. ptprna expression (A3, green) overlaps with pkd2l1 in both dorsolateral and ventral CSF-cNs. Scale bar, 20 µm. (A2i-A4i) Magnified view of two dorsal (triangle) and one ventral (arrowhead) pkd2l1+/ptprna+ CSF-cNs. Scale bar, 10 µm. (B) Sagittal section showing DAPI staining (B1) outlining dorsal and ventral boundaries (solid lines), the central canal (horizontal dashed lines). ptprna expression (B3, green) is found in ventral (arrowhead) and dorsolateral (triangle) pkd2l1+ CSF-cNs (B2, magenta) and in pkd2l1− cells near the central canal. Scale bar, 20 µm. (B2i-B4i) Magnified view of two ventral (arrowhead) and one dorsal pkd2l1+ ptprna+ CSF-cNs. Scale bar, 10 µm. (C) Schematic represents in 3D a transverse section of the spinal cord showing that both ventral and dorsolateral (magenta) CSF-cNs (pkd2l1+) express ptprna along with unknown cells around the central canal in the spinal cord (green asterisk). (D) Cell count per 100 µm of ptprna+ in CSF-cNs per larval (n = 3 larvae) are normalized per 100µm. In dorsolateral position to the central canal (CC): 10 ± 2.8 ptprna+/pkd2l1+ cells per 100 µm (grey); in ventral position to the central canal (CC): 15 ± 1.7 ptprna+/pkd2l1+ cells per 100 µm (grey). Cell count as pkd2l1+ cells (magenta): 14 ± 2.5 dorsolateral CSF-cNs per 100 µm and 19 ± 1.3 ventral CSF-cNs per 100 µm. Mean values are given ± s.e.m.

The receptor ldlrad2 is expressed in ventral and dorsolateral CSF-cNs as well as other cells in contacting the cerebrospinal fluid, most likely corresponding to ependymal radial glia.

(A) Sagittal optical section of the spinal cord showing DAPI staining (A1) outlining dorsal and ventral boundaries (solid lines), the central canal (horizontal dashed lines), and somite boundaries (oblique dashed lines). Orientation: dorsal (D), ventral (V), rostral (R), caudal (C). pkd2l1 labeling (A2, magenta) marks dorsolateral (triangle) and ventral (arrowhead) CSF-cNs. ldlrad2 expression (A3–A4, green) overlaps with pkd2l1 in both dorsolateral and ventral CSF-cNs, and is also detected in other cells surrounding the central canal. Scale bar, 20 µm. (A2i-A4i) Magnified view of one dorsolateral and one ventral pkd2l1+/ldlrad2+ CSF-cNs and a pkd2l1− in the ventral position to the central canal. (A2ii-A4ii) Magnified view of two dorsolateral pkd2l1+/ldlrad2+ CSF-cNs and pkd2l1− cells surrounding the central canal (asterisk). (from box in A2–A4; scale bar, 10 µm). (B) Schematic represents in 3D a transverse section of the spinal cord showing that both ventral and dorsolateral (magenta) CSF-cNs (pkd2l1+) express ldlrad2 along with unknown cells around the central canal and dorsally located in the spinal cord (green asterisk). (C) Cell counts of ldlrad2+ per 100 µm in CSF-cNs per larval (n = 3 larvae) are normalized per 100µm. In dorsolateral position to the central canal (CC): 14 ± 1.5 ldlrad2+/pkd2l1+ cells per 100µm (grey); in ventral position to the central canal (CC): 15 ± 0.6 ldlrad2+/pkd2l1+ cells per 100µm (grey). Total pkd2l1+ cells (magenta): 16 ± 1.9 dorsolateral CSF-cNs per 100µm and 17 ± 0.6 ventral CSF-cNs per 100µm. Mean values are given ± s.e.m.