Specific Connectivity and Unique Molecular Identity of MET Receptor Tyrosine Kinase Expressing Serotonergic Neurons in the Caudal Dorsal Raphe Nuclei

Abstract

Molecular characterization of neurons across brain regions has revealed new taxonomies for understanding functional diversity even among classically defined neuronal populations. Neuronal diversity has become evident within the brain serotonin (5-HT) system, which is far more complex than previously appreciated. However, until now it has been difficult to define subpopulations of 5-HT neurons based on molecular phenotypes. We demonstrate that the MET receptor tyrosine kinase (MET) is specifically expressed in a subset of 5-HT neurons within the caudal part of the dorsal raphe nuclei (DRC) that is encompassed by the classic B6 serotonin cell group. Mapping from embryonic day 16 through adulthood reveals that MET is expressed almost exclusively in the DRC as a condensed, paired nucleus, with an additional sparse set of MET+ neurons scattered within the median raphe. Retrograde tracing experiments reveal that MET-expressing 5-HT neurons provide substantial serotonergic input to the ventricular/subventricular region that contains forebrain stem cells, but do not innervate the dorsal hippocampus or entorhinal cortex. Conditional anterograde tracing experiments show that 5-HT neurons in the DRC/B6 target additional forebrain structures such as the medial and lateral septum and the ventral hippocampus. Molecular neuroanatomical analysis identifies fourteen genes that are enriched in DRC neurons, including 4 neurotransmitter/neuropeptide receptors and 2 potassium channels. These analyses will lead to future studies determining the specific roles that 5-HTMET+ neurons contribute to the broader set of functions regulated by the serotonergic system.

Distinct projection targets define subpopulations of mouse brainstem vagal neurons that express the autism‐associated MET receptor tyrosine kinase

“…In addition to the vagal motor nuclei, EGFP+ neurons were located in the caudal portion of the dorsal raphe nucleus, as reported by Kast et al (), the spinal trigeminal nucleus, and neurons situated in the nucleus of the solitary tract. There also were a few, scattered EGFP+ neurons located at the lateral borders of the hypoglossal nucleus of the 12th cranial nerve (n12), that were most apparent at perinatal ages (Figure c).…”

“…There was clear MET labeling of neuronal somata in the nAmb on GD123, but by GD151, immunostaining was reduced in intensity with more apparent neuropil labeling (Figure a, b). In addition to the DMV and nAmb, MET immunostained neurons and neuropil were evident in the nucleus of the solitary tract, the spinal trigeminal nucleus, and the caudal portion of the dorsal raphe (Figure d, e), consistent with patterns of MET expression described in the mouse (Kast et al, ; Wu & Levitt, ).…”

“…In the current study, we took advantage of a Met EGFP transgenic mouse line, which we show here in the vagal brainstem, and in other studies (Kast et al, 2017), exhibits high fidelity expression of an EGFP reporter to label MET expressing neurons. In the following discussion, A large body of work has described the viscerotopic organization of the vagal motor nuclei, providing context for understanding the putative function of MET1 vagal neurons.…”

“…Fresh frozen brains from Met EGFP mice were cryosectioned at 16 μm and processed according to a commercially available RNAscope Multiplex Fluorescent Assay Kit (Advanced Cell Diagnostics, Hayward, CA) as described previously (Kast et al, ). RNAscope probes used in this study include: Met (cat# 405301), Gfp (cat# 400281), and Chat (cat# 410071).…”

“…as described previously (Kast et al, 2017). RNAscope probes used in this study include: Met (cat# 405301), Gfp (cat# 400281), and Chat (cat# 410071).…”