Identifying functional and anatomic mechanisms of glucose mobilization by cholecystokinin b receptor containing neurons in the ventromedial nucleus of the hypothalamus

Abstract
Nutrient mobilization fuels the metabolic activity needed to fight or escape threats and is a critical
component of the central nervous system (CNS) response to stress. While mild stress-induced hyperglycemia
is associated with improved survival in critical illness, severe or chronic responses are associated with
development of diabetes. The sympathetic nervous system (SNS) plays a crucial role in coordinating the
response to stress; however, the specific mechanisms and neural circuits by which the brain mediates SNS-
dependent stress responses, including nutrient mobilization, remain poorly understood. Pre-autonomic neurons
in the ventromedial nucleus of the hypothalamus (VMN) modulate SNS outflow to multiple organs and can
mediate a diverse range of responses, including hepatic glucose production, glucose disposal, and energy
expenditure. We have recently identified a subset of VMN neurons (marked by cholecystokinin b receptor (Cckbr)
expression (VMNCCKBR neurons)) that are activated by stressors (including restraint and noxious stimuli) and
mediate glucose and lipid mobilization as well as defensive freezing behaviors, suggesting that VMNCCKBR cells
coordinate multiple responses to specific stressors. Single nucleus RNA-Sequencing (snRNA-Seq) analysis of
the VMN reveals that VMNCCKBR neurons distribute across multiple transcriptionally defined classes of neurons
within the VMN (VMN T-types) and they project to multiple brain regions, including the preoptic area (POA;
involved in metabolism) and the periaqueductal gray (PAG; implicated in the behavioral response to threats).
The goals of this proposal are to define the anatomic neural circuits and functional mechanisms of
VMNCCKBR neuron-mediated nutrient mobilization and to test the hypothesis that VMNCCKBR neurons
mediate metabolic and behavioral stress responses through independent neural pathways. We will
identify the target tissues of VMNCCKBR neurons using optogenetic-stimulated norepinephrine turnover and will
determine the mechanisms through which VMNCCKBR neurons regulate acute hepatic glucose production using
stable isotope fluxomics. We will then assess whether VMNCCKBR-dependent lipid mobilization contributes to
glucose production chronically. To identify which populations of VMNCCKBR neurons regulate nutrient mobilization
versus defensive behaviors, we will use retrograde tracing followed by snRNA-Seq to define clusters that project
to the POA and PAG respectively. We will then use optogenetic activation of VMNCCKBR neuron terminals
projecting to the POA and PAG to establish the functional outputs of these cell classes. This work will define the
anatomic and functional mechanisms driving SNS-dependent stress-mediated nutrient mobilization. Completion
of this project will provide training in liver and white adipose physiology, neuroanatomical tracing, behavioral
phenotyping and genomic bioinformatics, allowing me to become an expert in central nervous system regulation
of glucose homeostasis and providing the tools necessary to transition to an independent career as a physician-
scientist.

Grant Number: 5K08DK129722-04
NIH Institute/Center: NIH
Principal Investigator: Alison Affinati