A Forebrain‐Hypothalamic Circuit Mediates Hepatic Steatosis
Authors: Blackmore Katherine A. , Houchen Claire J. , Simonyan Hayk , Young Colin N. 
Published: June 2021

Non‐alcoholic fatty liver disease (NAFLD), characterized by an accumulation of hepatic triglycerides (i.e. steatosis), is a growing health epidemic. While the majority of investigations have taken a liver‐centric perspective, we recently demonstrated a role for the brain in NAFLD. In particular, disruptions in the subfornical organ (SFO) ‐ a small circumventricular forebrain nucleus ‐ appear to play a key role in the development of hepatic steatosis. However, the neural network(s) through which the SFO contributes to NAFLD remains unknown. The paraventricular nucleus of the hypothalamus (PVN) is a central regulator of peripheral autonomic and endocrine function, and the SFO has dense excitatory projections to the PVN. Taken together, we hypothesized that activation of excitatory SFO PVN‐projecting neurons would result in NAFLD development. Using adult male C57Bl/6 mice, we first confirmed that SFO PVN projecting neurons are excitatory by using retrograde viral labeling from the PVN (CAV2‐GFP) combined with SFO immunohistochemistry for the excitatory neuronal marker calcium‐calmodulin‐dependent kinase II (CAMKII). Consistent with previous findings, nearly 100% of the retrograde tracer co‐localized with CAMKII in the SFO (not shown, n=3). Subsequently, we employed an intersectional viral strategy (Figure) in male C57Bl/6 mice in which a retrograde transported canine adenovirus was targeted to the PVN to allow for expression of Cre‐recombinase in SFO PVN‐projecting neurons (CAV2‐Cre‐GFP), combined with SFO‐targeted delivery of a Cre‐inducible designer receptors engineered against designer drugs (DREADDs) excitatory construct (AAV2‐DIO‐hM3Gq‐mCherry). With this approach, the pharmacological ligand clozapine‐N‐oxide (CNO; 3 mg/kg i.p.) was administered once daily over 7 days to activate SFO PVN‐projecting neurons (n=4). Saline served as a vehicle control (n=4). Oil Red O staining of the liver demonstrated that, relative to control animals, one‐week activation of SFO PVN‐projecting neurons resulted in a clear and consistent development of hepatic steatosis (2.69±0.02 vs. 2.92±0.02 au × 107, saline vs. CNO, p<0.05, 9 liver sections averaged/animal). Taking into consideration that NAFLD is associated with insulin resistance and elevated hepatic gluconeogenesis, we also examined blood glucose upon sacrifice and found an ~35% increase in circulating glucose concentrations in CNO treated animals relative to saline controls (161±20 vs. 249±16 mg/dl, saline vs. CNO, p<0.05,). Importantly, these changes occurred independent of differences in body weight (25±1 vs. 25+1 g, saline vs. CNO, p<0.05) and food intake. Real time qPCR analysis further indicated that activation of SFO neurons that project to the PVN resulted in a marked upregulation of liver markers of de novo lipogenesis and gluconeogenesis (e.g. DGAT1: 3.6±0.3 fold saline, p<0.05; G6Pase: 2.6±1.0 fold saline, p=0.12). Viral targeting and activation of the SFO was confirmed using anatomical and functional analyses (i.e. cFOS staining). Collectively, these findings indicate that short‐term activation of excitatory SFO PVN‐projecting neurons results in a NAFLD phenotype characterized by elevated liver triglycerides, disruptions in liver metabolic markers, and hyperglycemia. Furthermore, these findings suggest that manipulating this forebrain‐hypothalamic network in the context of obesity may be a novel approach to target NAFLD.Support or Funding InformationHL116776This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.