A Subfornical Organ→Paraventricular Nucleus Neuronal Network Contributes to Non‐Alcoholic Fatty Liver Disease via Hepatic Sympathetic Outflow
Authors: Blackmore Katherine , Simonyan Hayk , Young Colin 
Published: June 2021

Non‐alcoholic fatty liver disease (NAFLD), characterized by hepatic steatosis, affects 1 in 3 adults and leads to an increased risk for metabolic and cardiovascular diseases. Our recent work indicates that obesity‐induced NAFLD is mediated by elevations in hepatic sympathetic nerve activity. However, the neural pathways that contribute to hepatic sympathetic overactivity and subsequent NAFLD development remain unknown. The subfornical organ (SFO) ‐ a forebrain circumventricular region that lies outside the blood brain barrier ‐ senses, integrates and responds to circulating stimuli. Anatomical and electrophysiological data support dense, excitatory projections from the SFO to the paraventricular nucleus of the hypothalamus (SFO→PVN), an integrative nucleus with direct hepatic spinal projections. Taken together, we hypothesized that an SFO→PVN excitatory neuronal network causes hepatic steatosis via elevations in liver sympathetic outflow. An intersectional viral strategy was used in which a retrograde transported canine adenovirus was targeted to the PVN to express Cre‐recombinase in SFO→PVN neurons (CAV2‐Cre‐GFP). This was 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 6 days to activate SFO→PVN neurons (n=4). Short‐term activation of SFO→PVN neurons resulted in hepatic steatosis (2.6±0.02 vs 2.9±0.02 density*107, saline vs CNO, p<0.05) that was paralleled by an elevation in liver tyrosine hydroxylase protein (1.8±0.4 CNO fold saline, p<0.05), the rate‐limiting enzyme for catecholamine synthesis within postganglionic nerve terminals. Based on this, we combined the above viral strategy with selective hepatic denervation (phenol application) or sham surgery (n=5–6). Oil Red O staining (Figure 1) demonstrated that hepatic denervation prevented liver lipid accumulation in response to 6‐day activation of SFO→PVN neurons (1.9±0.01 vs 1.7±0.01 density*107, CNO sham vs CNO denervation, p<0.05). Importantly, this occurred independent of changes in body weight and food intake. Hepatic denervation was confirmed by a reduction in liver tyrosine hydroxlase protein expression (0.3±0.1 vs 1.8±0.3 vs 0.4±0.1 fold saline sham, CNO sham vs saline denervation vs CNO denervation, all p<0.05). Lastly, male C57Bl/6 mice that were fed a high fat diet (10 wks) underwent intersectional viral targeting for expression of an inhibitory DREADDs construct (AAV2‐DIO‐hM4Gi‐mCherry) in SFO→PVN neurons. Remarkably, acute inhibition of SFO→PVN neurons in obese mice resulted in an approximate 45% reduction in hepatic steatosis (Figure 2, n=4/group). Collectively, these findings indicate that activation of SFO→PVN neurons causes liver triglyceride deposition via elevations in hepatic sympathetic outflow. Furthermore, in the context of obesity, inhibition of this forebrain‐hypothalamic‐autonomic circuit results in a marked reduction in NAFLD.Support or Funding InformationR01DK117007This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.