Do Ccka Receptor Containing Vagal Afferent Neurons Mediate Gut-Brain Inflammatory Signals?

Inflammatory responses are associated with chronic diseases and persistent fatigue and may contribute significantly to Gulf War Illness (GWI). The gut-brain axis is emerging as a conduit of cross-talk between central and peripheral inflammatory responses. However, research on the neuroimmune mechanisms involved in the gut-brain axis is lacking. Recent work has identified an important role for vagal afferent neurons (VANs) containing CALCA and TRPA1 receptors that regulate pro-inflammatory and anti-inflammatory responses, respectively. However, the contribution of cholecystokinin (CCK) A receptor-containing VANs to gut-brain immune signaling is unknown. CCKARs are G-protein-coupled receptors that are activated by the satiety-inducing gut peptide CCK and are ubiquitously expressed in VANS, whose cell bodies are located in vagal nodose ganglia. We hypothesized that selective ablation of CCKAR-containing VANs in nodose ganglia (ND) would dampen the inflammatory response after LPS challenge. C57BL/6N male mice were fasted overnight before undergoing bilateral vagal deafferentation via injection of the neurotoxin, CCK-saporin (CCK-SAP) (250nL ND; 250ng/uL) into the nodose ganglia. Controls received bilateral injections with Blank-SAP. Six days post surgery, mice were treated with ip injection of LPS (0.1 mg/kg) or 0.9% NaCl. At sacrifice, 2h later, mice were transcardially perfused and plasma, brains and nodose ganglia were collected. Fluorescent in-situ hybridization of Cckar mRNA expression using specific probe (313751, ACD) confirmed this preparation produces deafferentation of CCKAR-containing VANs. Cell counts of Cckar in VANs were markedly lower in CCK-SAP vs Blank-SAP (p<0.05, n=5). Cell counts measured in sham controls receiving Blank-SAP were no different than those obtained from WT (no surgery) (ns, n=3-5/group). Plasma was processed for Interleukin-6 (IL-6) ELISA (ThermoFisher), nodose ganglia for Cckar RNAcope and brains for c-FOS immunofluorescence. LPS increased IL-6 levels by 66% and 123.2% on average in both CCK- and Blank-SAP groups, respectively, compared to their saline control (p<0.01-0.0003, n=3-10/group). Mean normalized values of LPS-stimulated IL-6 expressed as a percent of corresponding saline control was significantly lower (46%) for CCK-SAP vs Blank-SAP (p<0.05, n=10/group). The percent increase in counts of c-fos-positive cells in solitary tract nucleus (NTS) in response to LPS stimulation was significantly lower (31%) in the CCK-SAP vs Blank-SAP group (p<0.05, n=4-6/group) indicating an interruption of gut signals arriving into the brainstem. Using a different cohort of mice the CCK-SAP method of vagal deafferentation abolished a significant decrease in food consumption in response to ip CCK (4ug/kg) (n=7; p<.05-.0001), indicating the correct targeting of gut VANs. Targeting gut-brain inflammatory signals via CCKAR VANs may help us provide novel strategies for treatment of GWI and other disorders with deregulated immune responses. Supported by DOD grant W81XWH-19-1-0802 (MCC), NIH F31ES034304 (EVK), Danone of North America Fellowship (EVK), UCR Undergraduate Minigrant (SA, JS) and RISE fellowship (JS) This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.