Characterization of plasma cytokine response to intraperitoneally administered LPS & subdiaphragmatic branch vagus nerve stimulation in rat model.

Vagus nerve stimulation (VNS) has been on the forefront of inflammatory disorder research and has yielded many promising results. Questions remain, however, about the biological mechanisms of such treatments and the inconsistencies in the methods used in research efforts. Here, we aimed to clarify the inflammatory response to intraperitoneal (IP) injections of lipopolysaccharide (LPS) in rats, while analyzing corresponding effects of electrical stimulation to subdiaphragmatic branches (anterior gastric, accessory celiac, and hepatic) of the left vagus nerve. We accomplished an in-depth characterization of the time-varying cytokine cascade response in the serum of 58 rats to an acute IP LPS challenge over a 330-minute period by utilizing curve-fitting and starting point-alignment methods. We then explored the post-LPS neuromodulation effects of electrically stimulating individually cuffed subdiaphragmatic branches. Through our analysis, we found there to be a consistent order of IP LPS cytokine response (IL-10, TNF-α, GM-CSF, IL-17F, IL-6, IL-22, INF-γ). Apart from IL-10, the IP cytokine cascade was more variable in starting time and occurred later than in previously recorded intravenous (IV) challenges. We also found distinct regulatory effects on multiple cytokine levels by each of the three subdiaphragmatic stimulation subsets. While the time-variability of IP LPS use in rats complicates its utility, we have shown it to be a practical, arguably more physiologically relevant method than IV in rats when our methods are used. More importantly, we have shown that selective subdiaphragmatic neurostimulation can be utilized to selectively induce specific effects on inflammation in the body.

Modulating the Inflammatory Reflex in Rats Using Low-Intensity Focused Ultrasound Stimulation of the Vagus Nerve.

Tumor necrosis factor α (TNF-α) is linked to several chronic inflammatory diseases. Electrical vagus nerve stimulation reduces serum TNF-α levels but may cause chronic nerve damage and requires surgery. Alternatively, we proposed focused ultrasound stimulation of the vagus nerve (uVNS), which can be applied non-invasively. In this study, we induced an inflammatory response in rats using lipopolysaccharides (LPS) and collected blood to analyze the effects of uVNS on cytokine concentrations. We applied one or three 5-min pulsed focused ultrasound stimulation treatments to the vagus nerve (250 kHz, ISPPA = 3 W/cm2). Animals receiving a single ultrasound application had an average reduction in TNF-α levels of 19%, similar to the 16% reduction observed in electrically stimulated animals. With multiple applications, uVNS therapy statistically reduced serum TNF-α levels by 73% compared with control animals without any observed damage to the nerve. These findings suggest that uVNS is a suitable way to attenuate TNF-α levels.

Chronic cuffing of cervical vagus nerve inhibits efferent fiber integrity in rat model.

OBJECTIVE: Numerous studies of vagal nerve stimulation (VNS) have been published showing it to be a potential treatment for chronic inflammation and other related diseases and disorders. Studies in recent years have shown that electrical stimulation of the vagal efferent fibers can artificially modulate cytokine levels and reduce systematic inflammation. Most VNS research in the treatment of inflammation have been acute studies on rodent subjects. Our study tested VNS on freely moving animals by stimulating and recording from the cervical vagus with nerve cuff electrodes over an extended period of time. APPROACH: We used methods of electrical stimulation, retrograde tracing (using Fluorogold) and post necropsy histological analysis of nerve tissue, flow cytometry to measure plasma cytokine levels, and MRI scanning of gastric emptying. This novel combination of methods allowed examination of physiological aspects of VNS previously unexplored. MAIN RESULTS: Through our study of 53 rat subjects, we found that chronically cuffing the left cervical vagus nerve suppressed efferent Fluorogold transport in 43 of 44 animals (36 showed complete suppression). Measured cytokine levels and gastric emptying rates concurrently showed nominal differences between chronically cuffed rats and those tested with similar acute methods. Meanwhile, results of electrophysiological and histological tests of the cuffed nerves revealed them to be otherwise healthy, consistent with previous literature. SIGNIFICANCE: We hypothesize that due to these unforeseen and unexplored physiological consequences of the chronically cuffed vagus nerve in a rat, that inflammatory modulation and other vagal effects by VNS may become unreliable in chronic studies. Given our findings, we submit that it would benefit the VNS community to re-examine methods used in previous literature to verify the efficacy of the rat model for chronic VNS studies.