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Objective.  Vagus nerve stimulation (VNS) is typically delivered at increasing stimulus intensity until a neurological or physiological response is observed (‘threshold’) for dose calibration, preclinically and therapeutically. Factors affecting VNS thresholds have not been studied systematically. In a rodent model of VNS we measured neural and physiological responses to increasing VNS intensity, determined neurological and physiological thresholds and examined the effect of implant- and anesthesia-related factors on thresholds.  Approach.  In acute and chronic vagus implants (45 and 20 rats, respectively) VNS was delivered under isoflurane, ketamine-xylazine, or awake conditions. Evoked compound action potentials (CAPs) were recorded and activation of different fiber types was extracted. Elicited physiological responses were registered, including changes in heart rate (HR), breathing rate (BR), and blood pressure (BP). CAP and physiological thresholds were determined.  Main results . The threshold for evoking discernable CAPs (&gt;10  µ V) (CAP threshold) is significantly lower than what elicits 5%–10% drop in heart rate (heart rate threshold, HRT) (25  µ A ± 1.8 vs. 80  µ A ± 5.1, respectively; mean ± SEM). Changes in BP and small changes in BR (bradypnea) occur at lowest intensities (70  µ A ± 8.3), followed by HR changes (80  µ A ± 5.1) and finally significant changes in BR (apnea) (310  μ A ± 32.5). HRT and electrode impedance are correlated in chronic (Pearson correlation  r = 0.47;  p &lt; 0.001) but not in acute implants ( r  = −0.34;  p  NS); HRT and impedance both increase with implant age ( r = 0.44;  p &lt; 0.001 and  r  = 0.64;  p  &lt; 0.001, respectively). HRT is lowest when animals are awake (200  µ A ± 35.5), followed by ketamine-xylazine (640  µ A ± 151.5), and isoflurane (1000  µ A ± 139.5). The sequence of physiological responses with increasing VNS intensity is the same in anesthetized and awake animals. Pulsing frequency affects physiological responses but not CAPs.  Significance . Implant age, electrode impedance, and type of anesthesia affect VNS thresholds and should be accounted for when calibrating stimulation dose.

Implant- and anesthesia-related factors affecting cardiopulmonary threshold intensities for vagus nerve stimulation