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. 2021 Jan 25;4:107. doi: 10.1038/s42003-020-01604-x

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — Spontaneously breathing rats under urethane anesthesia received an intravenous injection of fentanyl (30 mcg/kg) sufficient to cause lasting respiratory suppression. Following fentanyl dosing, animals received intramuscular TI stimulation or no intervention. Panel a shows an example of sustained restoration of respiratory airflow when the stimulation was initiated shortly after opioid overdose-induced apnea, and was delivered with a pattern of 60 s on and 3 s off until spontaneous breathing resumed. Note: The gaps in the blood pressure traces indicate when arterial blood samples were taken. b Average respiratory airflow (two-way ANOVA: group F(1) = 40.84, p < 0.0001; time F(3) = 41.86, p < 0.0001; interaction F(3) = 21.4, p < 0.0001) and (c) mean arterial blood pressure (MABP) (n = 4 animals/group, two-way ANOVA: group F(1) = 19.81, p = 0.0008; time F(3) = 45.95, p < 0.0001; interaction F(3) = 7.82 p = 0.0037). The gray bars indicate time periods selected for quantitative comparison (Baseline–BL, immediately Post Fentanyl–PF, five minutes–5 min, and ten minutes–10 min after fentanyl). The gray dots indicate data points from the animals used for the example traces in panel a, (mean + 1 standard deviation; asterisk (*) indicates p < 0.05). d TI stimulation restored ventilation and prevented fatal apnea in all animals (TI rescue). TI stimulation with a 1 Hz beat frequency produced a respiratory rate of 60 breaths per minute (bpm) during stimulation. When the beat frequency was targeted to the endogenous respiratory rate, there was no difference in the evoked vs. spontaneous respiratory rate (n = 4 animals, two-way ANOVA: group F(1) = 22.64, p = 0.0002; time F(3) = 3.85, p = 0.0407; interaction F(3) = 5.03, p = 0.0183). e Survival curve for no-intervention (n = 4 animals) and TI rescue (60 bpm) conditions (n = 4 animals).