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Differential effects of anaesthesia on the contractility of lymphatic vessels in vivo: authors’ reply

We thank Liu et al. (2020) for their kind appreciation of our work (Bachmann et al. 2019). As stated in their letter, the major aim of our study was to investigate the differential effects of commonly used anaesthesia regimens on the lymphatic contractility and function in mice. As indicated in our Table 1 (Bachmann et al. 2019), the study was motivated by the widely varying published data on the contraction frequency of collecting lymphatic vessels in mice, which we hypothesized might be related to the choice of anaesthesia regimens. Indeed, our study identified the specific effects of the most widely used anaesthesia regimens in rodents, explaining the disparity of previous results, and now serves as guidance for researchers in the field of lymphatic vessel biology. Although we agree with Liu et al. (2020) that evaluating single clinically used aneesthetic agents would be of great interest regarding their effects on lymphatic function in human patients, single agent anaesthesia regimens such as sevoflurane or propofol are not commonly used in mice where usually combination treatments are required. Our study will hopefully motivate future studies of the distinct effects of anaesthesia in the clinical setting. Liu et al. (2020) raise an important point regarding the effects of the depth of anaesthesia on the measured lymphatic vessel functions. Indeed, in our studies, all animals were deeply anaesthetized (and not only sedated), as required by regulatory authorities, and a righting reflex (for which the time is measured until an animal, placed on its back on a table top, rights itself) could thus not be evoked. To ensure a comparable anaesthetic depth between the regimens, all imaging procedures were started as soon as animals showed a relaxed muscle tone when lifted and no movement could be sensed when they were placed and taped for imaging. In case of invasive imaging methods, the preparation started as soon as no response to toe pinching could be evoked. Although it might be of interest to investigate the effects of different anaesthetic regimens over a longer period of time, our investigation was focused on the comparison of lymphatic collecting vessel contractility at the initial state. However, to provide additional information about the anaesthesia depth over time, we included the major physiological parameters over a 50 min time course in the report. Such data demonstrate that no major changes in breathing or heart rates occurred during this time interval.