Briefly, mechanical properties were assessed using organ bath and transducer systems, coupled to IOX software (EMKA Technologie).
Surface hydrophobicity was assessed using the bacterial adherence to hydrocarbons (BATH) assay, with xylene (BDH, VWR International, Leicestershire, UK) as the hydrocarbon of choice [ 11].
The voltage sensitivity of extracellularly applied JPW3027 was assessed using fast imaging and voltage-clamp recordings.
The effect of bath application of drugs on XII nerve inspiratory output was assessed using Clampfit software to measure the resulting average change in XII inspiratory burst amplitude, area, duration and frequency during a two minute period 15 minutes after the drug was applied to the bath.
Subjective sleepiness was assessed using the Stanford Sleepiness Scale.
Time trends were assessed using birth cohorts.
Intentions to use ecstasy, synthetic cannabis, synthetic stimulants ('bath salts') and NEDs in general will be assessed using eight items.
Statistical significance was assessed by applying Student's T-test using Microsoft Excel software.
AS activity was assessed by the bath ankylosing spondylitis disease activity index (BASDAI) [ 22], and we used the bath ankylosing spondylitis function index (BASFI) [ 23] to evaluate function.
A similar linear model was used for knock out temperature, also introducing water-bath as a covariate since Tko was assessed in different water-baths.
