The Effect of Heat Exposure on Physiological Markers

Primary outcomes

1. Change in plasma metabolites of the kynurenine pathway (μm)

   Time frame: Day 1 (every 0.5 °C until rectal temperature (Trec) reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   An ultra-performance liquid chromatography-tandem mass spectrometry system (UPLC-MS/MS) was used to measure venous plasma levels of tryptophan, kynurenine, kynurenic acid, 3-hydroxy-kynurenine, quinolinic acid, nicotinamide and picolinic acid (in μm). The UPLC-MS/MS system uses a Xevo TQ-XS triple quadrupole mass spectrometer (Waters) with a Z-spray electrospray interface, and the system operates in electrospray positive multiple reaction monitoring mode.

2. Rectal temperature (°C)

   Time frame: Day 1 (during both heating modalities until Tre reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   Rectal temperature (in °C) was measured using a thermocouple (Rectal Probe, Ellab, Denmark) inserted to a depth of 12 cm past the anal sphincter.

3. Change in heart rate (bpm)

   Time frame: Day 1 (during both heating modalities until Tre reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   Heart rate (in bpm) was recorded using a heart rate sensor with a chest strap (Polar, Finland).

4. Change in physiological strain index

   Time frame: Day 1 (every 0.5 °C until rectal temperature (Trec) reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   A physiological strain index (PSI) was used to indicate heat strain. PSI = 5 x (Tret - Tre0) x (39.5 - Tre0)^-1+ 5 x (HRt - HR0) x (180 - HR0)^-1, where rectal temperature (Tre) t and heart rate (HR) t are simultaneous measurements taken every 0.5°C of the heat exposure and Tre0 and HR0 are the initial measurements.

5. Change in glucose (mmol/L)

   Time frame: Day 1 (every 0.5 °C until rectal temperature (Trec) reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   The glucose concentration (in mmol/L) was measured using an automatic blood glucose meter Glucocard X-mini-Plus (Arkray Inc, Japan).

6. Change in lactate (mmol/L)

   Time frame: Day 1 (every 0.5 °C until rectal temperature (Trec) reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   The lactate concentration (in mmol/L) was measured using an Accutrend Plus System (Roche, Germany).

7. Changes in prolactin concentration (ng/mL)

   Time frame: Day 1 (every 0.5 °C until rectal temperature (Trec) reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   The venous prolactin (in ng/mL) was measured using enzyme-linked immunosorbent assay kit and a Spark multimode microplate reader (Tecan, Austria).

8. Changes in catecholamines concentration (ng/mL)

   Time frame: Day 1 (every 0.5 °C until rectal temperature (Trec) reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   The venous adrenaline and noradrenaline concentrations (in ng/mL) was measured using enzyme-linked immunosorbent assay kits and a Spark multimode microplate reader (Tecan, Austria).

9. Changes in salivary cortisol concentration (µg/dL)

   Time frame: Day 1 (every 0.5 °C until rectal temperature (Trec) reached 39°C, followed by a recovery phase until Trec reached 37.5°C)

   The saliva samples was collected to measure cortisol level (in µg/dL) using an ELISA kits and a Spark multimode microplate reader (Tecan, Austria).