Investigation of the Effects of Sleep Deprivation on Itch and Pain Sensitivity

Immediately following the itch provocations, participants will be instructed to rate the itch intensity for 10 minutes using a digital visual analogue scale (VAS; eVAS Software: Aalborg, University, Denmark), on a tablet. The scale will be measured from 0 to 100, where 0 represents 'no itch' and 100 'worst itch imaginable'

Immediately following the itch provocations, participants will be instructed to rate the itch intensity for 10 minutes using a digital visual analogue scale (VAS; eVAS Software: Aalborg, University, Denmark), on a tablet. The scale will be measured from 0 to 100, where 0 represents 'no itch' and 100 'worst itch imaginable'

Immediately following the itch provocations, participants will be instructed to rate the pain intensity for 10 minutes using a digital visual analogue scale (VAS; eVAS Software: Aalborg, University, Denmark), on a tablet. The scale will be measured from 0 to 100, where 0 represents 'no pain' and 100 'worst pain imaginable'.

Immediately following the itch provocations, participants will be instructed to rate the pain intensity for 10 minutes using a digital visual analogue scale (VAS; eVAS Software: Aalborg, University, Denmark), on a tablet. The scale will be measured from 0 to 100, where 0 represents 'no pain' and 100 'worst pain imaginable'.

Microvascular reactivity The evoked cutaneous inflammation (quantified by superficial blood perfusion) will be measured by full-field laser perfusion imaging

Microvascular reactivity The evoked cutaneous inflammation (quantified by superficial blood perfusion) will be measured by full-field laser perfusion imaging

Pleasant touch sensation measured using a standardized sensory brush (SENSELab Brush-05, Somedic AB, Hörby, Sweden) exerting a force of 200 to 400 mN.The stimulation consists of three strokes (2 cm in length) over the treated/control areas. The strokes will be applied perpendicularly to the skin and the subject will rate the sensation induced by the brush on a NRS scale labeled "very unpleasant" and "very pleasant" at the extremity and "neutral" at the center.

Pleasant touch sensation measured using a standardized sensory brush (SENSELab Brush-05, Somedic AB, Hörby, Sweden) exerting a force of 200 to 400 mN. The stimulation consists of three strokes (2 cm in length) over the treated/control areas. The strokes will be applied perpendicularly to the skin and the subject will rate the sensation induced by the brush on a NRS scale labeled "very unpleasant" and "very pleasant" at the extremity and "neutral" at the center.

MEI is measured by using mildly pruritic, non-painful von Frey nylon filaments of a predetermined intensity (generally 5-30 miliNewton of force). This stimulator is moved in intervals of 0.5 cm outside the area of itch provocation. The subjects will report a score on a NRS scale ranging 0-10.

MEI is measured by using mildly pruritic, non-painful von Frey nylon filaments of a predetermined intensity (generally 5-30 miliNewton of force). This stimulator is moved in intervals of 0.5 cm outside the area of itch provocation. The subjects will report a score on a NRS scale ranging 0-10.

The von-Frey filament that better evokes the itch sensation during the assessment of mechanically evoked itch as well as a template of soft plastic are used to map the area of hyperkinesis in the test areas (forearms). The subjects will report a score on a NRS scale ranging 0-10.

The von-Frey filament that better evokes the itch sensation during the assessment of mechanically evoked itch as well as a template of soft plastic are used to map the area of hyperkinesis in the test areas (forearms). The subjects will report a score on a NRS scale ranging 0-10.

This test is conducted using a pinprick set (Aalborg University, Aalborg). The set consists of 7 needles each with a diameter of 0.6 mm and different force applications: 8, 16, 32, 64, 128, 256, and 512 mN. The subjects will report a score on a NRS scale ranging 0-10.

This test is conducted using a pinprick set (Aalborg University, Aalborg). The set consists of 7 needles each with a diameter of 0.6 mm and different force applications: 8, 16, 32, 64, 128, 256, and 512 mN. The subjects will report a score on a NRS scale ranging 0-10.

This test is conducted with the same pinprick set used to test the MPT. Starting with the lightest, each stimulator is applied at a rate of 2 s on, 2 s off in an ascending order, and the subject will be asked to give a pain rating for each stimulus on a scale from 0-10 (where 0 indicates ''no pain'', and 10 indicates the ''worst imaginable pain'') on a NRS.

This test is conducted with the same pinprick set used to test the MPT. Starting with the lightest, each stimulator is applied at a rate of 2 s on, 2 s off in an ascending order, and the subject will be asked to give a pain rating for each stimulus on a scale from 0-10 (where 0 indicates ''no pain'', and 10 indicates the ''worst imaginable pain'') on a NRS.

Thermal cold and heat stimuli will be applied by a contact heat-evoked potential stimulator (PATHWAYS; Medoc Ltd, Ramat Yishai, Israel), placed 5 cm distal to the elbow on the right dorsal forearm

Thermal cold and heat stimuli will be applied by a contact heat-evoked potential stimulator (PATHWAYS; Medoc Ltd, Ramat Yishai, Israel), placed 5 cm distal to the elbow on the right dorsal forearm

Thermal cold and heat stimuli will be applied by a contact heat-evoked potential stimulator (PATHWAYS; Medoc Ltd, Ramat Yishai, Israel), placed 5 cm distal to the elbow on the right dorsal forearm

Thermal cold and heat stimuli will be applied by a contact heat-evoked potential stimulator (PATHWAYS; Medoc Ltd, Ramat Yishai, Israel), placed 5 cm distal to the elbow on the right dorsal forearm

The tests will be conducted by using a PATHWAY ATS (Medoc Ltd, Israel) thermal sensory testing device. A thermode stimulator of 3x3 cm will be placed on the treated/placebo areas and kept in place by means of Velcro tape. The subjects will rate the pain to three suprathreshold heat pain stimuli (starting and ending at 32°C with an increase and decrease of 5°C and 3 s plateau at 50°C).

The tests will be conducted by using a PATHWAY ATS (Medoc Ltd, Israel) thermal sensory testing device. A thermode stimulator of 3x3 cm will be placed on the treated/placebo areas and kept in place by means of Velcro tape. The subjects will rate the pain to three suprathreshold heat pain stimuli (starting and ending at 32°C with an increase and decrease of 5°C and 3 s plateau at 50°C).

Deep-tissue pain sensitivity will be evaluated by cuff pressure stimuli using a computer-controlled cuff algometer (Cortex Technology and Aalborg University, Denmark) including a 13-cm wide tourniquet cuff (VBM, Sulz, Germany) and an electronic visual analog scale (VAS) (Aalborg University, Denmark) for recording of the pain intensity.

Deep-tissue pain sensitivity will be evaluated by cuff pressure stimuli using a computer-controlled cuff algometer (Cortex Technology and Aalborg University, Denmark) including a 13-cm wide tourniquet cuff (VBM, Sulz, Germany) and an electronic visual analog scale (VAS) (Aalborg University, Denmark) for recording of the pain intensity.

The pressure will be increased by 1 kPa/s and the subject will be instructed to rate the pain intensity continuously on the electronic VAS until the tolerance level is reached.

The pressure will be increased by 1 kPa/s and the subject will be instructed to rate the pain intensity continuously on the electronic VAS until the tolerance level is reached.

A total of 10 repeated mechanical pressure stimuli at the PTT level will be delivered at 0.5 Hz (1 s stimulus duration and 1 s interval between stimuli) to the lower leg. A constant pressure of 5 kPa will be applied between the individual pressure stimuli to avoid movement of the cuff. During the 10 repeated stimuli, the subject will continuously rate the pain intensity on a 10 cm continuous VAS.

A total of 10 repeated mechanical pressure stimuli at the PTT level will be delivered at 0.5 Hz (1 s stimulus duration and 1 s interval between stimuli) to the lower leg. A constant pressure of 5 kPa will be applied between the individual pressure stimuli to avoid movement of the cuff. During the 10 repeated stimuli, the subject will continuously rate the pain intensity on a 10 cm continuous VAS.

The concept of CPM is that a tonic painful stimulus (conditioning stimulus) will inhibit pain evoked simultaneously from another site (test stimulus). The painful conditioned stimulus will be applied simultaneously with the assessment

• Page 4 of 6 [DRAFT] - stimulus. The conditioned stimulus will be terminated right after the subject presses the stop button. CPM will be defined as the difference between stimulus during and before the conditioned pain (i.e., "during" minus "before").

The concept of CPM is that a tonic painful stimulus (conditioning stimulus) will inhibit pain evoked simultaneously from another site (test stimulus). The painful conditioned stimulus will be applied simultaneously with the assessment

• Page 4 of 6 [DRAFT] - stimulus. The conditioned stimulus will be terminated right after the subject presses the stop button. CPM will be defined as the difference between stimulus during and before the conditioned pain (i.e., "during" minus "before").

The itch catastrophizing scale is a modified version of the pain catastrophizing scale (PCS), which is a validated questionnaire measuring pain-related thoughts and feelings

The itch catastrophizing scale is a modified version of the pain catastrophizing scale (PCS), which is a validated questionnaire measuring pain-related thoughts and feelings

The PSQI consists of 19 items generating seven component scores: subjective sleep quality, sleep latency, sleep duration, habitual sleep efficiency, sleep disturbances, use of sleeping medication, and daytime dysfunction.

The PSQI consists of 19 items generating seven component scores: subjective sleep quality, sleep latency, sleep duration, habitual sleep efficiency, sleep disturbances, use of sleeping medication, and daytime dysfunction.

The HADS consists of 14 items with two 7-item subscales measuring anxiety and depression symptoms, respectively

The HADS consists of 14 items with two 7-item subscales measuring anxiety and depression symptoms, respectively

This 20-item instrument is a psychometric self-report test to assess affect (10 items for positive affect and 10 items for negative affect) and validated for the use in adolescents and adults

This 20-item instrument is a psychometric self-report test to assess affect (10 items for positive affect and 10 items for negative affect) and validated for the use in adolescents and adults

Whole blood samples will be collected from the antecubital vein using a vacutainer blood collection device (SMonovette, Sarstedt) with a 21-gauge needle.

Whole blood samples will be collected from the antecubital vein using a vacutainer blood collection device (SMonovette, Sarstedt) with a 21-gauge needle.