Effects of Acute Intermittent Hypoxia on Sensory Function in Healthy Adults
Phase: NA
Conditions: Sensitivity
Sponsor: University of Florida

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Protocol 
1. Project Title 
Effects of acute intermittent hypoxia on sensory function in healthy adults 
2. Investigator(s): 
 
Mark Bishop 
Joel Bialosky 
Shakeel Ahmed 
3. Abstract: 
Exposure to acute intermittent hypoxia (AIH) has safely induced facilitation of motor output in 
human studies. Despite this study, the affects of AIH on sensory function has not been 
established. We will use a commercially-available hypoxicator that can be programmed to 
achieve a targeted level of arterial oxygen saturation. We plan to compare 3 previously 
published protocols that have generated motor facilitation (and a sham condition) to 
characterize and effects on sensory function in healthy adults. Primary outcomes will be 
measures of sensory function (thresholds and tolerance). 
4.  Background: 
 
Brief intermittent episodes of hypoxia (or acute intermittent hypoxia [AIH]) induce plasticity 
in muscle motor neurons [1]. AIH activates carotid chemo afferents that stimulate episodic 
serotonin release [2, 3] which subsequently strengthens synaptic input and motor output of 
motor nuclei [3-8]. The persistent increase in phrenic nerve burst amplitude following 
repeated exposure to AIH is termed long-term facilitation (LTF) [9]. For example, increases 
in breathing capacity as demonstrated by a significant increase in minute volume has been 
demonstrated after exposure to AIH in unanesthesized and anesthetized rats [10, 11].  
 
AIH has been demonstrated to be safe in healthy persons and has been routinely used in 
training athletes for many years. Similarly AIH is safe for people with neurological 
dysfunction (eg patients with chronic spinal cord injury (SCI)). In these patients, a single 
exposure to IH increased ankle plantar flexion torque by more than 80% [12]. Significant 
improvements in 10 meter walk times and 6 minute walk distances have been seen in 
patients with chronic SCI when IH was combined with walking at maximal sustainable 
exertion for 30 minutes [13]. 
Despite this promising work, the effect of AIH on sensory function has not been determined 
in humans, making this proposal a logical progression of the promising work done to date in 
improving motor function. 

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5. Specific Aims: 
Our aim is to characterize how exposure to AIH protocols shown to enhance motor function 
impacts sensory function. 
 
6. Research Plan: 
Participants who meet the eligibility criteria and provide written consent to participate will be 
enrolled.  This includes a pregnancy test for female volunteers that will be performed on the 
first day. Each participant will complete 3 different protocols of AIH delivery based on 
protocols derived from the literature that have been used to enhance motor function as well 
as a sham condition. Each delivery and testing session will last approximately 2 hours and 
be separated by at least 24 hours. 
 
Inclusion criteria 
Male and female participants between the ages of 18-40 years will be eligible to participate 
in the study unless they meet any of the following criteria. 
Exclusion criteria 
a. Diagnosis of cardiovascular disease (Hypertension, arrhythmias, coronary artery 
disease, congenital and valvar heart diseases) 
b. Diagnosis of neuromuscular disease 
c. Diagnosis of any neurological disease 
d. Presence of concurrent medical illness including infection, fractures 
e. Diagnosis of obstructive sleep apnea 
f. Diagnosis of obstructive/restrictive lung disease 
g. Diagnosis of exercise induced asthma 
h. FEV1/FVC<80% and/or FVC<80% of predicted value indicating airway obstruction 
i. Subjects on prednisolone therapy or selective serotonin reuptake inhibitor (SSRI) therapy 
will be excluded from the study as these pharmacological agents are known to amplify the 
effects of IH[22, 23] 
j. Diagnosis of epilepsy or history of seizures and attention deficit disorders 
k. Pregnancy 
l. Diabetes 
m. History of coagulation disorders 
n. History of chronic pain  
o. Body mass index(BMI)> 35kg/m2  
p. Subjects on prescription medicines such as beta blockers and other drugs that are 
prescribed in any of the exclusionary disorders listed above. 
 
Intervention 
We will use a commercially-available hypoxicator that can be programmed to achieve a 
targeted level of arterial oxygen saturation. The fraction of inspired oxygen (FiO2) during 
the hypoxic episodes will be adjusted to maintain pulse oximetry (SPO2) between 80%- 

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90%. In our experience, this can be achieved with a FiO2 in the range of 9%-11 %. FiO2 
levels of 9% and 11% correspond to O2 concentrations in room air at altitudes of 6500 
meters and 5000 meters respectively, both altitudes humans are exposed to while on 
trekking expeditions and also have permanent human habitation [29, 30]. We will use 
alternating phases of hypoxia (FiO2 :9%-11%) and normoxic or hyperoxic oxygen 
saturation with an FiO2 between 22%-38%,based on the previous protocols used in human 
studies of motor function. 
 
A. 15 bouts of hypoxia for 1 minute, normoxia to hyperoxia 1 minute (Trumbower et al 
[12]) 
B. 15 bouts of hypoxia for 2 minutes, normoxia to hyperoxia  1 minute (Hayes et al [13]) 
C. 8 bouts of hypoxia for 2 minutes, normoxia to hyperoxia  1 minute (Tester et al [14]) 
D. Sham hyperoxia (normoxia) for 2 minutes, normoxia to hyperoxia  1 minute 
 
The order of the protocol will be randomly assigned for each participant who will remain 
blinded to the actual protocol they are receiving. All measures will be collected by a 
research assistant blinded to protocol assignment. 
 
Outcomes 
We propose to collect standardized measures of sensory function. Measures will assessed 
at the hand and foot at baseline, immediately post- and every 10 minutes for 60 minutes 
after the AIH protocol.  
 
Tactile: 
Semmes-Weinstein filaments will be used to assess sensory thresholds. The patient in will 
be in sitting position with shoes and socks removed. Starting with the weakest plastic kept 
perpendicular to the skin surface force will slowly be applied until there is a slight bend in 
the filament. The participant will indicate if they feel pressure. If not, the next filament is 
used and testing proceeds until the thinnest fiber that the participant can perceive is 
reached. 
 
Pain sensitivity: 
Participants will undergo quantitative sensory testing (QST) to determine pain sensitivity to 
thermal and pressure stimuli, as per protocols in our preliminary studies.  
 
Study participants will undergo standard psychophysical pain testing using a contact 
thermode to deliver the evoked, thermal pain stimuli. The stimuli are to be applied by a 
research assistant who ensures proper thermode application and the range of stimulus 
intensities to be used (40-51°C) will be presented beforehand in ascending one-degree 
steps to each subject. We have found this procedure to be useful because it familiarizes 
subjects with the stimulus range, tends to obviate range effects in psychophysical scaling, 
and helps alleviate subject anxiety about the upper limit of stimulus intensities to be used. 
In order to standardize the scaling instructions and to clarify the distinction between the 
sensory intensity and affective dimensions of pain, a standardized instructional set will be 
used for all subjects. The research assistant will record numeric rating scale (NRS) 
response to each stimulus. The NRS will consist of a scale whose endpoints are 

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designated as '0 - no pain sensation' and '100 - the most intense pain sensation 
imaginable'.  In order to standardize the scaling instructions, standard instructions will be 
used for all participants.   
 
Threshold and Tolerance.  A continuous heat stimulus will be delivered to the subjects’ 
dominant arm.  The stimulus started at 35°C and will be increased at a rate of 0.5°C with 
subjects terminating the stimulus when the temperature reached pain threshold (“when the 
sensation first transitions from heat to pain”) and tolerance (“when the sensation becomes 
so strong you want to remove it from your skin”).  These procedures will be repeated three 
times and the average threshold and tolerance will be calculated. 
 
Threshold and tolerance measures will also be collected using mechanical pressure 
applied using a Fischer Dolorimeter (Pain Diagnostics, Great Neck, NY). The tip of the 
dolorimeter is equipped with a rubber foot-plate of 1-cm diameter. During Dolorimeter 
testing, force is slowly increased until the subject indicates that the sensation changes from 
pressure to pain. Subjects are asked to rate any pain they are experiencing using a 
numeric rating scale (NRS) anchored at 0 (no pain sensation at all) and 100 (worst pain 
imaginable). Dolorimetry measures are made in the hand (1st dorsal interosseus muscle) 
and between the first and second toe on the dorsal aspect of the foot. 
 
Temporal summation.  A train of 6 heat pulses will be applied to the glabrous skin of the 
foot.  To ensure temporal summation, an inter-stimulus interval of 2 seconds will be used 
with temperatures starting at 39°C and increasing to 50°C (the TSA medoc maintains 
10°C/sec rate in this range which is sufficient for maintaining the desired inter-stimulus 
interval).  The participants will be asked to rate the magnitude of their delayed (second) 
pain sensation following each heat pulse.  These response ratings are believed to be 
primarily C-fiber mediated.35,36  
 
We will test effects protocol using a two way (protocol and time) analysis of covariance with 
sensory measures as the dependent variables. Demographic variables that show an 
association with the sensory measures in our sample (eg gender differences have mixed 
evidence related to differences in sensory report) will be included as covariates. 
7. Possible Discomforts and Risks: 
 
Evoked pain: Risk of injury with this testing is minimal in that they are widely used and safe 
procedures.  While pain is produced, risk to the individual is minimal, because 1) the pain is 
transient in nature and generally subsides immediately after the procedure; 2) subjects are 
instructed that they may stop any procedure at any time with no adverse consequences; 3) 
the level of pain experienced by subjects is below their tolerance level; 4) this pain will only 
last as long as the stimulus is applied (which is under the control of the study participant); 
5) thermal stimulus intensity will not cause any damage to the study participant’s skin.  
Some study participants will feel pain after the painful stimulus is removed, but no long-
lasting pain perception (i.e. longer than 90s) is expected from quantitative sensory testing.  
Similarly a slight risk of burn is involved using thermal stimuli. The thermal stimulator 

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(Medoc TSA) has a built in shut of threshold that prevents the delivery of temperatures over 
51 deg C. Our group has completed more than 1000 testing sessions in the past 5 years 
without any adverse events. 
 
Acute intermittent hypoxia: Our experiment is modeled from past human studies that have 
delivered similar doses of hypoxia that was considered safe and sufficient to elicit an 
increase in physiological response [12, 13, 14]. Moderate reductions in inspired oxygen can 
cause light- headedness, dizziness, reduced vision, and euphoria. We will continuously 
monitor the subject’s heart rate, respiratory rate, and oxygen saturation (SPO2) to ensure 
that the administered oxygen levels remain within the targeted range (80- 85%). In an 
unlikely circumstance where in the heart rate drops below 50 BPM or rises more than 150 
BPM and/or systolic blood pressure drops below 85mm Hg or increases beyond 150mmHg, 
IH intervention will be stopped. If SPO2 falls below 75% or if subjects complain of light 
headedness, dizziness and reduced vision, the IH intervention will be terminated and 
supplemental O2 will be provided. In addition, the hypoxicator has a built in, feedback 
based, safety mechanism. In a circumstance of desaturation below a pre-set threshold of 
80%, the hypoxicator switches to deliver hyperoxic air to increase saturation above the 
minimum set threshold of 80%. 
 
8. Possible Benefits: 
There is no direct benefit expected for any of the participants. Potential future benefits to 
the public include an increased knowledge of the effects of IH, which may eventually lead 
to the future treatments for neuromotor diseases. 
9. Conflict of Interest: 
We declare no conflicts of interest. 
 
References 
1. Gonzalez-Rothi, E.J., et al., Intermittent Hypoxia and Neurorehabilitation. J Appl 
Physiol (1985), 2015: p. jap 00235 2015.   
2. MacFarlane, P.M. and G.S. Mitchell, Episodic spinal serotonin receptor activation 
elicits long-lasting phrenic motor facilitation by an NADPH oxidase-dependent 
mechanism. J Physiol, 2009. 587(Pt 22): p. 5469-81.   
3. Fuller, D.D., et al., Synaptic pathways to phrenic motoneurons are enhanced by 
chronic intermittent hypoxia after cervical spinal cord injury. J Neurosci, 2003. 23(7): 
p. 2993- 3000.   
4. Baker-Herman, T.L., et al., BDNF is necessary and sufficient for spinal respiratory 
plasticity following intermittent hypoxia. Nature neuroscience, 2004. 7(1): p. 48-55.   

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5. Lovett-Barr, M.R., et al., Repetitive intermittent hypoxia induces respiratory and 
somatic motor recovery after chronic cervical spinal injury. J Neurosci, 2012. 32(11): 
p. 3591-600.   
6. Satriotomo, I., et al., Repetitive Acute Intermittent Hypoxia Increases BDNF and 
TrkB Expression in Respiratory Motor Neurons: Dose Effects. Faseb Journal, 2010. 
24.   
7. Satriotomo, I., et al., Repetitive acute intermittent hypoxia increases expression of 
proteins associated with plasticity in the phrenic motor nucleus. Experimental 
neurology, 2012. 237(1): p. 103-115.   
8. Golder, F.J. and G.S. Mitchell, Spinal synaptic enhancement with acute intermittent 
hypoxia improves respiratory function after chronic cervical spinal cord injury. The 
Journal of neuroscience, 2005. 25(11): p. 2925-2932. 
9. Hayashi, F., et al., Time-dependent phrenic nerve responses to carotid afferent 
activation: intact vs. decerebellate rats. Am J Physiol, 1993. 265(4 Pt 2): p. R811-9. 
  
10. Fuller, D.D., et al., Long term facilitation of phrenic motor output. Respir Physiol, 
2000. 121(2-3): p. 135-46.   
11. Olson, E.B., Jr., et al., Ventilatory long-term facilitation in unanesthetized rats. J Appl 
Physiol (1985), 2001. 91(2): p. 709-16.   
12. Trumbower, R.D., et al., Exposure to acute intermittent hypoxia augments somatic 
motor function in humans with incomplete spinal cord injury. Neurorehabil Neural 
Repair, 2012. 26(2): p. 163-72.   
13. Hayes, H.B., et al., Daily intermittent hypoxia enhances walking after chronic spinal 
cord injury: a randomized trial. Neurology, 2014. 82(2): p. 104-13.   
14. Tester, N.J., et al., Long-term facilitation of ventilation in humans with chronic spinal 
cord injury. Am J Respir Crit Care Med, 2014. 189(1): p. 57-65.