Hyperthermia modulates regional differences in cerebral blood flow to changes in CO 2

Shigehiko Ogoh, Kohei Sato, Kazunobu Okazaki, Tadayoshi Miyamoto, Ai Hirasawa and Manabu Shibasaki J Appl Physiol 117:46-52, 2014. First published 1 May 2014; doi:10.1152/japplphysiol.01078.2013 You might find this additional info useful... This article cites 40 articles, 25 of which can be accessed free at: /content/117/1/46.full.html#ref-list-1 This article has been cited by 1 other HighWire hosted articles Forehead versus forearm skin vascular responses at presyncope in humans Daniel Gagnon, R. Matthew Brothers, Matthew S. Ganio, Jeffrey L. Hastings and Craig G. Crandall Am J Physiol Regul Integr Comp Physiol, October 1, 2014; 307 (7): R908-R913. [Abstract] [Full Text] [PDF] Updated information and services including high resolution figures, can be found at: /content/117/1/46.full.html

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J Appl Physiol 117: 46–52, 2014. First published May 1, 2014; doi:10.1152/japplphysiol.01078.2013.

Hyperthermia modulates regional differences in cerebral blood flow to changes in CO2 Shigehiko Ogoh,1 Kohei Sato,2 Kazunobu Okazaki,3 Tadayoshi Miyamoto,4 Ai Hirasawa,1 and Manabu Shibasaki5 1

Department of Biomedical Engineering, Toyo University, Saitama, Japan; 2Research Institute of Physical Fitness, Japan Women’s College of Physical Education, Tokyo, Japan; 3Department of Environmental Physiology for Exercise, Osaka City University Graduate School of Medicine, Osaka, Japan; 4Morinomiya University of Medical Sciences, Osaka, Japan; and 5 Department of Environmental Health, Nara Women’s University, Nara, Japan Submitted 25 September 2013; accepted in final form 30 April 2014

arterial pressure; cardiac output; hyperthermia; Doppler ultrasound; humans ORTHOSTATIC TOLERANCE is impaired in hyperthermic conditions (21, 34, 39). Various hyperthermia-induced alterations, including decreased peripheral vascular resistance and ventricular filling pressures, altered arterial baroreflex control, and reduced cerebral perfusion, are thought to contribute to this impairment (39). We recently demonstrated that blood flow distribution toward the head is altered during heat stress and found that external carotid artery (ECA) blood flow increased whereas cerebral blood flow, including internal carotid artery (ICA) and vertebral artery (VA) blood flows, decreased (4, 28). Any condition that jeopardizes cerebral blood flow (CBF) regulation can lead to syncope (36); thus the maintenance of cerebral perfusion is critical for preserving brain function. Therefore,

Address for reprint requests and other correspondence: M. Shibasaki, Laboratory for Exercise and Environmental Physiology, Dept. of Environmental Health, Nara Women’s Univ., Kita-uoya Nishi-machi, Nara 630-8506 Japan (e-mail: [email protected]). 46

CBF regulation during heat stress has been investigated (7, 8, 11, 22, 23, 25, 40), but the mechanism responsible for impaired CBF regulation during an orthostatic intolerance combined with heat stress remains unclear. CBF regulation is different from the regulation of other regional circulation, such as renal, splanchnic, muscle, and cutaneous blood flows (26), and has unique mechanisms: dynamic cerebral autoregulation and cerebrovascular response to carbon dioxide (CO2) (i.e., CO2 reactivity). Specific mechanisms in the cerebral circulation need to be identified to understand the characteristics of dynamic CBF regulation. It is recognized that hyperthermia induces hypocapnia following hyperventilation (37, 39). Therefore, the cerebrovascular response to CO2 is an important mechanism for dynamic CBF regulation during hyperthermia. Previous reports have clearly demonstrated that heat stress does not affect the slope of the relationship between changes in middle cerebral artery mean blood velocity (MCA Vmean) and end-tidal CO2 partial pressure (PETCO2) (11, 23). However, several studies demonstrated that despite clamped end-tidal CO2 (PETCO2) at preheat level, the heat stress-induced decreased in MCA Vmean was increased although it remained lower than the normothermic baseline (7, 13). These findings suggest that heat stress alters the cerebrovascular response to CO2. In contrast, it was reported recently that clamping PETCO2 at the preheat level returned ICA blood flow and MCA Vmean to the preheat baseline (4). Although methodological differences may be responsible for this discrepancy, small changes in PETCO2 (⬃5–7 mmHg) observed in the previous study (4) compared with the functional range of CO2 reactivity (20 –50 mmHg) (30) might influence cerebrovascular responses to CO2 during heat stress. Moreover, no study has investigated the cerebrovascular response of further hypocapnia during heat stress. Thus the influence of heat stress on the cerebrovascular response to CO2 remains unclear. We demonstrated that CO2 reactivity in intra- and extracranial arteries was not uniform with greater CO2 reactivity in ICA and lesser CO2 reactivity in the ECA relative to in the ICA and VA (30). Given these observations, coupled with the increased ECA blood flow during heat stress for thermoregulation, altered blood distribution due to heat stress may be associated with an alteration in CBF regulation. We hypothesized that heat stress modifies intra- and extracranial blood flow responsiveness to changes in CO2. To test our hypothesis, we measured blood flow and its response to changes in PETCO2 in the ICA, ECA, and VA, as well as MCA Vmean during normothermia and following passive heat stress.

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Ogoh S, Sato K, Okazaki K, Miyamoto T, Hirasawa A, Shibasaki M. Hyperthermia modulates regional differences in cerebral blood flow to changes in CO2. J Appl Physiol 117: 46 –52, 2014. First published May 1, 2014; doi:10.1152/japplphysiol.01078.2013.—The purpose of this study was to assess blood flow responses to changes in carbon dioxide (CO2) in the internal carotid artery (ICA), external carotid artery (ECA), and vertebral artery (VA) during normothermic and hyperthermic conditions. Eleven healthy subjects aged 22 ⫾ 2 (SD) yr were exposed to passive whole body heating followed by spontaneous hypocapnic and hypercapnic challenges in normothermic and hyperthermic conditions. Right ICA, ECA, and VA blood flows, as well as left middle cerebral artery (MCA) mean blood velocity (Vmean), were measured. Esophageal temperature was elevated by 1.53 ⫾ 0.09°C before hypocapnic and hypercapnic challenges during heat stress. Whole body heating increased ECA blood flow and cardiac output by 130 ⫾ 78 and 47 ⫾ 26%, respectively (P ⬍ 0.001), while blood flow (or velocity) in the ICA, MCA, and VA was reduced by 17 ⫾ 14, 24 ⫾ 18, and 12 ⫾ 7%, respectively (P ⬍ 0.001). Regardless of the thermal conditions, ICA and VA blood flows and MCA Vmean were decreased by hypocapnic challenges and increased by hypercapnic challenges. Similar responses in ECA blood flow were observed in hyperthermia but not in normothermia. Heat stress did not alter CO2 reactivity in the MCA and VA. However, CO2 reactivity in the ICA was decreased (3.04 ⫾ 1.17 vs. 2.23 ⫾ 1.03%/mmHg; P ⫽ 0.039) but that in the ECA was enhanced (0.45 ⫾ 0.47 vs. 0.95 ⫾ 0.61%/mmHg; P ⫽ 0.032). These results indicate that hyperthermia is capable of altering dynamic cerebral blood flow regulation.

Cerebral CO2 Reactivity during Heat Stress METHODS

Eleven healthy male subjects with a mean age of 21 ⫾ 1 yr (mean ⫾ SD), height of 173 ⫾ 7 cm, and weight of 67 ⫾ 12 kg participated in this study voluntarily. We originally enrolled 12 volunteers, but 1 subject could not complete the experiment. A part of this project has been published in another journal (28), but the data collected and represented have not been duplicated. Each subject provided written, informed consent after all procedures and potential risks were explained. All experimental procedures and protocols conformed to the Declaration of Helsinki and were approved by the Human Subjects Committee of Nara University. The subjects were free of any known cardiovascular or pulmonary disorders and were not using any prescribed or over-the-counter medications. In addition, they were not engaged in endurance training on a regular basis (⬍5 h/wk). Prior to the experiment, each subject gave written informed consent and visited the laboratory for familiarization with the techniques and procedures. Subjects were requested to abstain from caffeinated beverages for 12 h and strenuous physical activity and alcohol for at least 24 h before the day of the experiment. Experimental Protocol

Modulation of Respiration to Change End-Tidal PCO2 To evaluate CO2 reactivity in each artery (ICA, MCA, ECA, and VA), subjects underwent hypocapnic and hypercapnic challenges for ⬃8 min each. The subjects were allowed a few minutes of rest after both challenges. The three conditions (baseline, hypocapnic, and hypercapnic conditions) were performed in random order. For the hypocapnic challenge, each subject gradually increased their respiratory frequency by adjusting their respiration to the pace of a metronome. By monitoring with a respiratory gas analyzing system (ARCO2000-MET, Arcosystem, Chiba, Japan), the pace of the metronome was adjusted by a coordinator. For the hypercapnic challenge, the subjects respired through 500 ml of dead space. The coordinator indicated their respiratory frequency and depth. Following the 8 min required to reach steady state under each condition (baseline, hypocapnic, and hypercapnic conditions), blood flow was measured. CO2 reactivity was identified as the percent change in each blood flow per millimeter Hg change in PETCO2 by linear regression analysis. Instrumentation and Measurements Thermoregulatory and hemodynamic variables. Tes and skin temperature were measured using thermocouples. For the measurement of

Ogoh S et al.

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Tes, each subject inserted a thermocouple via the nasal passage to a distance equivalent to one-fourth of the subject’s height. Skin temperatures were measured at six sites (chest, upper back, abdomen, lower back, thigh, and lower leg), while mean skin temperature (Tsk) was calculated from the weighted average of these six points (35). Heart rate was obtained from an electrocardiogram (Biomulti 1000, NEC). Continuous beat-by-beat arterial blood pressure was recorded from a finger using the Penaz method (Portapres, Finapres Medical Systems). Intermittent arterial blood pressure was also measured by auscultation of the brachial artery via electrosphygmomanometry (STBP-780, Colin, Japan). Skin blood flux was measured via laserDoppler flowmetry using an integrating flow probe (moor VMSLDF2, Moor Instruments, UK) attached to the forehead (uncovered) and forearm (covered by the suit). Respiratory variables. The subject breathed through a mouthpiece attached to a pressure flowmeter. Respiratory and metabolic variables throughout the experiment were recorded using an automatic breathby-breath respiratory gas analyzing system (ARCO2000-MET, Arcosystem, Chiba, Japan). We digitized expired flow, CO2 and oxygen (O2) concentrations, and derived tidal volume (VT), respiratory rate ˙ E), and PETCO2. Flow signals were com(RR), minute ventilation (V puted to single breath data, and matched to gas concentrations identified as single breaths using PETCO2, after accounting for the time lag (350 ms) in gas concentration measurements. The corresponding O2 ˙ CO2) values for each breath were calculated uptake and CO2 output (V from inspired-expired gas concentration differences, and by expired ventilation, with inspired ventilation being calculated by nitrogen ˙ E and PETCO2 were recorded correction. During each protocol, V continuously at 200 Hz (ARCO2000-MET, Arcosystem). Intra- and extracranial blood flows. Right ICA, ECA, and VA blood flow was measured using two color-coded ultrasound systems (Vivid-e; GE Healthcare, Tokyo) equipped with a 10-MHz linear transducer. ICA and ECA measurements were performed ⬃1.0 –1.5 cm distal to the carotid bifurcation on the right side while the subject’s chin was slightly elevated. VA was measured between the transverse processes of the C3 and subclavian artery. An experienced sonographer measured all three sites in a random order, and first used the brightness mode to measure the mean diameter of each vessel in a longitudinal section, and the Doppler velocity spectrum was subsequently identified using pulsed-wave mode. When taking blood flow velocity measurements, care was taken to ensure that the probe position was stable, that the insonation angle did not vary (⬃60° in most cases), and that the sample volume was positioned in the center of the vessel and adjusted to cover the width of the vessel diameter. Middle cerebral artery mean blood velocity. Left MCA Vmean was measured continuously using a 2-MHz pulsed transcranial Doppler ultrasound system (WAKI, Atys Medical, France). A Doppler probe was adjusted over the temporal window until an optimal signal was identified. The probe was then fixed and held in place using a headband strap. Cardiac output. The best echocardiography image for each subject was selected at each stage for cardiac output (Q) analysis. Stroke volume (SV) was estimated from aortic flow, calculated as the product of beat-to-beat measurements of the Doppler velocity time integral (VTI) and aortic root diameter (Vivid-i; GE Healthcare, Tokyo, Japan). Aortic root diameter was determined using two-dimensional echocardiogram imaging obtained in the left parasternal long-axis view with the subject in the supine position, and SV was calculated as SV ⫽ ␲(D/2)2 ⫻ VTI. Q was calculated from the product of SV and HR. Data Analysis MCA blood velocity and thermoregulatory and hemodynamic variables were continuously measured, and sampled at 1 kHz via a data-acquisition system (MP150, BIOPAC Systems). Data from the

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Experiments were performed in a temperature-controlled laboratory (26 ⫾ 1°C). Subjects arrived at the laboratory at least 2 h after a light meal. Each subject, wearing underwear and short pants, was dressed in a tube-lined water-perfused suit (Med-Eng, Ottawa, Canada). The water-perfused suit covered the entire body, except the head, face, hands, and feet. The zipper of the upper suit was opened during measurement of cardiac echography. After instrumentation, the subjects laid supine on a bed for ⬃30 min under normothermic conditions, while breathing room air through a mouthpiece. During this period, 33°C water was perfused through the suit to maintain a thermoneutral condition. After this equilibration period, baseline data were collected for 10 min during spontaneous respiration, followed by cardiac output (Q) measurement via Doppler techniques. The subjects then performed hypocapnic and hypercapnic challenges in a random order (see below). Subjects were exposed to heat stress through perfusion of 50°C water through the suit for ⬃45–90 min, a time sufficient to increase esophageal temperature (Tes) ⬃1.5°C. Once Tes reached this target level, the temperature of the water perfusion suit was manipulated to prevent further increases in Tes during the ensuing data collection periods. As mentioned above, one subject was unable to undergo the hypocapnic and hypercapnic challenges during heat stress; thus 11 subjects were included in the final analysis.

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Cerebral CO2 Reactivity during Heat Stress

last 120 s of each period (hypo-, iso-, and hypercapnia at each thermal condition) were averaged for steady-state statistical analyses. For the data collection of intra- and extracranial blood flows, blood velocity in each artery was stored for at least 30 s, and mean blood velocities were calculated from ⬃10 –20 cardiac cycles. For intra- and extracranial blood flows calculation, the systolic and diastolic diameters were measured, and then the mean diameter (cm) was calculated in relation to the blood pressure curve: mean diameter ⫽ (systolic diameter ⫻ 1/3) ⫹ (diastolic diameter ⫻ 2/3). The time-averaged mean flow velocity obtained in pulsed-wave mode was defined as the mean blood flow velocity (cm/s). The measurements of blood flow velocity were determined from the average of ⬃10 –20 cardiac cycles to eliminate effects of the breathing cycle. Blood flow was calculated by multiplying the cross-sectional area [␲ ⫻ (mean diameter/2)2] with mean blood flow velocity; blood flow ⫽ mean blood flow velocity ⫻ area ⫻ 60 (ml/min). In addition, conductance was estimated from the ratio of MCA Vmean, ICA, ECA, or VA blood flow to MAP. Statistical Analysis

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Ogoh S et al.

⫹2.6 l/min (P ⬍ 0.001), respectively. Table 1 shows averaged responses of cardiovascular function and respiration during hypocapnia/hypercapnia challenges during normothermia and hyperthermia. Regardless of the thermal status, hypocapnia/ hypercapnia challenges did not alter cardiac output, although HR and MAP were changed. The hypocapnia/hypercapnia challenges did not change skin blood flux or cutaneous vascular conductance at both sites during normothermia. Heat stress increased skin blood flux from 83.4 ⫾ 33.7 AU to 215.6 ⫾ 55.1 AU (P ⬍ 0.001) on the forehead and from 16.0 ⫾ 11.3 AU to 153.1 ⫾ 60.2 AU (P ⬍ 0.001) on the forearm. During heat stress, hypocapnic challenge decreased skin blood flux on the forehead (215.6 ⫾ 55.1 AU to 202.0 ⫾ 58.5 AU, P ⫽ 0.011), but not on the forearm. In contrast, hypercapnic challenge increased skin blood flux at forearm (153.1 ⫾ 60.2 AU to 167.4 ⫾ 51.3 AU), but not on the forehead. Intra- and Extracranial Blood Flow Responses

RESULTS

Thermoregulatory, Respiratory, and Hemodynamic Variables During heat stress Tes and Tsk were increased from 37.02 ⫾ 0.23°C and 33.71 ⫾ 0.92°C to 38.55 ⫾ 0.26°C and 38.30 ⫾ 1.08°C, respectively (P ⬍ 0.001 for Tes and Tsk). During hypercapnia/hypocapnia challenges, Tes was maintained at the same level (hypocapnia trial, 1.51 ⫾ 0.10°C; hypercapnia trial, ˙ E ⫹3.1 1.54 ⫾ 0.10°C). Heat stress induced hyperventilation (V l/min, P ⫽ 0.013) and subsequent hypocapnia (PETCO2 ⫺5.9 mmHg, P ⬍ 0.001). MAP was maintained during heat stress, while HR and Q increased by ⫹31 beats/min (P ⬍ 0.001) and

In normothermic conditions, hypocapnic and hypercapnic challenges decreased and increased MCA Vmean, ICA, and VA blood flows, respectively (Fig. 1). However, neither challenge changed ECA blood flow. Although MAP was increased during the hypercapnic challenge, similar statistical results were obtained with these conductance data (Table 1). Heat stress decreased ICA (⫺17.2%, P ⬍ 0.001) and VA (⫺11.7%, P ⫽ 0.002) blood flows and MCA Vmean (⫺24.0%, P ⬍ 0.001). However, ECA blood flow was increased from 196 ⫾ 69 to 439 ⫾ 195 ml/min (⫹124.1%, P ⬍ 0.001). During heat stress, hypocapnic challenge decreased ECA blood flow, as well as MCA Vmean, and ICA and VA blood flows, while these were increased by the hypercapnic challenge (Fig. 1). However, CO2 reactivity in the MCA was unchanged during heat stress (Fig. 2A, P ⫽ 0.08) and decreased significantly in the ICA (Fig. 2B, P ⫽ 0.039). CO2 reactivity in the ECA was significantly lower than that in the ICA during both normothermic (P ⬍ 0.001) and heat-stressed conditions (P ⫽ 0.010), but CO2 reactivity in the ECA was significantly increased in the heat-stressed condition (Fig. 2C, P ⫽ 0.032). However, CO2 reactivity in the VA was unchanged during heat stress (Fig. 2D, P ⫽ 0.258).

Table 1. Cardiorespiratory and cerebrovascular hemodynamic responses during respiratory challenges in normothermia and hyperthermia Normothermia

Cardiac output, l/min Stroke volume, ml HR, beats/min MAP, mmHg PETCO2, mmHg ˙ E, l/min V MCA Vmean conductance, cm·s⫺1·mmHg⫺1 ICA conductance, ml·min⫺1·mmHg⫺1 ECA conductance, ml·min⫺1·mmHg⫺1 VA conductance, ml·min⫺1·mmHg⫺1

Hyperthermia

Hypocapnia

Control

Hypercapnia

Hypocapnia

Control

Hypercapnia

6.4 ⫾ 1.5 89.5 ⫾ 17.8 71.8 ⫾ 8.8 90.8 ⫾ 7.9 27.3 ⫾ 4.1 23.8 ⫾ 4.9

5.8 ⫾ 1.6 91.1 ⫾ 18.8 63.8 ⫾ 10.1† 89.5 ⫾ 8.9 40.6 ⫾ 2.9† 9.2 ⫾ 3.4†

5.9 ⫾ 1.4 88.7 ⫾ 16.5 66.8 ⫾ 10.7† 94.2 ⫾ 9.8†‡ 46.5 ⫾ 3.4†‡ 12.9 ⫾ 6.7†

8.6 ⫾ 2.2 83.0 ⫾ 17.7 104.4 ⫾ 14.3 82.9 ⫾ 5.0 23.9 ⫾ 3.1 26.8 ⫾ 7.3

8.4 ⫾ 2.2 87.8 ⫾ 19.8 95.2 ⫾ 11.1† 87.6 ⫾ 7.9† 34.7 ⫾ 5.7† 12.3 ⫾ 4.5†

8.1 ⫾ 2.0 81.9 ⫾ 17.6 99.5 ⫾ 13.1†‡ 86.0 ⫾ 6.6 43.5 ⫾ 3.4†‡ 16.0 ⫾ 7.4†

Thermal

P P P P P P

⬍ ⫽ ⬍ ⫽ ⬍ ⬍

0.001 0.110 0.001 0.009 0.001 0.001

Trial

P P P P P P

⫽ ⫽ ⬍ ⫽ ⬍ ⬍

0.117 0.095 0.001 0.038 0.001 0.001

Interaction

P P P P P P

⫽ ⫽ ⫽ ⫽ ⫽ ⫽

0.721 0.694 0.823 0.017 0.029 0.999

0.37 ⫾ 0.11 0.58 ⫾ 0.20† 0.67 ⫾ 0.22†‡

0.35 ⫾ 0.12 0.45 ⫾ 0.17† 0.56 ⫾ 0.21†‡ P ⫽ 0.002 P ⬍ 0.001 P ⫽ 0.006

2.36 ⫾ 0.77 3.26 ⫾ 0.90† 4.18 ⫾ 1.12†‡

2.55 ⫾ 0.70 2.75 ⫾ 0.88

3.65 ⫾ 1.14†‡ P ⫽ 0.014 P ⬍ 0.001 P ⫽ 0.002

1.99 ⫾ 0.68 2.20 ⫾ 0.80

4.70 ⫾ 1.98 5.00 ⫾ 2.19

5.43 ⫾ 2.38†‡ P ⬍ 0.001 P ⫽ 0.004 P ⫽ 0.025

0.83 ⫾ 0.40 0.92 ⫾ 0.44

1.07 ⫾ 0.45†‡ P ⫽ 0.207 P ⬍ 0.001 P ⫽ 0.124

2.12 ⫾ 0.81

0.80 ⫾ 0.40 1.02 ⫾ 0.46† 1.14 ⫾ 0.56†‡

˙ E, minute ventilation; MCA Vmean, middle cerebral Values are means ⫾ SD. HR, heart rate; MAP, mean arterial pressure; PETCO2, end-tidal carbon dioxide; V artery mean blood velocity; ICA, internal carotid artery; ECA, external carotid artery; VA, vertebral artery. †P ⬍ 0.05 vs. hypocapnia. ‡P ⬍ 0.05 vs. control. J Appl Physiol • doi:10.1152/japplphysiol.01078.2013 • www.jappl.org

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Analyses were conducted using SigmaStat (Jandel Scientific Software, SPSS, Chicago, IL). Normal distribution was confirmed with the Shapiro-Wilk test. Data were subsequently analyzed using a repeated-measures two-way analysis of variance (ANOVA) test and Student-Newman-Keul post hoc tests. The effect of heat stress on the CO2 reactivity at each artery was compared using a priori one-tailed paired t-test. A two-tailed paired t-test was used for comparisons between conditions (e.g., normothermic control vs. hyperthermic hypercapnia). Data are expressed as means ⫾ SD, and significance was indicated at P ⬍ 0.05.

Cerebral CO2 Reactivity during Heat Stress

Normothermia Hyperthermia

*

90 80

*

70 50 40 30

*

450 350

*

Control Hypercapnia

*

300 250 200

-1

C

Hypocapnia

700 600

*

500 400 300 200

-1

D

4 3 2 1

Hypocapnia

180 160 140 120 100 80 60 40 20 0

Control Hypercapnia

*

* Hypocapnia

*

C

*

DISCUSSION

This study demonstrated that passive heat stress modified CO2 reactivity in intra- and extracranial blood flows and that the heat stress-induced alterations differed among arteries. Heat stress reduced ICA and VA blood flows as well as MCA Vmean. Similar to previous CBF studies, CO2 reactivity in MCA

Hyperthermia

P=0.039

6 5 4 3 2 1

Normothermia

Hyperthermia

6 5 4

D

P=0.032

3 2 1 0 -1

Control Hypercapnia

Fig. 1. Middle cerebral artery mean blood velocity (MCA Vmean; A) and internal carotid artery (ICA; B), external carotid artery (ECA; C) and vertebral artery (VA; D) blood flows during hypocapnic/hypercapnic challenges in normothermia and hyperthermia. Values are means ⫾ SD. *Significant differences from hypocapnia (P ⬍ 0.05); †significant differences from control (P ⬍ 0.05).

Normothermia

7

0

-1

100

B

P=0.08

5

*

Control Hypercapnia

*

6

0

-1

100

7

Normothermia

Hyperthermia

7 6

P=0.258

5 4 3 2 1 0

Normothermia

Hyperthermia

Fig. 2. CO2 reactivity in the MCA (A), ICA (B), ECA (C), and VA (D) during normothermia and hyperthermia. Values are mean ⫾ SD.

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*

150

A

-1

400

Hypocapnia

CO2 reactivity at MCA

500

*

CO2 reactivity at ECA

B -1

10

CO2 reactivity at ICA

20

-1

MCAVmean

60

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Vmean did not change during heat stress. CO2 reactivity in the VA was also unchanged, but that in the ICA was reduced. However, ECA blood flow increased during heat stress and CO2 reactivity in the ECA was enhanced. Thus heat stress caused regional differences in blood flow regulation to change in PETCO2. This altered CO2 reactivity as well as blood flow distribution due to heat stress may contribute to CBF regulation in hyperthermic individuals.

CO2 reactivity at VA

-1

A

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Cerebral CO2 Reactivity during Heat Stress

Regional Differences in Cerebrovascular Responses to Changes in PETCO2

Ogoh S et al.

diameter of the MCA might change with arterial PCO2 (PaCO2). Since ICA blood flow is distributed primarily by the MCA, as well as the anterior and posterior cerebral arteries across the circle of Willis, altered regional metabolic demands during heat stress might affect the blood distribution. The latter speculation is more likely correct, because Fierstra et al. (12) recently suggested that MCA Vmean responses to changes in PETCO2 may be inaccurate as an index of global CBF. They evaluated cerebrovascular reactivity (CVR) by means of blood oxygen level-dependent (BOLD) magnetic resonance imaging (MRI), and tracked voxel BOLD signals at two different sites, while PETCO2 was manipulated by using a computer-controlled gas blender. They showed opposite responses to change in PETCO2 at two sites. Therefore, the different responsiveness between MCA Vmean and ICA might arise from the regional differences in CVR during heat stress. Vertebral arteries are the other branches that distribute blood to the cerebrovasculature. In addition to ICA blood flow, the volume of VA blood flow in normothermia was similar to that reported previously (4). Although VA blood flow was also reduced during heat stress in this study, the reduction was less than that reported previously (4). Therefore, the effect of clamping PETCO2 on VA blood flow might have been overemphasized in the previous study. In normothermia, CO2 reactivity in the VA was lower than that in the ICA (P ⬍ 0.001), which is consistent with our previous report (30). Although heat stress reduced CO2 reactivity in the ICA but not in the VA (Fig. 2D), the ICA slope remained steeper relative to that in VA during heat stress (2.23 ⫾ 1.03%/mmHg vs. 1.56 ⫾ 0.74%/mmHg, P ⫽ 0.03). We performed hypercapnic and hypocapnic challenges in this study. Regardless of the thermal status, a hypocapnic challenge decreased VA blood flow, and the ratio of changes in VA blood flow to changes in PETCO2 was similar between hypocapnic and hypercapnic challenges (1.11 ⫾ 1.07%/mmHg vs. 1.73 ⫾ 1.51%/mmHg, P ⫽ 0.14). Central blood volume or degree of cerebral hypoperfusion may influence cerebral CO2 reactivity (19, 24). Brothers et al. (6) demonstrated that heat stress induced a decline in central blood volume and CBF. In a follow-up study by this group, Schlader et al. (32) demonstrated that acute volume expansion did not reverse hyperthermia-induced reduction in cerebral perfusion. These findings suggest that cerebral CO2 reactivity may be due to cerebral perfusion. Indeed, in the present study, because of increased ECA conductance during heat stress, the reduction in ICA conductance would be greater than in the VA. In addition, altered CO2 reactivity in the ICA might be affected by the changes in vascular conductance in these arteries. These observations, coupled with the fact that the CO2 reactivity in VA did not change, suggest that a change in blood perfusion at each cerebral artery during heat stress might modify cerebral CO2 reactivity. The ICA provides blood to a large portion of the cerebral cortex via intracranial arteries, while the vertebrobasilar arteries supply blood to the occipital cortex, cerebellum, and brain stem. Therefore, this phenomenon may offer an advantage for maintaining function of the autonomic nervous system with preserved dynamic CBF regulation during passive heat stress. The difference in hyperthermia-induced alteration of CO2 reactivity between vertebrobasilar territories (VA) and the cerebral cortex (ICA) may be due to the diverse characteristics of the vasculature e.g., regional microvascular density (31), basal

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Blood distribution toward the cerebrovasculature comprises two pairs of arteries: internal carotid arteries and vertebral arteries. Since heat stress reduces CBF, dynamic CBF regulation (i.e., dynamic cerebral autoregulation and cerebral CO2 reactivity) becomes more important to maintain an adequate CBF for brain function. Heat stress decreased ICA and VA blood flows as well as MCA Vmean (Table 1), which are consistent with previous reports (4, 7, 8, 23, 25, 40). The percentage of reduction in MCA Vmean (⬃24%) due to heat stress was not significant but slightly greater than that of ICA blood flow (⬃17%), similar to a previous study (MCA ⬃22% and ICA ⬃18%) (4). Regardless of the thermal conditions in the present study, ICA blood flow and MCA Vmean increased during hypercapnic challenges and decreased during hypocapnic challenges. The results observed during hypercapnic challenges were consistent with Bain et al. (4). In previous studies, hyperthermia-induced reduced PETCO2 was restored to the preheating level (4, 7, 13), whereas Bain et al. (4) demonstrated that MCA Vmean was restored to the normothermic baseline level. However, other studies (7, 13) showed that MCA Vmean remained lower relative to the normothermic baseline. These previous studies did not identify cerebral CO2 reactivity, and PETCO2 was controlled using a computer-controlled gas blender, inducing the increased ventilation. In the present study, cerebral CO2 reactivity was identified in the hypercapnic challenge as each subject breathed air through 500 ml of dead space, and respiratory frequency and depth were controlled to minimize increased ventilation. Hypercapnic challenge during heat stress increased PETCO2 (43.5 ⫾ 3.4 mmHg) more than expected, to a level greater than that prior to preheating stress (40.6 ⫾ 2.9 mmHg, P ⫽ 0.014). Consequently, ICA blood flow was increased relative to the normothermic baseline level (310.4 ⫾ 81.7 vs. 285.2 ⫾ 52.8 ml/min, P ⫽ 0.06), whereas MCA Vmean restored but at a slightly lower level relative to the normothermic baseline level (preheat, 52.0 ⫾ 17.5 cm/s; hyperthermic hypercapnia, 47.3 ⫾ 16.3 cm/s). Similar to previous studies (11, 23), CO2 reactivity in the MCA, calculated by the response of MCA Vmean to changes in PETCO2, was unaffected during heat stress (Fig. 2A), whereas the heat stress attenuated CO2 reactivity in the ICA (Fig. 2B). These observations indicate that the slope of MCA Vmean to changes in PETCO2 was not changed, even though the relationship between MCA Vmean and PETCO2 was shifted downward during heat stress. Moreover, during the hypocapnic challenge, PETCO2 was decreased by ⬎10 Torr in both normothermic and hyperthermic conditions (Table 1). Because of the unaltered CO2 reactivity in the MCA, MCA Vmean was lower relative to the normothermic hypocapnic level (P ⫽ 0.016). However, ICA blood flow also decreased in the hypocapnic challenge during heat stress, but the absolute value was identical to the normothermic hypocapnic level. Since MCA diameter is unchanged under several physiological conditions (14, 33), MCA blood velocity has been used as an index of CBF, even during heat stress (2). However, the different responsiveness between ICA blood flow and MCA Vmean might compromise this premise under hyperthermic conditions. Willie et al. (38) raised the possibility that the

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Cerebral CO2 Reactivity during Heat Stress

vascular tone (1, 16), autonomic innervation (10, 15) and regional heterogeneity in ion channels or production of NO (17). Particularly, regional sympathetic innervation of intracranial arterioles is reportedly less extensive in the posterior cerebral cortex and cerebellum compared with arterioles in the anterior cerebral circulation (10). In the vertebrobasilar circulation, less autonomic-induced vascular tone might be related to a lower vasodilatation capacity during hypercapnia (29). Thus differential hyperthermia-induced alterations in CO2 reactivity between ICA and VA may be due to regional sympathetic innervation of intracranial arterioles. Extracranial Blood Flow Responses to Changes in PETCO2

Limitations Our findings regarding CO2 reactivity in the present study were based on the assumption that PETCO2 accurately reflects PaCO2. The difference between the two variables is influenced by metabolic CO2 production and tidal volume, but it is not altered by breathing frequency (18). In addition, estimated PaCO2 using PETCO2 has been validated under a relatively wide range of core temperatures and orthostatic stresses (5).

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These findings suggest that the observed changes in PETCO2 are reflective of changes in PaCO2 during heat stress. We used linear regression analysis to assess the steady-state blood flows/velocity and PETCO2 relationship. It should be noted that cerebral CO2 reactivity was identified using only three data points (baseline, hypo-, hypercapnia; range, ⬃13 Torr below and ⬃6 Torr above baseline values). Again, each subject performed hypocapnic/hypercapnic challenges while Tes remained elevated by ⬃1.5°C. Approximately 20 min was required to complete these challenges. Within the range of PETCO2 performed in this study, we confirmed a linear relationship between each blood flow measured (i.e., ICA, ECA, and VA) and changes in CO2 (30). To shorten the experiment, we decided on three stages. Thus, on the basis of the present findings, we are confident that passive heat stress affected cerebral CO2 reactivity within the evaluated ranges of PETCO2 change. We did not consider the diurnal cycle in this study. However, in other studies, circadian rhythm, as well as menstrual cycle, was found to modulate thermoregulatory responses (9), and cerebrovascular CO2 reactivity was influenced by circadian rhythm and sex difference (3, 20). We reported previously that CO2 reactivity in VA blood flow was greater in females rather than in males (30), but CO2 reactivity in ICA blood flow was greater than that in VA blood flow in both sexes. Therefore, it is unlikely to influence regional difference of CO2 reactivity, but the influence of these factors on other CBF regulations in hyperthermia warrants further investigation. In summary, hyperthermia decreased CO2 reactivity in the ICA but enhanced CO2 reactivity in the ECA, while heat stress decreased ICA blood flow and increased ECA blood flow. These results indicate that hyperthermia is capable of modifying the cerebrovascular response to changes in PETCO2 in the ICA and ECA. Despite the decrease in VA and ICA blood flows during passive heat stress, CO2 reactivity in the VA was preserved. This phenomenon may offer an advantage for maintaining autonomic nervous system function and preserving dynamic CBF regulation during passive heat stress. ACKNOWLEDGMENTS The authors appreciate the time and effort expended by the volunteer subjects. We also thank Ms. Aota, Sasaoka, Gonda, Imai, Ishihara, and Sakamoto for recruitment of subjects and support of this project. GRANTS This study was supported in part by Grant-in-Aid for Scientific Research B-23300265 (to M. Shibasaki) and B-80553841 (to S. Ogoh). DISCLOSURES No conflicts of interest, financial or otherwise, are declared by the author(s). AUTHOR CONTRIBUTIONS Author contributions: S.O. and M.S. conception and design of research; S.O., K.S., K.O., T.M., and M.S. performed experiments; S.O. drafted manuscript; S.O. and M.S. edited and revised manuscript; S.O., K.S., K.O., T.M., A.H., and M.S. approved final version of manuscript; K.S., K.O., T.M., and A.H. analyzed data; K.S., K.O., T.M., and A.H. interpreted results of experiments; A.H. and M.S. prepared figures. REFERENCES 1. Ackerman RH. The relationship of regional cerebrovascular CO2 reactivity to blood pressure and regional resting flow. Stroke 4: 725–731, 1973.

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CO2 reactivity in the ECA was evaluated in this study. Similar to a previous report (30), CO2 reactivity was very small relative to other blood flows in normothermic condition; however, the responsiveness increased significantly during heat stress (Fig. 2C). Thus it is possible that the large increase in ECA blood flow affected its CO2 reactivity, which is broadly comparable to the possible influence of blood volume on cerebral CO2 reactivity (19, 24). During heat stress, cutaneous vascular conductance was not changed during the hypocapnic challenge, despite a decrease in skin blood flux on the forehead when blood pressure increased. In contrast, ECA conductance was decreased during the hypocapnic challenge. We observed a clear relationship between ECA conductance and cutaneous vascular conductance on the forehead during whole body heating (28). However, responses to changes in PETCO2 were inconsistent. We measured skin blood flux at only one region (forehead), but ECA mainly distributes blood to cutaneous vessels in the extracranium. Therefore, we should have measured skin blood flux more widely using a laser-Doppler scanner or at several different areas. Further investigation is required to clarify this discrepancy between the measurements. Curiously, Bain et al. (4) reported that ECA blood flow increased during heat stress but did not change due to clamping CO2, which was inconsistent with our finding. Although they did not assess the effect of decreased CO2 on ECA blood flow, we observed increased and decreased ECA blood flow during hypercapnic and hypocapnic challenges, respectively. A similarity in ECA conductance was confirmed, which excluded the influence of blood pressure. Recently, we confirmed that the ECA vascular response affects ICA blood flow because both arteries are branches of the common carotid artery (27). Possible reasons for the discrepancy between the present study and Bain et al. may likely be attributable to methodological differences such as the degree of hyperthermia and control of respiration. As mentioned above, to prevent hypercapnia-induced hyperventilation, we used spontaneous breathing control with 500 ml of dead space. Further investigation is required to clarify this discrepancy.

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