839
Exp Physiol 100.7 (2015) pp 839–851
Research Paper
Steady-state tilt has no effect on cerebrovascular CO2 reactivity in anterior and posterior cerebral circulations Michael M. Tymko1,2 , Rachel J. Skow2,3 , Christina M. MacKay2,3 and Trevor A. Day2 1
Experimental Physiology
Centre for Heart, Lung, and Vascular Health, School of Health and Exercise Science, University of British Columbia, Kelowna, British Columbia, Canada 2 Department of Biology, Faculty of Science and Technology, Mount Royal University, Calgary, Alberta, Canada 3 Faculty of Physical Education and Recreation, University of Alberta, Edmonton, Alberta, Canada
New Findings r What is the central question of this study? We investigated the effects of superimposed tilt and hypercapnia-induced cerebral arteriolar dilatation on anterior and posterior cerebrovascular CO2 reactivity using hyperoxic rebreathing in human participants. r What is the main finding and its importance? The main findings are threefold: (i) cerebrovascular CO2 reactivity in the anterior and posterior cerebrovasculature is unchanged with tilt; (ii) cerebral autoregulation is unlikely responsible due to unchanging cerebrovascular resistance reactivity between positions; and (iii) cerebral blood flow is not pressure passive during tilt as it is with pharmacological or lower body negative pressure-induced changes in mean arterial pressure, suggesting that sympathetic activation or balanced transmural pressures during head-down tilt regulate cerebral blood flow.
Cerebral autoregulation is a protective feature of the cerebrovasculature that maintains relatively constant cerebral perfusion in the face of static and dynamic fluctuations in mean arterial pressure (MAP). However, the extent that the cerebrovasculature can autoregulate in the face of superimposed steady-state orthostasis-induced changes in MAP (e.g. head-up and head-down tilt; HUT and HDT) and CO2 -mediated arteriolar dilatation is unknown. We tested the effects of steady-state tilt on cerebrovascular CO2 reactivity in the middle and and posterior cerebral artery in the following five body positions: 90 deg HUT, 45 deg HUT, supine, 45 deg HDT and 90 deg HDT on a tilt table during a modified hyperoxic rebreathing test. Absolute and relative cerebrovascular CO2 reactivity [cerebral blood velocity (CBV)/CO2 ], cerebrovascular resistance (CVR) reactivity (CVR/CO2 ) and MAP reactivity (MAP/CO2 ) were quantified using linear regression. Mean arterial pressure was significantly elevated in 90 deg HDT compared with other positions during baseline steady-state tilt (P < 0.01). Absolute CBV/CO2 and CVR/CO2 were greater in the middle cerebral artery than the posterior cerebral artery (P < 0.01) in all body positions, but relative measures were not different (P = 0.143 and P = 0.360, respectively), nor was there any interaction with tilt position. In addition, there was no difference in absolute (P = 0.556) and relative MAP/CO2 (P = 0.308) between positions. Our data demonstrate that cerebral blood flow remains well regulated during superimposed steady-state orthostatic stress and dynamic changes in the partial pressure of end-tidal CO2 during rebreathing. Cerebral autoregulation is likely not the mechanism responsible, but rather sympathetic nervous system
C 2015 The Authors. Experimental Physiology C 2015 The Physiological Society
DOI: 10.1113/EP085084
840
M. M. Tymko and others
Exp Physiol 100.7 (2015) pp 839–851
activation or a balanced cerebrovascular transmural pressure with HDT maintains resting cerebral blood flow and cerebrovascular CO2 reactivity during rebreathing. (Received 18 January 2015; accepted after revision 11 May 2015; first published online 13 May 2015) Corresponding author T. A. Day: Department of Biology, Faculty of Science and Technology, Mount Royal University, 4825 Mount Royal Gate SW, Calgary, Alberta, Canada T3E 6K6. Email: [email protected]
Introduction Cerebral autoregulation (CA) is a protective feature of the cerebrovasculature that maintains relatively constant cerebral perfusion in the face of static and dynamic fluctuations in mean arterial pressure (MAP; Tiecks et al. 1995; Lucas et al. 2010). Cerebral autoregulation was initially thought to maintain constant cerebral perfusion across a broad range of perfusion pressures (Lassen, 1959; Willie et al. 2014). However, recent evidence suggests that the cerebrovasculature is more pressure passive, where even small changes in steady-state MAP are reflected in cerebral perfusion (Lucas et al. 2010). In addition, high-frequency oscillations in arterial pressure can easily penetrate the cerebrovasculature and are reflected in cerebral blood flow (Claassen et al. 2009; Smirl et al. 2014). The extent to which oscillations in MAP are dampened by the cerebrovasculature determines the degree of CA. Experimental perturbations in MAP can be used to interrogate the degree of CA in different conditions, which can then be quantified through transfer function analysis methods (Kuo et al. 1998; Panerai, 1998; Bellapart & Fraser, 2009; Tzeng et al. 2012). Alterations in MAP can be brought about via thigh-cuff release (e.g. Mahony et al. 2000), squat–stand manoeuvres (e.g. Claassen et al. 2009; Smirl et al. 2014), paced breathing (e.g. Lucas et al. 2013), the Valsalva manoeuvre (e.g. Hetzel et al. 1999), pharmacological manipulations (e.g. Lucas et al. 2010) or orthostatic challenges (e.g. Gelinas et al. 2012). There may be differences in the extent to which various experimental perturbations in MAP are reflected in the cerebral vasculature. Head-up tilt (HUT) and head-down tilt (HDT) induce gravity-dependent shifts in blood volume distribution, which may have profound effects on venous return, cardiac output (CO) and MAP (Bundgaard-Nielsen et al. 2009; Murrell et al. 2011; Gelinas et al. 2012). Regardless of the mechanism of increases in MAP in HDT, this change in cardiovascular driving force with orthostatic stress can potentially affect cerebral perfusion (Lucas et al. 2010; Tzeng et al. 2014). Indeed, transient drops in MAP and the resulting reduction in cerebral perfusion can be ameliorated by assuming sitting or supine postures (Rickards et al. 2007). However, in a recent study, Gelinas et al. (2012) showed that baseline cerebral blood velocity (CBV; an index of volumetric flow) was unchanged in the middle and posterior cerebral arteries (MCA and PCA)
despite increases in MAP (i.e. driving force) in HDT positions (Gelinas et al. 2012). In their study, the partial pressure of end-tidal CO2 (P ET CO2 ) was maintained at supine levels through respiratory coaching, and they did not test the cerebrovascular response to increases in CO2 with tilt (Gelinas et al. 2012). Cerebral blood flow is highly sensitive to the influence of changes in the partial pressure of arterial CO2 on arteriolar diameter. Carbon dioxide acts as a vasodilator of cerebral arterioles, where cerebral blood flow in intracranial vessels (e.g. MCA and PCA) and extracranial vessels (e.g. internal carotid and vertebral arteries) increases in response to increasing levels of CO2 , largely due to dilatation of downstream arterioles (Wolff et al. 1930; Kety & Schmidt, 1948; Willie et al. 2014). The relationship between cerebral blood flow and CO2 is termed cerebrovascular reactivity (Poulin et al. 1996; Battisti-Charbonney et al. 2011; Willie et al. 2011; Fierstra et al. 2013; Skow et al. 2013). Studies investigating resting CBV and cerebrovascular reactivity using transcranial Doppler ultrasound (TCD) generally insonate the MCA, assuming that it reflects global cerebral blood flow. Recent evidence from our research group (Skow et al. 2013) and others (Sato et al. 2012b; Willie et al. 2012) suggests that this assumption may be misleading. For example, we recently showed that the MCA has a higher absolute reactivity to increases in CO2 than the PCA in the supine position. However, these differences were not apparent in relative (i.e. normalized to baseline) values (Skow et al. 2013). Other groups have also demonstrated regional differences in CA (Sato et al. 2012a) and cerebrovascular reactivity to CO2 and O2 (Sato et al. 2012b; Willie et al. 2012). The effects of superimposed steady-state orthostatic stress and cerebrovascular CO2 reactivity in the anterior and posterior cerebral circulations are poorly understood, particularly in HDT. Measuring CBV and CO2 reactivity in the anterior and posterior cerebral circulations simultaneously during postural challenges could provide an important non-pharmacological comparison of relative blood flow responses between these two intracranial arterial circuits and the respective downstream cerebral regions that they perfuse. Thus, the primary purpose of this study was to characterize and compare the effects of steady-state tilt on resting CBV and CO2 reactivity in the MCA and PCA using TCD in the following five different body positions: 90 deg HUT, 45 deg HUT, supine, 45 deg HDT and 90 deg HDT. We hypothesized that owing to C 2015 The Authors. Experimental Physiology C 2015 The Physiological Society
Exp Physiol 100.7 (2015) pp 839–851
Tilt and cerebrovascular CO2 reactivity
gravity-dependent shifts in blood volume distribution and the resulting increase in MAP in head-down positions (i.e. increase in driving force), cerebrovascular CO2 reactivity would be larger in HDT positions compared with supine and HUT positions. Methods Ethical approval
This study received ethical approval from the Conjoint Human Research Ethics Board at the University of Calgary (protocol E-23655) and abided by the Declaration of Helsinki and the Canadian Government Tri-Council Policy Statement (TCPS2) for Integrity in Research. All participants provided written informed consent prior to participation. Participants
Sixteen healthy participants were recruited from Mount Royal University and University of Calgary student populations. Two of these participants were excluded from data analysis, in one case due to a participant requesting to stop the protocol early due to discomfort in HDT and in the other case due to our inability to obtain adequate CBV measurements. Participants were between 18 and 40 years old, non-smoking, non-obese (body mass index < 30 kg m−2 ), had no previous history of cardiovascular, cerebrovascular or respiratory diseases, and were not taking any medications during testing apart from oral contraceptives. Female participants were tested within their self-reported follicular phase (days 0–14) of their ovarian cycle. Prior to each experiment, all participants were asked to refrain from caffeine, alcohol and vigorous exercise for at least 12 h before participation in the study. The same participants also took part in the following studies: (i) Skow et al. (2014), where central respiratory chemoreflex sensitivity was compared with tilt; and (ii) Skow et al. (2013) Protocol B, where we initially reported differences in cerebrovascular CO2 reactivity between the MCA and PCA in the supine position only. Thus, the supine data within the present study were published previously by Skow et al. (2013); however, the cerebrovascular data collected during the other four body positions in the present manuscript (90 deg HDT, 45 deg HDT, 45 deg HUT and 90 deg HUT) have not been reported previously and are compared with the supine position, within subjects. Respiratory measurements
All respiratory parameters were collected at 200 Hz and digitally archived using a 16-channel analog-to-digital converter (Powerlab/16SP ML880; ADInstruments, C 2015 The Authors. Experimental Physiology C 2015 The Physiological Society
841
Colorado Springs, CO, USA) and commercially available software (LabChart V7.2; ADInstruments) and analysed offline. Participants breathed through a rebreathing system that incorporated a mouthpiece (with nose-clip), bacteriological filter and three-way valve to allow switching of airflow between room air and a 5 litre latex anesthesia bag (Teleflex Medical, Markham, Ontario, Canada) prefilled with 93% O2 and 7% CO2 . Respiratory flow was measured near the mouth by the use of a flow head (MLT1000L; ADInstruments) and a differential pressure amplifier (ML141; ADInstruments). Expired gas was sampled breath-by-breath distal to the flow head, dried with nafion tubing, and CO2 and O2 were measured using a dual gas analyser (ML206; ADInstruments). This gas analyser has been used to analyse CO2 and O2 on a breath-by-breath basis during exercise (Smith et al. 2014), modified CO2 rebreathing tests (Skow et al. 2013) and steady-state CO2 tests (Willie et al. 2012; Hoiland et al. 2015). The resolution, linearity and drift (over 8 h) are reported to be 0.1%. Gas measurements were calibrated daily and were converted to body temperature and pressure when saturated with water vapour (BTPS) using the daily atmospheric pressure. The average atmospheric pressure in Calgary is 660–665 mmHg. The laboratory room temperature was consistently 20–21°C and was quiet throughout the protocol to avoid participant distraction.
Cardiovascular and cerebrovascular measurements
Participants were instrumented non-invasively for cardiovascular and cerebrovascular parameters. Electrocardiogram electrodes were placed in lead II configuration (ADInstruments bioamp, ML132) to measure heart rate (HR). Beat-by-beat arterial pressure was measured by finger photoplethysmography (Finometer Pro; Finapres Medical Systems, Amsterdam, The Netherlands). Bilateral TCD was employed using a 2 MHz pulsed Doppler ultrasound system (PMD150B; Spencer Technologies, Redmond, WA, USA) to insonate and measure the CBV of the MCA and PCA. Systolic and diastolic arterial pressure and CBV were quantified from raw recordings. Mean arterial pressure and mean CBV were calculated using a standard weighted mean (⅓ systolic + ⅔ diastolic). Participants were positioned supine on a tilt table, and both the MCA and PCA were insonated and secured using a bilateral headpiece. We confirmed that we were insonating the correct vessels through previously reported techniques (Willie et al. 2011) and in an identical fashion to Skow et al. (2013). Cerebrovascular resistance (CVR) was calculated from the MAP and mean MCA and PCA velocity values (MAP/CBV). With regard to the repeatability of cerebrovascular responses to CO2 with multiple rebreathing tests, we calculated our intraclass correlation coefficient for CBV/CO2 for the MCA and
842
M. M. Tymko and others
PCA, which were 0.967 and 0.889, respectively (Boulet et al., unpublished data in 19 subjects). Protocol
This study was conducted using the following five body positions: 90 deg HUT (i.e. standing with knees locked), 45 deg HUT, supine, 45 deg HDT and 90 deg HDT. Each participant started the protocol in the supine position, and the order of subsequent positions was determined randomly. Participants were first instrumented in the supine position on the tilt table, wearing specifically designed boots that safely secured the participant to the tilt table (Teeter Hang-ups, Tacoma, WA, USA). They then underwent a baseline period for 10 min. Immediately after the baseline period, participants were coached to hyperventilate voluntarily for 1 min in order to decrease P ET CO2 to 20 Torr (BTPS). After the 1 min of hyperventilation, participants were asked to exhale fully, at which point they were switched to the rebreathing apparatus, whereupon they were instructed to inspire three deep breaths to equilibrate with the rebreathing circuit (93% O2 , 7% CO2 ; Duffin 2011; Skow et al. 2013). The participants were asked to breathe on the rebreathing circuit until one of the following conditions was met: (i) the participant’s P ET CO2 reached 55 Torr; (ii) the 5 litre rebreathing bag was deflated; or (iii) the participant indicated that they wished to discontinue the rebreathing protocol with a hand signal. At this time, the three-way circuit was then switched back to room air for a 2 min recovery period. The entire protocol (baseline, hyperventilation, rebreathing and recovery) was then repeated randomly in the other four positions, with each taking 20 min. Participants were given a 10 min rest period between body positions in the supine position.
Exp Physiol 100.7 (2015) pp 839–851
the figures are SEM. One-factor repeated measures (RM) ANOVAs were used to compare baseline data for HR, MAP and P ET CO2 . For absolute and relative MAP CO2 reactivity (MAP/CO2 ), one-factor RM ANOVA statistics was conducted on all five body positions, and on four body positions excluding 90 deg HDT, due to a much smaller n in that position. However, mean data ± SEM are reported for all five body positions in the figures. Two-factor RM ANOVAs were used to test for differences in baseline MCA and PCA CBV and CVR with tilt and absolute and relative cerebrovascular CO2 reactivity (CBV/CO2 ) with tilt across all five positions. For absolute and relative CVR CO2 reactivity (CVR/CO2 ), two-factor RM ANOVA statistics were conducted on all five body positons, and on four body positions excluding 90 deg HDT, due to a much smaller n in that position. However, mean data ± SEM are reported for all five body positions in the figures. When overall significance was detected, pairwise comparisons were made using Tukey’s post hoc test. Significance was assumed at P < 0.05. In figures 1, 2, 4 and 5, significance between body positions is represented between positions with no tick, compared to positions with a tick above them.
Results Participant characteristics
Fourteen healthy adults (seven female) were included in the data analysis for this study, with a mean age of 28.1 ± 1.3 years and body mass index of 24.3 ± 0.6 kg m−2 . Not all measures were obtained in all positions, so the relative number of participants is provided in each figure, if 20 min) in 90 deg HDT. We were also not able to obtain all measures in each position, because we had difficulty maintaining adequate finometer readings in 90 deg HDT, probably due to a combination
Exp Physiol 100.7 (2015) pp 839–851
of changes in blood flow to the finger and participant restlessness from being upside down for a considerable period of time. Thus, not only is our steady-state MAP sample size smaller in 90 deg HDT (n = 7; Fig. 1B), but also calculations of CVR, CVR/CO2 and MAP/CO2 are limited in this position (n = 5; Figs 2 and 5). Due to the inconsistent numbers between tilt position trials, we had lower than anticipated power (
Exp Physiol 100.7 (2015) pp 839–851
Research Paper
Steady-state tilt has no effect on cerebrovascular CO2 reactivity in anterior and posterior cerebral circulations Michael M. Tymko1,2 , Rachel J. Skow2,3 , Christina M. MacKay2,3 and Trevor A. Day2 1
Experimental Physiology
Centre for Heart, Lung, and Vascular Health, School of Health and Exercise Science, University of British Columbia, Kelowna, British Columbia, Canada 2 Department of Biology, Faculty of Science and Technology, Mount Royal University, Calgary, Alberta, Canada 3 Faculty of Physical Education and Recreation, University of Alberta, Edmonton, Alberta, Canada
New Findings r What is the central question of this study? We investigated the effects of superimposed tilt and hypercapnia-induced cerebral arteriolar dilatation on anterior and posterior cerebrovascular CO2 reactivity using hyperoxic rebreathing in human participants. r What is the main finding and its importance? The main findings are threefold: (i) cerebrovascular CO2 reactivity in the anterior and posterior cerebrovasculature is unchanged with tilt; (ii) cerebral autoregulation is unlikely responsible due to unchanging cerebrovascular resistance reactivity between positions; and (iii) cerebral blood flow is not pressure passive during tilt as it is with pharmacological or lower body negative pressure-induced changes in mean arterial pressure, suggesting that sympathetic activation or balanced transmural pressures during head-down tilt regulate cerebral blood flow.
Cerebral autoregulation is a protective feature of the cerebrovasculature that maintains relatively constant cerebral perfusion in the face of static and dynamic fluctuations in mean arterial pressure (MAP). However, the extent that the cerebrovasculature can autoregulate in the face of superimposed steady-state orthostasis-induced changes in MAP (e.g. head-up and head-down tilt; HUT and HDT) and CO2 -mediated arteriolar dilatation is unknown. We tested the effects of steady-state tilt on cerebrovascular CO2 reactivity in the middle and and posterior cerebral artery in the following five body positions: 90 deg HUT, 45 deg HUT, supine, 45 deg HDT and 90 deg HDT on a tilt table during a modified hyperoxic rebreathing test. Absolute and relative cerebrovascular CO2 reactivity [cerebral blood velocity (CBV)/CO2 ], cerebrovascular resistance (CVR) reactivity (CVR/CO2 ) and MAP reactivity (MAP/CO2 ) were quantified using linear regression. Mean arterial pressure was significantly elevated in 90 deg HDT compared with other positions during baseline steady-state tilt (P < 0.01). Absolute CBV/CO2 and CVR/CO2 were greater in the middle cerebral artery than the posterior cerebral artery (P < 0.01) in all body positions, but relative measures were not different (P = 0.143 and P = 0.360, respectively), nor was there any interaction with tilt position. In addition, there was no difference in absolute (P = 0.556) and relative MAP/CO2 (P = 0.308) between positions. Our data demonstrate that cerebral blood flow remains well regulated during superimposed steady-state orthostatic stress and dynamic changes in the partial pressure of end-tidal CO2 during rebreathing. Cerebral autoregulation is likely not the mechanism responsible, but rather sympathetic nervous system
C 2015 The Authors. Experimental Physiology C 2015 The Physiological Society
DOI: 10.1113/EP085084
840
M. M. Tymko and others
Exp Physiol 100.7 (2015) pp 839–851
activation or a balanced cerebrovascular transmural pressure with HDT maintains resting cerebral blood flow and cerebrovascular CO2 reactivity during rebreathing. (Received 18 January 2015; accepted after revision 11 May 2015; first published online 13 May 2015) Corresponding author T. A. Day: Department of Biology, Faculty of Science and Technology, Mount Royal University, 4825 Mount Royal Gate SW, Calgary, Alberta, Canada T3E 6K6. Email: [email protected]
Introduction Cerebral autoregulation (CA) is a protective feature of the cerebrovasculature that maintains relatively constant cerebral perfusion in the face of static and dynamic fluctuations in mean arterial pressure (MAP; Tiecks et al. 1995; Lucas et al. 2010). Cerebral autoregulation was initially thought to maintain constant cerebral perfusion across a broad range of perfusion pressures (Lassen, 1959; Willie et al. 2014). However, recent evidence suggests that the cerebrovasculature is more pressure passive, where even small changes in steady-state MAP are reflected in cerebral perfusion (Lucas et al. 2010). In addition, high-frequency oscillations in arterial pressure can easily penetrate the cerebrovasculature and are reflected in cerebral blood flow (Claassen et al. 2009; Smirl et al. 2014). The extent to which oscillations in MAP are dampened by the cerebrovasculature determines the degree of CA. Experimental perturbations in MAP can be used to interrogate the degree of CA in different conditions, which can then be quantified through transfer function analysis methods (Kuo et al. 1998; Panerai, 1998; Bellapart & Fraser, 2009; Tzeng et al. 2012). Alterations in MAP can be brought about via thigh-cuff release (e.g. Mahony et al. 2000), squat–stand manoeuvres (e.g. Claassen et al. 2009; Smirl et al. 2014), paced breathing (e.g. Lucas et al. 2013), the Valsalva manoeuvre (e.g. Hetzel et al. 1999), pharmacological manipulations (e.g. Lucas et al. 2010) or orthostatic challenges (e.g. Gelinas et al. 2012). There may be differences in the extent to which various experimental perturbations in MAP are reflected in the cerebral vasculature. Head-up tilt (HUT) and head-down tilt (HDT) induce gravity-dependent shifts in blood volume distribution, which may have profound effects on venous return, cardiac output (CO) and MAP (Bundgaard-Nielsen et al. 2009; Murrell et al. 2011; Gelinas et al. 2012). Regardless of the mechanism of increases in MAP in HDT, this change in cardiovascular driving force with orthostatic stress can potentially affect cerebral perfusion (Lucas et al. 2010; Tzeng et al. 2014). Indeed, transient drops in MAP and the resulting reduction in cerebral perfusion can be ameliorated by assuming sitting or supine postures (Rickards et al. 2007). However, in a recent study, Gelinas et al. (2012) showed that baseline cerebral blood velocity (CBV; an index of volumetric flow) was unchanged in the middle and posterior cerebral arteries (MCA and PCA)
despite increases in MAP (i.e. driving force) in HDT positions (Gelinas et al. 2012). In their study, the partial pressure of end-tidal CO2 (P ET CO2 ) was maintained at supine levels through respiratory coaching, and they did not test the cerebrovascular response to increases in CO2 with tilt (Gelinas et al. 2012). Cerebral blood flow is highly sensitive to the influence of changes in the partial pressure of arterial CO2 on arteriolar diameter. Carbon dioxide acts as a vasodilator of cerebral arterioles, where cerebral blood flow in intracranial vessels (e.g. MCA and PCA) and extracranial vessels (e.g. internal carotid and vertebral arteries) increases in response to increasing levels of CO2 , largely due to dilatation of downstream arterioles (Wolff et al. 1930; Kety & Schmidt, 1948; Willie et al. 2014). The relationship between cerebral blood flow and CO2 is termed cerebrovascular reactivity (Poulin et al. 1996; Battisti-Charbonney et al. 2011; Willie et al. 2011; Fierstra et al. 2013; Skow et al. 2013). Studies investigating resting CBV and cerebrovascular reactivity using transcranial Doppler ultrasound (TCD) generally insonate the MCA, assuming that it reflects global cerebral blood flow. Recent evidence from our research group (Skow et al. 2013) and others (Sato et al. 2012b; Willie et al. 2012) suggests that this assumption may be misleading. For example, we recently showed that the MCA has a higher absolute reactivity to increases in CO2 than the PCA in the supine position. However, these differences were not apparent in relative (i.e. normalized to baseline) values (Skow et al. 2013). Other groups have also demonstrated regional differences in CA (Sato et al. 2012a) and cerebrovascular reactivity to CO2 and O2 (Sato et al. 2012b; Willie et al. 2012). The effects of superimposed steady-state orthostatic stress and cerebrovascular CO2 reactivity in the anterior and posterior cerebral circulations are poorly understood, particularly in HDT. Measuring CBV and CO2 reactivity in the anterior and posterior cerebral circulations simultaneously during postural challenges could provide an important non-pharmacological comparison of relative blood flow responses between these two intracranial arterial circuits and the respective downstream cerebral regions that they perfuse. Thus, the primary purpose of this study was to characterize and compare the effects of steady-state tilt on resting CBV and CO2 reactivity in the MCA and PCA using TCD in the following five different body positions: 90 deg HUT, 45 deg HUT, supine, 45 deg HDT and 90 deg HDT. We hypothesized that owing to C 2015 The Authors. Experimental Physiology C 2015 The Physiological Society
Exp Physiol 100.7 (2015) pp 839–851
Tilt and cerebrovascular CO2 reactivity
gravity-dependent shifts in blood volume distribution and the resulting increase in MAP in head-down positions (i.e. increase in driving force), cerebrovascular CO2 reactivity would be larger in HDT positions compared with supine and HUT positions. Methods Ethical approval
This study received ethical approval from the Conjoint Human Research Ethics Board at the University of Calgary (protocol E-23655) and abided by the Declaration of Helsinki and the Canadian Government Tri-Council Policy Statement (TCPS2) for Integrity in Research. All participants provided written informed consent prior to participation. Participants
Sixteen healthy participants were recruited from Mount Royal University and University of Calgary student populations. Two of these participants were excluded from data analysis, in one case due to a participant requesting to stop the protocol early due to discomfort in HDT and in the other case due to our inability to obtain adequate CBV measurements. Participants were between 18 and 40 years old, non-smoking, non-obese (body mass index < 30 kg m−2 ), had no previous history of cardiovascular, cerebrovascular or respiratory diseases, and were not taking any medications during testing apart from oral contraceptives. Female participants were tested within their self-reported follicular phase (days 0–14) of their ovarian cycle. Prior to each experiment, all participants were asked to refrain from caffeine, alcohol and vigorous exercise for at least 12 h before participation in the study. The same participants also took part in the following studies: (i) Skow et al. (2014), where central respiratory chemoreflex sensitivity was compared with tilt; and (ii) Skow et al. (2013) Protocol B, where we initially reported differences in cerebrovascular CO2 reactivity between the MCA and PCA in the supine position only. Thus, the supine data within the present study were published previously by Skow et al. (2013); however, the cerebrovascular data collected during the other four body positions in the present manuscript (90 deg HDT, 45 deg HDT, 45 deg HUT and 90 deg HUT) have not been reported previously and are compared with the supine position, within subjects. Respiratory measurements
All respiratory parameters were collected at 200 Hz and digitally archived using a 16-channel analog-to-digital converter (Powerlab/16SP ML880; ADInstruments, C 2015 The Authors. Experimental Physiology C 2015 The Physiological Society
841
Colorado Springs, CO, USA) and commercially available software (LabChart V7.2; ADInstruments) and analysed offline. Participants breathed through a rebreathing system that incorporated a mouthpiece (with nose-clip), bacteriological filter and three-way valve to allow switching of airflow between room air and a 5 litre latex anesthesia bag (Teleflex Medical, Markham, Ontario, Canada) prefilled with 93% O2 and 7% CO2 . Respiratory flow was measured near the mouth by the use of a flow head (MLT1000L; ADInstruments) and a differential pressure amplifier (ML141; ADInstruments). Expired gas was sampled breath-by-breath distal to the flow head, dried with nafion tubing, and CO2 and O2 were measured using a dual gas analyser (ML206; ADInstruments). This gas analyser has been used to analyse CO2 and O2 on a breath-by-breath basis during exercise (Smith et al. 2014), modified CO2 rebreathing tests (Skow et al. 2013) and steady-state CO2 tests (Willie et al. 2012; Hoiland et al. 2015). The resolution, linearity and drift (over 8 h) are reported to be 0.1%. Gas measurements were calibrated daily and were converted to body temperature and pressure when saturated with water vapour (BTPS) using the daily atmospheric pressure. The average atmospheric pressure in Calgary is 660–665 mmHg. The laboratory room temperature was consistently 20–21°C and was quiet throughout the protocol to avoid participant distraction.
Cardiovascular and cerebrovascular measurements
Participants were instrumented non-invasively for cardiovascular and cerebrovascular parameters. Electrocardiogram electrodes were placed in lead II configuration (ADInstruments bioamp, ML132) to measure heart rate (HR). Beat-by-beat arterial pressure was measured by finger photoplethysmography (Finometer Pro; Finapres Medical Systems, Amsterdam, The Netherlands). Bilateral TCD was employed using a 2 MHz pulsed Doppler ultrasound system (PMD150B; Spencer Technologies, Redmond, WA, USA) to insonate and measure the CBV of the MCA and PCA. Systolic and diastolic arterial pressure and CBV were quantified from raw recordings. Mean arterial pressure and mean CBV were calculated using a standard weighted mean (⅓ systolic + ⅔ diastolic). Participants were positioned supine on a tilt table, and both the MCA and PCA were insonated and secured using a bilateral headpiece. We confirmed that we were insonating the correct vessels through previously reported techniques (Willie et al. 2011) and in an identical fashion to Skow et al. (2013). Cerebrovascular resistance (CVR) was calculated from the MAP and mean MCA and PCA velocity values (MAP/CBV). With regard to the repeatability of cerebrovascular responses to CO2 with multiple rebreathing tests, we calculated our intraclass correlation coefficient for CBV/CO2 for the MCA and
842
M. M. Tymko and others
PCA, which were 0.967 and 0.889, respectively (Boulet et al., unpublished data in 19 subjects). Protocol
This study was conducted using the following five body positions: 90 deg HUT (i.e. standing with knees locked), 45 deg HUT, supine, 45 deg HDT and 90 deg HDT. Each participant started the protocol in the supine position, and the order of subsequent positions was determined randomly. Participants were first instrumented in the supine position on the tilt table, wearing specifically designed boots that safely secured the participant to the tilt table (Teeter Hang-ups, Tacoma, WA, USA). They then underwent a baseline period for 10 min. Immediately after the baseline period, participants were coached to hyperventilate voluntarily for 1 min in order to decrease P ET CO2 to 20 Torr (BTPS). After the 1 min of hyperventilation, participants were asked to exhale fully, at which point they were switched to the rebreathing apparatus, whereupon they were instructed to inspire three deep breaths to equilibrate with the rebreathing circuit (93% O2 , 7% CO2 ; Duffin 2011; Skow et al. 2013). The participants were asked to breathe on the rebreathing circuit until one of the following conditions was met: (i) the participant’s P ET CO2 reached 55 Torr; (ii) the 5 litre rebreathing bag was deflated; or (iii) the participant indicated that they wished to discontinue the rebreathing protocol with a hand signal. At this time, the three-way circuit was then switched back to room air for a 2 min recovery period. The entire protocol (baseline, hyperventilation, rebreathing and recovery) was then repeated randomly in the other four positions, with each taking 20 min. Participants were given a 10 min rest period between body positions in the supine position.
Exp Physiol 100.7 (2015) pp 839–851
the figures are SEM. One-factor repeated measures (RM) ANOVAs were used to compare baseline data for HR, MAP and P ET CO2 . For absolute and relative MAP CO2 reactivity (MAP/CO2 ), one-factor RM ANOVA statistics was conducted on all five body positions, and on four body positions excluding 90 deg HDT, due to a much smaller n in that position. However, mean data ± SEM are reported for all five body positions in the figures. Two-factor RM ANOVAs were used to test for differences in baseline MCA and PCA CBV and CVR with tilt and absolute and relative cerebrovascular CO2 reactivity (CBV/CO2 ) with tilt across all five positions. For absolute and relative CVR CO2 reactivity (CVR/CO2 ), two-factor RM ANOVA statistics were conducted on all five body positons, and on four body positions excluding 90 deg HDT, due to a much smaller n in that position. However, mean data ± SEM are reported for all five body positions in the figures. When overall significance was detected, pairwise comparisons were made using Tukey’s post hoc test. Significance was assumed at P < 0.05. In figures 1, 2, 4 and 5, significance between body positions is represented between positions with no tick, compared to positions with a tick above them.
Results Participant characteristics
Fourteen healthy adults (seven female) were included in the data analysis for this study, with a mean age of 28.1 ± 1.3 years and body mass index of 24.3 ± 0.6 kg m−2 . Not all measures were obtained in all positions, so the relative number of participants is provided in each figure, if 20 min) in 90 deg HDT. We were also not able to obtain all measures in each position, because we had difficulty maintaining adequate finometer readings in 90 deg HDT, probably due to a combination
Exp Physiol 100.7 (2015) pp 839–851
of changes in blood flow to the finger and participant restlessness from being upside down for a considerable period of time. Thus, not only is our steady-state MAP sample size smaller in 90 deg HDT (n = 7; Fig. 1B), but also calculations of CVR, CVR/CO2 and MAP/CO2 are limited in this position (n = 5; Figs 2 and 5). Due to the inconsistent numbers between tilt position trials, we had lower than anticipated power (
