Human femoral artery and estimated muscle capillary blood flow kinetics following the onset of exercise

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1 Exp Physiol 914 pp Experimental Physiology Human femoral artery and estimated muscle capillary blood flow kinetics following the onset of exercise Allison J Harper, Leonardo F Ferreira, Barbara J Lutjemeier, Dana K Townsend and Thomas J Barstow Department of Kinesiology, Kansas State University, Manhattan, KS , USA The purpose of this study was to compare the kinetics of estimated capillary blood flow ( Q cap ) to those of femoral artery blood flow ( Q FA ) and estimated muscle oxygen uptake ( V O2 m) Nine healthy subjects performed a series of transitions from rest to moderate (below estimated lactate threshold, 6 min bouts) knee extension exercise Pulmonary oxygen uptake ( V O2 ) was measured breath by breath, Q FA was measured continuously using Doppler ultrasound, and deoxyhaemoglobin ([HHb]) was estimated by near-infrared spectroscopy over the rectus femoris throughout the tests The time course of Q cap was estimated by rearranging the Fick equation (ie Q cap = V O2 m/(a v)o 2 ), (arterio venous O 2 difference) using the primary component of V O2 to represent V O2 m and [HHb] as a surrogate for (a v)o 2 The overall kinetics of Q FA (mean response time, MRT, 137 ± 70 s), V O2 m (τ, 278 ± 90 s) and Q cap (MRT, 414 ± 190 s) were significantly (P < 005) different from each other We conclude that for moderate intensity knee extension exercise, conduit artery blood flow ( Q FA ) kinetics may not be a reasonable approximation of blood flow kinetics in the microcirculation ( Q cap ), the site of gas exchange This temporal dissociation suggests that blood flow may be controlled differently at the conduit artery level than in the microcirculation (Received 29 November 2005; accepted after revision 16 March 2006; first published online 23 March 2006) Corresponding author T J Barstow: 1A Natatorium, Kansas State University, Manhattan, KS , USA tbarsto@ksuedu Following the onset of exercise, an increase in O 2 delivery to the working muscles is necessary to facilitate increases in muscle oxygen consumption ( V O2 m) However, there is controversy regarding whether the kinetics of V O2 m are limited by oxygen delivery or by some other intrinsic (intramuscular) mechanism (Grassi, 2001; Hughson et al 2001) Adequacy of oxygen delivery has previously been assessed by a comparison of the temporal profiles of limb or muscle blood flow ( Q m ) and V O2 m (Shoemaker et al 1994; Grassi, 2000) Previous research has shown the kinetics of Q m to be either faster than (Macdonald et al 1997; Grassi et al 1998; Behnke et al 2002a) or similar to V O2 m (Grassi et al 1996; Koga et al 2005) Owing to technical limitations, however, this conclusion has been based on measurement of blood flow either through conduit arteries (Hughson et al 1996; Bangsbo et al 2000) or through the capillary bed of isolated muscles (Behnke et al 2002a) It is presently unknown whether the characteristics of limb (conduit artery) blood flow following exercise onset are representative of blood flow in the microcirculation where gas exchange actually occurs Therefore, clarifying the relationship between Q m and capillary blood flow ( Q cap ) is critical to furthering our understanding of the mechanisms that govern the balance between O 2 delivery and V O2 m in health and disease Estimates of the kinetics of Q cap can be made by rearranging the Fick equation to solve for blood flow ( Q cap = V O2 m/(a v)o 2 ) (arterio venous O 2 difference) and using the primary component of pulmonary oxygen uptake ( V O2 ) during phase 2 and [HHb] derived from near-infrared spectroscopy (NIRS) to represent V O2 m and (a v)o 2, respectively (Ferreira et al 2005a,d) These estimates have yielded a temporal response of Q cap that is similar to that of V O2 m following the onset of cycling exercise (Ferreira et al 2005d) Estimates of Q cap have not been compared with limb blood flow measurements collected simultaneously Thus, it is presently unknown whether conduit artery blood flow can be used to estimate Q cap and, by inference, the kinetics of capillary oxygen delivery Any discrepancy that may exist between the kinetic responses of blood flow in the limb and in the microcirculation could have serious DOI: /expphysiol

2 662 A J Harper and others Exp Physiol 914 pp implications for the use of limb blood flow kinetics to evaluate the adequacy of oxygen delivery at the site of gas exchange during transitional phases of exercise in health and disease Therefore, the purpose of this study was to describe the relationship between the temporal profiles of leg blood flow ( Q FA ), estimated Q cap and estimated V O2 m in healthy human subjects following the onset of moderate intensity knee extension exercise, and to determine whether Q FA kinetics are a reasonable representation of Q cap kinetics We hypothesized that the kinetics of (1) Q cap and (2) Q FA would be faster than those of V O2 m, and that (3) the kinetics of Q cap would be similar to those of Q FA Methods Subjects Nine subjects (6 male, 3 female), age 313 ± 123 years (mean ± sd), height 172 ± 001 m, and weight 674 ± 75 kg, participated in this study The subjects adipose tissue thickness over the rectus femoris was 078 ± 022 cm, and mean femoral artery diameter was 101 ± 019 cm Experimental procedures and all benefits and risks were explained to each subject, and written informed consent was obtained before any testing began All procedures were approved by the Institutional Review Board for Research Involving Human Subjects at Kansas State University and followed the principles outlined in the Declaration of Helsinki Protocol Each subject completed a minimum of five visits to the laboratory, with at least 1 day of rest between each visit All visits consisted of two-leg dynamic knee extension exercise at a rate of approximately 40 contractions per minute (cpm), with legs extended simultaneously for each kick (ie as a dolphin kick ) On the first visit, an incremental test to volitional fatigue was completed on the knee extension ergometer to estimate lactate threshold and peak pulmonary oxygen consumption ( V O2 peak) for this mode of exercise On each subsequent visit the subjects completed three 6 min constant work rate exercise bouts at a moderate work rate selected to elicit a metabolic rate of approximately 80% lactate threshold, each separated by 6 min rest This protocol was performed four to six times by each subject Measurements During each test, pulmonary oxygen consumption and other ventilatory and gas exchange variables were measured using an open-circuit breath-by-breath system (CardiO 2, Medical Graphics, St Paul, MN, USA) The volume signal was calibrated before each test using a 3 l syringe, and the O 2 and CO 2 analysers were calibrated using two gases of known composition Blood velocity through the right femoral artery ( V FA ) was measured continuously using Doppler ultrasound (Model 500-V, Multigon Industries, Mt Vernon, NY, USA) with the transducer operating at a frequency of 4 MHz and the isonation angle fixed at 45 deg The probe was positioned flat against the skin above and parallel to the common femoral artery, proximal ( 2 cm) to the bifurcation, as confirmed using a 2-D ultrasound imaging system (Vivid 3-Pro, GE, Rochester, MN, USA) Femoral artery diameter (D FA ) at this position was measured at rest in each subject, also using 2-D ultrasound imaging Femoralarterybloodflow( Q FA ) was calculated as follows: Q FA = V FA π(d FA /2) 2 (1) Electrocardiogram was obtained using a modified lead I Blood velocity, ECG and the ergometer data (force and displacement) were digitized at 200 Hz and stored for offline analysis Muscle capillary O 2 extraction (as deoxyhaemoglobin concentration, [HHb]) was determined using a frequencydomain multidistance NIRS system (OxiplexTS, ISS, Champaign, IL, USA) during the incremental exercise tests and each subject s first two constant work rate test sessions This device operated at two wavelengths (690 and 830 nm) with light source detector separation distances of 20, 25, 30 and 35 cm for each wavelength Data were stored at 3125 Hz The probe was placed longitudinally along the belly of the right rectus femoris, which has been shown to produce electromyogram activity similar to that of the vastus medialis and vastus lateralis during concentric knee extension exercise (Pincivero et al 2006) It was then bound to the skin, which had previously been shaved and dried, with skin cement (Skin-Bond, Smith & Nephew, Largo, FL, USA), and secured using a Velcro elastic strap around the thigh Probe position was marked to ensure accurate repositioning on each test day The NIRS probe was calibrated each day according to the manufacturer s recommendations Data analysis The breath-by-breath V O2 data were first converted to second-by-second values; then the V O2 and Q FA (at 200 Hz) were each time aligned to the onset of exercise for each exercise bout and ensemble averaged across bouts for each subject to generate a single data set for each variable These averaged responses were then filtered using a lowpass filter with cut-off frequencies of 0075 Hz for V O2 and 02 Hz for Q FA (SigmaPlot 2001, Jandel Scientific; Ferreira et al 2006) Kinetic analysis was conducted

3 Exp Physiol 914 pp Capillary versus femoral artery blood flow kinetics 663 using non-linear regression with a least squares technique (Marquardt Levenberg and SigmaPlot 2001) Pulmonary oxygen uptake responses were fitted as follows: V O2 (t) = V O2 (r) + A I 1 e -(t-td I )τ I (phase 1; initial component) + A P 1 e -(t-td P )τ P (phase 2; primary component) (2) where the subscripts r, I and P refer to rest, the initial component and the primary component, respectively A represents the amplitude, TD the time delay and τ the time constant of each exponential response The initial component of V O2 was described up to TD P, where the amplitude of the response (A I ) was calculated as A I = A I (1 e -(TD P)τ I ) The overall amplitude of the response was calculated as A P = A I + A P Femoral artery blood flow data were also fitted in this manner, while [HHb] was described as a monoexponential response up to its peak value, in a manner similar to the initial component of V O2 in eqn (2) In three subjects, the phase 1 response of V O2 was such a high percentage of the total increase (mean = 63%) that accurate determination of the phase 2 time constant was precluded Previous studies using a cycle ergometer have shown that the phase 2 time constant is similar for transitions from a baseline of rest or very light exercise (Whipp et al 1982) Based on this, these three subjects subsequently performed four to five bouts of exercise with a baseline of unloaded knee extension (estimated to be approximately 7 W) followed by an immediate transition to the preset resistance on the knee extension ergometer In each case, this reduced the relative amplitude of the phase 1 response of V O2 and increased the relative amplitude for phase 2 Subsequent curve fitting produced more physiologically realistic time constants The estimated Q cap response following the onset of exercise was calculated from the kinetics of V O2 and the [HHb] data as described in detail previously (Ferreira et al 2005a,d) The kinetics of the primary component of V O2 have been predicted (Barstow et al 1990) and shown (Grassi et al 1996; Rossiter et al 1999) to closely approximate those of muscle V O2 ( V O2 m) The [HHb] response determined by NIRS has been used to estimate the dynamic response of muscle capillary oxygen extraction (ie C (a v)o2 ;DeLoreyet al 2003; Grassi et al 2003) By rearranging the Fick equation, the temporal characteristics of Q cap were estimated using the ratio of V O2 m to [HHb] as shown below (Ferreira et al 2005d): Q cap (t) = V O2 m(t) (C ao2 C vo2 )(t) V O2 (phase 2)(t) [HHb](t) (3) The temporal characteristics of this Q cap response were described using a three component exponential equation: Q cap (t) = Q cap(r) + A I 1 e -(t-td I )τ I (phase 1; initial component) + A P 1 e -(t-td P )τ P (phase 2; primary component) + A S 1 e -(t-td S )τ S (phase 3; slow component) (4) where subscript S represents the slow component For subsequent comparison among responses, the mean response time (MRT), time to approximately 63% of the primary response, for both Q cap and Q FA was determined as: A MRT = I AP (TD I + τ I ) + (TD P + τ P ) (5) A P where the parameters in eqns (4) and (5) are the same as those described following eqn (2) (Swanson & Hughson, 1988) Note that for MRT (eqn (5)), only the initial and primary components were used to describe the early, predominant portion of the response Statistical analysis Significant differences between means were determined using repeated measures analysis of variance, followed when appropriate with the Tukey Kramer post hoc test for pairwise comparisons Relationships between two variables were analysed using the Pearson product moment correlation Statistical tests were performed using NCSS 2000 software (NCSS Statistical Software, Kaysville, UT, USA) For all comparisons, significance was declared when P < 005 Results Subjects reached a mean (± sd) V O2 peak of 1460 ± 390 ml min 1 (216 ± 51 ml kg 1 min 1 ) and a peak heart rate of 166 ± 21 beats min 1 during the incremental knee extension exercise test, at a peak work rate of 537 ± 94 W Their estimated lactate threshold occurred at a V O2 of 1040 ± 260 ml min 1, or 715 ± 78% of V O2 peak Work rates for the constant work rate tests averaged 238 ± 57 W, or approximately 832 ± 136% lactate threshold heart rate 980 ± 133%, and contraction frequency 392 ± 07% Average pulmonary V O2, Q FA, [HHb], tissue oxygenation ([HbO 2 ]) and total haemoglobin for the three exercise transitions performed during each constant work rate test by a representative subject are shown in Fig 1 Estimated V O2 m, [HHb] and the resultant estimated Q cap for a representative subject are shown in Fig 2, while the comparison between Q FA and Q cap for the same subject is shown in Fig 3 Noticeable overshoots A P

4 664 A J Harper and others Exp Physiol 914 pp A V O2 (l min 1 ) B Q FA (ml min 1 ) C [HHb] (μm) D [HbO 2 ] (μm) E Total Hb (μm) Time (s) Figure 1 Responses to bout-1 (black line), bout-2 (grey line) and bout-3 (dotted line) of moderate exercise by a representative subject A, pulmonary V O2 (in l min 1 ) after filtering in the frequency domain B, limb blood flow ( Q FA, in ml min 1 ) after filtering in the frequency domain C, deoxyhaemoglobin concentration ([HHb], μm) D, oxyhaemoglobin concentration ([HbO 2 ], μm) E, total haemoglobin (Hb, μm)

5 Exp Physiol 914 pp Capillary versus femoral artery blood flow kinetics 665 Figure 2 Representative data from one subject A, V O2 m (in l min 1 ) estimated from kinetic parameters of pulmonary V O2 B, deoxyhaemoglobin concentration ([HHb], μm) C, estimated Q cap profile obtained from V O2 m divided by [HHb] The amplitude of Q cap is displayed in arbitrary units (au; Ferreira et al 2005d)

6 666 A J Harper and others Exp Physiol 914 pp Table 1 Kinetics parameters of pulmonary V O2, Q cap, Q FA and [HHb] for moderate exercise Parameter V O2 Q cap Q FA [HHb] Baseline 036 ± 014 a 0042 ± ± ± 310 A b I 021 ± ± ± 373 τ I (s) 224 ± ± ± 16 A b P 031 ± ± ± ± 157 TD P (s) 206 ± ± ± ± 352 τ P (s) 278 ± ± ± ± 331 A b S ± 0009 c TD S (s) 174 ± 48 τ S (s) 333 ± 52 MRT (s) d 312 ± ± ± ± 57 Values are means ± SD A, amplitude; TD, time delay; and τ, time constant of each component I, initial component; P, primary component; and S, slow component MRT is the time to approximately 63% of the primary response (see Methods for calculation) a Baseline values for V O2 reflect a combination of resting (6 subjects) and unloaded exercise data (3 subjects) b Units for V O2,lmin 1 ; Q cap, arbitrary; Q FA,mlmin 1 ; and [HHb], μm c For the slow component of Q cap, n = 4 d Comparison of the overall kinetics is presented in Fig 4 were seen in four (of 9) subjects for phase 2 of Q FA and in six subjects for [HHb] (see Fig 2) Kinetic parameters for pulmonary V O2, Q cap, Q FA and [HHb] are presented in Table 1 The MRT of Q FA was significantly faster than V O2 m and Q cap kinetics, while the kinetics of V O2 m were also significantly faster than the MRT of Q cap (Fig 4) The relationships between kinetic parameters [MRT of Q cap versus τ of V O2 m;mrtof Q FA versus τ of V O2 m (Fig 5) and MRT of Q cap versus MRT of Q FA (Fig 6)] were not significant (P > 005) Discussion This study was conducted in order to compare the temporal profiles of leg blood flow ( Q FA ), estimated Q cap and estimated V O2 m in healthy human subjects following the onset of upright knee extension exercise, and to determine whether Q FA kinetics are a reasonable representation of Q cap kinetics The major findings are as follows Consistent with our first hypothesis, the kinetics (as MRT) of Q FA for moderate exercise were significantly faster than those of V O2 (τ for V O2 m) In contrast to our second and third hypotheses, however, the kinetics of Q cap were significantly slower than those of V O2 m and Q FA While measurements of capillary blood flow have been made in isolated animal muscle preparations (Kindig et al 1999, 2002), direct measurement in humans has been problematic Recently, Ferreira and coworkers (Ferreira et al 2005a,b,c,d) have proposed a non-invasive method to estimate Q cap kinetics in exercising humans using [HHb] derived from NIRS During cycling exercise, the MRT of Q cap was found to be similar to τ of V O2 m for both moderate and heavy exercise (Ferreira et al 2005d) Similar results were generally observed in the present study for subjects whose τ of V O2 m values were relatively fast (Fig 5A) However, unlike the results for cycling (Ferreira 18 MRT QFA = 17 s Q FA (L min 1 ) MRT QCAP = 42 s Qcap (au) Time (s) 004 Figure 3 Q FA and Q cap responses for the subject shown in Fig 2 Overall kinetics are described by the mean response time (MRT) Note the slower increase in Q cap compared to Q FA

7 Exp Physiol 914 pp Capillary versus femoral artery blood flow kinetics 667 et al 2005d), subjects with slower V O2 m kinetics displayed disproportionally longer MRT of Q cap The reason(s) for this discrepancy are unclear at present Ferreira and coworkers examined oxygenation of the vastus lateralis during cycling, which may have different patterns of motor unit recruitment, fibre type distribution, blood flow and/or oxygen extraction than that of the rectus femoris during knee extension exercise as studied here In addition, the cycle exercise protocol used by Ferreira and coworkers (Ferreira et al 2005b,d) used an unloaded-toloaded exercise transition, while in the present study restto-exercise transitions were performed, with the exception of three subjects who also performed unloaded-to-loaded exercise solely for determination of V O2 kinetics The underlying mechanisms for the observed discrepancies must await further investigation The relationship between the kinetics of Q FA and V O2 m seen in the present study is similar to previous results in which conduit artery blood flow has been shown to increase more rapidly than V O2 (Hughson et al 1996; MacDonald et al 1998) These observations, as well as results indicating a similar temporal profile of Q FA and V O2 m (Hughson et al 1996; Koga et al 2005), have been interpreted as evidence that bulk O 2 delivery Q O2 does not limit V O2 m kinetics following the onset of exercise However, the temporal discrepancy between Q FA and Q cap shown here, and predicted by DeLorey et al (2003), suggests it may not be appropriate to use Q FA to represent the kinetics of oxygen delivery to the microcirculation in assessing the dynamic adequacy of Q O2 to V O2 m matching As the kinetics of Q cap approach or become slower relative to those of V O2 m, it is possible that there is a smaller reserve of capillary O 2 available to the muscle, so that at some point O 2 delivery would transiently limit (slow) the increase in V O2 m following the onset of exercise At present it is not possible to determine from the time course of Q cap alone whether its slower rate of adjustment in some subjects may limit the increase in V O2 m following the onset of exercise, since the amplitudes of both responses are also critical to this assessment, and the units of Q cap are arbitrary (see assumptions below) In the present study, overshoots were seen in several subjects for [HHb] and Q FA An overshoot in the [HHb] response was seen in six subjects This overshoot has been shown previously, and may be eliminated either by B 40 MRT (s) A B 10 0 V O2m Q cap Q FA Figure 4 Overall kinetics of V O2 m, Q cap and Q FA For V O2 m, there is no time delay, so τ is used as MRT Data are means ± SEM A Significantly different from MRT of Q cap (P < 005) B Significantly different from MRT of Q FA (P < 005) Figure 5 Relationships between the kinetics of Q cap (A) and Q FA (B) with τ of V O2 m for moderate exercise Dashed line is the line of identity for each plot Neither of the correlations was significant (P > 005)

8 668 A J Harper and others Exp Physiol 914 pp prior exercise or by application of a topical vasodilating ointment (Maehara et al 1997) An overshoot in [HHb] is equivalent to an undershoot in microvascular partial pressure of O 2, which has been seen in rat models of both heart failure and diabetes (Behnke et al 2002b; Diederich et al 2002) Since subjects for the present study were generally healthy, this overshoot may have been caused by regional differences in [HHb] or the mode of exercise employed here Overshoots in Q FA have been reported following, but not prior to, a short-term training programme (Shoemaker et al 1994) The underlying cause of the Q FA overshoots seen in four subjects in the present study is unclear, but the response was consistent between transitions Assumptions The assumptions associated with estimation of Q cap have been summarized previously (Ferreira et al 2005a,b,c,d) Pertinent to the present study, the relative contributions to [HHb] from the arterioles, capillaries and venules are not clear, nor is it clear whether this weighted distribution remains constant throughout the transition from rest to exercise (McCully & Hamaoka, 2000; Ferreira et al 2005d) However, it has been estimated that in the rat diaphragm capillaries comprise 84% of the vascular space (Poole et al 1995), indicating that the majority of the NIRS signal is likely to reflect capillary oxygen extraction Evidence also shows that, in the brain, the fractional contribution of arterial and venous blood values to tissue saturation remains relatively constant throughout the physiological MRT - Q cap (s) MRT - Q FA (s) Figure 6 Relationship between the kinetics of Q cap and Q FA for moderate exercise Dashed line is the line of identity The correlation was not significant (P > 005) rangeofo 2 saturation values for the tissues (Chance et al 2003) Another consideration is the relative contribution of [HHb] from cutaneous and subcutaneous adipose tissue microvasculature, and whether this contribution is constant (Maehara et al 1997) However, the vascular volume of these tissues is substantially less than that of skeletal muscle, so the relative contribution is likely to be small, although contribution by these tissues to the overshoot cannot be ruled out (Maehara et al 1997) The small sampling volume of the NIRS probe determines the oxygenation state of only a small (relatively superficial) portion of the working muscle Owing to potential regional differences in fibre type distribution, motor unit recruitment and muscle blood flow, it is unclear whether the area sampled is representative of the entire muscle (Boushel et al 2001) However, concentric knee extension exercise has been shown to elicit similar recruitment patterns for the rectus femoris, vastus medialis and vastus lateralis (Pincivero et al 2006), suggesting that the rectus femoris is representative of the muscle group involved in the exercise task Furthermore, phosphocreatine kinetics of the knee extensor group as a whole reflect those of pulmonary V O2 during knee extension exercise (Rossiter et al 1999) Thus, examination of microvascular oxygen exchange and calculation of Q cap in the rectus femoris is likely to be kinetically representative of the other heads of the vastus during knee extension exercise It is also unclear whether the onset of movement accompanying rest-toexercise transitions introduces error into the NIRS signal following the onset of exercise As illustrated in Fig 1, the baseline of [HHb] is slightly elevated during subsequent bouts of exercise, but the kinetic response following the onset of exercise appears similar in all three bouts Since V O2 is indistinguishable among the three bouts, the individual estimates of Q cap would be very similar Note, in contrast, that [HbO 2 ] appears to be quite sensitive to warm-up exercise, possibly owing, in part, to changes in skin blood flow, which make it less informative in assessing the relationship between Q cap and V O2 m (Maehara et al 1997; DeLorey et al 2003; Grassi et al 2003) Discrepancy between Q FA and Q cap Assuming the discrepancy between conduit artery ( Q FA ) and microcirculation ( Q cap ) kinetics to be real (but see discussion of assumptions above), the fundamental question emerges, Where is the blood flow seen in the femoral artery going, if not through the capillaries of the contracting muscle? Since the circulation below the knee was not occluded in the present study, Q FA is a measurement of blood flow to the entire leg, including active muscle and inactive tissues in both the upper and lower leg

9 Exp Physiol 914 pp Capillary versus femoral artery blood flow kinetics 669 During phase 1, the increase in blood flow is thought to be mediated through a combination of the muscle pump, which probably results from pressure changes within the intramuscular vasculature with each contraction cycle, and rapid vasodilatation Evidence of vasodilatation has been seen within the first 5 s of exercise (Wunsch et al 2000; Saunders & Tschakovsky, 2004; Tschakovsky & Sheriff, 2004; Tschakovsky et al 2004), but the mechanism(s) for this has not been determined (Shoemaker et al 1997; Brock et al 1998; Radegran & Saltin, 1999; Hamann et al 2004) The muscle pump would, and rapid vasodilatation might, cause an indiscriminate increase in blood flow to all parts of the contracting muscle Phase 2 of the blood flow increase is tightly coupled to metabolic activity, and may be controlled in part by H +, adenosine, ATP, nitric oxide, potassium, prostaglandins and/or a number of other metabolites (Delp & Laughlin, 1998) It appears that, like phase 1, vasodilatation during phase 2 is controlled by multiple mechanisms in combination (Clifford & Hellsten, 2004) Blood flow to the contracting muscles (and more specifically the contracting motor units) is also achieved by a functional sympatholysis, which directs the increasing blood flow to the sites of active metabolism (Buckwalter & Clifford, 2001; Tschakovsky & Hughson, 2003; Wray et al 2004) The time course for the increase in α-adrenergic vasoconstriction of the vasculature of inactive tissue could potentially cause a delay before redistribution of flow towards contracting muscle and motor units was fully achieved The decrease in renal blood flow following the onset of mild exercise seen in conscious baboons, which may reflect the time course of sympathetic vasoconstrictor outflow, can take up to 15 min (Hohimer & Smith, 1979) Given the systemic nature of sympathetic vasoconstrictor outflow (Wray et al 2004), a similar time course is possible in the inactive tissue of the legs Consistent with this, direct measurement of muscle sympathetic nerve activity (MSNA) showed a lag of at least 1 min before it increased following exercise onset in both exercising and non-exercising muscles (Hansen et al 1994) However, muscle sympathetic nerve activity, as evidenced by plasma noradrenaline levels, does not increase until 50% V O2 max (for review see Laughlin et al 1996) Therefore, it is at present unclear whether muscle sympathetic nerve activity (ie α-adrenergic vasoconstriction) is activated during moderate intensity exercise such as that performed in the present study, and, if it is, whether its temporal profile could contribute to the difference seen here between Q cap and Q FA Conclusions We have shown that for moderate intensity exercise: (1) the kinetics of Q FA are significantly faster than those of V O2 m; (2) the kinetics of Q cap are slower than those of V O2 m; and (3) the kinetics of Q cap are slower than those of Q FA Our finding regarding the difference in Q cap and Q FA kinetics for this mode of exercise, as predicted by DeLorey et al (2003), indicates that the time course of adjustment of limb blood flow may not be a reasonable representation of blood flow in the microcirculation for conditions such as those used here References Bangsbo J, Krustrup P, González-Alonso J, BoushelR&Saltin B (2000) Muscle oxygen kinetics at onset of intense dynamic exercise in humans Am J Physiol Regul Integr Comp Physiol 279, R899 R906 Barstow TJ, Lamarra N & Whipp BJ (1990) Modulation of muscle and pulmonary O 2 uptakes by circulatory dynamics during exercise J Appl Physiol 68, Behnke BJ, Barstow TJ, Kindig CA, McDonough P, Musch TI & Poole DC (2002a) Dynamics of oxygen uptake following exercise onset in rat skeletal muscle Respir Physiol Neurobiol 133, Behnke BJ, Kindig CA, McDonough P, Poole DC & Sexton WL (2002b) Dynamics of microvascular oxygen pressure during rest-contraction transition in skeletal muscle of diabetic rats Am J Physiol Heart Circ Physiol 283, H926 H932 Boushel R, Langberg H, Olesen J, Gonzales-Alonzo J, Bulow J & Kjaer M (2001) Monitoring tissue oxygen availability with near infrared spectroscopy (NIRS) in health and disease Scand J Med Sci Sports 11, Brock RW, Tschakovsky ME, Shoemaker JK, Halliwill JR, Joyner MJ & Hughson RL (1998) Effects of acetylcholine and nitric oxide on forearm blood flow at rest and after a single muscle contraction J Appl Physiol 85, Buckwalter JB & Clifford PS (2001) The paradox of sympathetic vasoconstriction in exercising skeletal muscle ExercSportSciRev29, Chance B, Ma HY & Nioka S (2003) Quantitative brain tissue oximetry, phase spectroscopy and imaging the range of homeostasis in piglet brain In Oxygen Transport to Tissue XXIV, ed JF Dunn & HM Swartz, pp New York: Kluwer Academic/Plenum Publishers Clifford PS & Hellsten Y (2004) Vasodilatory mechanisms in contracting skeletal muscle J Appl Physiol 97, DeLorey DS, Kowalchuk JM & Paterson DH (2003) Relationship between pulmonary O 2 uptake kinetics and muscle deoxygenation during moderate-intensity exercise J Appl Physiol 95, Delp MD & Laughlin MH (1998) Regulation of skeletal muscle perfusion during exercise Acta Physiol Scand 162, Diederich ER, Behnke BJ, McDonough P, Kindig CA, Barstow TJ, Poole DC & Musch TI (2002) Dynamics of microvascular oxygen partial pressure in contracting skeletal muscle of rats with chronic heart failure Cardiovasc Res 56, Ferreira LF, Harper AJ & Barstow TJ (2006) Frequency-domain characteristics and filtering of blood flow following the onset of exercise: implications for kinetics analysis J Appl Physiol 100,

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