The effect of the menstrual cycle on human cerebral blood flow: studies using Doppler ultrasound

Transcription

1 Ultrasound Obstet Gynecol 1999;14:52 57 The effect of the menstrual cycle on human cerebral blood flow: studies using Doppler ultrasound K. J. Brackley*, M. M. Ramsay*, F. Broughton Pipkin* and P. C. Rubin Departments of *Obstetrics and Gynaecology and Therapeutics, University of Nottingham Medical School, Queen s Medical Centre, Nottingham, UK Key words: DOPPLER ULTRASOUND, CEREBRAL BLOOD FLOW, MENSTRUAL CYCLE ABSTRACT Objective Previous studies have demonstrated hemodynamic changes at different phases in the menstrual cycle, but the cerebral circulation has not been investigated. Our aim was to study carotid and cerebral blood flow during the menstrual cycle using Doppler ultrasound. Two different techniques of Doppler waveform analysis were used: standard Doppler indices and Laplace transform analysis (LTA), which may provide additional hemodynamic information. Design This was a prospective study of healthy volunteers who were providing pre-conception data for a subsequent longitudinal study set in the Department of Obstetrics and Gynaecology, Nottingham University Hospital. Nineteen women were studied in the mid-follicular and mid-luteal phases of 27 ovulatory menstrual cycles. Doppler recordings were obtained from the internal and external carotid and middle cerebral arteries. The standard Doppler indices (systolic/diastolic ratio, pulsatility index and resistance index) and LTA parameters were calculated. Results The standard Doppler indices were all significantly higher in the luteal compared to the follicular phase in the right middle cerebral artery (p < 0.05). However, no changes were seen in the standard indices in the carotid arteries or in any of the LTA parameters in any artery. Using the LTA, vessel wall stiffness was greater and absolute velocity of flow lower in the middle cerebral compared to the carotid arteries. Conclusions Increased ventilation and a subsequent lowering of alveolar CO 2 pressure secondary to a raised progesterone level in the mid-luteal phase could account for the observed changes within the middle cerebral artery. Under the conditions of this study the LTA appears less sensitive at detecting alterations in downstream resistance compared to the standard Doppler indices. INTRODUCTION The changing levels of estradiol and progesterone in the follicular and luteal phases of the menstrual cycle are likely to have vasoactive effects 1 5. Several investigators have explored systemic circulatory changes during the menstrual cycle, including variation in blood pressure and heart rate 4,6 11. Results have been conflicting, possibly because of differences in study design and inconsistent determination of cycle phase. Doppler ultrasound has been used increasingly to describe changes in ovarian and uterine blood flow during the menstrual cycle 12,13. However, there is a paucity of information concerning changes in other circulations. The existing reports in the literature have measured skin blood flow using Doppler techniques 4,14,15 and limb blood flow (mainly muscle bed vasculature) using venous occlusion plethysmography 11,16. The effect of the menstrual cycle on cerebral hemodynamics has not been investigated. An understanding of hormonal influences on cerebral blood flow is relevant when considering the importance of cerebral blood flow changes in certain pregnancy complications such as pre-eclampsia/eclampsia. The non-invasive technique of Doppler ultrasonography is ideally suited to provide serial measurements of cerebral blood flow in healthy volunteers. Laplace transform analysis (LTA) of Doppler signals may provide additional and more readily interpretable information regarding flow conditions compared to standard Doppler parameters such as the pulsatility index (PI) or the resistance index (RI) 17. The LTA is a mathematical method of Doppler waveform shape analysis which is based on an electrical analog model of the arterial circulation Previous work in our departments has demonstrated that the LTA parameters obtained using this technique relate to separate features of the arterial circulation, i.e. upstream and downstream flow conditions and vessel wall stiffness or tone 17,22. The LTA parameters are not affected by heart rate because calculations are Correspondence: Dr K. J. Brackley, Department of Fetal Medicine, Birmingham Women s Hospital, Egbaston, Birmingham, B15 2TG, UK ORIGINAL PAPER 52 Received Revised Accepted

2 performed in the frequency domain, the original Doppler signal having been described in terms of its normalized Fourier transform 23. This differs from the standard Doppler indices which are all affected by the duration of each cardiac cycle, i.e. heart rate. The normal ranges for LTA parameters within the external and internal carotid arteries in 26 non-pregnant women were determined in a cross-sectional study by Ramsay 22, but repeated measurements within the same individual at different phases of the menstrual cycle were not performed. Recently, we have reported the normal ranges for the LTA parameters during pregnancy and the puerperium in a variety of maternal vessels including the carotid and middle cerebral arteries 24. The current study was performed in order to obtain pre-conception data for this longitudinal study. The aims of this study were to investigate menstrual cycle changes in blood flow within the cerebral circulation using standard Doppler indices and the LTA technique. We anticipated that extra hemodynamic information would be available concerning cerebral blood flow using LTA. Compared to the carotid arteries, we considered the middle cerebral artery to be the vessel most likely to demonstrate any hemodynamic differences secondary to cerebral effects of hormones, as it is an end-organ vessel. METHODS Nineteen women of childbearing age were recruited. All subjects were healthy with no known cardiovascular, renal or metabolic disease. All of the subjects were trying to conceive and had therefore stopped taking any oral contraceptives at least 3 months prior to the study. Written informed consent was obtained and the study was approved by the University Hospital Ethics Committee. Recordings were made in the mid-follicular (days 6 8 from the start of the last menstrual period) and the mid-luteal (days 20 22) phases of the menstrual cycle. An ovulatory cycle was confirmed by a serum progesterone level of > 40 nmol/l on day 21. Studies were performed for a maximum of three consecutive menstrual cycles. Subjects were in a relaxed semi-recumbent position for the Doppler recordings. Doppler signals were obtained from the external and internal carotid arteries and middle cerebral arteries as previously described 24. At least waveforms were recorded from each vessel onto videotape for later identification and analysis. Recordings from both right and left sides were made if time permitted, each study lasting approximately 30 min. A Kontron Sigma 44 HVCD ultrasound scanner was used, the power output of which complies with the limits required by the Food and Drug Administration (FDA) in the imaging and Doppler modes. An annular array 7.5-MHz sector scanning probe with 8-MHz pulsed wave Doppler was used to image and record signals from the external and internal carotid arteries, aided by color flow imaging. A 2-MHz pencil probe was used to obtain middle cerebral artery signals using the technique described by Aaslid and co-workers 25. Pulsed wave Doppler was used to record signals from a depth of approximately 50 mm. Mean blood pressure was determined over the period of each min study visit, using 10-min interval measurements with an automatic sphygmomanometer (Dinamap, Critikon Inc., Tampa, Florida, USA). A stabilization period of 10 min was allowed. Mean heart rate was obtained from the analysis of the Doppler signals. The Doppler waveforms were analyzed using a Dopstation computer (Scimed, Fishponds, Bristol, UK) as previously described 22,24,26. The Dopstation digitizes the audio signal of the tape. Manual signal modification is performed and then several optimal Doppler signals can be frozen on the computer screen for automatic analysis. Depending on the maternal heart rate, the number of waveforms on the screen varied between six and nine. The averaged waveform from the maximum frequency envelopes was generated, and the standard waveform indices were calculated from this. Fourier transform analysis of the averaged signal was performed and the harmonic frequency content was displayed by the computer as a plot of normalized amplitude against angular frequency (up to 40 radians/s). The equivalent Laplace transform was calculated using a curve-fitting technique. The LTA parameters were determined from the roots of the Laplace transform equation. The LTA parameters obtained include the following. (1) Alpha On theoretical grounds, this is inversely proportional to the upstream vessel radius 17 and therefore relates to upstream flow conditions. Alpha increases in the presence of an upstream occlusion. Alpha has been shown to be less affected by changes in arterial pressure compared to the closely related damping factor (d) 17,22. (2) Natural frequency of oscillation (ω o or omega) This is theoretically proportional to the square root of Young s modulus (E) of elasticity of the vessel wall and is therefore concerned with vessel wall stiffness or tone. (3) Real pole This is inversely proportional to the square of the distal vessel radius and is related to changes in downstream resistance. Increasing downstream resistance is associated with larger values of real pole. (4) C-coefficient This is the most dominant coefficient in the Laplace transform equation, and differs from the above LTA parameters in that it is not related on a theoretical basis to a specific feature of the circulation. However, it has been shown to demonstrate an inverse relationship to changes in downstream resistance 22,27. Data were transferred to the university mainframe computer. Statistical analysis was performed using SPSSX-3 (Statistics Package for the Social Sciences, SPSSX, USA). Median and interquartile ranges for the LTA parameters and the standard Doppler indices were determined for each vessel for the follicular and luteal phases of the ovulatory menstrual cycle. There are some missing data for reasons Ultrasound in Obstetrics and Gynecology 53

3 that included limited study time, inability to obtain satisfactory signals (particularly of the middle cerebral arteries) in certain patients or signals being too weak when later analysis on the Dopstation was attempted. The Mann Whitney U test was used to make comparisons between groups, i.e. between different vessels and between right- and left-sided vessels. The Wilcoxon matched-pairs signed-rank test was used to compare consecutive follicular and luteal recordings within each menstrual cycle. Short- and long-term variabilities within a single individual were determined by calculating the percentage coefficients of variation (CV) for the standard Doppler indices and the LTA parameters. Repeated Doppler signals were recorded from the vascular territories under investigation over a period of 60 min to assess short-term variability and at for 10 consecutive days to assess longer-term variability. Table 1 Systemic blood pressure and heart rate during the menstrual cycle. Values are medians, with interquartile ranges in parentheses Systolic blood pressure (mmhg) Diastolic blood pressure (mmhg) Heart rate (beats/min) (n = 27) 107 ( ) 66 (61 71) 67 (58 70) Luteal phase (ovulatory) (n = 27) 108 (99 111) 65 (61 68) 66 (60 75) RESULTS The age range of the 19 subjects was years. Sixteen of the subjects were non-smokers and three were smokers. The subjects were all white Caucasians except one, who was of Afro-Caribbean origin. Data from a total of 27 paired follicular- and luteal-phase studies in ovulatory cycles were available for analysis from the 19 subjects. Four subjects provided two sets of consecutive data and two subjects provided three sets. Median and interquartile ranges for systolic and diastolic blood pressures and heart rate for the different cycle phases are shown in Table 1. No significant differences were observed in blood pressure or heart rate between follicular or luteal phases of the menstrual cycle. Median and interquartile ranges for the LTA parameters (damping, alpha, ω o, real pole and C-coefficient), systolic/ diastolic ratio (S/D ratio), PI and RI from right and left external and internal carotid arteries and middle cerebral arteries at different phases of the menstrual cycle are displayed in Tables 2 4. Real pole values are displayed as positive numbers in the tables. There were no significant differences between right- and left-sided vessels in any Doppler parameter at any cycle phase. Comparisons between cycle phase There were no obvious visual changes in waveform shape between cycle phase. There were no significant differences in LTA parameters between the follicular and luteal phases of the menstrual cycle from any artery under study. Table 2 In non-pregnant subjects: standard Doppler indices and Laplace transform analysis parameters during the menstrual cycle in the external carotid artery. Values are medians, with interquartile ranges in parentheses S/D ratio PI RI Damping Alpha ω o Real pole C-coefficient Luteal phase (ovulatory) Right (n = 24) Left (n = 24) Right (n = 24) Left (n = 24) 8.66 ( ) 2.52 ( ) 0.87 ( ) 0.18 ( ) 1.94 ( ) 21.8 ( ) 3.66 ( ) 0.30 ( ) 8.72 ( ) 2.78 ( ) 0.88 ( ) 0.17 ( ) 1.94 ( ) 23.0 ( ) 3.40 ( ) 0.32 ( ) 8.24 ( ) 2.48 ( ) 0.86 ( ) 0.16 ( ) 1.89 ( ) 21.9 ( ) 3.52 ( ) 0.31 ( ) S/D ratio, systolic/diastolic ratio; PI, pulsatility index; RI, resistance index; ω o, natural frequency of oscillation 9.26 ( ) 2.73 ( ) 0.88 ( ) 0.18 ( ) 1.99 ( ) 22.6 ( ) 3.42 ( ) 0.32 ( ) Table 3 In non-pregnant subjects: standard Doppler indices and Laplace transform analysis parameters during the menstrual cycle in the internal carotid artery. Values are medians, with interquartiles ranges in parentheses S/D ratio PI RI Damping Alpha ω o Real pole C-coefficient Luteal phase (ovulatory) Right (n = 22) Left (n = 21) Right (n = 22) Left (n = 21) 2.28 ( ) 0.87 ( ) 0.55 ( ) 0.15 ( ) 2.06 ( ) 27.5 ( ) 3.64 ( ) 0.33 ( ) 2.18 ( ) 0.81 ( ) 0.53 ( ) 0.13 ( ) 1.83 ( ) 27.1 ( ) 3.28 ( ) 0.36 ( ) 2.40 ( ) 0.90 ( ) 0.56 ( ) 0.15 ( ) 1.98 ( ) 25.8 ( ) 3.48 ( ) 0.34 ( ) S/D ratio, systolic/diastolic ratio; PI, pulsatility index; RI, resistance index; ω o, natural frequency of oscillation 54 Ultrasound in Obstetrics and Gynecology 2.26 ( ) 0.85 ( ) 0.54 ( ) 0.10 ( ) 1.80 ( ) 24.9 ( ) 3.39 ( ) 0.35 ( )

4 Table 4 In non-pregnant subjects: standard Doppler indices and Laplace transform analysis parameters during the menstrual cycle in the middle cerebral artery. Values are medians, with interquartile ranges in parentheses S/D ratio PI RI Damping Alpha ω o Real pole C-coefficient Luteal phase (ovulatory) Right (n = 15) Left (n = 10) Right (n = 15) Left (n = 10) 2.10* ( ) 0.72* ( ) 0.51* ( ) 0.11 ( ) 1.79 ( ) 31.2 ( ) 3.72 ( ) 0.34 ( ) 2.02** ( ) 0.68 ( ) 0.48 ( ) 0.12 ( ) 2.03 ( ) 33.2 ( ) 3.75 ( ) 0.36 ( ) 2.29 ( ) 0.81 ( ) 0.54 ( ) 0.08 ( ) 1.47 ( ) 30.0 ( ) 3.63 ( ) 0.35 ( ) 2.24 ( ) 0.83 ( ) 0.53 ( ) 0.10 ( ) 1.82 ( ) 31.1 ( ) 3.48 ( ) 0.34 ( ) S/D ratio, systolic/diastolic ratio; PI, pulsatility index; RI, resistance index; ω o, natural frequency of oscillation; *, follicular < luteal (ovulatory) phase p < 0.05; **, follicular < luteal (ovulatory) phase p = Increases in the standard Doppler indices were noted in the luteal phase compared to the follicular phase in the middle cerebral arteries but not in the external and internal carotid arteries. The S/D ratio was significantly higher in the luteal phase in both right middle cerebral artery (n = 15; p < 0.05) and left middle cerebral artery (n = 10; p = 0.005). The PI and RI were also higher in the luteal phase in the middle cerebral arteries but significance was only reached on the right side (p < 0.05). Comparisons between vessels ω 0 was significantly higher in the middle cerebral arteries compared to the external carotid artery (p < ). Values were also greater in the middle cerebral arteries compared to the internal carotid artery, but significance was only reached in the left-sided vessels. Alpha and damping values were generally lower in the middle cerebral arteries compared to the internal carotid artery and particularly when compared to the external carotid artery. Real pole and C-coefficient were generally comparable within the internal carotid and middle cerebral arteries and within the external carotid and middle cerebral arteries. However, the S/D ratio, PI and RI were consistently lower in the middle cerebral arteries compared to the external carotid artery. The standard Doppler indices were significantly lower in the middle cerebral arteries compared to the internal carotid artery during the follicular phase on the right but not on the left side. Coefficients of variation Damping was the most unstable of the LTA parameters at all sites. The CV for damping was highest within the internal carotid artery over the long term (29.9%) but was more acceptable (< 15%) for the other arteries. The other LTA parameters had satisfactory CVs (< 11%) within each artery under investigation over both short- and long-term periods. ω o was the most stable LTA parameter within each vessel. The standard Doppler indices had acceptable CVs: < 8% within all arteries except for the S/D ratio in the external carotid artery which was very unstable (CVs > 20%). DISCUSSION The normal ranges for the LTA parameters and standard Doppler indices have now been determined for the internal and external carotid and middle cerebral arteries in a group of healthy non-pregnant women according to the phase of the menstrual cycle. The different vessels have distinctive Doppler signals because of the particular flow conditions present. Similarly, each artery has been shown to have characteristic normal ranges for the Doppler indices consistent with the typical hemodynamic conditions. The LTA parameters provide extra information regarding these flow conditions which are not available with the standard Doppler indices, i.e. PI, RI and S/D ratio. Our data are comparable to the normal non-pregnant ranges quoted by Ramsay 22 and the postnatal data we have previously published 24. The smaller cerebral vessels have the greatest stiffness or tone (higher values for ω o) but lower absolute velocity of flow (alpha) despite being of a narrower caliber compared to the carotid vessels. There is a low downstream resistance (small real pole and high C-coefficient values). In postmortem studies, intracranial arteries from patients below middle age have been shown to be significantly stiffer compared to extracranial arteries 28. These findings were mainly attributed to the thin wall (or low thickness/radius ratio) as well as the high elastic modulus of the vessel wall material. Doppler parameters within the external carotid artery reflect the more elastic vessel walls (lower ω o ), higher flow velocity and smaller upstream vessel diameter (increased alpha) and higher downstream resistance (lower C-coefficient and increased PI, RI and S/D ratio). Using Doppler ultrasound, a significant change in flow conditions has been demonstrated within the intracranial cerebral circulation according to the phase of the menstrual cycle. The standard Doppler indices (S/D ratio, PI and RI) are generally considered to reflect downstream resistance to flow. The higher values within the middle cerebral arteries in the luteal phase compared to the follicular phase suggest an increased downstream resistance to flow in this vascular bed. In contrast, no significant change was detected in the internal carotid artery, i.e. the extracranial cerebral circulation. The increase in standard Doppler indices in the luteal phase could not be attributed to a change in central Ultrasound in Obstetrics and Gynecology 55

5 circulatory conditions as no change in heart rate or blood pressure was detected. The PI can change if an upstream stenosis is present in the circulation 17,29 but there were no differences noted in damping or alpha, the LTA parameters reflecting upstream flow conditions. One of the many mechanisms involved in cerebral blood flow regulation is a change in arterial carbon dioxide (CO 2) pressure 30.CO 2 has a strong vasodilatory effect on cerebral vessels, particularly on smaller pial arteries and arterioles. A simple linear relationship has been demonstrated between alveolar CO 2 pressure and average blood velocity in the internal carotid and vertebral arteries using Doppler velocimetry 31. An increase in ventilation and a subsequent decrease in alveolar CO 2 pressure are known to occur in the luteal phase of the menstrual cycle, secondary to a progestogenic effect Vasoconstriction leading to a decrease in cerebral blood flow would be expected, and this is consistent with the changes we have described in the standard Doppler indices in the middle cerebral arteries. Transcranial Doppler ultrasound measurements performed on anesthetized men have previously demonstrated increases in PI and RI during hypocapnia that are consistent with the observed reduction in cerebral blood flow measured using the Kety Schmidt technique 35. Interestingly, an increase in the standard Doppler indices from pre-pregnant follicular phase levels was detectable by 4 7 weeks gestation in both the internal carotid and middle cerebral arteries 24. Progesterone is also known to stimulate ventilation in early pregnancy 36. The physiological fluctuations within the normal menstrual cycle would be expected to be small and this may explain why changes are seen in only the more sensitive narrower cerebral vessels rather than the larger internal carotid arteries. In addition, we had previously hypothesized that the middle cerebral artery, being an end-organ vessel, would be more likely to demonstrate any cerebral effects of hormones compared to the carotid arteries. The real pole and C-coefficient did not change significantly in the middle cerebral arteries during the menstrual cycle. An increase in real pole and a decrease in C-coefficient values would be expected with a rise in downstream resistance. The reason for the inconsistent findings between these LTA parameters and the standard Doppler indices is not clear. Previous validation studies involving the LTA technique have suggested that the standard Doppler indices were less reliable at detecting changes in flow conditions compared to the LTA parameters. For example, both the PI and the RI were shown to increase in the presence of upstream or downstream occlusions 22.Inin vivo experiments using the brachial artery, the RI increased in size in the presence of a downstream occlusion as well as during a Valsalva maneuver which lowered blood pressure 17. Similarly, the S/D ratio and PI were not significantly altered during the Valsalva maneuver even though the shape of the Doppler waveform obviously changed. In contrast, the various LTA parameters were able to detect separate changes in arterial pressure and upstream and downstream occlusions. However, the ability of the standard Doppler indices to reflect changes in downstream resistance alone was confirmed by these last experiments 17. There is considerable experimental evidence, particularly in the fetoplacental circulation, that S/D ratio, PI and RI reflect downstream hemodynamic changes However, this relationship is dependent on steady-state conditions in the circulation. It should be noted that the degree of downstream occlusions generated in the validation experiments 17,22 would be relatively marked. The LTA technique may therefore lack the sensitivity required to detect minor alterations in downstream flow conditions in the middle cerebral arteries which occur during the menstrual cycle. During controlled ovarian hyperstimulation following pituitary suppression, Shamma and associates 41 were able to demonstrate a significant increase in peak systolic velocity and PI in the middle cerebral arteries associated with the marked rise in estrogen levels, but there was no statistically significant association with serum progesterone levels. It is recognized that some variability in results during the mid-follicular phase can be accounted for by individual variability in endocrine status, especially circulating estradiol levels, although the majority of our subjects reported regular day cycles. An alternative explanation for the discordant findings in our study is that the LTA parameters reflect the real situation in that there is no change in the cerebral circulation with cycle phase, i.e. the standard waveform analysis techniques are falsely positive (a type I error). We acknowledge the possibility of a type I error in view of multiple comparisons that are made in the study, but point out that there were a priori reasons for expecting differences to be more apparent in the middle cerebral arteries. Larger numbers of women would need to be investigated with measurement of arterial/alveolar CO 2 pressures to clarify the situation. In the present study, the effects of the menstrual cycle phase on cerebral hemodynamics have been investigated for the first time using Doppler ultrasound. Standard Doppler indices suggest that there is an increased downstream resistance to blood flow within the middle cerebral artery in the luteal compared to the follicular phase, a phenomenon that could be attributed to decreased alveolar CO 2 pressure subsequent to the progestogenic stimulus to ventilation. Blood flow within the external and internal carotid arteries is not affected by the menstrual cycle. The Laplace transform analysis technique provides additional Doppler data concerning influences on the circulation including vessel wall tone and upstream flow conditions. Real pole, the Laplace transform parameter pertaining to downstream resistance to flow, appears to be less sensitive compared to the standard Doppler indices at detecting the potentially minor alterations in flow conditions in the middle cerebral artery during the menstrual cycle. Future work with this alternative Doppler waveform analysis technique will investigate its use in normal and hypertensive pregnancies. ACKNOWLEDGEMENT This study was funded by the Sir Jules Thorn Charitable Trust. 56 Ultrasound in Obstetrics and Gynecology

6 REFERENCES 1. McCalden TA. The inhibitory action of oestradiol-17-β and progesterone on venous smooth muscle. Br J Pharmacol 1975; 53: Rylance PB, Brincat M, Lafferty K, De Trafford JC, Brincat S, Parsons V. Natural progesterone and antihypertensive action. Br Med J 1985;290: Magness RR, Rosenfeld CR. Local and systemic estradiol- 17β: effects on uterine and systemic vasodilation. Am J Physiol 1989;256:E Hassan AAK, Garter G, Tooke JE. Postural vasoconstriction in women during the normal menstrual cycle. Clin Sci 1990; 78: Van Buren GA, Da-seng Yang, Clark KE. Estrogen-induced uterine vasodilatation is antagonized by L-nitroarginine methyl ester, an inhibitor of nitric oxide synthesis. Am J Obstet Gynecol 1992;1667: Littler WA, Bojorges-Bueno R, Banks J. Cardiovascular dynamics in women during the menstrual cycle and oral contraceptive therapy. Thorax 1974;29: Greenberg G, Imeson JD, Thompson SG, Meade TW. Blood pressure and the menstrual cycle. Br J Obstet Gynaecol 1985; 92: Manhem K, Jern S. Influence of daily-life activation on pulse rate and blood pressure changes during the menstrual cycle. J Hum Hypertens 1994;8: Kelleher C, Joyce C, Kelly G, Ferriss JB. Blood pressure alters during the normal menstrual cycle. Br J Obstet Gynaecol 1986;93: Dunne FP, Barry DG, Ferriss JB, Grealy G, Murphy D. Changes in blood pressure during the normal menstrual cycle. Clin Sci 1991;81: Bartelink ML, Wollersheim H, Theeuwes A, v. Duren D, Theien T. Changes in skin blood flow during the menstrual cycle: the influence of the menstrual cycle on the peripheral circulation in healthy female volunteers. Clin Sci 1990;78: Scholtes MCW, Wladimiroff JW, van Rijen HJM, Hop WCJ. Uterine and ovarian flow velocity waveforms in the normal menstrual cycle: a transvaginal study. Fertil Steril 1989;52: Sladkevicius P, Valentin L, Marsal K. Blood flow velocity in the uterine and ovarian arteries during the normal menstrual cycle. Ultrasound Obstet Gynecol 1993;3: Lafferty K, De Trafford JC, Potter C, Roberts VC, Cotton LT. Reflex vascular responses in the finger to contralateral thermal stimuli during the normal menstrual cycle: a hormonal basis to Raynaud s phenomenon? Clin Sci 1985;68: Bungum L, Kvernebo K, Oian P, Maltau JM. Laser dopplerrecorded reactive hyperaemia in the forearm skin during the menstrual cycle. Br J Obstet, Gynaecol 1996;103: Plismane SO, Ozolin PP, Merten AA. Effect of the ovarian cycle on blood supply to the limbs during exertion. Hum Physiol 1984;10: Ramsay MM, Broughton Pipkin F, Rubin PC, Skidmore R. Waveform shape analysis: extraction of physiologically relevant information from Doppler recordings. Clin Sci 1994;86: Skidmore R. The use of the transcutaneous ultrasonic flowmeter in the dynamic assessment of blood. PhD thesis, University of Bristol, Skidmore R, Woodcock JP. Physiological interpretation of Doppler-shift waveforms. I. Theoretical considerations. Ultrasound Med Biol 1980;6: Skidmore R, Woodcock JP. Physiological interpretation of Doppler-shift waveforms. II. Validation of the Laplace transform method for characterisation of the common femoral blood-velocity/time waveform. Ultrasound Med Biol 1980;6: Skidmore R, Woodcock JP, Wells PNT, Bird D, Baird RN. Physiological interpretation of Doppler-shift waveforms. III. Clinical results. Ultrasound Med Biol 1980;6: Ramsay MM. Cardiovascular studies in pregnant and nonpregnant women using Doppler ultrasound. MD thesis, University of Cambridge, Milnor WR. Hemodynamics. Baltimore: Williams and Wilkins, Brackley KJ, Ramsay MM, Broughton Pipkin F, Rubin PC. A longitudinal study of maternal bloodflow in normal pregnancy and the puerperium: analysis of Doppler waveforms using Laplace transform techniques. Br J Obstet Gynaecol 1998; 105: Aaslid R, Markwalder T-M, Nornes H. Noninvasive transcranial Doppler ultrasound recording of flow velocity in basal cerebral arteries. J Neurosurg 1982;57: Brackley KJ. Maternal blood pressure and the fetus: investigations using Doppler ultrasound. DM thesis, University of Nottingham, Stone PR, MacPhail S, Bull SB, Morrow RJ. Use of Laplace transform analysis to describe the effect of placental embolisation on umbilical arterial Doppler waveforms in fetal sheep. Ultrasound Med Biol 1994;20: Hayashi K, Handa H, Nagasawa S, Okumura A, Moritake K. Stifness and elastic behaviour of human intracranial and extracranial arteries. J Biomechanics 1980;13: Evans DH, MacPherson DS, Bentley S, Asher MJ, Bell PRF. The effect of proximal stenosis on Doppler waveforms: a comparison of three methods of waveform analysis in an animal model. Clin Phys Physiol Meas 1981;2: Ursino M. Mechanisms of cerebral blood flow regulation. Crit Rev Biomed Eng 1991;18: Hauge A, Thoresen M, Walloe L. Changes in cerebral blood flow during hyperventilation and CO2-breathing measured transcutaneously in humans by a bidirectional, pulsed, ultrasound doppler blood velocitymeter. Acta Physiol Scand 1980; 110: Milne JA, Pack AI, Coutts JRT. Gas exchange and acid-base status during the normal human menstrual cycle and in subjects taking oral contraceptives. Proc Soc Endocrinol 1977;75: 17P 18P 33. England SJ, Farhi LE. Fluctuations in alveolar CO2 and in base excess during the menstrual cycle. Respir Physiol 1976; 26: Dutton K, Blanksby BA, Morton AR. CO 2 sensitivity during the menstrual cycle. J Appl Physiol 1989;67: Weyland A, Stephan H, Grune F, Weyland W, Sonntag H. Effect of ketanserin on global cerebral blood flow and middle cerebral artery velocity. Anesth Analg 1995;80: Spatling L, Fallenstein F, Huch A, Huch R, Rooth G. The variability of cardiopulmonary adaptation to pregnancy at rest and during exercise. Br J Obstet Gynaecol 1992;99(Suppl 8): Trudinger BJ, Stevens D, Connelly A, Hales JRS, Alexander G, Bradley L, Fawcett A, Thompson RS. Umbilical artery flow velocity waveforms and placental resistance: the effects of embolization of the umbilical circulation. Am J Obstet Gynecol 1987;157: Giles WB, Trudinger BJ, Baird PJ. Fetal umbilical artery flow velocity waveforms and placental resistance: pathological correlation. Br J Obstet Gynaecol 1985;92: Irion GL, Clark KE. Direct determination of the ovine fetal umbilical artery blood flow waveform. Am J Obstet Gynecol 1990;162: Morrow RJ, Adamson SL, Bull SB, Knox Ritchie JW. Effect of placental embolization on the umbilical arterial velocity waveform in fetal sheep. Am J Obstet Gynecol 1989;161: Shamma FN, Fayad P, Brass L, Sarrel P. Middle cerebral artery blood velocity during controlled ovarian hyperstimulation. Fertil Steril 1992;57: Ultrasound in Obstetrics and Gynecology 57