Venous occlusion plethysmography in cardiovascular research: methodology and clinical applications (2001)

The use of venous occlusion plethysmography to measure blood flow in humans was first described over 90 years ago by Hewlett & van Zwaluwenburg [1]. Although initially limited by the need for cumbersome equipment, the advent of mercury-in-silastic strain gauges over 50 years ago greatly increased its applicability, and the technique has remained essentially unchanged since except for the introduction of computerization. Venous occlusion plethysmography has been used extensively to study human vascular physiology in vivo, and is at its most powerful when combined with intra-arterial drug administration, usually into the forearm vascular bed. This permits detailed study of both vascular pharmacology, usually by infusion of receptor agonists, and physiology, by infusion of receptor antagonists. Indeed, forearm venous occlusion plethysmography with local brachial artery infusion has become one of the ‘gold-standards’ in the assessment of vascular function in health and disease, and an accurate, reproducible and convenient method with which to assess the effect of new vasoactive drugs and hormones in humans in vivo[2]. Besides being minimally invasive, the major advantage of this approach is that drugs can be administered at subsystemic doses minimizing disturbances to systemic physiology. Although venous occlusion plethysmography can be equally well applied to the lower limb, this review will focus on the forearm vascular bed. The underlying principle of forearm venous occlusion plethysmography is simple: when venous drainage from the arm is briefly interrupted, arterial inflow is unaltered and blood can enter the forearm but cannot escape. This results in a linear increase in forearm volume over time, which is proportional to arterial blood inflow, until venous pressure rises towards the occluding pressure [3]. Under resting conditions, ∼70% of total forearm blood flow (FBF) is through skeletal muscle, with skin blood flow accounting for most of the remainder [4]. However, the hand contains a high proportion of arterio-venous shunts and, because skin blood flow is highly dependent on temperature [5] and has a different basic pharmacology and physiology than muscle blood flow, it is standard practice to exclude the hands from the circulation during measurement of forearm blood flow. Indeed, if the hand is not excluded then blood flow is nonlinear [6]. The basic methodology has changed little since its first description by Hewlett & Zwaluwenburg over 90 years ago [1]. Venous return from the forearm is briefly interrupted by inflating a cuff, placed around the upper arm, to well above venous pressure but below diastolic pressure. Typically, an inflation pressure of around 40 mmHg is used for intervals of 10 s, followed by 5 s of deflation, which does not alter arterial inflow and allows venous emptying [7]. The forearm must be positioned above the level of the heart to ensure adequate venous emptying during the period of deflation, which is usually achieved by resting the elbows on foam pads and supporting the hands with pillows (Figure 1). The hands are excluded from the circulation during measurements by initial rapid inflation of a smaller cuff, placed around the wrist, to well above systolic pressure (220 mmHg for normotensive subjects). The wrist-cuffs must be inflated at least 60 s before starting measurements of flow in order to allow FBF to stabilize [8]. As this manoeuvre renders the hand ischaemic, the measurement period is limited, but periods of up to 13 min have been employed safely [9]. Automated venous occlusion plethysmography equipment has been developed [10] and is of particular use when on-line measurement of flow is required, as discussed later. Assessment of forearm blood flow using venous occlusion plethysmography. Changes in forearm volume are measured by a plethysmograph. Initially, air and then water-filled jackets were used, but these have been largely replaced by mercury-in-rubber (or silastic) strain gauges [11], which may themselves be ultimately replaced by indium-gallium gauges due to concerns over the potential toxicity of mercury. The strain gauges should be placed around the widest part of the forearm, and simply act as resistors connected as one arm of a Wheatstone bridge [6]. Changes in forearm volume result in a corresponding change in arm circumference and thus strain gauge length, which can be detected as an alteration in electrical resistance of the gauge, and thus potential difference (Figure 2) [11]. If the gauge length is made equal to the resting circumference of the limb, then changes in limb volume are directly proportional to the changes in resistance [12]. Effect of intra-arterial substance P on forearm blood flow. 2 pmol min−1 substance P produces a marked increase in blood flow in the infused arm, as illustrated by the increase in the slope of the tracing. Venous occlusion plethysmography provides a measure of blood flow to that part of the forearm enclosed by the two cuffs. This is usually expressed as ml per 100 ml of forearm volume per minute, when electronic calibration is employed [6, 12]. Actual forearm volume can be estimated by calculation, assuming the forearm is a simple truncated cone, or by simple water displacement. When studies are conducted in a quiet, temperature-controlled room (22–26 ° C), with the subject resting in a comfortable supine position, measurement of FBF by strain gauge venous occlusion plethysmography compares favourably with values obtained by other established techniques [13, 14]. More recently, strain gauges employing a nonelastic loop attached to a passive inductive transducer, which acts as an electromagnetic sensor, have been developed and appear to be at least as accurate as mercury gauges [15]. Blood flow can also be measured using ultrasound. Usually this involves combining estimates of mean blood velocity with the cross sectional area of the vessel. Doppler ultrasound is used to estimate the mean blood velocity, and B-mode ultrasound used to estimate the vessel diameter. Measurement of mean velocity is most accurate when the ultrasound beam-width is greater than the vessel diameter; with modern ultrasound scanners the 3dB beam width is typically 1–2 mm, so that this condition is not met in most vessels of interest, leading to an overestimation of mean velocity. Assessment of vessel diameter relies on the ability to image the anterior and posterior vessel walls and depends on the angle of the probe to the artery. As the area is obtained by squaring the diameter, any error in diameter estimation results in doubling of the error for area; in general it is difficult to measure area in vessels less than about 3–4 mm in diameter, though this is dependent on the location of the vessel and the frequency of the transducer used, with greater accuracy being achieved if high frequency transducers are used for scanning superficial vessels. Typically, in vessels of 5–10 mm in diameter, the flow may be measured at best with an accuracy of ∼20–30%. In routine practice, using the above technique, it is easy to generate very large errors of ∼50 or even 100% in flow estimation, but in the experimental setting improved accuracy, below 10%, may be possible [16]. Overall, the use of ultrasound does not confer any particular advantage over plethysmography in assessing blood flow, although it does allow conduit vessel diameter to be determined at the same time. Moreover, ultrasonic methods require expensive equipment and highly skilled operators and can be difficult to use with intra-arterial needles. Following limb ischaemia there is a rapid increase in forearm blood flow, which slowly returns to baseline values and is termed reactive hyperaemia. The maximum flow and area under the curve for this increase in blood flow are directly related to the duration of ischaemia [17] until ∼13 min, after which lengthening the period of ischaemia does not increase the maximum recorded FBF further [9]. Indeed, the maximal post-ischaemic FBF is not influenced by sympathetic tone [18] or administration of vasodilators [9] and seems to represent the minimum forearm vascular resistance, which is a function of the average wall:lumen ratio of the resistance vessels [19]. Therefore, minimum vascular resistance provides indirect information concerning the structure of resistance vessels in the forearm. Moreover, minimum forearm vascular resistance correlates well with more direct measurements of resistance vessel structure, such as the media:lumen ratio [20]. Increased minimum vascular resistance has been demonstrated in individuals with established hypertension [19, 21], young men with ‘borderline’ hypertension [18], and those with ‘white-coat’ hypertension [22]. Interestingly, such structural changes can be reversed with antihypertensive medication [23–25]. Minimum vascular resistance can be assessed reproducibly in the forearm following 13 min of ischaemia, induced by inflating a cuff sited around the upper arm to 300 mmHg for 13 min, and asking subjects to perform 20–30 hand contractions during the last 1 min of ischaemia [9]. As is usual, the hands should be excluded from the circulation before starting the study and measurements of FBF, using venous occlusion plethysmography, should be made at 15 s intervals for 3 min following ischaemia. 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