Ocular Hemodynamics during Isometric Exercise

Microvascular Research 61, 1–13 (2001) doi:10.1006/mvre.2000.2269, available online at http://www.idealibrary.com on

Ocular Hemodynamics during Isometric Exercise Barbara Kiss,* Susanne Dallinger,* Kaija Polak,* Oliver Findl,† Hans-Georg Eichler,* and Leopold Schmetterer* ,‡ *Department of Clinical Pharmacology, ‡Institute of Medical Physics, and †Department of Ophthalmology, University of Vienna, Vienna, Austria Received November 30, 1999; published online November 14, 2000

The autoregulatory capacity of the human retina is well documented, but the pressure–flow relationship of the human choroid is still a matter of controversy. Recent data, using laser Doppler flowmetry to measure choroidal blood flow, indicate that the choroid has some autoregulatory potential, whereas most data using other techniques for the assessment of choroidal hemodynamics indicate that the choroidal pressure–flow curve is linear. We used a new laser interferometric technique to characterize choroidal blood flow during isometric exercise. Twenty healthy subjects performed squatting for 6 min during normocapnia and during inhalation of 5% CO 2 and 95% air. Ocular fundus pulsation amplitude, flow velocities in the ophthalmic artery, intraocular pressure, and systemic hemodynamics were measured in 2-min intervals. To gain information on choroidal blood flow fundus pulsation amplitude was corrected for changes in flow pulsatility using data from the ophthalmic artery and for changes in pulse rate. Ocular perfusion pressure was calculated from mean arterial pressure and intraocular pressure. The ocular pressure–flow relationship was calculated by sorting data according to ascending ocular perfusion pressure values. In a pilot study in 6 healthy subjects comparable ocular pressure flow relationships were obtained when choroidal blood flow was assessed with the method described above and with laser Doppler flowmetry. In the main study isometric exercise caused a significant increase in mean arterial pressure (56%, P < 0.001), pulse rate (84%, P < 0.001), and intraocular pres0026-2862/00 $35.00 Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

sure (37%, P 0.004), but decreased fundus pulsation amplitude (ⴚ36%, P < 0.001). Significant deviations from baseline choroidal blood flow were observed only at ocular perfusion pressures >69% during normocapnia and 70% during hypercapnia. Our data indicate that during isometric exercise the choroid has a high capacity to keep blood flow constant despite changes in perfusion pressure and that this pressure–flow relationship is not altered by moderate changes in arterial carbon dioxide levels. © 2000 Academic Press Key Words: choroidal blood flow; isometric exercise; autoregulation; ocular fundus pulsation; ocular blood flow; human.

INTRODUCTION Autoregulation is the ability of a vascular bed to maintain blood flow despite changes in perfusion pressure. Much effort has been directed to study the autoregulatory capacity of the ocular vasculature. There is evidence from a variety of studies that the human retina shows some autoregulatory capacity in response to both increase and decrease in ocular perfusion pressure (Riva et al., 1986; Robinson et al., 1986; Dumsky et al., 1996). By contrast, the autoregulatory capacity of the choroidal vasculature is still a matter of controversy. Early studies using the radioactively labeled micro-

1

2

sphere technique indicated that the choroid is a passive vascular bed (Alm and Bill, 1972, 1973b). A few years ago, evidence for an autoregulatory capacity of the choroid came from studies in rabbits during both experimental increase and decrease in ocular perfusion pressure (Kiel, 1994; Kiel and van Heuven, 1995). More recently, Riva and co-workers (Riva et al., 1997a) have shown that the human choroid exhibits some autoregulation in response to moderate decreases in perfusion pressure. Results of the same group (Riva et al., 1997b) indicate that choroidal blood flow remains constant when an isometric exercise-induced increase in ocular perfusion pressure does not exceed 67%. The above mentioned results (Kiel, 1994; Kiel and van Heuven, 1995; Riva et al., 1997a,b) were obtained from experiments using laser Doppler flowmetry (LDF) for quantification of choroidal blood flow. Kiel (1994) discussed possible explanations for the discrepancies between LDF studies and studies assessing ocular blood flow with other techniques in detail. However, it is still a matter of discrepancy whether the choroid is autoregulated. Hence, we decided to study human choroidal blood flow during isometric exercise with an alternative method for the assessment of choroidal hemodynamics, namely, laser interferometric measurement of fundus pulsation. A pilot study was done to investigate whether this approach to study choroidal hemodynamics provides results comparable with those of LDF. In the main study subjects performed isometric exercise in the absence and the presence of hypercapnia to gain insight into the mechanisms behind choroidal blood flow regulation.

MATERIALS AND METHODS Subjects After approval from the Ethics Committee of Vienna University School of Medicine was obtained, 6 healthy male volunteers (age range, 20 –27 years; mean ⫾SD, 23.2 ⫾ 2.6 years) participated in the pilot study and 20 healthy male volunteers (age range, 19 –34 years; mean ⫾SD, 26.1 ⫾ 3.1 years) participated in the main study. The nature of the study was explained and all subjects

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

Kiss et al.

gave written consent to participate. Each subject passed a screening examination that included medical history and physical examination, 12-lead electrocardiogram, complete blood count, activated partial thromboplastin time, thrombin time, fibrinogen, clinical chemistry (sodium, potassium, creatinine, uric acid, glucose, cholesterol, triglycerides, alanine aminotransferase, aspartate aminotransferase, ␥-glutamyltransferase, alkaline phosphatase, total bilirubin, total protein), hepatitis A, B, C and HIV serology, urine analysis, and a urine drug screen. Subjects were excluded if any abnormality was found as part of the screening unless the investigators considered an abnormality clinically irrelevant. Furthermore, an ophthalmic examination, including slit lamp biomicroscopy and indirect funduscopy, was performed. Inclusion criteria were normal ophthalmic findings and a refractive error of less than 3 diopters in either eye.

Study Design On the trial day subjects arrived after an overnight fast. After steady-state conditions were reached, which was ensured by repeated blood pressure measurements during the resting period, hemodynamic baseline measurements were taken. Thereafter the subjects squatted for 6 min in a position where the upper and the lower leg were as close to a right angle as possible, without leaning against a wall or support. Systemic and ocular hemodynamics were assessed every 2 min during isometric exercise. Blood gas analysis was performed at baseline and at the end of the squatting period. Thereafter a resting period was scheduled. When systemic hemodynamics had returned to baseline, an additional 6-min squatting period was performed by the subjects under study. During this period intraocular pressure (IOP) and systemic hemodynamics were measured in 2-min intervals. Data were only included for analyses if the deviation between exercise-induced changes in blood pressure from the two squatting periods were within 10%. After another resting period subjects repeated the procedure during inhalation of 5% CO 2 ⫹ 95% air. The two inhalation periods lasted for 10 min each. Squatting was performed during the last 6 min of 5% CO 2 ⫹

Ocular Hemodynamics in Man

95% air breathing. Blood gas analysis was done at baseline, just before the start of isometric exercise after 4 min of gas inhalation, and at the end of the squatting period. This study design was chosen because it was technically not possible to measure all parameters simultaneously. In the pilot study three squatting periods were performed by each subject. The first two were identical to those in the main study. During the third squatting period LDF was performed continuously and systemic hemodynamics were assessed every 2 min.

Methods of Evaluation Blood pressure and pulse rate. Systolic, diastolic, and mean blood pressures (SBP, DBP, MAP) were measured on the upper arm by an automated oscillometric device (HP-CMS patient monitor, Hewlett– Packard, Palo Alto, CA). Pulse rate (PR) was automatically recorded from a finger pulse oxymetric device (HP-CMS patient monitor). Fundus pulsations. Pulse synchronous pulsations of the eye fundus (Fercher, 1984) were assessed by laser interferometry on the subject’s right eye. The method is described in detail by Schmetterer et al. (1995). Briefly, the eye is illuminated by the beam of a single mode laser diode with a wavelength (␭) of 783 nm. The laser beam has a diameter of approximately 1 mm at the cornea and is focused to a spot of approximately 50 ␮m at the fundus. The light is reflected at both the front side of the cornea and the retina. The two reemitted waves produce interference fringes from which the distance changes between cornea and retina during a cardiac cycle can be calculated. Distance changes between cornea and retina lead to a corresponding variation of the interference order (⌬N(t)). This change in interference order can be evaluated by counting the fringes moving inward and outward during the cardiac cycle. Changes in optical distance (⌬L(t)), corresponding to the cornea–retina distance changes, can then be calculated by ⌬L(t) ⫽ ⌬N(t) 䡠 ␭ / 2. The maximum distance change is called the fundus pulsation amplitude (FPA). FPA was calculated as the mean of at least five cardiac cycles. The short-term and day-to-day variability of the method is small, which allows detection of even small changes in

3

local pulsatile blood flow following pharmacological stimulation (Schmetterer et al., 1997a). To obtain information on the choroidal blood flow, the macula, where the retina lacks vasculature, was chosen for measurements. Doppler sonography. Mean blood flow velocity (MFV), peak systolic flow velocity (PSV), and end diastolic flow velocity (EDV) were determined in the right ophthalmic artery with color Doppler ultrasound (Guthoff et al., 1991). MFV was measured manually as time mean of the spectral outline. Measurements were performed with a 7.5-MHz probe (CFM 750, Vingmed Sound, Horten, Norway). The ophthalmic artery was measured anteriorly, at the point where it crosses the optic nerve. The sample volume marker was placed approximately 25 mm posterior to the globe. The resistive index (Pourcelot, 1976) in the ophthalmic artery was calculated as RI ⫽ (PSV-EDV)/(PSV). All parameters were determined as mean values over at least three cardiac cycles. Laser Doppler flowmetry. Application of this technique for the assessment of choroidal blood flow in humans has been described in detail (Riva et al., 1994). For this purpose the vascularized tissue is illuminated by coherent laser light. Scattering by moving red blood cells (RBCs) leads to a frequency shift in the scattered light. In contrast, static scatterers in tissue do not change light frequency, but lead to randomization of light directions impinging on RBCs. Hence, RBCs receive light from numerous random directions. As the frequency shift is dependent not only on the velocity of the moving RBC, but also on the angle between the wave vectors of the incident and the scattered light, scattering of the light in tissue affects the Doppler shift power spectrum (DSPS). This process leads to a broadening of the spectrum of scattered light, from which mean RBC velocity, the blood volume, and the blood flow (FLOW) can be calculated in relative units (Bonner and Nossal, 1990). Applanation tonometry. IOP was measured by Goldmann applanation tonometry (Haag-Streit, Bern, Switzerland). Ocular perfusion pressure (OPP) was calculated as 32 MAP-IOP (Robinson et al., 1986). Blood gas analysis. Blood gas analysis was done using capillary blood samples of the earlobe. After spreading the earlobe with nicotinate ⫹ nonylvanill-

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

4

amid paste (Finalgon, Thomae, Biberach, Germany) to induce capillary vasodilation, a lancet incision was made. The arterialized blood was drawn into a thin glass capillary tube. Arterial pH, pCO 2, and pO 2 were determined with an automatic blood gas analysis system (AVL 995-Hb, Graz, Austria). This technique accurately measures arterial blood gas tensions (Pitkin et al., 1994). Estimation of total choroidal blood flow. A method for the estimation of pulsatile ocular blood flow from measurement of changes in IOP during the cardiac cycle has been proposed previously (Silver and Farrell, 1994). This method is based on a theoretical model eye. It is assumed that venous outflow from the eye is nonpulsatile. In addition, the ocular rigidity, which is used to calculate ocular volume changes from changes in IOP, is assumed to be standard for all subjects. Whereas pneumotonometry measures the IOP change over time, ocular fundus pulsation is a point measure of the ocular volume change over time. We have previously shown that there is a significant correlation between FPA and pulsatile ocular blood flow as assessed with pneumotonometry (Schmetterer et al., 1998) at baseline and during isometric exercise. Due to the fact that the laser beam on the retina is only 50 ␮m in diameter FPA is only a point measure of the ocular volume changes during the cardiac cycle (Schmetterer et al., 2000). Hence, one needs to know the area of the retina to calculate total choroidal blood flow. In the following we present an approach for the calculation of total choroidal blood flow based on our data. We assume two spheres, an outer scleral sphere with a radius r s and a choroidal/retinal sphere with the radius r c . The difference between the radii of the two spheres is the choroidal thickness (t c ): r s ⫺ r c ⫽ t c . The volume of the outer scleral sphere is V s ⫽ 34 ⴱ ␲ ⴱ r s3 . Likewise, the volume of the choroidal/retinal sphere is V c ⫽ 34 ⴱ ␲ ⴱ (r s ⫺ t c ) 3 . To estimate choroidal volume we assume that the choroid extends over only two thirds of the sphere volumes (Kiel, 1994). In addition we introduce a constant factor ␣ which accounts for the thinning of the choroid anteriorly. Using these corrections the choroidal volume becomes

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

Kiss et al.

V c ⫽ ␣ ⴱ 23 ⴱ 共V s ⫺ V c兲 ⫽ ␣ ⴱ 89 ⴱ ␲ ⴱ 共r s3 ⫺ 共r s ⫺ t c兲 3 兲. (1) Furthermore, we assume FPA represents a uniform increase in choroidal thickness. However, FPA is an optical distance and needs to be converted into a geometrical distance, FPA g ⫽ FPA ⴱ n g(eye), where n g(eye) is the group refractive index of the eye, which is approximately 1.3456 (Schmetterer et al., 2000). Hence, r c varies from r c (diast) ⫽ r s ⫺ t c during diastole to r c (syst) ⫽ r s ⫺ t c ⫺ FPA g during systole. Accordingly the volume change during the cardiac cycle can be calculated as ⌬V c ⫽ V c共syst兲 ⫺ V c共diast兲 ⫽ ␣ ⴱ 98 ⴱ ␲ ⴱ 共共r s ⫺ t c ⫺ FPAg兲 3 ⫺ 共r s ⫺ t c兲 3 兲.

(2)

To calculate choroidal blood flow from these data one has to account for the pulse rate and the fraction of pulsatile blood flow. The change in flow pulsatility during squatting was assumed to be equal in the ophthalmic artery and in the choroid. The pulsatile fraction of blood flow in the ophthalmic artery was calculated as (MFV-EDV)/MFV (Krejcy et al., 1997). This calculation is based on the relation between pulsatile, steady, and total blood flow. Total blood flow through an organ or a vessel of diameter d is the sum of pulsatile and steady blood flow. In a vessel total blood flow during a heart cylce is given by MFV ⴱ ␲ ⴱ (d 2 )/4 and steady blood flow is EDV ⴱ ␲ ⴱ (d 2 )/4 if the vessel diameter is considered constant over the heart cycle. Consequently, the pulsatile component of blood flow through this artery is (MFV-EDV) ⴱ ␲ ⴱ (d 2 )/4. Hence, the pulsatile fraction of blood flow, which is (total blood flow ⫺ steady blood flow)/total blood flow, was estimated as (MFV-EDV)/MFV. Finally, the following formula is derived: CHBF ⫽ MFV ⴱ PR/共MFV-EDV兲 ⴱ ␣ ⴱ 98 ⴱ ␲ ⴱ 共共r s ⫺ t c ⫺ FPAg兲 3 ⫺ 共r s ⫺ t c兲 3 兲.

(3)

In the pilot study we used an additional approach to estimate the exercise-induced change in choroidal blood flow pulsatility. Laser Doppler flowmetry provides information on choriocapillaris blood flow with high temporal resolution. Hence, maximum flow dur-

5

Ocular Hemodynamics in Man

TABLE 1 Effect of Isometric Exercise on Systemic and Ocular Hemodynamic Variables and Intraocular Pressure (Pilot Study) Squatting period (min)

SBP (mm Hg) DBP (mm Hg) MAP (mm Hg) PR (beats/min) PSV (cm/s) EDV (cm/s) MFV (cm/s) RI FPA (␮m) Flow (arbitrary units) (MFV-EDV)/MFV CHBF (␮l/min)

Baseline

2

4

6

123 ⫾ 6 70 ⫾ 8 87 ⫾ 5 68 ⫾ 5 45 ⫾ 9 9⫾2 21 ⫾ 4 0.80 ⫾ 0.05 4.1 ⫾ 1.3 158 ⫾ 41 0.57 ⫾ 0.07 448 ⫾ 218

167 ⫾ 15* 97 ⫾ 14* 121 ⫾ 13* 102 ⫾ 7* 61 ⫾ 9* 12 ⫾ 2* 28 ⫾ 4* 0.80 ⫾ 0.05 2.8 ⫾ 0.9* 173 ⫾ 46 0.59 ⫾ 0.04 427 ⫾ 155

172 ⫾ 15* 101 ⫾ 14* 125 ⫾ 13* 105 ⫾ 6* 60 ⫾ 8* 12 ⫾ 2* 28 ⫾ 4* 0.81 ⫾ 0.03 2.7 ⫾ 0.8* 186 ⫾ 57 0.58 ⫾ 0.05 447 ⫾ 162

179 ⫾ 18* 104 ⫾ 14* 129 ⫾ 14* 113 ⫾ 10* 61 ⫾ 8* 12 ⫾ 3* 28 ⫾ 4* 0.81 ⫾ 0.04 2.7 ⫾ 0.7* 190 ⫾ 59* 0.58 ⫾ 0.05 482 ⫾ 199

Note. Data are presented as means ⫾ SD (n ⫽ 6); an asterisk indicates significant differences from baseline.

ing the systole (FLOW syst), minimum flow during the diastole (FLOW diast), and mean blood flow during the cardiac cycle (FLOW mean) can be extracted as described previously (Riva et al., 1994). Based on these data we calculated CHBFP ⫽ FLOWmean ⴱ PR/共FLOWmean ⫺ FLOWdiast兲 ⴱ ␣ 98 ⴱ ␲ ⴱ 共共r s ⫺ t c ⫺ FPAg兲 3 ⫺ 共r s ⫺ t c兲 3 兲.

(4)

Data Analysis All statistical analyses were done using the Statistica software package (Release 4.5, StatSoft Inc., Tulsa, OK). All outcome variables were calculated for each subject individually and then averaged. The effect of exercise on the outcome parameters was assessed with repeated measures ANOVA. The effect of hypercapnia on these parameters was characterized using paired t tests. The relative change in hemodynamic parameters induced by isometric exercise was calculated. For the experiments during 5% CO 2 and 95% air breathing the value during hypercapnia alone was taken as baseline. To gain information on the pressure–flow relationship relative data were sorted according to ascending OPP values (Riva et al., 1997b). In the pilot study we obtained a total of 18 OPP and blood flow data. These were divided into three groups of 6 OPP and CHBF,

CHBFP, or FLOW values. Hence, the first group consisted of the data with the lowest relative OPP values (n ⫽ 6) and the third group of the data with the highest relative OPP values (n ⫽ 6). Data from the main study were analyzed in the same way. Accordingly, data were divided into six groups of 10 OPP and CHBF values each. The mean values from these groups were used to determine the OPP at which the CHBF significantly deviated from baseline using ANOVA and Bonferroni corrections for multiple analysis. Data are presented as means ⫾SD. A two-tailed P ⬍ 0.05 was considered the level of significance.

RESULTS Pilot Study Hemodynamic responses as obtained in the pilot study are presented in Table 1. Isometric exercise caused a significant increase in blood pressure, PR, and flow velocities in the ophthalmic artery (P ⬍ 0.001 each). FPA was significantly reduced during squatting (P 0.004), whereas CHBF and CHBFP did not change. By contrast, FLOW as assessed with LDF was significantly different from baseline (P 0.024), but only at the end of the squatting period. The pressure–flow curves as obtained in the six

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

6

Kiss et al.

FIG. 1. Pressure-flow relationship using the categorized ocular perfusion pressure (OPP)-choroidal blood flow (CHBF; using pulsatility data from the ophthalmic artery) data (solid up triangles), ocular perfusion pressure (OPP)— choroidal blood flow (CHBFP; using pulsatility data from laser Doppler flowmetry) data (solid circles), and the ocular perfusion pressure (OPP)—FLOW data (open squares) as obtained in the pilot study (n ⫽ 6). Relative data were sorted into groups of 6 values according to ascending OPP values. Asterisks indicate significant changes in CHBF and FLOW from baseline.

healthy subjects participating in the pilot study are presented in Fig. 1. No difference was observed between the curves for ocular perfusion pressure and CHBF, ocular perfusion pressure and CHBFP, and ocular perfusion pressure and FLOW. A significant deviation from baseline was observed at a 66% increase in OPP.

Main Study As expected, isometric exercise induced a strong increase in SBP, DBP, MAP, and PR (Fig. 2, P ⬍ 0.001 each). At the end of the squatting period MAP was, on average, 56% higher than at baseline and PR was increased by 84%. Hypercapnia did not induce any

FIG. 2. Effect of isometric exercise on mean arterial pressure, pulse rate, systolic blood pressure, and diastolic blood pressure in the absence (solid up triangles) and presence of hypercapnia (open down triangles; n ⫽ 20). Data are presented as means ⫾ SD. Asterisks indicate significant exercise-induced changes versus pre-exercise levels.

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

Ocular Hemodynamics in Man

7

FIG. 3. Effect of isometric exercise on peak systolic velocity, end diastolic velocity, mean flow velocity, and resistive index in the absence (solid up triangles) and presence of hypercapnia (open down triangles; n ⫽ 20). Data are presented as means ⫾ SD. Asterisks indicate significant exercise-induced changes versus pre-exercise levels.

changes in systemic hemodynamic parameters. The increases of 59% in MAP and 95% in PR during isometric exercise under hypercapnia were comparable to those under normocapnia. FPA significantly decreased during isometric exercise (⫺36%; P ⬍ 0.001; Fig. 3). Breathing 5% CO 2 ⫹ 95% air significantly increased FPA by ⫹22% (P ⬍ 0.001) under resting conditions. However, the decrease in FPA during isometric exercise was comparable during hypercapnia (⫺42%; P ⬍ 0.001) and normocapnia (n.s. between periods). IOP was markedly elevated during isometric exercise (Fig. 3). This increase was slightly lower during normocapnia (37%; P 0.004) than during hypercapnia (50%, P ⬍ 0.001). Hypercapnia itself, however, did not change IOP. Isometric exercise increased flow velocities in the ophthalmic artery (Fig. 4; MFV: 33%, P 0.002). Breathing 5% CO 2 ⫹ 95% air tended to increase MFV in the ophthalmic artery (10%), but this effect was not significant. The increase in MFV during squatting was slightly smaller during hypercapnia (⫹24%; P 0.006) than during normocapnia. In contrast, neither isomet-

ric exercise nor breathing of 5% CO 2 ⫹ 95% air altered RI in the ophthalmic artery. The pressure–flow relationship is presented in Fig. 5. No difference in the association between calculated CHBF and OPP was observed in the absence or the presence of hypercapnia. CHBF was significantly different from baseline at a 69% increase in OPP during normocapnia and at a 70% increase in OPP during hypercapnia. Squatting induced no significant change in blood gas values (Table 2). By contrast, breathing 5% CO 2 ⫹ 95% air caused the expected increase in pCO 2 and decrease in pH. When squatting was performed during the inhalation period no additional increase in pCO 2 was observed. To calculate CHBF based on Eq. 3 it is necessary to choose a value for ␣, r s, and t c according to Eq. 3. Assuming ␣ ⫽ 0.65, r s ⫽ 11.5 mm, and t c ⫽ 0.3 mm (Kiel, 1994) and taking baseline values of FPA (4.1 ␮m), MFV (21 cm/s), EDV (9 cm/s), and PR (68 beats/ min) CHBF ⫽ 448 ␮l/min. Data as calculated according to Eq. 3 are only slightly dependent on r s and t c.

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

8

FIG. 4. Effect of isometric exercise on fundus pulsation amplitude and intraocular pressure in the absence (solid up triangles) and presence of hypercapnia (open down triangles; n ⫽ 20). Data are presented as means ⫾ SD. Asterisks indicate significant exerciseinduced changes versus pre-exercise levels. # indicates a significant change in fundus pulsation amplitude during hypercapnia.

Assuming a choroidal thickness of 390 ␮m (Cristini et al., 1991) the corresponding value for CHBF at baseline is 441 ␮l/min. The results are, however, more dependent on the value selected for the scleral radius. Assuming a scleral radius of 10.5 mm and a choroidal thickness of 300 ␮m, baseline CHBF becomes 372 ␮l/ min. Obviously the choroidal blood flow values are strongly dependent on ␣, a factor which is difficult to estimate due to the lack of in vivo data.

DISCUSSION The data of the present study indicate that blood flow in the human choroid is constant over a wide range of perfusion pressures during isometric exercise. This is in keeping with a recent study using laser Doppler flowmetry for quantification of choroidal he-

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

Kiss et al.

modynamics (Riva et al., 1997b). In extension of these previous findings our study indicates that hypercapnia does not substantially influence the choroidal pressure–flow relationship. During isometric exercise under normocapnic and hypercapnic conditions choroidal blood flow increased significantly versus baseline only when MAP exceeded the preexercise value by more than 70%. Our study therefore supports the concept that the choroid does not behave like a passive vascular bed and has a nonlinear pressure–flow relationship (Kiel, 1994; Kiel and van Heuven, 1995). Interpreting our results as evidence of choroidal autoregulation, however, should be done with caution. In its strict sense autoregulation can only be investigated in an isolated organ where perfusion pressure can be varied experimentally, but in humans such experiments are obviously not possible. In particular the neural input to the choroidal vessels has to be considered interpreting the results of the present study. There is evidence from several animal experiments that ocular sympathetic vasomotor nerves play a key role in choroidal vasoconstriction elicited by an increase in ocular perfusion pressure (Alm, 1977; Alm and Bill, 1973a; Ernest, 1977). It is well established that hypercapnia narrows the autoregulatory plateau in the cerebral vasculature (Ekstro¨m-Jodal et al., 1972; Raichle and Stone, 1972). This phenomenon has been interpreted as an indicator of a metabolic mechanism underlying cerebral blood flow autoregulation (Aaslid et al., 1989). In addition, hypercapnia induces cerebral vasodilation. At high cerebral blood flow levels elicited by hypercapnia the resistance vessels are dilated and do not have enough dilatory capacity to keep blood flow at a constant level to the normal lower limit of autoregulation. This is obviously not the case in the choroid. Hypercapnia significantly increased FPA and therefore altered choroidal tone and it obviously altered the metabolic status of the choroid. Nevertheless, breathing 5% CO 2 ⫹ 95% air did not influence the choroidal pressure flow relation in our healthy subjects. Our results therefore indicate that metabolic mechanisms are not responsible for the stable choroidal blood flow despite the strong increase in ocular perfusion pressure. It may well be that higher degrees of hypercapnia would

9

Ocular Hemodynamics in Man

FIG. 5. Pressure-flow relationship using the categorized ocular perfusion pressure (OPP)-choroidal blood flow (CHBF) data in the absence (upper panel) and presence (lower panel) of hypercapnia (n ⫽ 20). Relative data were sorted into groups of 10 values according to ascending OPP values. Asterisks indicate significant changes in CHBF from baseline.

steepen the relation between pressure and flow, but such experiments cannot be performed in humans for ethical reasons. Hypercapnia induced only a small increase in ophthalmic artery flow velocity, which is in keeping with previous findings (Harris et al., 1996a; Schmetterer et al., 1997b). By contrast, arterial carbon dioxide tension appears to be an important regulator of basal choroidal blood flow, which also has also been shown in previous human studies (Riva et al., 1994; Schmetterer et al., 1996, 1997b). We have previously shown that even a slight change in carbon dioxide tension may alter choroidal blood flow in response to isometric exercise (Schmetterer et al., 1998). Hence, it is impor-

tant to sufficiently control pCO 2 when choroidal pressure–flow relationships are investigated. We have asked our subjects under study not to change their breathing pattern during exercise. Special care was taken to ensure that subjects did not involuntarily perform a Valsalva maneuver. Obviously both laser Doppler flowmetry in the subfoveal choroid as well as the method we employed in the present study have considerable limitations. In laser Doppler flowmetry vascularized tissue is illuminated by laser light. Light scattered at moving red blood cells is shifted in frequency, which depends on particle velocity as well as on the angle between the wave vectors of the incident and the scattered light.

TABLE 2 Blood Gas Values at Baseline and during Isometric Exercise

Baseline Isometric exercise 5% CO 2 ⫹ 95% air 5% CO 2 ⫹ 95% air ⫹ isometric exercise

pH

pCO 2

pO 2

7.40 ⫾ 0.02 7.39 ⫾ 0.02 7.33 ⫾ 0.02* 7.31 ⫾ 0.03*

38.4 ⫾ 3.3 39.2 ⫾ 3.7 47.9 ⫾ 4.6* 48.4 ⫾ 4.4*

95.6 ⫾ 7.5 96.9 ⫾ 8.4 101.1 ⫾ 7.3 100.4 ⫾ 9.2

Note. Data are presented as means ⫾ SD (n ⫽ 20); an asterisk indicates significant differences from baseline.

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

10

Calculation of red blood cell flux from the Doppler shift power spectrum is based on the theory of Bonner and Nossal (1990). This model assumes scattering of the laser light by stationary tissue and multiple scattering leads to a complete randomization in the direction of the wave vectors of the incident and the scattered laser beam. That this prerequisite is fulfilled in the subfoveal choroid has, however, not yet been shown. In addition, it is not clear from which depth the Doppler spectrum obtained with laser Doppler flowmetry arises. The use of red or infrared lasers ensures adequate penetration of the laser light, but the relative contribution of different vascular layers to the Doppler signal remains unclear. One limitation of the technique we used in the present study is that FPA does not directly measure pulsatile blood flow in the choroid. However, we have previously shown that there is a high association between FPA as measured with laser interferometry and pulsatile ocular blood flow as measured with pneumotonometry during isometric exercise (Schmetterer et al., 1998). An important limitation of this study is that we corrected pulsatile blood flow in the choroid by using pulsatility data from the ophthalmic artery. It is obvious that any change in flow pulsatility in the supplying artery, as induced, for instance, by a change in blood pressure profile, will lead to a concomitant change in flow pulsatility in distal vascular beds. However, isometric exercise may induce additional local changes in flow pulsatility due to local vasoconstriction which are not taken into account with the present method. For instance, blood flow in the choroid and the ophthalmic artery can easily be uncoupled when an artificial increase in IOP is applied (Harris et al., 1996b; Findl et al., 1997). The error induced by this method of calculating changes in flow pulsatility from color Doppler imaging data in the ophthalmic artery is difficult to calculate, because the ratio of pulsatile to nonpulsatile blood flow in the human choroid at baseline is not known. Estimates of the fraction of nonpulsatile blood flow in the human choroid vary between 20% (Langham et al., 1989) and 50% (Krakau, 1995). Hence, a deviation from squatting induced changes in flow pulsatility of 20% between the ophthalmic artery and the choroid would lead to only 4 to 10%

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

Kiss et al.

error in our estimations of total blood flow. Our data obtained during the pilot study indicate that squatting-induced changes in flow pulsatility are not significantly different in the ophthalmic artery and the choroid. An important question is whether alterations in blood flow pulsatility are adequately reflected by alterations in velocity pulsatility in the ophthalmic artery. A simple calculation indicates that the error introduced by this assumption is small. According to Wetterer et al. (1971) the magnitude of the diameter change of an artery during the cardiac cycle may be calculated as ⌬r/r ⫽ ⌬p/(2 ⴱ ␳ ⴱ c 2 ), where ⌬r is the diameter change during the cardiac cycle, r is the diameter of the artery, ⌬p is the pulse pressure amplitude, ␳ is the density of blood, and c is the pulse wave velocity. For the ophthalmic artery we assume a vessel diameter of 0.57 mm (Michelson and Schuierer, 1991). The baseline pulse pressure amplitude in the present study was 57 mm Hg (7598 Pa), and the density of blood is 1065 kg/m 3. Currently there are no data for pulse wave velocities in the ophthalmic artery available. Hence, we chose data from the cerebral circulation. By simultaneous measurement of velocity waveforms from the cervical carotid artery and the ipsilateral middle cerebral artery, Giller and Aaslid (1994) determined a value of 12.8 m/s for cerebral pulse wave velocity. Using these values the diameter change during the cardiac cycle in the ophthalmic artery (⌬r) is as small as 0.012 mm or 2.1% (⌬r/r). For the estimation of the error introduced by diameter fluctuations of the ophthalmic artery during the cardiac cycle we used a more conservative estimate of 5% for the diameter change. Taking this value, the flow pulsatility at baseline will be approximately 10% higher than calculated from velocity data. During squatting, pulse pressure amplitude increased by approximately 50% versus baseline. Consequently, vessel diameter change during the cardiac cycle in the ophthalmic artery during squatting may be in the order of 7.5% (this does not account for changes in arterial elasticity due to vasoconstriction or vasodilation). Hence, flow pulsatility during squatting will be approximately 15% higher than calculated from velocity data. As only relative changes in flow pulsatility are taken in the present study the error introduced is

11

Ocular Hemodynamics in Man

in the order of 5%. Importantly, our method of quantifying CHBF changes during isometric exercise requires neither that flow velocities in the ophthalmic artery adequately reflect blood flow through this vessel nor that baseline pulsatility in the ophthalmic artery and the choroid are comparable. In the present study we observed good agreement between pressure–flow relationships assessed with the two methods in a pilot study in six healthy subjects. FLOW as well as CHBF and CHBFP showed a significant deviation from baseline at a 66% increase of OPP. At higher OPP values FLOW values tended to be higher than CHBF and CHBFP values, as evidenced from the pressure flow curve as well as from Table 1. At the end of the squatting period FLOW was significantly different from baseline, whereas CHBF and CHBFP were not. Nevertheless, our results indicate good agreement between the two approaches to assess choroidal blood flow, although additional studies will be required using other types of pharmacological or physiological interventions to confirm these results. Interestingly, we observed a small, but significant, increase in blood flow velocity in the ophthalmic artery during squatting. Whether this is indicative of increased blood flow through this artery is unclear, because no information on vessel diameter was available. A reduction in ophthalmic artery vessel diameter due to enhanced sympathetic activity during isometric exercise could theoretically counteract the increased flow velocity (Kontos, 1989). However, blood flow in the eye is only a small portion of ophthalmic artery blood flow. Total blood flow through the human ophthalmic artery has been determined as approximately 6750 ␮l/min (Michelson and Schuierer, 1991). Absolute values for retinal blood flow are available from laser Doppler velocimetry studies and are in the order of 35 ␮l/min (Riva et al., 1985), which is less than 1% of ophthalmic artery blood flow. Absolute blood flow data for the human choroid are not available. In the present study we estimated a choroidal blood flow of approximately 450 ␮l/min. This is slightly lower than the results of Alm and Bill (1973b), who measured a choroidal blood flow of 600 ␮l/min in primates. The main reason for this difference may be a scleral

outward movement during the cardiac cycle, which would lead to a more pronounced change in choroidal thickness than estimated in the present study. Values of pulsatile ocular blood flow as obtained with pneumotonometry are, however, considerably higher (830 ␮l/min; Schmetterer et al., 1998). Nevertheless, these considerations indicate that ocular blood flow is at maximum 25% of total ophthalmic artery blood flow. An increased blood flow through this artery is therefore not necessarily associated with an increase in choroidal perfusion. Interestingly, neither hypercapnia nor squatting had a significant impact on RI. This is in contrast to previous findings, where a decrease in RI has been observed during hand gripping (Beck et al., 1995; Schmetterer et al., 1998). This is most likely caused by the different blood pressure profiles induced by squatting and handgripping. We have previously shown that interpretation of RI as a measure of distal vascular resistance is not valid when changes in blood pressure profile occur (Krejcy et al., 1997; Schmetterer et al., 1998). In conclusion, our data indicate that the pressure volume relationship during isometric exercise is nonlinear. A significant change in choroidal blood flow was observed only when ocular perfusion pressure was approximately 70% higher than baseline. The mechanism underlying this regulatory response in the human choroid is likely to be different from that of the cerebral circulation.

ACKNOWLEDGMENT The authors thank an anonymous reviewer for help in calculating choroidal blood flow.

REFERENCES Aaslid, R., Lindegaard, K. R., Sorteberg, W., and Nornes, H. (1989). Cerebral autoregulation dynamics in humans. Stroke 20, 45–52.

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

12 Alm, A. (1977). The effect of sympathetic stimulation on blood flow through the uvea, retina, and optic nerve in monkeys. Exp. Eye Res. 25, 19 –25. Alm, A., and Bill, A. (1972). The oxygen supply to the retina. II Effects of high intraocular pressure and of increased carbon dioxide tension on uveal and retinal blood flow in cats. Acta Physiol. Scand. 84, 306 –319. Alm, A., and Bill, A. (1973a). The effect of stimulation of the sympathetic chain on retinal oxygen tension and uveal, retinal, and cerebral blood flow in cats. Acta Physiol. Scand. 88, 84 –96. Alm, A., and Bill, A. (1973b). Ocular and optic nerve blood flow at normal and increased pressure in monkeys (macaca irus): A study with radioactively labeled microspheres including flow determinations in the brain and some other tissues. Exp. Eye Res. 15, 15–29. Beck, D., Harris, A., Evans, D., and Martin, B. (1995). Ophthalmic arterial hemodynamics during isometric exercise. J. Glaucoma 4, 317–321. Bonner, R., and Nossal, R. (1990). Principles of laser-Doppler flowmetry in laser-Doppler blood flowmetry. In “Developments in ¨ berg, Eds.), Cardiovascular Medicine” (A. P. Shepard and A. P. O pp. 17– 45. Kluwer Academic, Boston. Cristini, G., Gennamo, G., and Daponte, P. (1991). Choroidal thickness in primary glaucoma. Ophthalmologica 202, 81– 85. Dumsky, M. J., Eriksen, J. E., Dore, C. J., and Kohner, E. M. (1996). Autoregulation in the human retinal circulation: Assessment using isometric exercise, laser Doppler velocimetry, and computer assisted image analysis. Microvasc. Res. 51, 378 –392. Ekstro¨m-Jodal, B., Ha¨ggendal, E., Linder, L. E., and Nilsson, N. J. (1972). Cerebral blood flow autoregulation at high arterial pressures and different levels of carbon dioxide tension. Eur. Neurol. 6, 6 –10. Ernest, J. T. (1977). The effect of systolic hypertension on rhesus monkey eyes after ocular sympathectomy. Am. J. Ophthalmol. 84, 341–344. Fercher, A. F. (1984). In vivo measurement of fundus pulsations by laser interferometry. IEEE J. Q. El. 20, 1469 –1471. Findl, O., Strenn, K., Wolzt, M., Menapace, R., Vass, C., Eichler, H. G., and Schmetterer, L. (1997). Effects of changes in intraocular pressure on human ocular haemodynamics. Curr. Eye Res. 16, 1024 –1029. Giller, C. A., and Aaslid, R. (1994). Estimates of pulse wave velocity and measurement of pulse transit time in the human cerebral circulation. Ultrasound Med. Biol. 20, 101–105. Guthoff, R. E., Berger, R. W., Winkler, P., Helmke, K., and Chumbley, L. C. (1991). Doppler ultrasonography of the ophthalmic and central retinal vessels. Arch. Ophthalmol. 109, 532–536. Harris, A., Tippke, S., Sievers, C., Picht, G., Lieb, W., and Martin, B. (1996a). Acetazolamide and CO 2: Acute effects on cerebral and retrobulbar hemodynamics. J. Glaucoma 5, 39 – 45. Harris, A., Joos, K., Kay, M., Evans, D., Shetty, R., Sponsel, W. E., and Martin, B. (1996b). Acute IOP elevation with scleral suction: effects on retrobulbar haemodynamics. Br. J. Ophthalmol. 80, 1055–1059.

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.

Kiss et al.

Kiel, J. W. (1994). Choroidal myogenic autoregulation and intraocular pressure. Exp. Eye Res. 58, 529 –543. Kiel, J. W., and van Heuven, W. A. J. (1995). Ocular perfusion pressure and choroidal blood flow in the rabbit. Invest. Ophthalmol. Vis. Sci. 36, 579 –585. Kontos, H. A. (1989). Validity of vertebral arterial blood flow calculations from velocity measurements. Stroke 20, 1–3. Krakau, C. E. T. (1995). A model for pulsatile and steady ocular blood flow. Graefes Arch. Clin. Exp. Ophthalmol. 233, 112–118. Krejcy, K., Wolzt, M., Kreuzer, C., Breiteneder, H., Schu¨tz, W., Eichler, H. G., and Schmetterer, L. (1997). Characterisation of Angiotensin-II effects on cerebral and ocular circulation by noninvasive methods. Br. J. Clin. Pharmacol. 43, 501–508. Langham, M. E., Farrell, R. A., O’Brien, V., Silver, D. M., and Schilder, P. (1989). Blood flow in the human eye. Acta Ophthalmol. 67(Suppl. 191), 9 –13. Michelson, G., and Schuierer, G. (1991). Absoluter Blutfluß in der Arteria Ophthalmica. Fortschr. Ophthalmol. 88, 687– 689. Pitkin, A. D., Roberts, C. M., and Wedzicha, J. A. (1994). Arterialised earlobe blood gas analysis: An underestimated technique. Thorax 49, 364 –366. Pourcelot, L. (1976). Diagnostic ultrasound for cerebral vascular diseases. In “Present and Future of Diagnostic Ultrasound” (I. Donald and S. Levi, Eds.), pp. 141–147. Kooyker, Rotterdam, The Netherlands. Raichle, M. E., and Stone, H. L. (1972). Cerebral blood flow autoregulation and graded hypercapnia. Eur. Neurol. 6, 1–5. Riva, C. E., Grunwald, J. E., Sinclair, S. H., and Petrig, B. L. (1985). Blood velocity and volumetric flow rate in human retinal vessels. Invest. Ophthalmol. Vis. Sci. 26, 1124 –1132. Riva, C. E., Grunwald, J. E., and Petrig, B. L. (1986). Autoregulation of human retinal blood flow. Invest. Ophthalmol. Vis. Sci. 27, 1706 –1712. Riva, C. E., Cranstoun, S. D., Grunwald, J. E., and Petrig, B. L. (1994). Choroidal blood flow in the foveal region of the human ocular fundus. Invest. Ophthalmol. Vis. Sci. 35, 4273– 4281. Riva, C. E., Titze, P., Hero, M., and Petrig, B. L. (1997a). Effect of acute decreases of perfusion pressure on choroidal blood flow in humans. Invest. Ophthalmol. Vis. Sci. 38, 1752–1760. Riva, C. E., Titze, P., Hero, M., Movaffaghy, A., and Petrig, B. L. (1997b). Choroidal blood flow during isometric exercises. Invest. Ophthalmol. Vis. Sci. 38, 2338 –2343. Robinson, F., Riva, C. E., Riva, J. E., Petrig, B. L., and Sinclair, S. H. (1986). Retinal blood flow autoregulation in response to an acute increase in blood pressure. Invest. Ophthalmol. Vis. Sci. 27, 722–726. Schmetterer, L., Lexer, F., Unfried, C., Sattmann, H., and Fercher, A. F. (1995). Topical measurement of fundus pulsations. Opt. Eng. 34, 711–716. Schmetterer, L., Lexer, F., Graselli, U., Findl, O., Eichler, H. G., and Wolzt, M. (1996). The effect of different mixtures of O 2 and CO 2 on ocular fundus pulsations. Exp. Eye Res. 63, 351–355. Schmetterer, L., Strenn, K., Findl, O., Breiteneder, H., Graselli, U., Agneter, E., Eichler, H. G., and Wolzt, M. (1997a). Effects of

Ocular Hemodynamics in Man

antiglaucoma drugs on ocular hemodynamics in healthy volunteers. Clin. Pharmacol. Ther. 61, 583–595. Schmetterer, L., Krejcy, K., Findl, O., Strenn, K., Graselli, U., Kastner, J., Eichler, H. G., and Wolzt, M. (1997b). The role of nitric oxide in the regional O 2 and CO 2 responsiveness of cerebral and ocular circulation in man. Am. J. Physiol. 42, R2005–R2012. Schmetterer, L., Dallinger, S., Findl, O., Strenn, K., Graselli, U., Eichler, H. G., and Wolzt, M. (1998). Noninvasive investigations of the normal ocular circulation in humans. Invest. Ophthalmol. Vis. Sci. 39, 1210 –1220.

13 Schmetterer, L., Dallinger, S., Findl, O., Schmetterer, K., Lexer, F., Eichler, H. G., and Wolzt, M. (2000). A comparison between laser interferometric measurement of fundus pulsation and pneumotonometric measurement of pulsatile ocular blood flow. 1. Baseline considerations. Eye 14, 39 – 45. Silver, D. M., and Farrell, R. A. (1994). Validity of pulsatile ocular blood flow measurements. Surv. Ophthalmol. 38(Suppl.), S72–S80. Wetterer, E., Bauer, R. D., and Pasch, T. (1971). Arteriensystem. In “Physiologie des Kreislaufs” (E. Bauercisen, Ed.), pp. 1– 66. Springer-Verlag, Berlin.

Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.