| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Departments of 1Medicine and 2Cardiology, Cardiovascular Research Institute Maastricht, and University Hospital, 6202 AZ Maastricht, The Netherlands
Submitted 30 January 2003 ; accepted in final form 29 April 2003
 |
ABSTRACT |
|---|
 TOP ABSTRACT METHODSRESULTSDISCUSSIONREFERENCES |
|---|
microcirculation; function; blood pressure; heart rate; variability
|
METHODS |
|---|
|
TOPABSTRACTMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
Experiments were performed on 12 healthy volunteers. During the week before the measurements, all subjects adhered to a diet that contained 175 mmol Na+ so as to minimize variations in results due to salt intake. Compliance with the diet was checked by measuring Na+ and creatinine output in 24-h urine collections obtained during the last 24 h before the first experimental day. None of the subjects used any medication (including nonsteroidal anti-inflammatory drugs) during the 2 wk before the measurements. In addition, subjects had to refrain from smoking and drinking caffeine- or alcohol-containing beverages for at least 12 h before the experiments, which started at 8:30 AM after an overnight fast. The Medical Ethics Committee of Maastricht University Hospital approved the study, and all participants gave written informed consent. The investigations conformed to the principles outlined in the Declaration of Helsinki (11).
Experimental Design
All volunteers were studied on two separate occasions (at least 2 days apart), during which they received in random order (double blind) an intravenous infusion of either BNP (4 pmol · kg–1 · min–1) or vehicle (5% glucose). This dose of BNP was chosen to allow comparison with data from the literature (5, 7). Experiments were performed in a quiet, temperature-controlled room (mean temperature, 24.2 ± 0.3°C). Precautions were taken to minimize external disturbances. Except during the microcirculatory measurements (which had to be performed in a sitting position), subjects remained supine throughout the experiments. A 20-gauge catheter was inserted into the antecubital vein of both arms. One was connected to a three-way tap for infusion of BNP (Clinalfa Ethifarma Nederland, The Netherlands) and p-aminohippurate (PAH) or inulin (for measuring renal hemodynamics), whereas the other was used for blood sampling. To ensure adequate diuresis, subjects consumed 200 ml of water every hour until the last blood samples had been drawn. At time (t) = 0 min, the PAH or inulin infusion was started. Between t = 0 and t = 60 min, baseline measurements of the conjunctiva and nailfold microcirculation, skin blood flow (SBF), and total skin oxygen capacity were obtained. At t = 60 min, an echocardiogram was taken to assess CO. At t = 120 min, the intravenous infusion of either BNP or placebo (5% glucose) was started. At t = 180 min (i.e., after 1 h of BNP or placebo infusion), a second echocardiogram was performed, which was followed by a second set of microvascular measurements. Blood pressure and HR were measured before and after the microvascular measurements with subjects in a sitting position and at 10-min intervals during the infusion with them in the supine position. Blood samples were drawn at t = 0, 120, and 180 min for PAH and inulin and at t = 120 and 180 min for determination of cGMP and BNP. Urine samples for measurement of Na+ and K+ levels were collected at t = 60, 120, and 180 min (immediately after blood sampling).
Measurements
Systemic hemodynamics. Systolic blood pressure (SBP), diastolic blood pressure (DBP), mean arterial blood pressure (MAP), and HR were measured using a semiautomatic oscillometric device (Dinamap model 1846 vital signs monitor, Critikon). Echocardiography to assess CO was performed in the semi-left lateral position using a cross-sectional, phased-array (2.5 MHz) echocardiographic Doppler system (Sonos 2000 and 2500, Hewlett-Packard). Stroke volume (SV) was determined from the flow velocity across the aortic valve (apical approach) and the diameter of the aortic orifice during systole. CO was calculated as SV x HR (expressed as l/min). Total peripheral resistance (TPR) was calculated as (MAP/CO) x 80 (expressed as dyn · s/cm5).
Microcirculatory measurements. The microcirculation of the lateral part of the bulbar conjunctiva of the right eye was studied with a custom-built horizontal microscope as previously described (3). For microvascular density measurements, recordings were made on videotape with a standard achromatic objective x2.5 (numeric aperture, 0.10). Arterioles, capillaries, and venules were classified in several video frames using image-analysis software (Optimas version 5.0; Breda, The Netherlands). As a measure of density, the total length of each microvascular class per square millimeter of conjunctiva was determined and averaged.
Measurement of nailfold capillary density was also performed as described earlier (4). For the capillary density measurements, recordings were made a few millimeters proximal to the terminal row of capillaries. Baseline skin-capillary density was defined as the amount of erythrocyte-filled capillaries in one video screen (1.6 mm2 of skin). The recruitment of functionally available capillaries was defined as the increase in the number of erythrocyte-filled capillaries after 4 min of arterial occlusion (by cuff inflation of 200 mmHg at the wrist).
SBF, which predominantly reflects thermoregulatory flow, was determined simultaneously with nailfold capillary density using laser-Doppler fluxmetry (Periflux PF3, Perimed; Järfälla, Sweden) using probe PF-308, a wide-band (12 kHz) mode, and a time constant of 0.2 s. Total skin oxygen capacity, which is a measure of nutritive blood flow, was determined using transcutaneous oxygen tension measurements (TcPO2, Radiometer) with the probe heated to 44°C. Probes were placed on the dorsum of the interphalanx of the finger and the hand between digits IV and V, respectively, of the same hand in which the nailfold capillary density was measured. Flux values are expressed as arbitrary perfusion units calibrated against an external standard. Total skin oxygen capacity is defined in millimeters of mercury. SBF and total skin oxygen capacity were measured before and during reactive hyperemia after 4 min of arterial occlusion (200 mmHg).
Renal function. Renal hemodynamics, i.e., effective renal plasma flow (ERPF) and glomerular filtration rate (GFR), were measured as the clearance of PAH (MSD; West Point, PA) and inulin (Inutest, Laevosan Gesellschaft; Linz, Austria), respectively, during continuous infusion of these substances (1). Both GFR and ERPF measurements were corrected for body surface area [expressed as ml/(min x 1.73 m2)]. Effective renal blood flow (ERBF) was calculated by the formula ERBF = ERPF/(1 – hematocrit). Filtration fraction (FF) was calculated as FF = GFR/ERPF. Renal vascular resistance (RVR, expressed in dyn · s/cm5) was calculated according to the formula RVR = (MAP/ERBF) x 80,000. Renal fraction (RF) was calculated as (ERBF/CO) x 100%.
The filtered load of Na+ (FLNa, expressed in mmol/h) was calculated as GFR x 60 x [Na+]plasma, where [Na+]plasma is the plasma concentration of Na+. Fractional tubular reabsorption of Na+ (FTRNa) was calculated as [(FLNa – UNaV)/FLNa] x 100%, where UNa and V represent the urinary Na+ concentration and volume, respectively. Tubular rejection is defined as 1 – FTRNa.
Assay Methods
PAH and inulin levels were measured by means of a spectrophotometer. BNP and cGMP levels were measured using a competitive protein-binding RIA (RIK 9086, Peninsula Laboratories and RE 29071, IBL Hamburg, respectively). Before assay, plasma samples of BNP were acidified and extracted using a SEP-Pak C-18 column (Waters-Millipore). In our hands, the intra- and interassay variabilities of all assays were <10%. All samples from the same individual were assayed in a single run.
Statistics
Data are presented as medians with interquartile ranges (IQRs). The Wilcoxon paired-sign test for paired analysis (both within one visit and between the BNP and placebo infusions) and Friedman's test for analysis of multiple related measurements of MAP and HR (within one visit) were used. P values < 0.05 denote statistical significance. On the basis of previous experiments, we calculated that this study is able to demonstrate in 12 experimental subjects a 10% difference in any of the test variables with a power of 85%.
|
RESULTS |
|---|
|
TOPABSTRACTMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
|
Plasma Levels of BNP and cGMP
At baseline, BNP plasma levels did not differ between visits (Table 2). BNP infusion significantly increased plasma levels of this peptide to 191 (IQR, 172–367) pg/ml (P < 0.01), whereas placebo infusion had no effect on BNP levels [21 (IQR, 11–68) pg/ml after infusion].
|
In parallel with the increase in BNP, plasma levels of cGMP also significantly increased to 19.5 (IQR, 15.6–25.6) pmol/ml (P < 0.01) after BNP infusion, whereas placebo infusion did not change levels of cGMP [6.6 (IQR, 5.7–7.5) pmol/ml after infusion]. No differences between cGMP values at baseline were observed (Table 2).
Systemic Hemodynamic Effects
Systemic hemodynamics with supine position. At baseline, no differences in blood pressure or HR were observed between BNP and placebo experiments (Table 2). SBP and DBP were not altered by infusion of either BNP or placebo and neither was MAP (Fig. 1). During BNP infusion, supine HR measurements tended to increase, but differences from baseline just failed to reach statistical significance (P = 0.058); moreover, HR responses did not differ between the two infusion experiments (Fig. 1; P = 0.686).
|
Baseline values of SV and CO did not differ either (Table 2). SV significantly decreased after BNP infusion [to 73 (IQR, 66–83) ml; P = 0.007], although it did not change during placebo infusion [83 (IQR, 76–93) ml; P = 0.594]. This difference between the effects of BNP and placebo was statistically significant (P = 0.015; Fig. 2). CO significantly decreased to 4.3 (IQR, 3.7–5.2) l/min during BNP infusion (P = 0.013), whereas placebo did not induce any significant change [to 4.6 (4.2–5.2) l/min; P = 0.182]. Expressed as percent change, BNP had no significant effect on CO compared with placebo (P = 0.082; Fig. 2). TPR was not influenced by infusion of either BNP [1,880 (IQR, 1,453–2,049) dyn · s/cm5] or placebo [1,755 (IQR, 1,426–2,004) dyn · s/cm5].
|
Influence of posture on BNP effects: MAP and HR measurements with sitting position. As expected, the transition from the lying to the sitting position was associated with a decrease in SBP and DBP and an increase in HR. However, during BNP infusion, postural changes were not tolerated well by four subjects. Two of these became markedly hypotensive in a sitting position 15 min after completion of the BNP infusion and were unconscious for a few minutes. The other subjects "only" appeared pale and experienced dizziness and sweating in a sitting position at the same moment during the protocol.
Before the infusions, sitting MAP and HR values did not differ between the BNP and the placebo experiments. As shown in Fig. 3, BNP infusion decreased the sitting MAP value from 92 (IQR, 86–103) to 80 (IQR, 72–93) mmHg (P = 0.05). This BNP-induced decrease in pressure was significantly greater than that observed during placebo infusion (P = 0.028). Sitting HR values increased significantly after BNP infusion [from 59 (IQR, 56–63) to 68 (IQR, 60–78) beats/min; P = 0.011], whereas these values did not increase after placebo administration.
|
Renal Effects
Renal hemodynamics. Baseline renal hemodynamics did not differ between BNP and placebo (Table 2). BNP significantly increased GFR compared with placebo (P = 0.007, BNP vs. placebo; Fig. 4), whereas ERPF did not change significantly during either infusion. FF increased during BNP infusion to 29% (IQR, 26–31%), but it tended to decrease during placebo infusion [to 25% (IQR, 21–28%); P = 0.022, BNP vs. placebo; Fig. 4].
|
BNP tended to increase RVR to 11,526 (IQR, 7,612–12,916) dyn · s/cm5, but the difference just failed to reach statistical significance (P = 0.063). However, there was no difference in the percent change compared with placebo infusion (P = 0.465). RF increased during placebo infusion to 18% (IQR, 12–20%); P = 0.033; however, no difference was observed in RF responses between placebo and BNP infusion (P = 0.674).
Natriuresis and diuresis. At baseline, UNaV values were slightly lower and FTRNa values were slightly higher on the BNP than on the placebo infusion day (Table 2). Compared with placebo infusion, BNP infusion resulted in significant natriuresis and diuresis. As shown in Fig. 5, urinary Na+ excretion in a 1-h urine collection increased to 24.1 (IQR, 15.9–28.2) mmol during BNP infusion (P = 0.002), whereas it did not change during placebo infusion [10.3 (IQR, 7.3–13.0) mmol/h after infusion; P = 0.308]. Urinary volume of the 1-h urine collection significantly increased both after BNP [to 408 (IQR, 360–584) ml] and after placebo [to 225 (IQR, 176–314) ml] infusion as a result of water supplementation during the renal clearance study protocol (Fig. 5). However, the increase in urinary volume after BNP infusion was significantly greater than after placebo infusion (P = 0.002).
|
FLNa values increased significantly from 855 (IQR, 753–939) to 894 (IQR, 803–983) mmol/h during BNP infusion, whereas placebo infusion did not influence FLNa. Furthermore, BNP significantly decreased FTRNa values [to 97.4% (IQR, 97.1–98.2%)] compared with placebo infusion (P = 0.005), which resulted in a greater tubular rejection during BNP infusion (Fig. 6).
|
Microcirculation
Skin blood flow. The effects of BNP on both basal and hyperemic peak (thermoregulatory) SBF are shown in Table 3. Compared with placebo, BNP infusion did not change any of the variables. Also, time to peak and duration of hyperemia measurements were not influenced by infusion of either BNP or placebo infusion. Furthermore, no differences in total skin oxygen capacity at 44°C were observed (Table 3).
|
Nailfold microcirculation. Although small changes in capillary density or postocclusive recruitment were observed during either placebo or BNP infusion, overall, no significant changes between the two experiments could be observed (Table 3).
Conjunctival microcirculation. Arteriolar, capillary, and venular densities did not change significantly during either BNP or placebo infusion (Table 4).
|
|
DISCUSSION |
|---|
|
TOPABSTRACTMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
When subjects were in the supine position, BNP infusion significantly reduced SV and tended to decrease CO. However, MAP, TPR, and HR did not change significantly. Other investigators who obtained plasma levels of BNP similar to ours also observed decreases in SV without changes in CO and MAP, whereas HR only increased after infusion of higher doses of BNP (7, 9). These observations suggest that BNP primarily reduces preload, possibly by lowering venous return. This hypothesis is supported by our observations of the hemodynamic pattern with subjects in a sitting position. Indeed, when subjects rose, they experienced orthostatic symptoms and a significant decrease in MAP. At the same time, HR increased rapidly, probably due to activation of the baroreceptor reflex. In the literature, no such influence of body position on BNP effects has yet been described. However, given the age of our study group, some impairment in baroreceptor function may have exaggerated the orthostatic responses.
To the best of our knowledge, our study is the first to report that low-dose infusion of BNP has no effects on skin and conjunctival microcirculation in humans. In all likelihood, this lack of effect is not due to methodological problems such as large variability of the measurements, because by using the same setup, we were able to demonstrate microvascular effects of atrial natriuretic peptide (ANP; Ref. 3). In that study, ANP caused vasoconstriction of the microcirculation mainly on the venular side. When we combine our observations of the systemic and peripheral vasculatures, it seems that the most probable site of action for BNP is the venous system, where it may increase the "unstressed" volume. This would also explain, at least in part, the beneficial effects of this peptide in patients with congestive heart failure (2, 10). Additional studies with measurements of cardiac filling pressures and venous compliance are necessary to confirm or refute this hypothesis.
Of particular interest are the effects of BNP administration on the kidney. In previous studies, ERPF was found to decrease (5), increase (7), or remain unchanged (6, 8) during infusion of BNP. However, these studies are difficult to compare with the present one because of differences in design, BNP doses, and infusion times. For instance, concurrent changes in GFR and ERPF only occurred when the dose of BNP exceeded 2 pmol · kg–1 · min–1 (5, 7). With lower doses of BNP, La Villa et al. (8) observed a natriuretic effect of BNP in the absence of changes in ERPF and GFR. Our data are in line with the latter observations, although the dose of BNP that we employed was similar to that in the studies of Jensen et al. (5) and La Villa et al. (7). However, Jensen et al. did not compare placebo and BNP in the same subjects, which may have introduced bias. The difference between our findings and those of La Villa et al. can probably be explained by the small number of (younger) subjects and the lower salt intake in their study compared with ours. Although RVR tended to increase during BNP infusion, there were no differences in RVR or RF compared with placebo. Thus also in one of the major target organs for BNP, the peptide did not markedly influence arteriolar tone.
Besides the expected increases in plasma cGMP, urinary Na+ excretion, and urinary volume, we observed a significant increase in GFR, FF, and the filtered load of Na+ during BNP. This suggests that BNP has a direct effect on postglomerular vessels that causes vasoconstriction and an increase in FF. Although our results do not allow us to draw definite conclusions, they are compatible at least with the hypothesis that this vasoconstriction occurs at the level of the peritubular vessels rather than at the level of the efferent arterioles. Indeed, an increase in FF due to increased efferent arteriolar resistance would tend to enhance proximal tubular reabsorption of Na+. In fact, others have demonstrated that proximal reabsorption of Na+ may be reduced by BNP (5, 6), which would be expected if the site of increased resistance was located further down the nephron and intrarenal physical factors raised peritubular hydrostatic pressure.
Although we did not perform a head-to-head comparison of BNP and ANP in this study, data from a previous study by our laboratory suggest that both peptides have different effects not only on microcirculatory but also on central and renal hemodynamics. In that study, we found that ANP had no effect on MAP and HR measurements even with subjects in a sitting position (3). Furthermore, in that study, ANP infusion decreased GFR and ERPF and increased RVR values to stimulate natriuresis and diuresis (3). Finally, ANP caused venular vasoconstriction in the microcirculation. Thus BNP and ANP seem to have differential actions on the vascular system. Unfortunately, it is not yet possible to explain the differences between such actions on the basis of known mechanisms, because data in the literature on ANP and BNP are too diverse with respect to dosing, duration of administration of the peptides, and general study design.
In conclusion, this study suggests that low-dose intravenous infusion of BNP in healthy subjects has predominantly central and renal hemodynamic effects, whereas it does not influence peripheral microcirculation.
|
FOOTNOTES |
|---|
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
|
REFERENCES |
|---|
|
TOPABSTRACTMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
This article has been cited by other articles:
|
|
 |
|
  I. George, B. Morrow, K. Xu, G.-H. Yi, J. Holmes, E. X. Wu, Z. Li, A. A. Protter, M. C. Oz, and J. Wang Prolonged effects of B-type natriuretic peptide infusion on cardiac remodeling after sustained myocardial injury Am J Physiol Heart Circ Physiol, August 1, 2009; 297(2): H708 - H717. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V. Mitrovic, P. M. Seferovic, D. Simeunovic, A. D. Ristic, M. Miric, V. S. Moiseyev, Z. Kobalava, K. Nitsche, W.-G. Forssmann, H. Luss, et al. Haemodynamic and clinical effects of ularitide in decompensated heart failure Eur. Heart J., December 1, 2006; 27(23): 2823 - 2832. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. W. Vogelsang, C. C. Yoshiga, M. Hojgaard, A. Kjaer, J. Warberg, N. H. Secher, and S. Volianitis The plasma atrial natriuretic peptide response to arm and leg exercise in humans: effect of posture Exp Physiol, July 1, 2006; 91(4): 765 - 771. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Visit Other APS Journals Online |
) or brain
natriuretic peptide (BNP; 
) infusion. Data are presented as medians and
interquartile ranges. No significant changes were observed.
