| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Departments of 1 Physiology and Pharmacology and 2 Medicine (Cardiology), Oregon Health and Science University, Portland, Oregon 97201
 |
ABSTRACT |
|---|
 TOP
 ABSTRACT
 INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Pregnancy produces marked
systemic vasodilation, but the mechanism is unknown. Experiments
were performed in conscious rabbits to test the hypotheses that
increased nitric oxide (NO) production contributes to the increased
vascular conductance, but that the contribution varies among vascular
beds. Rabbits were instrumented with aortic and vena caval catheters
and ultrasonic flow probes implanted around the ascending aorta,
superior mesenteric artery, terminal aorta, and/or a femoral artery.
Hemodynamic responses to intravenous injection of
N
-nitro-L-arginine
(L-NA; 20 mg/kg or increasing doses of 2, 5, 10, 15, and 20 mg/kg) were determined in rabbits first before pregnancy (NP) and then
at the end of gestation (P). L-NA produced similar
increases in arterial pressure between groups, but the following
responses were larger (P < 0.05) when the rabbits were pregnant: 1) decreases in total peripheral conductance
[
3.7 ± 0.3 (NP), 5.0 ± 0.5 (P)
ml · min
1 · mmHg1],
2) decreases in mesenteric conductance [0.47 ± 0.05 (NP), 0.63 ± 0.07 (P)
ml · min1 · mmHg1],
3) decreases in terminal aortic conductance [0.43 ± 0.05 (NP), 0.95 ± 0.19 ml · min1 · mmHg1 (P)],
and 4) decreases in heart rate [41 ± 4 (NP),
62 ± 5 beats/min (P)]. Nevertheless, total peripheral and
terminal aortic conductances remained elevated in the pregnant rabbits
(P < 0.05) after L-NA. Furthermore,
decreases in cardiac output and femoral conductance were not different
between the reproductive states. We conclude that the contribution of
NO to vascular tone increases during pregnancy, but only in some
vascular beds. Moreover, the data support a role for NO in the
pregnancy-induced increase in basal heart rate. Finally, unknown
factors in addition to NO must also underlie the basal vasodilation
observed during pregnancy.
heart rate; cardiac output; arterial pressure; mesenteric flow; femoral flow; terminal aortic flow; total peripheral resistance; conductance
| |
INTRODUCTION |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
MARKED SYSTEMIC VASODILATION is a well-described feature of normal mammalian pregnancy (15, 20), yet the mechanism for this response is unknown. The discovery of the potent endothelial vasodilator, nitric oxide (NO), initiated significant investigation into the potential role of this factor. Indeed, considerable indirect evidence has accumulated suggesting that basal NO production is elevated during pregnancy (for reviews, see Refs. 18, 25, and 26). For example, increased levels of metabolic degradation products of NO, nitrate and nitrite, as well as the NO second messenger, cGMP, have been found in plasma and urine of pregnant animals. Moreover, uterine arteries exhibit increased nitric oxide synthase (NOS) activity and protein expression during pregnancy.
In contrast, a more direct assessment of NO activity fails to support the hypothesis that increased NO production underlies the systemic vasodilation of pregnancy. More specifically, it has been repeatedly found that acute administration of NOS antagonists produces the same increase in arterial pressure in pregnant and virgin animals (25). However, NOS blockade may have produced greater vasoconstriction in pregnant animals, but this action was not evident from the pressor response because of a simultaneously greater decrease in cardiac output. Surprisingly, no studies have been performed that determine the effect of acute NOS blockade on cardiac output and peripheral resistance (or conductance, the inverse of resistance) in pregnant and nonpregnant conscious animals, to address this issue. Therefore, one aim of the present study was to test the hypothesis that increased NO production mediates the pregnancy-induced increase in vascular conductance by determining if acute NOS blockade decreases the flow per unit driving force, or total peripheral conductance, more in pregnant compared with nonpregnant conscious rabbits.
Another possible explanation for the inability of acute NOS blockade to
elevate pressure more during pregnancy is that NOS activity is
increased in selective vascular beds, but decreased in others. Only
limited studies of acute NOS blockade in vivo on regional vascular
resistance have been performed. Because
N-nitro-L-arginine methyl ester
(L-NAME) or
NG-monomethyl-L-arginine
treatment equalized renal vascular resistance in conscious midterm
pregnant and virgin rats, Danielson and Conrad (7)
concluded that NO contributes to renal vasodilation during pregnancy in
the rat. Infusion of L-NAME into the uterine artery of
pregnant sheep decreased uterine blood flow in one study
(19), suggesting that NO contributes to basal tone,
although nonpregnant sheep were not studied. In another study,
intrauterine L-NAME did not consistently alter uterine
vascular resistance in pregnant sheep (24). Therefore, the
role of NO in maintaining increased uterine basal flow is unclear.
Therefore, a second aim of this study is to test the hypothesis that the contribution of NO to increased vascular conductance differs among vascular beds. To test this hypothesis, changes in mesenteric, terminal aortic, and femoral flows were determined before and after NOS blockade. A key feature of these experiments is that each rabbit was studied both before pregnancy and at the end of gestation, so that potentially subtle within-animal differences could be detected.
| |
METHODS |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
All studies were conducted in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals and were approved by the Oregon Health and Science University Animal Use and Care Committee.
Surgical Preparation
Female (n = 17) New Zealand White rabbits weighing 3.7 ± 0.1 kg (nonpregnant) were used for these experiments. The rabbits were received when they were 14-15 wk old and allowed to habituate at least 1 wk. Surgery was then performed to implant nonocclusive abdominal aortic and vena caval catheters, for arterial pressure measurement and drug administration, and ultrasonic flow probes as previously described (10). Briefly, anesthesia was induced with a cocktail (1 ml/kg im) containing 5:2.5:1 of ketamine (100 mg/ml), xylazine (20 mg/ml), and acepromazine (10 mg/ml). A surgical plane of anesthesia was maintained with 1:10 ketamine-0.9% NaCl solution administered intravenously as needed. Three rabbits were then implanted with an ultrasonic flow probe (2SB, Transonic Systems) around the left femoral artery just below the inguinal ligament. A midline abdominal incision was then made, and indwelling polyethylene catheters with Silastic tips were implanted in the abdominal aorta (one) and vena cava (two). The catheters were tunneled from the abdominal cavity, subcutaneously, and were exteriorized at the nape of the neck. During the laparotomy, some rabbits received flow probes (2SB or 3SB, Transonic Systems) around the terminal aorta, just proximal to the bifurcation and/or the superior mesenteric artery. Collectively, three rabbits received a terminal aortic and mesenteric probe, and one rabbit received a femoral and a terminal aortic probe. Nine rabbits received only one probe. Flow probe leads were also tunneled subcutaneously to the nape of the neck. After a minimum 2-wk recovery period, a second surgery was performed in four additional rabbits to implant an ultrasonic flow probe (6SB, Transonic Systems) around the ascending aorta. Rabbits were initially anesthetized with the ketamine cocktail at one-half the normal dose (0.5 ml/kg); they were intubated, and a surgical plane of anesthesia was maintained with isoflurane (2%). Rabbits were then placed on a respirator, and the flow probe was implanted around the ascending aorta via a right thoracotomy through the second intercostal space. The probe leads were exteriorized at the nape of the neck, and the incision was closed in layers. All probe leads were protected in a 3.5-cm plastic pillbox that was sutured to the rabbits' skin. In addition, all probes except the aortic probes were wrapped with sterile silicon sheeting to prevent fat invagination and to lengthen probe life span. The rabbits were given penicillin G procaine (60,000 U im) just before and the day after the surgery. The animals were also injected with Buprenex (0.09 mg im) 2-3 h after surgery and again the next day. The neck incision was treated with topical Nitrofurazone antibacterial dressing for 1 wk after surgery. Catheters were flushed immediately after surgery and then three times weekly using sterile 0.9% NaCl and filled with heparin (1,000 U/ml) to maintain patency.Animals were allowed at least 2 wk for recovery from all surgery.
Beginning before surgery, the rabbits were slowly moved from the normal
high-fiber diet (Ralston Purina, 5326) to a high-protein diet (Ralston
Purina, 5321), increasing 10% high-protein/day for 10 days. The
rabbits were then maintained on 150 g/day of the high-protein diet
(0.25% sodium and 16.2% protein) to enhance breeding efficiency. All
animals were allowed free access to distilled water. During surgical
recovery, the rabbits were also conditioned to the experimental
environment by placing them at least five times (
4 h per occasion) in
a specially designed opaque Plexiglas box that was used for restraint
during experiments. Room temperature was kept between 64 and 68°F,
and a 16-h light cycle was maintained for optimum breeding.
Experimental Protocol
Initial experiments were first performed in virgin rabbits before pregnancy. Afterward, the animals were bred with noninstrumented proven male breeder rabbits, and this was considered day 1 of pregnancy. Experiments were then repeated in each animal near term (after 28-30 days of pregnancy; term is 31 days), since many of the pregnancy-induced changes in the cardiovascular system are maximized at this time (15, 20).N-nitro-L-arginine
(L-NA, Sigma) was used to block NOS. The drug was first
dissolved in about 5-8 ml of 0.5% Na2CO3
in a warm water bath, and then the pH of the solution was adjusted
close to 7.4 with 1 M HCl.
On the day of the experiment, the rabbits were placed in the Plexiglas box and allowed 30-45 min to equilibrate. Arterial pressure and heart rate were measured continuously via the aortic catheter using a Statham pressure transducer, a Grass tachometer, and a Grass polygraph. Flow probes were connected to a model T206 Transonic flowmeter, and output was displayed on the polygraph.
After collecting control data, one of the following two protocols was performed.
Protocol 1: 20 mg/kg L-NA (n = 17). This experiment was performed to determine if NO production is greater when the rabbits are pregnant. The NOS inhibitor was injected intravenously slowly over ~3 min, and hemodynamic variables were measured for the subsequent 90 min. Mesenteric, terminal aortic, femoral, and total peripheral conductances were calculated as flow/arterial pressure. Blood samples were collected just before and 30, 60, and 90 min after injection of L-NA for measurement of hematocrit and plasma protein concentration. Samples were replaced with an equal volume of saline.
Protocol 2: L-NA dose response (n = 4). The purpose of this experiment was to establish that equivalent, maximal levels of NOS blockade were achieved when the rabbits were both pregnant and nonpregnant. Four of the rabbits used in protocol 1 were bred again after the first pregnancy, and a dose-response study was performed when the rabbits were again near term. The rabbits were also studied in the nonpregnant state before the first pregnancy. The following doses of L-NA were injected at 20-min intervals: 2, 3, 5, 5, and 5 mg/kg. Because L-NA is only slowly metabolized, it was assumed that the cumulative doses were 2, 5, 10, 15, and 20 mg/kg. Data collected at the end of each 20-min period were used to assess dose-response relationships.
At the end of each experiment, L-arginine (150 mg/kg) was injected intravenously to counteract the effects of L-NA.Data and Statistical Analysis
Hematocrit was determined from triplicate blood-filled capillary tubes that were centrifuged and read with an Adams microhematocrit reader. The tubes were subsequently broken, and the plasma was used for determination of plasma protein concentration with a Hitachi refractometer (National Instruments, Baltimore, MD).Differences in basal values in the rabbits before and during pregnancy
were determined with the paired t-test (27).
Between-group differences in the hemodynamic responses to
L-NA injection were determined using two-way ANOVA for
repeated measures (randomized block) and the post hoc Tukey-Kramer
procedure (17, 27). All statistics were performed using
GB-STAT (Dynamic Microsystems, Silver Spring, MD). For Figs. 1-7,
~30-s averages of arterial pressure, heart rate, and flows
were obtained from the polygraph recordings every 5 min beginning with
5 min before injection of L-NA.
|
|
|
|
|
|
|
| |
RESULTS |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Basal Values
Pregnancy produced several significant changes in the cardiovascular system at rest (Table 1). Arterial pressure was lower, and heart rate, cardiac output, total peripheral conductance, as well as mesenteric, femoral, and terminal aortic conductances, were higher. While terminal aortic blood flow was elevated, neither mesenteric nor femoral flows were significantly increased.
|
Effects of 20 mg/kg L-NA
Acute blockade of NOS increased arterial pressure (Fig. 1; P < 0.0001) in the rabbits in both the pregnant and nonpregnant state. While the small but statistically significant pregnancy-induced reduction in arterial pressure observed at rest was absent after L-NA, the pressure elevations produced by L-NA were not different when the rabbits were pregnant (19.4 ± 1.5 mmHg) compared with before pregnancy (17.6 ± 1.3 mmHg).Heart rate was dramatically decreased (Fig. 1) in both groups.
Interestingly, however, the fall in heart rate [41 ± 4 beats/min (nonpregnant); 62 ± 5 beats/min (pregnant) after
10-15 min] was significantly greater (P < 0.05)
when the rabbits were pregnant. Another difference exhibited by the
rabbits during pregnancy was that heart rate increased (by 12 ± 3 beats/min; P < 0.05) back toward basal values over the
90-min observation period, whereas the L-NA-induced
suppression in heart rate remained stable in virgin rabbits.
Cardiac output also rapidly decreased after L-NA
administration (Fig. 2), but the
decreases were not different between groups [191 ± 15 ml/min
(nonpregnant); 218 ± 18 ml/min (pregnant)]. L-NA
decreased stroke volume in nonpregnant rabbits (2.6 ± 0.2 to
2.3 ± 0.3 ml; P < 0.05) but not in pregnant
rabbits (2.9 ± 0.2 to 2.8 ± 0.3). As a result, stroke
volume was significantly higher in pregnant rabbits after
L-NA (P < 0.05).
Total peripheral conductance was markedly suppressed after acute
blockade of NOS in both groups (Fig. 3;
P < 0.0001). However, despite the fact that the
absolute changes in arterial pressure and cardiac output were not
different between pregnant and nonpregnant rabbits, the decrease in
flow as a function of the perfusion pressure, or total peripheral
conductance, was significantly greater during pregnancy [3.7 ± 0.3 ml · min1 · mmHg1
(nonpregnant); 5.0 ± 0.5 ml · min1 · mmHg1
(pregnant); P < 0.05]. Nevertheless, after
L-NA, total peripheral conductance remained higher in
pregnant compared with nonpregnant rabbits (Fig. 3).
L-NA decreased mesenteric flow (P < 0.05)
from 71 ± 3 to 51 ± 3 ml/min before pregnancy and from
78 ± 6 to 56 ± 4 ml/min at the end of gestation, and flow
was not different between groups after L-NA. Similarly,
L-NA decreased mesenteric conductance in both pregnant and
nonpregnant rabbits (Fig. 4;
P < 0.05). Whereas basal mesenteric conductance was
elevated during pregnancy compared with before pregnancy (Table 1),
this difference was eliminated by L-NA (Fig. 4). Thus
L-NA decreased mesenteric conductance more when the rabbits
were pregnant [0.47 ± 0.05 (nonpregnant), 0.63 ± 0.07 ml · min1 · mmHg1
(pregnant); P < 0.05].
Terminal aortic flow was decreased (P < 0.01) by acute
NOS blockade from 59 ± 6 to 41 ± 6 ml/min in nonpregnant
rabbits and from 114 ± 14 to 75 ± 7 ml/min during
pregnancy. Nevertheless, flow remained elevated in the pregnant rabbits
after L-NA (P < 0.05). Terminal aortic
conductance also decreased in both groups (Fig.
5; P < 0.001), but the
decrease in pregnant rabbits (0.95 ± 0.19 ml · min1 · mmHg1) was
larger (P < 0.05) than in nonpregnant rabbits
(0.43 ± 0.05 ml · min1 · mmHg1). As with
flow, conductance remained higher after L-NA administration in the rabbits during pregnancy compared with before gestation (P < 0.01).
In the rabbits both before and during pregnancy, femoral flow [19 ± 2.5 to 12 ± 1 ml/min (nonpregnant); 20.5 ± 2.3 to
15.3 ± 1.8 ml/min (pregnant)] and conductance (Fig.
6) decreased (P < 0.05) after acute NOS blockade, but the decreases in conductance 20 min after L-NA injection were not different between groups [0.15 ± 0.02 (nonpregnant) vs. 0.17 ± 0.01 ml · min1 · mmHg1
(pregnant)]. Moreover, the higher conductance of pregnant
rabbits observed at rest (before L-NA) generally
persisted after blockade of NOS (Fig. 6).
Both hematocrit [36.0 ± 0.6% (nonpregnant); 33.6 ± 0.9% (pregnant)] and plasma protein concentration [6.7 ± 0.1 g/dl (nonpregnant); 6.2 ± 0.2 g/dl (pregnant)] were lower during pregnancy (P < 0.01). ANOVA of the effect of L-NA on both hematocrit and protein revealed a significant interaction (P < 0.02), because both variables tended to rise after L-NA during pregnancy [hematocrit 90 min after L-NA to 35.2 ± 0.9%; protein to 6.4 ± 0.2 g/dl; not significant (NS)] but did not change before pregnancy (hematocrit 90 min after L-NA to 35.9 ± 0.8%; protein to 6.6 ± 0.2 g/dl). As a result, neither protein nor hematocrit were different between groups after L-NA.
Effects of Increasing Doses of L-NA
In four rabbits, increasing doses of L-NA were administered to confirm that maximal NOS blockade had been achieved with the 20 mg/kg dose (Table 2). Increasing the cumulative dose of L-NA produced stepwise increases in arterial pressure and decreases in mesenteric conductance and heart rate (P < 0.0001). More importantly, there were no differences in the effects of 10 vs. 20 mg/kg L-NA in each group, indicating that the 20 mg/kg dose produced maximal blockade in both pregnant and nonpregnant rabbits.
|
| |
DISCUSSION |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
This study tested the hypothesis that NO mediates pregnancy-induced increases in vascular conductance in conscious rabbits. The study also tested the hypothesis that the contribution of NO to increased vascular conductance differs among vascular beds. We confirm that acute blockade of NOS with L-NA produces similar increases in arterial pressure in pregnant and nonpregnant rabbits, which previously has been interpreted to suggest that NO production is not elevated (25, 26). However, major new findings include the observation that after injection of L-NA, the following responses were larger during pregnancy: 1) decreases in total peripheral conductance, 2) decreases in mesenteric conductance, 3) decreases in terminal aortic conductance, and 4) decreases in heart rate. It was also found that the following effects of L-NA were not different between pregnant and nonpregnant rabbits: 1) decreases in cardiac output and 2) decreases in femoral conductance. Collectively, these results suggest that the contribution of NO to vascular tone is increased during pregnancy, but only in some vascular beds. Moreover, it appears that NO mediates in part the increase in resting heart rate observed during pregnancy.
Role of NO in the Global Vasodilation During Pregnancy
Early studies showing that NOS inhibition produces similar pressor responses in pregnant and nonpregnant animals seemed to suggest that NO is not responsible for the marked vasodilation that occurs during pregnancy (25). However, by itself, this observation is inconclusive, since a greater degree of vasoconstriction after NOS blockade could be masked by a simultaneously greater decrease in cardiac output. The results of the present study confirm the similar pressor responses but further show that NOS blockade produces a slight but significantly greater decrease in total peripheral conductance in pregnant compared with nonpregnant conscious rabbits. Thus we conclude that NO does appear to contribute to the global vasodilation of pregnancy.Only limited previous studies of the potential differential effects of NOS blockade on cardiac output and therefore total peripheral conductance or resistance have been performed. Studying acutely prepared, anesthetized rabbits, Losonczy et al. (16) reported that L-NAME produced smaller absolute and percent increases in total peripheral resistance in pregnant rabbits and concluded that NO does not mediate the low vascular tone of pregnancy. Fiol et al. (8) studied the effects of L-NAME in conscious ganglion-blocked pregnant and nonpregnant rats but failed to statistically compare between groups the effects of L-NAME on cardiac output or total peripheral resistance. Nevertheless, inspection of the average values indicates that while absolute increases in total peripheral resistance by L-NAME were less during pregnancy, the percent increases were similar. Again, these data do not appear to support an important role for NO as a mediator in the vasodilation. Differences in experimental preparation can explain the conflicting results of the previous and the present study; in particular, it is noteworthy that the present study was performed using conscious animals and within-animal comparisons that allowed small differences to be detected. Nevertheless, one result is clearly replicated among the three studies, that is, after NOS blockade, total peripheral conductance remains higher during pregnancy (or total peripheral resistance remains lower). Thus it appears that the contribution of NO to the overall vasodilation is at most small and that other unidentified factors must also be important.
Role of NO in Regional Vasodilation During Pregnancy
One explanation for the relatively small differential effect of NOS blockade on arterial pressure and peripheral conductance is that the effect of pregnancy on the contribution of NO to basal tone could differ among vascular beds. For this reason, a number of vascular beds were also studied in the present study. One such bed is the mesenteric. We observed that mesenteric conductance was elevated when the rabbits were pregnant and that this relative increase was abolished after NOS blockade. Therefore, we conclude that NO is a major contributor to the increased mesenteric vascular conductance observed during pregnancy in conscious rabbits.To our knowledge, this is the first study performed in conscious animals suggesting that NO is important in pregnancy-induced vasodilation in the mesenteric vascular bed. However, several previous in vitro studies have been performed. The finding that neither endothelium removal nor NOS blockade produces a greater increase in basal tone of resistance vessels from pregnant rats in vitro seems to indicate that NO production is not greater (18); however, these studies have been conducted in the absence of flow and endogenous hormones, which should enhance NO release (18). Nevertheless, studies of in vitro-perfused mesentery also generally fail to document increased basal NO production in mesenteric vessels from pregnant rats (4, 13, 23), although in one study, a greater percent increase in tone was observed after L-NA in preconstricted mesentery from pregnant rats compared with rats in diestrus or proestrus (5). Thus either the ability to detect an effect of NO in the mesentery requires an in vivo preparation or there are species differences.
During pregnancy in the present study, conductance of the terminal aorta was markedly increased and L-NA produced a greater decrease in conductance, supporting a major contribution of NO to the vasodilation of pregnancy in this vascular bed. The terminal aorta perfuses both the skin and muscle of the hindlimb as well as the uterus. Therefore, further experiments were performed to assess the role of NO in the hindlimb alone by determining the differential effects of L-NA on femoral conductance. Because L-NA failed to produce a greater decrease in femoral conductance during pregnancy, the results indicate that the elevated conductance in this bed is not dependent on NO. Similar conclusions were drawn from in vitro studies of femoral microvessels (28) and perfused hindlimbs (1) of rats. Moreover, by deduction, the results further suggest that the increased role of NO in the terminal aortic bed is due to an action in the uteroplacental unit. This conclusion agrees with multiple studies of the uterine vascular bed in vitro (for reviews, see Refs. 25 and 26). On the other hand, again, only limited in vivo studies of the effects of NOS blockade on uterine vascular tone have been performed, both in conscious pregnant sheep. Rosenfeld et al. (24) reported an inconsistent effect of intrauterine L-NAME on uterine blood flow, whereas Miller et al. (19) observed significant decreases in uterine flow and resistance. Methodological differences were cited to explain this conflict (19). Nevertheless, neither study compared the result obtained in pregnant animals with those in nonpregnant. Importantly, the present study documents a clear difference between reproductive states and indirectly suggests that at least part of the increased uterine flow during pregnancy is due to NO. Thus it appears that NO contributes not only to the generalized vasodilation of pregnancy but also to increased flow to specific vascular beds. This conclusion is supported by the study of Kassab et al. (11), who examined in pregnant and nonpregnant rats the effects of chronic L-NAME treatment on arterial pressure, cardiac output, and regional flows quantified using radioactive microspheres.
Importance of Method of Data Analysis
A major conclusion of the present study, that NO contributes to the vasodilation of pregnancy, was based primarily on the finding that the absolute decreases in total or regional conductances after L-NA were greater when the rabbits were pregnant. However, it must be acknowledged that several other methods of data analysis were possible, some of which would have led to different conclusions.The primary factor that leads to potential variance in conclusions is
that the basal values of pressure, flow, conductance, and therefore
resistance are significantly different between pregnant and nonpregnant
rabbits. Another complicating factor is that resistance is the inverse
of conductance, and the relationship between a variable and its inverse
is not linear. Because of these two factors, different conclusions
would have been made if resistance were calculated in the present study
rather than conductance. For example, the absolute increase in total
peripheral resistance after L-NA was, if anything, less
when the rabbits were pregnant [0.20 ± 0.04 (nonpregnant) vs.
0.13 ± 0.02 (pregnant)
mmHg · ml1 · min, NS; Fig.
7]. Similar trends can be found in the
other previous acute studies of cardiac output (8, 16); in
both cases, different conclusions would have been made depending on whether resistance or conductance changes were assessed.
One common strategy that is used to circumvent differences in baseline is to compare the percent changes. Indeed, in the present study, the percent decreases in terminal aortic conductance after L-NA were essentially identical between groups [decreased to 52.1 ± 3.2% of control (nonpregnant) and 51.3 ± 2.6% of control (pregnant) 20 min after L-NA], despite much larger absolute decreases observed during pregnancy. However, this strategy assumes that the relationship between NO production and vessel diameter, conductance, or resistance is always linear, which most likely is not a valid assumption. Another complicating factor is that basal conductance may be elevated during pregnancy because of increases in the radii of individual vessels present before pregnancy (e.g., mesentery) or because of the addition of vessels in parallel (e.g., uterus and placenta).
In the present study, we elected to perform between-group comparisons of the absolute changes in conductance primarily because we feel that this measure best represents the physiological contribution of NO to the increased flow of blood to the tissues, which is the primary change during pregnancy. In other words, the calculation of conductance gives the flow per unit of driving force, and it is the level of flow that is critical to tissue function. On the other hand, the calculation of resistance describes the amount of pressure produced for a given flow and is a better variable to examine if blood pressure regulation is the major interest.
The absolute level of conductance is clearly elevated during pregnancy. Because the difference in basal conductance observed in the mesenteric bed was completely eliminated after L-NA, it appears that NO is a major or the only mediator of the increased conductance during pregnancy in this bed. A similar situation has been described for the renal vascular bed of the rat (7). However, when a consideration of the total peripheral vascular tree or the terminal aortic bed is made, it can only be concluded that NO contributes to the higher absolute level of conductance, because L-NA did not eliminate differences in conductance between groups. In addition, the exact mechanism by which NO contributes is not revealed from these studies. For example, these results do not definitely prove that NO production is higher, since it could be that a change in the vascular architecture allows for a greater level of conductance for the same amount of NO produced. Indeed, this scenario may apply to the uterine bed, in which the increased conductance of pregnancy is clearly due in part to the addition of vessels in parallel. In the present study, one interpretation of the finding that the percent decreases in conductance were the same in pregnant and nonpregnant rabbits is that the NO production per vessel is the same, but the number of vessels is increased during pregnancy. Therefore, while these studies do support a role for NO in the vasodilation of pregnancy, the exact mechanism of action of NO remains to be determined. Moreover, the results suggest that other mediators must also be involved.
Role of NO in the Elevated Heart Rate During Pregnancy
Another novel observation in the present study was that L-NA decreased heart rate more when the rabbits were pregnant. Indeed, heart rate became similar in the two gestational states soon after NOS blockade, suggesting that NO production in some way contributes to the basal relative tachycardia of pregnancy. When these data are compared with our previous studies of the effect of pregnancy on baroreflex control of heart rate (2, 22), it becomes apparent that the L-NA-induced bradycardia in nonpregnant rabbits can be completely explained by the pressor effect of L-NA. On the other hand, we have consistently found that pregnancy elevates the minimum reflex heart rate produced during acute hypertension (2, 22). Because the decrease in heart rate after L-NA in the pregnant rabbits is greater than that produced by phenylephrine infusion to produce a similar pressor effect (suppressed to 125 vs. 150 beats/min), these data suggest that NO also contributes to the elevated baroreflex minimum heart rate. In addition, our previous studies indicate that the elevated minimum heart rate is due to a blunted ability to increase vagal tone (2). The enhanced bradycardia after NOS blockade in conscious rabbits appears to be due to a central effect on neuronal NOS (3, 14, 29). Therefore, it is tempting to speculate that NO acts to increase heart rate during pregnancy at least in part via an action in the brain to inhibit vagal tone.Residual Pregnancy-Induced Vasodilation After NOS Blockade
It is noteworthy that after L-NA, total peripheral conductance as well as terminal aortic and femoral conductances of pregnant rabbits remained elevated compared with conductances measured in rabbits before pregnancy. These data suggest that factors in addition to NO must contribute to the basal vasodilation of pregnancy. Alternatively, other mechanisms may counteract the vasoconstriction produced by local vascular blockade of NOS and NO production, especially because NO is known to act in numerous sites within the cardiovascular system. One possibility is that less blockade of NOS was achieved when the rabbits were pregnant. However, the dose-response study verifies that maximal blockade was produced in both groups.Another potential counteracting mechanism to the vasoconstriction produced by acute NOS blockade could be the baroreflex because, as described above, reflex decreases in heart rate are enhanced after L-NA when the rabbits are pregnant. Thus the reflex may counteract the pressor effect of L-NA more in pregnant compared with nonpregnant animals, which could lead to smaller increases in pressure and/or smaller decreases in conductances. However, it seems unlikely that enhanced baroreflex activity could completely explain the large difference in conductance that remains between groups after L-NA. Indeed, the decreases in cardiac output after L-NA were similar between groups. Moreover, lower levels of arterial pressure and total peripheral resistance persist after administration of L-NAME in ganglion-blocked, pregnant compared with nonpregnant conscious rats, in which reflex function is blocked (8). Collectively, these observations suggest that the baroreflex may quantitatively reduce the ability of L-NA to decrease conductances. However, because the absolute decreases in conductances after L-NA were greater in pregnant animals, it appears that qualitatively NO does contribute to total peripheral conductance as well as to some regional conductances. Furthermore, additional unidentified factors must also contribute.
In summary, the present studies of conscious rabbits in both the pregnant and nonpregnant state suggest that NO contributes to the increased vascular conductance and therefore the generalized vasodilation of pregnancy, in particular in the mesenteric and uterine beds. We also demonstrated a role for NO in the relative tachycardia exhibited by late pregnant rabbits. Finally, the data indicate that factors in addition to NO underlie the increased vascular conductance observed during pregnancy.
Perspectives
A major conclusion of this study is that factors in addition to NO must contribute to the enhanced vasodilation of pregnancy. While the specific mediators are unknown, possible contributors include the increased vascularity of the uterus and placenta and increased activity of an endothelium-derived hypopolarizing factor (5, 9). One reason why the identity of these additional factors has been illusive is that the hormonal milieu of pregnancy is exceedingly complex. Nevertheless, whether the vasodilation, either via NO or another factor, is initiated by a signal from the uteroplacental unit or from another site, such as the ovary, can be indirectly assessed from correlations between the number or size of the conceptus and the hemodynamic changes. The number of offspring delivered varies widely in rabbits; therefore, a role for a uteroplacental signal can be indirectly determined from the relationship between the number of kits born vs. the changes in regional conductances. When we performed such correlations, including rabbits in the present study as well as other studies, we found that there was no correlation between the number of kits and the increases from the nonpregnant to the pregnant state in cardiac output, total peripheral conductance, and mesenteric conductances. This finding is consistent with the lack of a temporal correlation between the increases in cardiac output and total systemic conductance, which occur early in pregnancy, to the increase in uterine vascular conductance, which follows much later (21). On the other hand, the pregnancy-induced increases in terminal aortic flow and conductances in the present study were correlated to the number of offspring (r = 0.53, P < 0.05, n = 11). Similarly, while the decrease in terminal aortic conductance produced by L-NA in the pregnant animals, relative to the decrease produced before pregnancy, was correlated to the number of kits born (r = 0.73, P < 0.05, n = 6), the action of L-NA on total peripheral conductance was not related to the number of kits. Finally, we found that after L-NA, the residual increases in terminal aortic and total peripheral conductances in pregnant compared with nonpregnant rabbits, which are an index of the non-NO component of the vasodilation, were not correlated to the number of offspring. Collectively, this analysis indirectly suggests that the increase in conductance to the uterus and the role of NO in this increase do appear to depend on the fetus and/or placenta. On the other hand, the global vasodilation of pregnancy appears to depend in part on an unknown factor originating from a site other than the fetal-placental unit, such as ovarian-derived gonadal steroids or relaxin. This idea agrees with previous considerations (6, 12, 21). Nevertheless, the identification of this factor(s) awaits further investigation.| |
ACKNOWLEDGEMENTS |
|---|
We are grateful for the surgical assistance of Dr. B. Ogden and for the suggestions made by Dr. K. Gamperl during preparation of the manuscript.
| |
FOOTNOTES |
|---|
This study was supported in part by National Heart, Lung, and Blood Institute Grant HL-39923.
Address for reprint requests and other correspondence: V. L. Brooks, Dept. of Physiology and Pharmacology, L-334, Oregon Health Sciences Univ., Portland, OR 97201 (E-mail: brooksv{at}ohsu.edu).
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.
Received 8 February 2001; accepted in final form 20 July 2001.
| |
REFERENCES |
|---|
|
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
1.
Ahokas, RA,
and
Sibai BM.
Endothelium-derived relaxing factor inhibition augments vascular angiotensin II reactivity in the pregnant rat hind limb.
Am J Obstet Gynecol
167:
1053-1058,
1992[ISI][Medline].
2.
Brooks, VL,
Kane CM,
and
Van Winkle DM.
Altered baroreflex control of heart rate during pregnancy: role of sympathetic and parasympathetic nervous system.
Am J Physiol Regulatory Integrative Comp Physiol
273:
R960-R966,
1997
3.
Chowdhary, S,
and
Townend JN.
Role of nitric oxide in the regulation of cardiovascular autonomic control.
Clin Sci
97:
5-17,
1999[Medline].
4.
Chu, ZM,
and
Beilin LJ.
Mechanisms of vasodilatation in pregnancy: studies of the role of prostaglandins and nitric-oxide in changes of vascular reactivity in the in situ blood perfused mesentery of pregnant rats.
Br J Pharmacol
109:
322-329,
1993[ISI][Medline].
5.
Dalle Lucca, JJ,
Adeagbo ASO,
and
Alsip NL.
Influence of oestrus cycle and pregnancy on the reactivity of the rat mesenteric vascular bed.
Hum Reprod
15:
961-968,
2000
6.
Danielson, LA,
Sherwood OD,
and
Conrad KP.
Relaxin is a potent renal vasodilator in conscious rats.
J Clin Invest
103:
525-533,
1999[ISI][Medline].
7.
Danielson, LA,
and
Conrad KP.
Acute blockade of nitric oxide synthesis inhibits renal vasodilation and hyperfiltration during pregnancy in chronically instrumented conscious rats.
J Clin Invest
96:
482-490,
1995.
8.
Fiol, G,
Machado F,
Hernandez I,
Ingles AC,
Abad L,
Parrilla JJ,
Meseguer J,
Quesada T,
and
Carbonell LF.
Role of nitric oxide on the central hemodynamic response to acute volume expansion in the pregnant rat.
Am J Obstet Gynecol
178:
823-829,
1998[ISI][Medline].
9.
Gerber, RT,
Anwar MA,
and
Poston L.
Enhanced acetylcholine induced relaxation in small mesenteric arteries from pregnant rats: an important role for endothelium-derived hyperpolarizing factor (EDHF).
Br J Pharmacol
125:
455-460,
1998[ISI][Medline].
10.
Gronan, RJ,
Schadt JC,
and
York DH.
Routine, direct measurement of aortic pressure in the conscious rabbit.
Physiol Behav
30:
719-722,
1983[Medline].
11.
Kassab, S,
Miller MT,
Hester R,
Novak J,
and
Granger JP.
Systemic hemodynamics and regional blood flow during chronic nitric oxide synthesis inhibition in pregnant rats.
Hypertension
31:
315-320,
1998
12.
Kristiansson, P,
and
Wang JX.
Reproductive hormones and blood pressure during pregnancy.
Hum Reprod
16:
13-17,
2001
13.
Le Marquer-Domagala, F,
and
Finet M.
Roles of NO-synthase and cyclooxygenase in sex- and pregnancy-dependent arterial and venous pressures in the rat.
J Cardiovasc Pharmacol
30:
205-213,
1997[ISI][Medline].
14.
Liu, JL,
Murakami H,
and
Zucker IH.
Effects of NO on baroreflex control of heart rate and renal nerve activity in conscious rabbits.
Am J Physiol Regulatory Integrative Comp Physiol
270:
R1361-R1370,
1996
15.
Longo, LD.
Maternal blood volume and cardiac output during pregnancy: a hypothesis of endocrinologic control.
Am J Physiol Regulatory Integrative Comp Physiol
245:
R720-R729,
1983
16.
Losonczy, G,
Mucha I,
Müller V,
Kriston T,
Ungvári Z,
Tornóci L,
Rosivall L,
and
Venuto R.
The vasoconstrictor effects of L-NAME, a nitric oxide synthase inhibitor, in pregnant rabbits.
Br J Pharmacol
118:
1012-1018,
1996[ISI][Medline].
17.
Ludbrook, J.
On making comparisons in clinical and experimental pharmacology and physiology.
Clin Exp Pharmacol Physiol
18:
379-392,
1991[ISI][Medline].
18.
McLaughlin, MK,
and
Conrad KP.
Nitric oxide biosynthesis during pregnancy: implications for circulatory changes.
Clin Exp Pharmacol Physiol
22:
164-171,
1994.
19.
Miller, SL,
Jenkin G,
and
Walker DW.
Effect of nitric oxide synthase inhibition on the uterine vasculature of late-pregnant sheep.
Am J Obstet Gynecol
180:
1138-1145,
1999[ISI][Medline].
20.
Morton, MJ.
Maternal hemodynamics in pregnancy.
In: Exercise and Pregnancy, edited by Artal R,
and Wiswell RA.. Baltimore, MD: Williams and Wilkins, 1990, p. 61-70.
21.
Morton, MJ.
Cardiovascular adaptation to pregnancy.
In: Sex Steroids and the Cardiovascular System, edited by Ramwell P,
Rubanyi G,
and Schillinger E.. New York: Springer-Verlag, 1992, p. 53-64.
22.
Quesnell, RR,
and
Brooks VL.
Alterations in the baroreflex occur late in pregnancy in conscious rabbits.
Am J Obstet Gynecol
176:
692-694,
1997[ISI][Medline].
23.
Ralevic, V,
and
Burnstock G.
Mesenteric arterial function in the rat in pregnancy: role of sympathetic and sensory-motor perivascular nerves, endothelium, smooth muscle, nitric oxide and prostaglandins.
Br J Pharmacol
117:
1463-1470,
1996[ISI][Medline].
24.
Rosenfeld, CR,
Cox BE,
and
Magness RR.
Nitric oxide contributes to estrogen-induced vasodilation of the ovine uterine circulation.
J Clin Invest
98:
2158-2166,
1996[ISI][Medline].
25.
Sladek, SM,
Magness RR,
and
Conrad KP.
Nitric oxide and pregnancy.
Am J Physiol Regulatory Integrative Comp Physiol
272:
R441-R463,
1997
26.
Weiner, CP,
and
Thompson LP.
Nitric oxide and pregnancy.
Semin Perinatol
21:
367-380,
1997[ISI][Medline].
27.
Winer, BJ.
Statistical Principles in Experimental Design. New York: McGraw-Hill, 1971.
28.
Yamasaki, M,
Lindheimer MD,
and
Umans JG.
Effects of pregnancy on femoral microvascular responses in the rat.
Am J Obstet Gynecol
175:
730-736,
1996[ISI][Medline].
29.
Zanzinger, J.
Role of nitric oxide in the neural control of cardiovascular function.
Cardiovasc Res
43:
639-649,
2000.
This article has been cited by other articles:
|
|
 |
|
  D. L. Daubert, D. Liu, I. H. Zucker, and V. L. Brooks Roles of nitric oxide and angiotensin II in the impaired baroreflex gain of pregnancy Am J Physiol Regulatory Integrative Comp Physiol, June 1, 2007; 292(6): R2179 - R2187. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. J. Stark, L. Dierkx, V. L. Clifton, and I. M. R. Wright Alterations in the Maternal Peripheral Microvascular Response in Pregnancies Complicated by Preeclampsia and the Impact of Fetal Sex Reproductive Sciences, December 1, 2006; 13(8): 573 - 578. [Abstract] [PDF]
|
|
|
|
 |
|
 K. A. Clow, G. D. Giraud, B. E. Ogden, and V. L. Brooks Pregnancy alters hemodynamic responses to hemorrhage in conscious rabbits Am J Physiol Heart Circ Physiol, April 1, 2003; 284(4): H1110 - H1118. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. P. Granger Maternal and fetal adaptations during pregnancy: lessons in regulatory and integrative physiology Am J Physiol Regulatory Integrative Comp Physiol, December 1, 2002; 283(6): R1289 - R1292. [Full Text] [PDF]
|
|
|
|
|
|
P. B. Persson Nitric oxide in the kidney Am J Physiol Regulatory Integrative Comp Physiol, November 1, 2002; 283(5): R1005 - R1007. [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Visit Other APS Journals Online |
) and nonpregnant (
) rabbits
(n = 17).
