This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Khalil, R. A. Articles by Granger, J. P. Search for Related Content PubMed PubMed Citation Articles by Khalil, R. A. Articles by Granger, J. P.

(Hypertension. 1998;31:1065-1069.)
© 1998 American Heart Association, Inc.

Scientific Contributions

Enhanced Vascular Reactivity During Inhibition of Nitric Oxide Synthesis in Pregnant Rats

Raouf A. Khalil; Janice K. Crews; Jacqueline Novak; Salah Kassab; ; Joey P. Granger

From the Department of Physiology and Biophysics and Center for Excellence in Cardiovascular-Renal Research, University of Mississippi Medical Center (Jackson).

	Abstract

Top Abstract Introduction Methods Results Discussion References

 
Abstract—Pregnancy-induced hypertension has been suggested to be mediated by several mechanisms, including reduced nitric oxide (NO) synthesis. In this study, the effects of chronic treatment with the NO synthase inhibitor N^G-nitro-L-arginine methyl ester (L-NAME) on blood pressure and the underlying changes in vascular reactivity were investigated in virgin and late-pregnancy Sprague-Dawley rats. The systolic blood pressure was 120±2, 124±5, 116±4, and 171±5 mm Hg in untreated virgin, virgin treated with L-NAME, untreated pregnant, and pregnant treated with L-NAME rats, respectively. The rats were killed, and the thoracic aorta was cut into strips for measurement of active stress in response to ₁-adrenergic stimulation with phenylephrine and membrane depolarization by high KCl. In pregnant rats, the maximal active stress to phenylephrine (0.31±0.03x104 N/m²) and the high-KCl–induced active stress (0.55±0.09x10⁴ N/m²) were smaller than those in virgin rats. By contrast, in the L-NAME–treated pregnant rats, the maximal phenylephrine-induced active stress (0.76±0.1x10⁴ N/m²) was greater than that in virgin rats (0.52±0.1x10⁴ N/m²), whereas the high-KCl–induced active stress (1.08±0.14x10⁴ N/m²) was indistinguishable from that in virgin rats (1.03±0.14x10⁴ N/m²). Treatment with L-NAME did not affect the phenylephrine-releasable Ca²⁺ stores in pregnant rats and had minimal effect on active stress in virgin rats. Thus, reduction of NO synthesis during late pregnancy is associated with a significant increase in blood pressure and vascular responsiveness to -adrenergic stimulation, which can possibly be explained in part by enhanced Ca²⁺ entry from extracellular space. However, other mechanisms such as increased myofilament force sensitivity to Ca²⁺ and/or activation of a completely Ca²⁺-independent mechanism cannot be excluded.

Key Words: blood pressure • calcium • muscle, smooth • contraction

	Introduction

Top Abstract Introduction Methods Results Discussion References

 
Normal pregnancy is associated with many hemodynamic changes such as increased heart rate and cardiac output,¹ increased plasma volume, and an increase in uterine^{2 3 4} and renal blood flow.⁵ Despite the increase in heart rate, blood volume, and cardiac output, normal pregnancy is usually associated with a significant decrease in arterial blood pressure and total peripheral resistance.⁶ This normal pregnancy-associated decrease in peripheral resistance has been explained by several mechanisms, including an increase in the metabolic requirements of both maternal and fetoplacental tissues and/or a decrease in vascular reactivity.⁷ Several explanations have been proposed for the decrease in vascular reactivity during normal pregnancy, such as decreased pressor response to vasoconstrictors,^{8 9 10} specific alterations within the vascular wall,¹¹ and an increase in nitric oxide (NO) synthesis.^{12 13} Also, Conrad and Vernier¹⁴ have found that the plasma level, metabolic production, and urinary excretion of cGMP, a second messenger of NO and a cellular mediator of vascular smooth muscle relaxation, are increased during pregnancy.

In 5% to 7% of pregnancies, women develop a condition called pregnancy-induced hypertension.¹⁵ In contrast to normal pregnancy, pregnancy-induced hypertension is characterized by increased arterial blood pressure, generalized vasoconstriction, increased systemic vascular resistance, increased capillary permeability, decreased plasma volume, severe edema, increased intravascular coagulation, reduced tissue perfusion, decreased glomerular filtration rate, proteinuria, and widespread vascular endothelial damage.^{15 16}

Although pregnancy-induced hypertension is a leading cause of maternal and fetal mortality, the exact mechanism of this disorder has not yet been clearly identified. Several mechanisms have been suggested, including reduction of NO synthesis^{17 18} and/or changes in the vasoconstrictor receptor affinity or density.⁹ Consistent with these suggested mechanisms are reports that chronic NO synthase blockade during mid to late gestation in rats results in many pathological changes similar to those found in women with pregnancy-induced hypertension, such as increased blood pressure, proteinuria, thrombocytopenia, and intrauterine growth retardation.^{19 20 21 22} These observations have led investigators to suggest the use of pregnant rats chronically treated with NO synthase blockers as a model to study pregnancy-induced hypertension.^{19 20 21 22} Although chronic NO synthase blockade in pregnant rats has been shown to enhance the pressor response to vasoconstrictor substances,⁸ it is not clear whether these pressor responses reflect changes in the vascular reactivity. The present study was designed to investigate whether chronic NO synthase blockade in pregnant rats is associated with changes in vascular reactivity. Pregnant rats were treated chronically with N^G-nitro-L-arginine methyl ester (L-NAME) during mid to late gestation. L-NAME is a structural analogue of L-arginine and is known to inhibit the synthesis of NO. The L-NAME–induced changes in blood pressure were measured, and the underlying changes in vascular reactivity were investigated for possible alterations in the Ca²⁺-handling mechanisms.

	Methods

Top Abstract Introduction Methods Results Discussion References

 
Animals
Female Sprague-Dawley rats (10 to 12 weeks of age) were purchased from Harlan Sprague-Dawley, Inc (Indianapolis, Ind). Rats were divided into four groups: (1) virgin, (2) virgin treated with L-NAME, (3) pregnant, and (4) pregnant treated with L-NAME. The first day of pregnancy was verified by the presence of sperm in vaginal smears (full term is 21 to 22 days). All procedures were performed in accordance with the guidelines of the Institutional Animal Care and Use Committee at the University of Mississippi Medical Center and the American Physiological Society.

Measurement of Blood Pressure
Systolic blood pressure was measured in the four groups of rats with an automated sphygmomanometer with a tail-cuff device (IITC Life Science Inc). On the day of the experiment (typically 18 to 20 days of gestation in pregnant rats or the equivalent period in virgin rats), pretrained rats were placed in a warm chamber maintained at 30°C for 1 hour, then each rat was transferred to an individual Plexiglas restrainer. The pneumatic tail-cuff device was placed around the rat's tail; at least three pressure measurements were recorded for each rat, and the average systolic blood pressure was calculated.

Tissue Preparation
Rats were terminally anesthetized by inhalation of chloroform. The thoracic aorta was rapidly removed, placed in oxygenated Krebs' solution, and cleaned of connective tissue. The aorta was cut transversely into 3-mm-wide rings. The endothelium was removed by rubbing of the vessel interior with forceps. Rings were cut open into strips. One end of the strip was attached to a glass hook using a thread loop, and the other end was connected to a Grass force transducer (FT03, Astro-Med Inc).

Isometric Tension
Aortic strips were stretched to 2 g of tension and allowed to equilibrate for 1 hour in an organ bath filled with 50 mL Krebs' solution continuously bubbled with 95% O₂/5% CO₂ at 37°C. The changes in isometric tension were recorded on a Grass polygraph (model 7D, Astro-Med Inc). Removal of the endothelium was routinely verified by the absence of acetylcholine (10^-6 mol/L)–induced vasorelaxation in aortic strips precontracted with phenylephrine (3x10^-7 mol/L).

Three different protocols were followed in this study. In the first protocol, after the rat aortic strip was allowed to equilibrate for 1 hour, a control contraction was elicited by applying 10^-5 mol/L phenylephrine to the organ bath solution. Once the phenylephrine contraction reached a plateau, the tissue was rinsed with Krebs' solution three times for a duration of 10 minutes each. The whole procedure of contraction and washing was repeated two times. Phenylephrine was then added cumulatively to the organ bath at concentrations ranging from 10^-9 mol/L to 10^-4 mol/L, and the changes in isometric tension were recorded. For each phenylephrine concentration, the contraction was allowed to reach a plateau before the addition of the next concentration.

In the second protocol, control phenylephrine (10^-5 mol/L) contractions were elicited; then the tissue was incubated in Ca²⁺-free (2 mmol/L EGTA) Krebs' solution for 10 minutes. Phenylephrine (10^-5 mol/L) was added, and the transient phenylephrine contraction in Ca²⁺-free solution was recorded.

In the third protocol, control contractions were elicited using 96 mmol/L KCl solution, and the tissue was rinsed with Krebs' solution three times for a duration of 10 minutes each. This procedure was repeated two times. Increasing concentrations of KCl solution (16, 24, 36, 51, 66, 78, 86, and 96 mmol/L) were added to the organ bath separately. Contractions were allowed to reach a steady plateau before the solution was drained and the next concentration of KCl was added.

Solutions
The Krebs' solution contained (in mmol/L) NaCl 120, KCl 5.9, NaHCO₃ 25, NaH₂PO₄ 1.2, dextrose 11.5, MgCl₂ 1.2, and CaCl₂ 2.5. The solution was bubbled with 95% O₂/5% CO₂ to adjust the pH to 7.4. For the Ca²⁺-free Krebs' solution, CaCl₂ was omitted and replaced with 2 mmol/L EGTA. The high-KCl depolarizing solution was prepared as Krebs' solution but with equimolar substitution of NaCl with KCl.

Drugs and Chemicals
The stock solution of phenylephrine (L-phenylephrine hydrochloride; Sigma Chemical Co) was prepared as 10^-1 mol/L in distilled water. Diluted phenylephrine solutions were also made in distilled water. The L-NAME (Sigma) solution was prepared by adding 50 mg to 1 L of the rat's drinking water. This L-NAME concentration resulted in a daily intake of approximately 1 mg/d. This dose of L-NAME was chosen based on studies of Molnar and coworkers²¹ that showed that a dose of approximately 1 mg/d resulted in significant elevation of blood pressure in pregnant rats while having minimal effect in virgin rats. The L-NAME–treated rats were allowed to drink the water containing L-NAME for 4 to 6 days before blood pressure measurements were taken or excision of the aorta. All other chemicals were of reagent grade or better.

Statistical Analysis
The developed force was corrected for the cross-sectional area of each individual strip and expressed as active stress (N/m²) using the following equation: stress=force/cross-sectional area, where cross-sectional area equals wet weight/(tissue densityxlength of the strip), and tissue density equals 1.055 g/cm³. Data were analyzed and expressed as mean±SEM. Data were compared using one-way ANOVA with Scheffé's test and unpaired Student's t test. Differences <.05 were considered statistically significant.

	Results

Top Abstract Introduction Methods Results Discussion References

 
Fig 1 shows the systolic blood pressure recorded in the four groups of rats. In the virgin rats, the systolic blood pressure was 120±2 mm Hg (n=12). The systolic blood pressure of the pregnant rats was slightly lower (116±4 mm Hg, n=17) when compared with the virgin rats. In contrast, the systolic blood pressure of the pregnant rats treated with L-NAME (171±5 mm Hg, n=14) was significantly higher than that in the virgin rats. On the other hand, the systolic blood pressure of the virgin rats treated with L-NAME (124±5 mm Hg, n=12) was not significantly different from that in the virgin rats.

View larger version (14K): [in this window] [in a new window]   Figure 1. Systolic blood pressure in virgin and pregnant rats untreated or treated with L-NAME. Bars represent the mean±SEM of individual measurements in 12 to 17 rats. *Significant (P<.05).

Fig 2 shows the contractile response to phenylephrine in rat aortic strips isolated from the four groups of rats. All groups showed a concentration-dependent increase in active stress to phenylephrine. In the virgin rats, phenylephrine (10^-5 mol/L) produced an active stress of 0.52±0.10x10⁴ N/m² (n=7). The active stress in the pregnant rats was markedly reduced. In the pregnant rats, phenylephrine (10^-5 mol/L) produced an active stress of 0.31±0.03x10⁴ N/m² (n=8). In contrast, the active stress in the pregnant rats treated with L-NAME was markedly increased. In the pregnant rats treated with L-NAME, phenylephrine (10^-5 mol/L) produced an active stress of 0.76±0.10x10⁴ N/m² (n=8). On the other hand, active stress in the virgin rats and the virgin rats treated with L-NAME showed no significant difference. In the virgin rats treated with L-NAME, phenylephrine (10^-5 mol/L) produced an active stress of 0.57±0.11x10⁴ N/m² (n=8).

View larger version (19K): [in this window] [in a new window]   Figure 2. Steady-state active stress induced by phenylephrine in aortic strips isolated from virgin and pregnant rats untreated or treated with L-NAME. Rat aortic strips were incubated in normal Krebs' solution (2.5 mmol/L Ca2+). Active stress was recorded at increasing concentrations of phenylephrine. The data points represent the mean±SEM of measurements in individual aortic strips from 7 to 8 rats.

Fig 3 shows that when the contractile response to increasing concentrations of phenylephrine was presented as a percentage of the maximal phenylephrine contraction, no significant difference among any of the four groups was observed.

View larger version (18K): [in this window] [in a new window]   Figure 3. Steady-state contraction induced by phenylephrine in aortic strips isolated from virgin and pregnant rats untreated or treated with L-NAME. Contraction was measured at increasing concentrations of phenylephrine and presented as percentage of maximal (10-5 mol/L) phenylephrine-induced contraction. The data points represent the mean±SEM of measurements in individual aortic strips from 7 to 8 rats.

Phenylephrine-induced contraction in Ca²⁺-free solution is often used as a measure of the phenylephrine-releasable intracellular Ca²⁺ stores. Fig 4 shows the phenylephrine contraction of rat aortic strips in Ca²⁺-free (2 mmol/L EGTA) Krebs' solution. In all groups of rats, phenylephrine showed a transient increase in active stress in Ca²⁺-free solution. However, there was no significant difference in the active stress among any of the four groups.

View larger version (15K): [in this window] [in a new window]   Figure 4. Transient active stress induced by phenylephrine in aortic strips isolated from virgin and pregnant rats untreated or treated with L-NAME. Rat aortic strips were incubated in Ca2+-free (2 mmol/L EGTA) Krebs' solution for 10 minutes. Active stress in response to phenylephrine (10-5 mol/L) was recorded. The data points represent the mean±SEM of measurements in individual aortic strips from 6 to 8 rats.

Membrane depolarization by high-KCl solution is known to activate Ca²⁺ entry through voltage-gated Ca²⁺ channels. Fig 5 shows the effect of increasing concentrations of extracellular KCl on contraction. In the virgin rats, 96 mmol/L KCl produced an active stress of 1.03±0.14x10⁴ N/m² (n=6). The 96 mmol/L KCl active stress in the pregnant rats was reduced to 0.55±0.09x10⁴ N/m² (n=8). On the other hand, the 96 mmol/L KCl–induced active stress in the virgin rats treated with L-NAME (0.94±0.12x10⁴ N/m,² n=17) and pregnant rats treated with L-NAME (1.08±0.14x10⁴ N/m,² n=9) was not significantly different from that in the virgin rats.

View larger version (17K): [in this window] [in a new window]   Figure 5. Depolarization-induced active stress in aortic strips isolated from virgin and pregnant rats untreated or treated with L-NAME. Rat aortic strips were incubated in normal Krebs' solution (2.5 mmol/L Ca2+). Active stress was recorded at increasing extracellular concentrations of KCl. The data points represent the mean±SEM of measurements in individual aortic strips from 6 to 17 rats.

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
The major findings of this study are that (1) the systolic blood pressure of pregnant rats treated with L-NAME was significantly greater than that of the pregnant rats, the virgin rats, and the virgin rats treated with L-NAME; (2) the vascular reactivity to phenylephrine was decreased in the pregnant rats and increased in the pregnant rats treated with L-NAME when compared with the virgin rats and the virgin rats treated with L-NAME; (3) the transient phenylephrine contraction in Ca²⁺-free solution, a measure of the phenylephrine-releasable Ca²⁺ stores, showed no difference among any of the four groups; and (4) an observed reduced vascular reactivity to membrane depolarization by KCl in pregnant rats was corrected only to the virgin levels when the pregnant rats were treated with L-NAME.

Our results show that systolic blood pressure is slightly lower in pregnant rats than in virgin rats. These results are consistent with the findings of other laboratories.^{7 8 9 11 12 23} This normal pregnancy-induced drop in blood pressure could be explained by changes in the function of the kidneys, metabolic changes, and/or changes in vascular reactivity. On the other hand, chronic NO synthase blockade with L-NAME caused severe hypertension in pregnant rats. The same dose of L-NAME had minimal effect on blood pressure in virgin rats. These data are consistent with the findings of Molnar and colleagues²¹ and further support a role for NO synthase blockade in pregnancy-induced hypertension.

The present study showed that the active stress in response to phenylephrine was greater in the pregnant rats treated with L-NAME than in the pregnant rats. However, since the pregnant rats had a lower blood pressure than the virgin control animals, and the pregnant rats treated with L-NAME showed a significant increase in blood pressure, the possible role of vascular wall remodeling should be considered. Several studies have shown that the increase in blood pressure is often associated with an increase in the thickness of the vascular wall.^{24 25} If this is the case, then the vascular wall thickness is predicted to be greater in the pregnant rats treated with L-NAME than in the untreated pregnant rats. Because the wall thickness was part of the denominator in the "active stress" calculation (N/m2), then the observed increase in active stress in the pregnant rats treated with L-NAME is probably underestimated.

It has been hypothesized that an increase in the production of NO during pregnancy leads to a decrease in peripheral resistance and a decrease in vascular reactivity.^{7 13} If this is the case, one would expect that blocking the formation of NO during pregnancy would bring the vascular reactivity back to the level observed in virgin rats. However, our data show that the vascular reactivity to phenylephrine in pregnant rats treated with L-NAME is greater than that in virgin rats. These results suggest that treatment of pregnant rats with L-NAME not only blocks the synthesis of NO by endothelial cells but may also cause an increase in the synthesis of or sensitivity to other vasoactive compounds that would increase vascular reactivity. It has been suggested that the reduction in the placental blood flow during pregnancy may be associated with placental release of cytotoxic factors that alter the endothelial cell function, leading to reduction in the synthesis of vasodilators such as NO or prostacyclin or, more importantly, increased production of vasoconstrictor factors such as endothelin.^{19 20 21 22} This is consistent with a recent study showing that long-term inhibition of NO synthesis during mid to late gestation in rats is associated with increased blood pressure and elevated plasma levels of endothelin-1.²⁶

The observed increase in vascular reactivity to phenylephrine in pregnant rats treated with L-NAME could be explained by an increase in the sensitivity to phenylephrine at the -adrenergic receptor level. However, our results showed no significant difference between the four groups of rats in the phenylephrine concentration-response curve when the contraction was presented as a percentage of the maximum. These results suggest that the change in the -adrenergic receptor sensitivity to phenylephrine may not be responsible for the observed increase in vascular reactivity in pregnant rats treated with L-NAME and that the increased vascular reactivity could be due to activation of a signaling mechanism downstream from receptor activation.

It is generally accepted that -adrenergic agonists, such as phenylephrine, react with -adrenergic receptors, causing activation of phospholipase C and increased hydrolysis of phosphatidylinositol 4,5-bisphosphate into inositol 1,4,5-trisphosphate (IP₃) and diacylglycerol.²⁷ IP₃ stimulates Ca²⁺ release from intracellular stores,²⁸ and diacylglycerol stimulates protein kinase C.²⁹ In addition, -adrenergic agonists cause plasma membrane Ca²⁺ channels to open.³⁰

We tested whether the amount of Ca²⁺ released from intracellular stores in response to phenylephrine is different in the four groups of rats. We found that the transient phenylephrine contraction in Ca²⁺-free solution, which is often used as a measure of phenylephrine-releasable intracellular Ca²⁺ stores, is not different in the four groups of rats. This suggests that the enhanced vascular reactivity observed in pregnant rats treated with L-NAME is not due to changes in Ca²⁺ uptake to or Ca²⁺ release from intracellular stores.

To test the possible role of plasma membrane Ca²⁺ channels in the observed enhanced vascular reactivity to phenylephrine in pregnant rats treated with L-NAME, we tested the effect of KCl, which exclusively stimulates Ca²⁺ entry through voltage-gated Ca²⁺ channels. We found that the KCl-induced contraction in the pregnant rats was significantly reduced when compared with that in the virgin rats. These results suggest that normal pregnancy is associated with decreased Ca²⁺ entry through voltage-gated Ca²⁺ channels. On the other hand, in the pregnant rats treated with L-NAME, we found that the pregnancy-induced decrease in vascular reactivity to KCl was corrected but only to the level observed in the virgin rats. These results can possibly be explained in part by an increase in the permeability of voltage-gated Ca²⁺ channels in blood vessels of pregnant rats treated with L-NAME. However, because direct measurements of Ca²⁺ or myosin light chain phosphorylation were not performed in the present study, we cannot make a definite conclusion that L-NAME treatment increases Ca²⁺ entry into smooth muscle cells, and other possible mechanisms cannot be excluded. These mechanisms may involve an increase in the myofilament force sensitivity to Ca²⁺ entry or possibly activation of a completely Ca²⁺-independent mechanism.

The rebound increase in vascular reactivity to phenylephrine in the pregnant rats treated with L-NAME above that in the virgin rats can then be explained by the following: (1) Phenylephrine may activate an additional group of Ca²⁺ channels that have a different sensitivity to activation by phenylephrine when compared with the voltage-gated Ca²⁺ channels. These Ca²⁺ channels have been found in several smooth muscle preparations and have been designated "receptor-operated Ca²⁺ channels."^{31 32} (2) Phenylephrine may further activate different pathways that increase the myofilament force sensitivity to Ca²⁺ or possibly activate a completely Ca²⁺-independent pathway. For example, phenylephrine may activate protein kinase C through increased formation of diacylglycerol.³³

In conclusion, the increased blood pressure in late-pregnancy rats treated with L-NAME is associated with an increased vascular reactivity to sympathetic amines. This increased vascular reactivity can possibly be explained in part by an enhancement of Ca²⁺ entry through Ca²⁺ channels. However, other mechanisms such as an increase in the myofilament force sensitivity to Ca²⁺ entry or activation of a completely Ca²⁺-independent pathway may also be involved. Further studies are needed to investigate these possible mechanisms.

	Acknowledgments

 
This work was supported by grants HL-51971 and HL-33849 from the National Institutes of Health to (Dr Granger) and grants from the University of Mississippi Medical Center, the American Health Assistance Foundation, the American Heart Association (Grant-in-Aid, Mississippi Affiliate), and the National Institutes of Health (HL-52696) (Dr Khalil).

	Footnotes

 
Reprint requests to Raouf A. Khalil, MD, PhD, Department of Physiology and Biophysics, University of Mississippi Medical Center, 2500 N State St, Jackson, MS 39216.

Received October 28, 1997; first decision November 21, 1997; accepted December 18, 1997.

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Clark SL, Cotton DB, Lee W, Bishop C, Hill T, Southwick J, Spillman T, DeVore GR, Phelan J. Central hemodynamic assessment of normal term pregnancy. Am J Obstet Gynecol. 1989;161:1439–1442.[Medline] [Order article via Infotrieve]

2. Bruce NW. The distribution of blood flow to the reproductive organs of rats near term. J Reprod Fertil. 1976;46:359–362.[Abstract/Free Full Text]

3. Dowell RT, Kauer CD. Uteroplacental blood flow at rest and during exercise in late gestation conscious rats. J Appl Physiol. 1993;74:2079–2085.[Abstract/Free Full Text]

4. Magness RR, Zheng J. Maternal cardiovascular alterations during pregnancy. In: Gluckman PD, Heymann MA, eds. Pediatric and Perinatal Perspectives: The Scientific Basis. London, UK: Arnold; 1996:762–772.

5. Davison JM, Dunlop W. Renal hemodynamics and tubular function in normal human pregnancy. Kidney Int. 1980;18:152–161.[Medline] [Order article via Infotrieve]

6. Duvekott JJ, Peeters LL. Renal hemodynamics and volume homeostasis in pregnancy. Obstet Gynecol Surv. 1994;49:830–839.[Medline] [Order article via Infotrieve]

7. St-Louis J, Sicotte B. Prostaglandin- or endothelium-mediated vasodilation is not involved in the blunted responses of blood vessels to vasoconstrictors in pregnant rats. Am J Obstet Gynecol. 1992;166:684–692.[Medline] [Order article via Infotrieve]

8. Molnar M, Hertelendy F. N-Nitro-L-arginine, an inhibitor of nitric oxide synthesis, increases blood pressure in rats and reverses the pregnancy induced refractoriness to vasopressor agents. Am J Obstet Gynecol. 1992;166:1560–1567.[Medline] [Order article via Infotrieve]

9. Nathan L, Cuevas J, Chaudhuri G. The role of nitric oxide in the altered vascular reactivity of pregnancy in the rat. Br J Pharmacol. 1995;114:955–960.[Medline] [Order article via Infotrieve]

10. Williams DJ, Vallance PJT, Neild GH, Spencer JAD, Imms FJ. Nitric oxide-mediated vasodilation in human pregnancy. Am J Physiol. 1997;272:H748–H752.[Abstract/Free Full Text]

11. Parent A, Schiffrin EL, St-Louis J. Role of the endothelium in adrenergic responses of mesenteric artery rings of pregnant rats. Am J Obstet Gynecol. 1990;163:229–234.[Medline] [Order article via Infotrieve]

12. Davidge ST, McLaughlin MK. Endogenous modulation of the blunted adrenergic responses in resistance-sized mesenteric arteries from the pregnant rat. Am J Obstet Gynecol. 1992;167:1691–1697.[Medline] [Order article via Infotrieve]

13. Sladek SM, Magness RR, Conrad KP. Nitric oxide and pregnancy. Am J Physiol. 1997;272:R441–R463.[Abstract/Free Full Text]

14. Conrad KP, Vernier KA. Plasma levels, urinary excretion and metabolic production of cGMP during gestation in rats. Am J Physiol. 1989;257:R847–R853.[Abstract/Free Full Text]

15. National High Blood Pressure Education Program Working Group. Report on high blood pressure in pregnancy: consensus report. Am J Obstet Gynecol. 1990;163:1689–1712.

16. Cabrera C, Bohr D. The role of nitric oxide in the central control of blood pressure. Biochem Biophys Res Commun. 1995;206:77–81.[Medline] [Order article via Infotrieve]

17. Friedman SA, Lubarsky SL, Ahokas RA, Nova A, Sibai BM. Pre-eclampsia and related disorders: clinical aspects and relevance of endothelin and nitric oxide. Clinic Perinatol. 1995;2:343–355.

18. Morris NH, Eaton BM, Dekker G. Nitric oxide, the endothelium, pregnancy and pre-eclampsia. Br J Obstet Gynecol. 1996;103:4–15.[Medline] [Order article via Infotrieve]

19. Baylis C, Engels K. Adverse interactions between pregnancy and a new model of systemic hypertension produced by chronic blockade of endothelial derived relaxing factor (EDRF) in the rat. Clin Exp Hypertens. 1992;B11:117–129.

20. Baylis C, Suto T, Conrad K. Importance of nitric oxide in control of systemic and renal hemodynamics during normal pregnancy: studies in the rat and implications for preeclampsia. Hypertens Pregnancy. 1996;15:147–169.

21. Molnar M, Suto T, Toth T, Hertelendy F. Prolonged blockade of nitric oxide synthesis in gravid rats produces sustained hypertension, proteinuria, thrombocytopenia, and intrauterine growth retardation. Am J Obstet Gynecol. 1994;170:1458–1466.[Medline] [Order article via Infotrieve]

22. Novak J, Cockrell K, Kassab S, Reckelhoff JF, Granger JP. Chronic nitric oxide synthesis inhibition during pregnancy increases thromboxane excretion in rats. FASEB J. 1997;11:A78.

23. Ahokas RA, Mercer BM, Sibai BM. Enhanced endothelium-derived relaxing factor activity in pregnant, spontaneously hypertensive rats. Am J Obstet Gynecol. 1991;165:801–807.[Medline] [Order article via Infotrieve]

24. Corson MA, Berk BC. Growth factors and the vessel wall. Heart Dis Stroke. 1993;2:166–170.[Medline] [Order article via Infotrieve]

25. Owens GK, Vernon SM, Madsen CS. Molecular regulation of smooth muscle cell differentiation. J Hypertens. 1996;14:S55–S64.

26. Edwards DL, Arora CP, Bui DT, Castro LC. Long-term nitric oxide blockade in the pregnant rat: effects on blood pressure and plasma levels of endothelin-1. Am J Obstet Gynecol. 1996;175:484–488.[Medline] [Order article via Infotrieve]

27. Berridge MJ, Irvine R. Inositol trisphosphate, a novel second messenger in cellular signal transduction. Nature. 1984;312:315–321.[Medline] [Order article via Infotrieve]

28. Suematsu E, Hirata M, Hashimoto T, Kuriyama H. Inositol 1,4,5-trisphosphate releases Ca²⁺ from intracellular storage sites in skinned single cells of porcine coronary artery. Biochem Biophys Res Commun. 1984;120:481–485.[Medline] [Order article via Infotrieve]

29. Nishizuka Y. Intracellular signaling by hydrolysis of phospholipids and activation of protein kinase C. Science. 1992;258:607–614.[Abstract/Free Full Text]

30. Khalil RA, van Breemen C. Mechanisms of calcium mobilization and homeostasis in vascular smooth muscle and their relevance to hypertension. In: Laragh JH, Brenner BM, eds. Hypertension: Pathophysiology, Diagnosis, and Management. 2nd ed. New York, NY: Raven Press; 1995:523–540.

31. Bolton TB. Mechanisms of action of transmitters and other substances on smooth muscle. Physiol Rev. 1979;59:606–718.[Free Full Text]

32. van Breemen C, Aaronson P, Loutzenhiser R. Na-Ca interactions in mammalian smooth muscle. Pharmacol Rev. 1979;30:167–208.[Medline] [Order article via Infotrieve]

33. Khalil RA, Morgan KG. Phenylephrine-induced translocation of protein kinase C and shortening of two types of vascular cells of the ferret. J Physiol (Lond). 1992;455:585–599.[Abstract/Free Full Text]

This article has been cited by other articles:

				A. K. Stennett, X. Qiao, A. E. Falone, V. V. Koledova, and R. A. Khalil Increased vascular angiotensin type 2 receptor expression and NOS-mediated mechanisms of vascular relaxation in pregnant rats Am J Physiol Heart Circ Physiol, March 1, 2009; 296(3): H745 - H755. [Abstract] [Full Text] [PDF]

				W. Chen and R. A. Khalil Differential [Ca2+]i signaling of vasoconstriction in mesenteric microvessels of normal and reduced uterine perfusion pregnant rats Am J Physiol Regulatory Integrative Comp Physiol, December 1, 2008; 295(6): R1962 - R1972. [Abstract] [Full Text] [PDF]

				Y. Lu, H. Zhang, N. Gokina, M. Mandala, O. Sato, M. Ikebe, G. Osol, and S. A. Fisher Uterine artery myosin phosphatase isoform switching and increased sensitivity to SNP in a rat L-NAME model of hypertension of pregnancy Am J Physiol Cell Physiol, February 1, 2008; 294(2): C564 - C571. [Abstract] [Full Text] [PDF]

				H. Huang, H.-H. Chang, Y. Xu, D. S. Reddy, J. Du, Y. Zhou, Z. Dong, J. R. Falck, and M.-H. Wang Epoxyeicosatrienoic Acid Inhibition Alters Renal Hemodynamics During Pregnancy Experimental Biology and Medicine, December 1, 2006; 231(11): 1744 - 1752. [Abstract] [Full Text] [PDF]

				H. Huang, Y. Zhou, V. T. Raju, J. Du, H.-H. Chang, C.-Y. Wang, M. W. Brands, J. R. Falck, and M.-H. Wang Renal 20-HETE inhibition attenuates changes in renal hemodynamics induced by L-NAME treatment in pregnant rats Am J Physiol Renal Physiol, November 1, 2005; 289(5): F1116 - F1122. [Abstract] [Full Text] [PDF]

				Y. Zhou, H.-H. Chang, J. Du, C.-Y. Wang, Z. Dong, and M.-H. Wang Renal epoxyeicosatrienoic acid synthesis during pregnancy Am J Physiol Renal Physiol, January 1, 2005; 288(1): F221 - F226. [Abstract] [Full Text] [PDF]

				J. P. Granger Inflammatory cytokines, vascular function, and hypertension Am J Physiol Regulatory Integrative Comp Physiol, June 1, 2004; 286(6): R989 - R990. [Full Text] [PDF]

				J. M. Orshal and R. A. Khalil Interleukin-6 impairs endothelium-dependent NO-cGMP-mediated relaxation and enhances contraction in systemic vessels of pregnant rats Am J Physiol Regulatory Integrative Comp Physiol, June 1, 2004; 286(6): R1013 - R1023. [Abstract] [Full Text] [PDF]

				J. M. Orshal and R. A. Khalil Reduced Endothelial NO-cGMP-Mediated Vascular Relaxation and Hypertension in IL-6-Infused Pregnant Rats Hypertension, February 1, 2004; 43(2): 434 - 444. [Abstract] [Full Text] [PDF]

				M.-H. Wang, J. Wang, H.-H. Chang, B. A. Zand, M. Jiang, A. Nasjletti, and M. Laniado-Schwartzman Regulation of renal CYP4A expression and 20-HETE synthesis by nitric oxide in pregnant rats Am J Physiol Renal Physiol, August 1, 2003; 285(2): F295 - F302. [Abstract] [Full Text] [PDF]

				J. G. Murphy, J. N. Herrington, J. P. Granger, and R. A. Khalil Enhanced [Ca2+]i in renal arterial smooth muscle cells of pregnant rats with reduced uterine perfusion pressure Am J Physiol Heart Circ Physiol, January 1, 2003; 284(1): H393 - H403. [Abstract] [Full Text] [PDF]

				R. A. Khalil and J. P. Granger Vascular mechanisms of increased arterial pressure in preeclampsia: lessons from animal models Am J Physiol Regulatory Integrative Comp Physiol, July 1, 2002; 283(1): R29 - R45. [Abstract] [Full Text] [PDF]

				J. B. Giardina, G. M. Green, K. L. Cockrell, J. P. Granger, and R. A. Khalil TNF-alpha enhances contraction and inhibits endothelial NO-cGMP relaxation in systemic vessels of pregnant rats Am J Physiol Regulatory Integrative Comp Physiol, July 1, 2002; 283(1): R130 - R143. [Abstract] [Full Text] [PDF]

				M.-H. Wang, B. A. Zand, A. Nasjletti, and M. Laniado-Schwartzman Renal 20-hydroxyeicosatetraenoic acid synthesis during pregnancy Am J Physiol Regulatory Integrative Comp Physiol, February 1, 2002; 282(2): R383 - R389. [Abstract] [Full Text] [PDF]

				J. R. Davis, J. B. Giardina, G. M. Green, B. T. Alexander, J. P. Granger, and R. A. Khalil Reduced endothelial NO-cGMP vascular relaxation pathway during TNF-alpha -induced hypertension in pregnant rats Am J Physiol Regulatory Integrative Comp Physiol, February 1, 2002; 282(2): R390 - R399. [Abstract] [Full Text] [PDF]

				J. B. Giardina, K. L. Cockrell, J. P. Granger, and R. A. Khalil Low-Salt Diet Enhances Vascular Reactivity and Ca2+ Entry in Pregnant Rats With Normal and Reduced Uterine Perfusion Pressure Hypertension, February 1, 2002; 39(2): 368 - 374. [Abstract] [Full Text] [PDF]

				L. A. Barron, J. B. Giardina, J. P. Granger, and R. A. Khalil High-Salt Diet Enhances Vascular Reactivity in Pregnant Rats With Normal and Reduced Uterine Perfusion Pressure Hypertension, September 1, 2001; 38(3): 730 - 735. [Abstract] [Full Text] [PDF]

				J. G. Murphy, J. B. Fleming, K. L. Cockrell, J. P. Granger, and R. A. Khalil [Ca2+]i signaling in renal arterial smooth muscle cells of pregnant rat is enhanced during inhibition of NOS Am J Physiol Regulatory Integrative Comp Physiol, January 1, 2001; 280(1): R87 - R99. [Abstract] [Full Text] [PDF]

				O. D. Sherwood, L. M. Olson, S. Zhao, and H. R. Little Inhibition of Nitric Oxide Synthase Activity Diminishes the Acute Effects of Relaxin on Growth, But Not Softening, of the Cervix in the Rat Endocrinology, July 1, 2000; 141(7): 2458 - 2464. [Abstract] [Full Text] [PDF]

				C. A. Kanashiro, K. L. Cockrell, B. T. Alexander, J. P. Granger, and R. A. Khalil Pregnancy-associated reduction in vascular protein kinase C activity rebounds during inhibition of NO synthesis Am J Physiol Regulatory Integrative Comp Physiol, February 1, 2000; 278(2): R295 - R303. [Abstract] [Full Text] [PDF]

				J. K. Crews, J. N. Herrington, J. P. Granger, and R. A. Khalil Decreased Endothelium-Dependent Vascular Relaxation During Reduction of Uterine Perfusion Pressure in Pregnant Rat Hypertension, January 1, 2000; 35(1): 367 - 372. [Abstract] [Full Text] [PDF]

				C. A. Kanashiro, B. T. Alexander, J. P. Granger, and R. A. Khalil Ca2+-Insensitive Vascular Protein Kinase C During Pregnancy and NOS Inhibition Hypertension, October 1, 1999; 34(4): 924 - 930. [Abstract] [Full Text] [PDF]

				J. K. Crews, J. Novak, J. P. Granger, and R. A. Khalil Stimulated mechanisms of Ca2+ entry into vascular smooth muscle during NO synthesis inhibition in pregnant rats Am J Physiol Regulatory Integrative Comp Physiol, February 1, 1999; 276(2): R530 - R538. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Khalil, R. A. Articles by Granger, J. P. Search for Related Content PubMed PubMed Citation Articles by Khalil, R. A. Articles by Granger, J. P.