AJP - Heart AJP: Renal Physiology
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Am J Physiol Heart Circ Physiol 291: H413-H420, 2006. First published February 24, 2006; doi:10.1152/ajpheart.00103.2006
0363-6135/06 $8.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
291/1/H413    most recent
00103.2006v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Xiao, D.
Right arrow Articles by Zhang, L.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Xiao, D.
Right arrow Articles by Zhang, L.

Regulation of baseline Ca2+ sensitivity in permeabilized uterine arteries: effect of pregnancy

Daliao Xiao, Xiaohui Huang, Lawrence D. Longo, William J. Pearce, and Lubo Zhang

Department of Physiology and Pharmacology, Center for Perinatal Biology, Loma Linda University School of Medicine, Loma Linda, California

Submitted 25 January 2006 ; accepted in final form 20 February 2006


   ABSTRACT
TOP
 ABSTRACT
METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
The adaptation of contractile mechanisms of the uterine artery to pregnancy is not fully understood. The present study examined the effect of pregnancy on the uterine artery baseline Ca2+ sensitivity. In beta-escin-permeabilized arterial preparations, Ca2+-induced concentration-dependent contractions were significantly decreased in uterine arteries from pregnant animals compared with those of nonpregnant animals. Time-course studies showed that Ca2+ increased phosphorylation of 20-kDa myosin light chain (MLC20), which preceded the tension development in vessels from both pregnant and nonpregnant animals. When compared with vessels from nonpregnant animals, there was a significant increase in the protein level of MLC20 and an accordance increase in the level of Ca2+-induced phosphorylated MLC20 (MLC20-P) in uterine arteries during pregnancy. Simultaneous measurements of MCL20-P levels and contractions stimulated with Ca2+ in the same tissues demonstrated a significant attenuation in the tension-to-MLC20-P ratio in uterine arteries during pregnancy. Activation of PKC with phorbol 12,13-dibutyrate (PDBu) potentiated Ca2+-induced contractions in uterine arteries from nonpregnant but not pregnant animals. Accordingly, inhibition of PKC attenuated Ca2+-induced contractions in uterine arteries from nonpregnant but not pregnant animals. PDBu produced contractions in the presence or absence of Ca2+ in the beta-escin-permeabilized arteries, which were significantly decreased in uterine arteries from pregnant compared with nonpregnant animals. The results suggest that pregnancy upregulates the thick-filament regulatory pathway by increasing MLC20 phosphorylation but downregulates the thin-filament regulatory pathway by decreasing the contractile sensitivity of MLC20-P, resulting in attenuated baseline Ca2+ sensitivity in the uterine artery. In addition, PKC plays an important role in the regulation of basal Ca2+ sensitivity, which is downregulated during pregnancy.

protein kinase C; thick filament; thin filament; myosin light chain; phosphorylation

DURING PREGNANCY, the uterine artery delivers oxygen and substrates to the developing fetus. Previous studies (5, 10, 35, 50, 51) have demonstrated a complex adaptation of uterine artery contractile and relaxation mechanisms to pregnancy. The mechanisms regulating vascular smooth muscle contraction include a variety of intracellular signaling pathways that integrate a multitude of stimuli into a coordinated contractile response. Ca2+ mobilization and the Ca2+ sensitivity of the contractile apparatus are the two major regulatory mechanisms for smooth muscle contraction. An increase in intracellular Ca2+ concentration ([Ca2+]i) leads to activation of Ca2+-dependent myosin light chain kinase, which in turn phosphorylates the 20-kDa regulatory myosin light chain (MLC20), and thereby triggers contraction (17, 38). Ca2+ sensitivity, defined by force production per unit changes in [Ca2+]i, is another important mechanism in the regulation of vascular contractility. Regulation of the Ca2+ sensitivity of the contractile apparatus involves inhibition of myosin light chain phosphatase (MLCP), resulting in an increase in the level of phosphorylated MLC20 (MLC20-P) (12, 15, 20, 34). Moreover, many studies have demonstrated dissociation between MLC20-P and cross-bridge cycling rates/tension development, suggesting a thin-filament regulatory pathway of Ca2+ sensitivity (4, 29, 37).

Recently, we have demonstrated that the Ca2+ sensitivity is a key mechanism in the regulation of uterine artery contractility (46, 48, 50, 51). We have shown that pregnancy increases {alpha}1-adrenoceptor-induced Ca2+ mobilization but inhibits {alpha}1-adrenoceptor-mediated Ca2+ sensitivity in the uterine artery (51). Moreover, {alpha}1-adrenoceptor-stimulated contractions were regulated through both thick- and thin-filament pathways, with the thick-filament regulatory pathway, i.e., MLC20 phosphorylation, predominating (51). In contrast, the PKC-mediated contraction was regulated predominantly through thin-filament pathways, i.e., independent of changes in either Ca2+ concentrations or MLC20-P (48, 51). More recently, we have shown that PKC plays a key role in the regulation of myogenic tone of the uterine artery, which is significantly decreased in uterine arteries during pregnancy (45). Myogenic contraction is an important physiological mechanism that regulates basal vascular tone and is a significant contributor to the modulation of blood flow. Our studies suggested that the Ca2+ sensitivity was a key component in the regulation of uterine artery myogenic tone (45). The differences in the baseline myofilament Ca2+ sensitivity and/or its alteration by stimuli have been shown to determine the variations in basal vascular tone and are associated with the differences in animal species, developmental age, artery size, and physiological and pathophysiological variations (1, 2, 11, 40). However, it is unknown whether or to what extent pregnancy alters the basal Ca2+ sensitivity in the uterine artery.

In the present study, we determined the effect of pregnancy on the baseline Ca2+ sensitivity in uterine artery smooth muscle and tested the hypothesis that pregnancy inhibited the basal Ca2+ sensitivity. By clamping Ca2+ concentrations, the Ca2+ sensitivity of myofilaments can be determined directly in permeabilized arteries (6, 23, 43). To investigate the potential role of thick- and/or thin-filament pathways in the regulation of the baseline Ca2+ sensitivity in the uterine artery and its alteration by pregnancy, we measured Ca2+-induced MLC20 phosphorylation and tension simultaneously in the same tissues and determined the phosphorylated MLC20-to-Ca2+ and tension-to-phosphorylated MLC20 ratios. In addition, we also investigated the role of PKC in the regulation of the basal Ca2+ sensitivity in the uterine artery and its adaptation to pregnancy.


   METHODS
TOP
 ABSTRACT
 METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Tissue preparation. Uterine arteries were isolated from nonpregnant and pregnant (~140 days gestation) sheep, as described previously (51). Briefly, animals were anesthetized with thiamylal (10 mg/kg), administered via the external left jugular vein. The ewes were then intubated, and anesthesia was maintained on 1.5% to 2.0% halothane in O2 throughout the surgery. An incision in the abdomen was made and the uterus exposed. The third (from nonpregnant sheep) and fourth (from pregnant sheep) branches of the main uterine arteries with a similar external diameter were separated from the surrounding tissue and cut into 2-mm ring segments in Krebs solution (pH 7.4) of the following composition (in mM): 115.21 NaCl, 4.7 KCl, 1.80 CaCl2, 1.16 MgSO4, 1.18 KH2PO4, 22.14 NaHCO3, 0.03 EDTA, and 7.88 dextrose, oxygenated with a mixture of 95% O2-5% CO2. After the tissues were removed, animals were euthanized with solution (T-61, Hoechst-Roussel; Somervile, NJ). All procedures and protocols used in the present study were approved by the Animal Research Committee of Loma Linda University and followed the guidelines in the National Institutes of Health's Guide for the Care and Use of Laboratory Animals.

Ca2+ buffer solutions. Two main buffer solutions were used in the present study, as previously described (1, 2, 30). The one was zero Ca2+ relaxing solution that contained (in mM) 110 K-acetate, 5 EGTA, 5 ATP, 6 Mg-acetate, 1 DTT, 0.01 leupeptin, 20 imidazole, and 20 HEPES, at pH 6.8 (titrated with KOH). The other contained 100 µM Ca2+ in addition to the same components in the zero Ca2+ buffer. As described in the previous study (1), Ca2+ buffer solutions were prepared by solving the multiequilibrium equations for interactions among the different ions, and solutions containing intermediate free Ca2+ concentrations were prepared by mixing appropriate amounts of zero Ca2+ relaxing solution and the maximum Ca2+ (100 µM) buffer solutions, titrated to pH 7.0 with 1 M KOH. As demonstrated in our previous studies, the accuracy of calculated free Ca2+ concentrations in Ca2+ buffer solutions was determined directly with dual-wavelength measurements of bound-to-unbound fura-2 ratios, and the calculated concentrations of free Ca2+ correlated well (r2 = 0.97) with the concentrations measured with fura-2 (1).

Permeabilization procedure. The arterial rings were attached to isometric force transducers and bathed in Krebs solution at 37°C. Isometric tensions were measured as described previously (46, 51). After 60 min of equilibration in the tissue bath, each ring was stretched to the optimal resting tension as determined by the tension developed in response to 120 mM KCl added at each stretch level. The arterial rings were then placed into the zero Ca2+ relaxing solution. The permeabilization procedure was modified from previous studies (1, 6, 30). Briefly, chemical permeabilization was achieved by adding 40 µM beta-escin to the relaxing solution and by allowing the rings to incubate for 20 min at 25°C. The permeabilized solution was then replaced with the relaxing solution containing 1 µM A-23187 to deplete Ca2+ from the sarcoplasmic reticulum. Concentration-response curves to Ca2+ were obtained by cumulative increases of Ca2+ concentrations in approximate one-half log increments in beta-escin-permeabilized arterial rings. Prism software (GraphPat; San Diego, CA) was used to fit the curve and determine the pD2 values (–log EC50) and the maximum response. In certain experiments, tissues were pretreated with the PKC activator or inhibitor (or vehicle DMSO for 20 min) and then stimulated with increasing concentrations of Ca2+.

Measurements of MLC20 phosphorylation. MLC20 phosphorylation levels and contractile tensions were determined simultaneously in the same tissues, as previously described (51). Tensions developments were continuously recorded with an online computer, and arterial rings were snap frozen with liquid N2-cooled clamps at the indicated times and rapidly immersed in a dry ice-acetone slurry containing 10% trichloroacetic acid (TCA), 5 mM NaF, and 10 mM DTT mixture. Tissues were then stored at –80°C until the analysis of MLC20-P. To measure MLC20-P, tissues were brought to room temperature in a dry ice-acetone-TCA-DTT mixture and then washed three times with ether to remove the TCA. Tissues were then extracted in 100 µl of sample buffer containing 20 mM Tris base and 23 mM glycine (pH 8.6), 8.0 M urea, 10 mM DTT, 10% glycerol, and 0.04% bromophenol blue, as previously described. Samples (20 µl) were electrophoresed at 12 mA for 2.5 h after a 30 min prerun in 1.0 mm minipolyacrylamide gels containing 10% arcelamide-0.27% bisacrylamide, 40% glycerol, 8.0 M urea, and 20 mM Tris base (pH 8.8). Proteins were transferred to nitrocellulose membranes and subjected to Western blot analysis with a specific MLC20 antibody (1:500). Goat anti-mouse IgG conjugated with horseradish peroxidase was used as a secondary antibody (1:2,000). Bands were detected with enhanced chemiluminescence (ECL), visualized on Hyperfilm, and analyzed with the Kodak 1D image analysis software. Moles of inorganic phosphate per mole of MLC20 (fractional light chain phosphorylation) were calculated by dividing the density of the phosphorylated band by the sum of the densities of the phosphorylated plus the unphosphorylated bands. In some experiments, the apparent levels of MLC20-P were calculated by multiplying the fractional MLC20 phosphorylation (in mol Pi/mol MLC20) by the total levels of MLC20 determined by Western blot analysis as described below.

Western blot analysis. Uterine arteries from both nonpregnant and pregnant animals were isolated as described above and homogenized in the lysis buffer containing 150 mM NaCl, 50 mM Tris·HCl, 10 mM EDTA, 0.1% Tween 20, 0.1% beta-mercaptoethanol, 0.1 mM phenylmethylsulfonyl fluoride, 5 µg/ml leupeptin, and 5 µg/ml aprotinin, pH 7.4. Homogenates were then centrifuged at 4°C for 5 min at 6,000 g, and the supernatants were collected. Protein was determined in the supernatant. Samples with equal protein were loaded on a 10% polyacrylamide gel with 0.1% SDS and were separated by electrophoresis at 100 V for 2 h. Proteins were then transferred to a nitrocellulose membrane. Nonspecific binding sites in the membranes were blocked by an overnight incubation at 4°C in a Tris-buffered saline solution containing 5% dry milk. The membranes were incubated with a specific MLC20 antibody, followed by a secondary antibody. Proteins were visualized with ECL reagents, and the blots were exposed to Hyperfilm. Results were quantified with the Kodak electrophoresis documentation and analysis system and Kodak 1D image analysis software.

Materials. Phorbol 12,13-dibutyrate (PDBu), staurosporine, monoclonal antimyosin (20-kDa light chains) antibody, beta-escin, and other chemicals were obtained from Sigma (St. Louis, MO). Goat anti-mouse IgG conjugated with horseradish peroxidase was from Calbiochem (La Jolla, CA). Electrophoretic and immunoblotting reagents were from Bio-Rad (Hercules, CA).

Data analysis. Data were expressed as means ± SE. Differences were evaluated for statistical significance (P < 0.05) by one-way ANOVA and Student's t-test.


   RESULTS
TOP
 ABSTRACT
 METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Effect of pregnancy on Ca2+-force relationship in permeabilized uterine arteries. To test the permeabilization efficiency of beta-escin in the uterine arteries of each experiment, the maximum contractile tension induced by 120 mM KCl before permeabilization was compared with the tension induced by the buffer solution containing 10 µM free [Ca2+] after permeabilization in the same tissue. As shown in Fig. 1, the tension induced by 10 µM Ca2+ after permeabilization was not significantly different from that induced by 120 mM KCl before permeabilization. Furthermore, 120 mM KCl produced no contractile tension in the uterine artery after permeabilization (data not shown). The results suggested complete permeabilization of the tissues by beta-escin.


Figure 1
View larger version (9K):
[in this window]
[in a new window]
  		Fig. 1. KCl- (top, left) and Ca2+-mediated (top, right) contractions before and after permeabilization of uterine arteries. Uterine arteries were contracted with 120 mM KCl and 10 µM Ca2+ before and after permeabilization with beta-escin as described in METHODS (bottom). Data are means ± SE of tissues from 14 pregnant and 8 nonpregnant animals.

 
Figure 2 shows that cumulative increases of Ca2+ produced concentration-dependent contractions of permeabilized uterine arteries from both nonpregnant and pregnant animals. The pD2 values were not significantly different between uterine arteries (P > 0.05) from nonpregnant (5.62 ± 0.14) and pregnant (5.60 ± 0.16) sheep. However, the maximum response of Ca2+-induced contractions [expressed as %120 mM KCl-induced contractions before permeabilization in the same tissue (%KCl)] was significantly decreased in uterine arteries from pregnant compared with nonpregnant sheep (87.9 ± 7.0 vs. 139.7 ± 9.4 %KCl; P < 0.05).


Figure 2
View larger version (8K):
[in this window]
[in a new window]
  		Fig. 2. Effect of pregnancy on basal Ca2+ sensitivity in beta-escin-permeabilized uterine arteries. Concentration-response curves of Ca2+-induced contractions were measured in beta-escin-permeabilized uterine arteries from nonpregnant and pregnant animals. Data are expressed as percentage of maximal KCl-induced contractions of the same tissue before permeabilization to normalize tissue size. Data are means ± SE of tissues from pregnant (n = 11) and nonpregnant (n = 7) animals. pD2 values and maximal response are presented in RESULTS.

 
Effect of pregnancy on Ca2+-induced MLC20 phosphorylation. Figure 3 shows the time courses of Ca2+ (10 µM)-induced increases in MLC20 phosphorylation and contractions in permeabilized uterine arteries. In the uterine arteries from both pregnant and nonpregnant sheep, Ca2+ produced time-dependent increases in MLC20 phosphorylation, which preceded contractions. Total protein levels of MLC20 were determined by Western blot analysis. Figure 4 shows that the MLC20 levels were significantly increased in uterine arteries from pregnant compared with nonpregnant sheep (19.23 ± 1.45 vs. 5.77 ± 1.35, arbitrary density units; P < 0.05). As shown in Fig. 5, Ca2+ produced dose-dependent increases in the levels of MLC20-P (Fig. 5, top) in the uterine arteries. The apparent levels of MLC20-P induced by Ca2+ were significantly higher in uterine arteries from pregnant versus nonpregnant sheep at the doses of 10 µM Ca2+ (11.2 ± 0.56 vs. 2.28 ± 0.25 mol Pi; P < 0.05) and 30 µM Ca2+ (9.32 ± 1.33 vs. 2.57 ± 0.18 mol Pi; P < 0.05), respectively. However, the ratios of tension to MLC20-P (g/MLC20-P), obtained from the simultaneous measurements of MLC20-P and tension in the same tissues, were significantly decreased in uterine arteries from pregnant compared with nonpregnant sheep at the doses of 10 µM Ca2+ (0.55 ± 0.11 vs. 1.63 ± 0.38; P < 0.05) and 30 µM Ca2+ (0.74 ± 0.10 vs. 1.89 ± 0.44; P < 0.05), respectively (Fig. 5, bottom).


Figure 3
View larger version (17K):
[in this window]
[in a new window]
  		Fig. 3. Ca2+-induced time courses of 20-kDa myosin light chain (MLC20) phosphorylation and contractions in beta-escin-permeabilized uterine arteries. beta-escin-permeabilized uterine arteries from nonpregnant (top) and pregnant (bottom) animals were stimulated with 10 µM Ca2+. Contractile tension ({circ}) and phosphorylated MLC20 (MLC20-P; bullet) were measured simultaneously in the same tissues at indicated time points. Data are means ± SE of tissues from pregnant (n = 5) and nonpregnant (n = 5) animals.

 

Figure 4
View larger version (20K):
[in this window]
[in a new window]
  		Fig. 4. Effect of pregnancy on total MLC20 protein levels in uterine arteries. MLC20 protein levels in uterine arteries from pregnant and nonpregnant animals were determined with Western blot analysis as described in METHODS. Data are means ± SE of tissues from pregnant (n = 4) and nonpregnant (n = 4) animals. *P < 0.05 vs. nonpregnant animals.

 

Figure 5
View larger version (8K):
[in this window]
[in a new window]
  		Fig. 5. Effect of pregnancy on Ca2+-induced MLC20 phosphorylation in beta-escin-permeabilized uterine arteries. beta-escin-permeabilized uterine arteries were stimulated with increasing concentrations of Ca2+. MLC20 phosphorylation and contractions were measured simultaneously in the same tissues. Top: apparent tissue levels of MLC20-P (in arbitrary units) were calculated by multiplying the fractional MLC20 phosphorylation (mol Pi/mol MLC20) by the total levels of MLC20 determined in Fig. 4. Bottom: tension-to-MLC20-P ratios. Data are means ± SE of tissues from pregnant (n = 9) and nonpregnant (n = 6) animals. *P < 0.05 vs. nonpregnant animals.

 
Effect of PKC activation on Ca2+ sensitivity. Figure 6 shows the effect of the PKC activator PDBu on tension development in permeabilized uterine arteries in the absence and presence of submaximal Ca2+ concentrations (1 µM). In the presence of Ca2+, the maximal concentration of PDBu (10 µM) induced tension development that was significantly greater in uterine arteries from nonpregnant sheep than during pregnancy (30.8 ± 2.8 vs. 18.1 ± 3.2 %KCl; P < 0.05) (Fig. 6). In the absence of Ca2+ in the Ca2+ free relaxing solution, PDBu-induced contractions were significantly decreased, but the difference between uterine arteries from nonpregnant and pregnant sheep remained (14.5 ± 1.9 vs. 7.3 ± 2.0 %KCl; P < 0.05) (Fig. 6). To determine the effect of PKC activation on Ca2+-force relations, we compared [Ca2+]-dependent contractions in the absence and presence of PDBu in permeabilized uterine arteries. As shown in Fig. 7, pretreatment with the subthreshold concentration of PDBu (0.3 µM) did not change the pD2 (5.68 ± 0.12 vs. 5.56 ± 0.06, P > 0.05) but significantly potentiated the maximum response of Ca2+-induced contractions (168.9 ± 8.4 vs. 129.2 ± 3.9 %KCl; P < 0.05) in uterine arteries from nonpregnant sheep (Fig. 7, top). In contrast, neither the pD2 (5.79 ± 0.12 vs. 5.70 ± 0.16, P > 0.05) nor the maximum response (106.9 ± 5.6 vs. 89.1 ± 6.7 %KCl, P > 0.05) of Ca2+-induced contractions was significantly changed in the presence of PDBu in uterine arteries during pregnancy (Fig. 7, bottom).


Figure 6
View larger version (8K):
[in this window]
[in a new window]
  		Fig. 6. Effect of pregnancy on phorbol 12,13-dibutyrate (PDBu)-induced contractions in beta-escin-permeabilized uterine arteries. beta-escin-permeabilized uterine arteries were contracted with 10 µM PDBu in the presence of submaximal (1 µM) or zero concentrations of Ca2+. Data are expressed as percentage of maximal KCl-induced contractions of the same tissue before permeabilization to normalize tissue size. Data are means ± SE of tissues from pregnant (n = 8) and nonpregnant (n = 5) animals. *P < 0.05 vs. nonpregnant animals.

 

Figure 7
View larger version (10K):
[in this window]
[in a new window]
  		Fig. 7. Effect of PKC activation on Ca2+-force relation in beta-escin-permeabilized uterine arteries. Concentration-response curves of Ca2+-induced contractions were measured in beta-escin-permeabilized uterine arteries in the absence ({circ}) or presence (bullet) of PDBu (0.3 µM, pretreatment for 20 min). Data are expressed as percentage of maximal KCl-induced contractions of the same tissue before permeabilization to normalize tissue size. Data are means ± SE of tissues from nonpregnant (top; n = 4) and pregnant (bottom; n = 7) animals. pD2 values and maximal response are presented in RESULTS.

 
Accordingly, in the presence of the PKC inhibitor staurosporine, both the pD2 value (5.10 ± 0.10 vs. 5.60 ± 0.07; P < 0.05) and the maximum response (66.7 ± 3.9 vs. 145.8 ± 5.3 %KCl; P < 0.05) of Ca2+-induced concentration-dependent contractions were significantly decreased in uterine arteries from nonpregnant sheep (Fig. 8, top). However, staurosporine did not significantly affect the pD2 value (5.30 ± 0.11 vs. 5.62 ± 0.20; P > 0.05) and the maximum response (85.0 ± 5.1 vs. 116.6 ± 12.3 %KCl; P > 0.05) in uterine arteries during pregnancy (Fig. 8, bottom).


Figure 8
View larger version (10K):
[in this window]
[in a new window]
  		Fig. 8. Effect of PKC inhibition on Ca2+-force relation in beta-escin-permeabilized uterine arteries. Concentration-response curves of Ca2+-induced contractions were measured in beta-escin-permeabilized uterine arteries in the absence ({circ}) or presence (bullet) of staurosporine (0.1 µM, pretreatment for 20 min). Data are expressed as percentage of maximal KCl-induced contractions of the same tissue before permeabilization to normalize tissue size. Data are means ± SE of tissues from nonpregnant (top; n = 4) and pregnant (bottom; n = 3) animals. pD2 values and maximal response are presented in RESULTS.

 

   DISCUSSION
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
The major findings of the present study are that in permeabilized uterine arteries, 1) the baseline Ca2+ sensitivity was significantly attenuated in uterine arteries during pregnancy; 2) MLC20 phosphorylation preceded Ca2+-induced contractions in uterine arteries from both nonpregnant and pregnant sheep; 3) the Ca2+ sensitivity of MLC20 phosphorylation was significantly increased in uterine arteries during pregnancy; 4) the sensitivity of MLC20-P in contractions was significantly decreased in uterine arteries during pregnancy; and 5) PKC modulated the baseline Ca2+ sensitivity in the uterine arteries, which was significantly attenuated in pregnant animals.

It has been well demonstrated that the Ca2+ sensitivity is a key mechanism in the regulation of vascular contractility. In the present study, we examined the effect of pregnancy on the baseline Ca2+ sensitivity in the uterine artery using beta-escin-permeabilized arterial preparations, the method that has been widely used to study the myofilament Ca2+ sensitivity by many investigators in a variety of smooth muscles (24, 33, 43), including our own in cerebral and coronary arteries (1, 2, 11, 30). In agreement with previous studies (1, 2), the present study showed that, in the beta-escin-permeabilized uterine arteries, the buffer solution containing 10 µM free [Ca2+] produced the equivalent level of contractions to that of the maximum KCl contractions in the same intact vessels before permeabilization. This suggests that the beta-escin permeabilization was complete and that smooth muscle contractile apparatus remained intact. The finding that the Ca2+-force relationship was significantly depressed in uterine arteries during pregnancy suggests that pregnancy decreased the baseline Ca2+ sensitivity in the uterine artery. This is consistent with our previous findings in the intact arterial preparations that pregnancy attenuated {alpha}1-adrenoceptor-mediated myofilament Ca2+ sensitivity in the uterine artery (51). The idea that the baseline Ca2+ sensitivity is regulated both physiologically and pathophysiologically is further supported by previous findings of differences in basal Ca2+ sensitivity associated with the artery type, arterial size, developmental age, and oxygen tension (1, 2, 11, 40). Our recent studies demonstrated that the Ca2+ sensitivity played an important role in the regulation of uterine artery myogenic tone (45). The present finding of attenuated baseline Ca2+ sensitivity in uterine arteries during pregnancy suggests a key mechanism in the decreased myogenic tone in the same arteries (41, 45).

The myofilament Ca2+ sensitivity is regulated through multiple mechanisms. Both thick- and thin-filament regulatory pathways contribute to the regulation of Ca2+ sensitivity (4, 15, 29, 48). In the present study, we measured Ca2+-induced MLC20 phosphorylation and contractions simultaneously in the same tissues of permeabilized uterine arteries. By examining the relationships of Ca2+ and MLC20 phosphorylation as well as MLC20 phosphorylation and contractions, we were able to separate the thick-filament mechanism, i.e., Ca2+ sensitivity of MLC20 phosphorylation, and the thin-filament mechanism, i.e., the sensitivity of MLC20 phosphorylation of contractions, in the regulation of baseline Ca2+ sensitivity in the uterine artery, and their adaptations to pregnancy. Consistent with previous studies (3), we found significantly increased protein levels of MLC20 in uterine arteries during pregnancy. In addition, the apparent amount of MLC20-P stimulated by Ca2+ was significantly increased in the uterine artery during pregnancy. These findings suggest that pregnancy increases the baseline Ca2+ sensitivity of MLC20 phosphorylation in the uterine artery. Previous studies in ovine uterine arteries demonstrated that the phenylephrine-stimulated total amount of MLC20-P was increased in uterine arteries from pregnant, compared with nonpregnant, animals (3, 51). This was primarily due to the increased Ca2+ mobilization by activation of {alpha}1-adrenoceptors in uterine arteries during pregnancy (51). When the {alpha}1-adrenoceptor-stimulated [Ca2+]i and MLC20-P relationship was assessed, it was shown that pregnancy was associated with a significant decrease in the Ca2+ sensitivity of MLC20 phosphorylation in response to phenylephrine, i.e., less MLC20 phosphorylation occurred at a given [Ca2+]i stimulated by phenylephrien in uterine arteries during pregnancy (51).

These apparent contradictory findings, i.e., increased baseline Ca2+ sensitivity of MLC20 phosphorylation and decreased {alpha}1-adrenoceptor-mediated Ca2+ sensitivity of MLC20 phosphorylation in uterine arteries during pregnancy, are intriguing and suggest different mechanisms in the agonist-induced Ca2+ and basal Ca2+ sensitivity. It has been well documented that the activity of MLCP plays a key role in the regulation of the Ca2+ sensitivity of MLC20 phosphorylation (39). MLCP consists of three subunits: 110- to 130-kDa regulatory subunit [myosin phosphate target protein-1 (MYPT-1)], ~37-kDa catalytic subunit (PP1c), and 20-kDa subunit of unknown function (13, 39). One of the major mechanisms in inhibition of MLCP is through phosphorylation of MYPT-1 (16). A key event in agonist-induced Ca2+ sensitization is G protein-dependent phosphorylation of MYPT-1, which leads to inhibition of MLCP (16, 24). At least two phosphorylation sites of MYPT-1 have been identified at Thr696 and Thr850 (39). MYPT-1/Thr696 is constitutively phosphorylated by kinases other than Rho-kinase, and phosphorylation of MYPT-1/Thr696 does not respond to agonists (22, 32, 39). In contrast to Thr696, MYPT-1/Thr850 is phosphorylated in response to agonists (9, 22, 23, 39). Our recent studies of the uterine artery demonstrated that phenylephrine stimulated phosphorylation of MYPT-1/Thr850 and that the time course of MYPT-1/Thr850 phosphorylation resembled that of phenylephrine-induced MLC20 phosphorylation and preceded contractions, suggesting that phenylephrine-mediated Ca2+ sensitization was regulated though phosphorylation of MYPT-1/Thr850, resulting in an inhibition of MLCP activity in the uterine artery (46, 48). In contrast, the basal levels of phosphorylated MYPT-1/Thr696 in the uterine artery were not altered by phenylephrine (46). Taken together, these findings suggest the exciting hypothesis that the different phosphorylation sites of MYPT-1 may regulate MLCP activities differently at basal and agonist-stimulated states in the adaptation of the uterine artery to pregnancy.

The finding that Ca2+-mediated contractions at given levels of MLC20 phosphorylation were significantly decreased in uterine arteries from pregnant compared with nonpregnant sheep suggests that pregnancy attenuates the thin-filament pathway in the regulation of baseline Ca2+ sensitivity in the uterine artery. Given that pregnancy upregulates the thick-regulatory pathway, i.e., the Ca2+ sensitivity of MLC20 phosphorylation, the data suggest that the downregulation of the thin-filament regulatory pathway in response to pregnancy dominates in the uterine artery, resulting in decreased baseline Ca2+ sensitivity of contractions. Consistent with the present findings, our previous studies demonstrated that pregnancy significantly depressed the thin-filament regulatory pathway in response to the activation of {alpha}1-adrenoceptors and PKC in the uterine artery (51). In contrast to the thick-filament pathway, the effect of pregnancy on the thin filaments was the same in both basal and agonist-mediated Ca2+ sensitivities in the uterine artery, suggesting that its downregulation is a major mechanism in the adaptation of decreased uterine artery contractility to pregnancy.

It has been demonstrated that PKC is able to modulate the Ca2+ sensitivity via the thin-filament pathway in vascular smooth muscle (19, 36). Our previous studies in the intact uterine arteries demonstrated that PKC-induced contractions were independent of changes in intracellular Ca2+ concentrations and MLC20 phosphorylation, indicating that PKC-induced contractions were mediated predominately through thin-filament regulatory pathways in the uterine artery (46, 51). In the present study, we demonstrated that activation of PKC with PDBu enhanced Ca2+-mediated contractions in membrane-permeabilized uterine arteries. In addition, inhibition of PKC with staurosporine attenuated Ca2+-induced contractions, suggesting an important role of endogenous PKC in the regulation of baseline Ca2+ sensitivity in the uterine artery. Our previous studies demonstrated that PDBu stimulated PKC activity, which was inhibited by staurosporine in the uterine artery (50). In the present study, the finding that PDBu induced contractions in the absence of Ca2+ is intriguing and suggests a Ca2+-independent isozyme of PKC in the regulation of basal Ca2+ sensitivity in the uterine artery. This contrasts with the previous findings in ovine cerebral arteries, in which PDBu failed to produce contractions in permeabilized arteries at zero Ca2+ (2). In agreement with the present finding, phorbol ester-induced Ca2+-independent contractions were demonstrated in ferret and rabbit aorta (28, 42). These studies suggest a tissue specificity of PKC isozymes in the regulation of basal Ca2+ sensitivity. It remains to be determined which isozyme of PKC is involved in the regulation of basal Ca2+ sensitivity in the uterine artery. Consistent with our recent finding of a primary role of PKC in pregnancy-associated decrease in uterine artery myogenic tone (45), the present study demonstrated that PKC-mediated modulation of baseline Ca2+ sensitivity was significantly attenuated in uterine arteries during pregnancy. The effects of PDBu and staurosporine on [Ca2+]-response curves did not reach the significant levels in uterine arteries during pregnancy, albeit they showed the similar patterns of tissue responses as those in vessels from nonpregnant sheep. Nevertheless, it is apparent that the effects of PDBu and staurosporine are much greater in uterine arteries from nonpregnant sheep, suggesting a key role of the decreased PKC pathway in the adaptation of basal Ca2+ sensitivity and, hence, vascular tone of the uterine artery during pregnancy. In agreement with the present findings, it has been demonstrated that pregnancy attenuates the PKC activity and decreases PKC-mediated contractions of the uterine artery (8, 26, 50, 51).

In conclusion, we have demonstrated that pregnancy downregulates the baseline Ca2+ sensitivity in the uterine artery, which is mediated primarily through the inhibition of thin-filament regulatory pathways. PKC plays an important role in the adaptation of uterine artery thin-filament function and baseline Ca2+ sensitivity to pregnancy. Phosphorylation of the actin-associated proteins calponin and caldesmon and actin polymerization have been proposed in the mechanisms of PKC in the regulation of Ca2+ sensitivity independent of MLC20 phosphorylation, and, hence, their involvement in the adaptation of basal Ca2+ sensitivity of the uterine artery during pregnancy presents an intriguing area of further investigation. Given that basal Ca2+ sensitivity plays a key role in the regulation of myogenic tone and, hence, basal vascular resistance and blood flow, decreased baseline Ca2+ sensitivity is likely to play an important role in the adaptation of uterine vascular hemodynamics during pregnancy.


   GRANTS
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
This work was supported in part by National Institutes of Health Grants HL-57787, HL-67745, and HD-31226 and by grants from the Loma Linda University School of Medicine. D. Xiao is a recipient of Postdoctoral Fellowship Award from The Regents of the University of California Tobacco-Related Disease Research Program (Award No. 14FT-0075).


   FOOTNOTES
 

Address for reprint requests and other correspondence: L. Zhang, Center for Perinatal Biology, Dept. of Physiology and Pharmacology, Loma Linda Univ. School of Medicine, Loma Linda, CA 92350 (e-mail lzhang{at}llu.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.


   REFERENCES
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. Akopov SE, Zhang L, and Pearce WJ. Physiological variations in ovine cerebrovascular calcium sensitivity. Am J Physiol Heart Circ Physiol 272: H2271–H2281, 1997.[Abstract/Free Full Text]
  2. Akopov SE, Zhang L, and Pearce WJ. Regulation of Ca2+ sensitization by PKC and rho proteins in ovine cerebral arteries: effects of artery size and age. Am J Physiol Heart Circ Physiol 275: H930–H939, 1998.[Abstract/Free Full Text]
  3. Annibale DJ, Rosenfeld CR, Stull JT, and Kamm KE. Protein content and myosin light chain phosphorylation in uterine arteries during pregnancy. Am J Physiol Cell Physiol 259: C484–C489, 1990.[Abstract/Free Full Text]
  4. Arner A and Pfitzer G. Regulation of cross-bridge cycling by Ca2+ in smooth muscle. Rev Physiol Biochem Pharmacol 134: 63–146, 1999.[Medline]
  5. Bird IM, Zhang L, and Magness RR. Possible mechanisms underlying pregnancy-induced changes in uterine artery endothelial function. Am J Physiol Regul Integr Comp Physiol 284: R245–R258, 2003.[Abstract/Free Full Text]
  6. Bonnevier J and Arner A. Actions downstream of cyclic GMP/protein kinase G can reverse protein kinase C-mediated phosphorylation of CPI-17 and Ca2+ sensitization in smooth muscle. J Biol Chem 279: 28998–29003, 2004.[Abstract/Free Full Text]
  7. Chatterjee M and Tejada M. Phorbol ester-induced contraction in chemically skinned vascular smooth muscle. Am J Physiol Cell Physiol 251: C356–C361, 1986.[Abstract/Free Full Text]
  8. Farley DB and Ford SP. Evidence for declining extracellular calcium uptake and protein kinase C activity in uterine arterial smooth muscle during gestation in gilts. Biol Reprod 46: 315–321, 1992.[Abstract]
  9. Feng J, Ito M, Ichikawa K, Isaka N, Nishikawa M, Harshorne DJ, and Nakano T. Inhibitory phosphorylation site for Rho-associated kinase on smooth muscle myosin phosphatase. J Biol Chem 274: 37385–37390, 1999.[Abstract/Free Full Text]
  10. Ford SP. Control of blood flow to the gravid uterus of domestic livestock species. J Anim Sci 73: 1852–1860, 1995.[Abstract]
  11. Garcia FC, Stiffel VM, Pearce WJ, Zhang L, and Gilbert RD. Ca2+ sensitivity of fetal coronary arteries exposed to long-term, high-altitude hypoxia. J Soc Gynecol Investig 7: 161–166, 2000.[CrossRef][Web of Science][Medline]
  12. Haeberle JR, Hott JW, and Hathaway DR. Regulation of isometric force and isotonic shortening velocity by phosphorylation of the 20,000 dalton myosin light chain of rat uterine smooth muscle. Pflügers Arch 403: 215–219, 1985.[CrossRef][Web of Science][Medline]
  13. Hartshorne DJ, Ito M, and Erdodi F. Myosin light chain phosphatase: subunit composition, interactions and regulation. J Muscle Res Cell Motil 19: 325–341, 1998.[CrossRef][Web of Science][Medline]
  14. Hill MA, Falcone JC, and Meininger GA. Evidence for protein kinase C involvement in arteriolar myogenic reactivity. Am J Physiol Heart Circ Physiol 259: H1586–H1594, 1990.[Abstract/Free Full Text]
  15. Horowitz A, Menice CB, Laporte R, and Morgan KG. Mechanisms of smooth muscle contraction. Physiol Rev 76: 967–1003, 1996.[Abstract/Free Full Text]
  16. Ichikawa K, Ito M, and Hartshrne DJ. Phosphorylation of the large subunit of myosin phosphatase and inhibition of phosphatase activity. J Biol Chem 271: 4733–4740, 1996.[Abstract/Free Full Text]
  17. Ito M and Hartshorne DJ. Phosphorylation of myosin as a regulatory mechanism in smooth muscle. Prog Clin Biol Res 327: 57–72, 1990.[Medline]
  18. Itoh T, Suzuki A, and Watanabe Y. Effect of a peptide inhibitor of protein kinase C on G-protein-mediated increase in myofilament Ca2+-sensitivity in rabbit arterial skinned muscle. Br J Pharmacol 111: 311–317, 1994.[Web of Science][Medline]
  19. Jiang MJ and Morgan KG. Agonist-specific myosin phosphorylation and intracellular calcium during isometric contractions of arterial smooth muscle. Pflügers Arch 413: 637–643, 1989.[CrossRef][Web of Science][Medline]
  20. Kamm KE and Stull JT. The function of myosin and myosin light chain kinase phosphorylation in smooth muscle. Annu Rev Pharmacol Toxicol 25: 593–620, 1985.[CrossRef][Web of Science][Medline]
  21. Kanashiro CA, Cockrell KL, Alexander BT, Granger JP, and Khail RA. Pregnancy-associated reduction in vascular protein kinase C activity rebounds during inhibition of NO synthesis. Am J Physiol Regul Integr Comp Physiol 278: R295–R303, 2000.[Abstract/Free Full Text]
  22. Kitazawa T, Eto M, Woodsome TP, and Khalequzzaman M. Phosphorylation of the myosin phosphatase targeting subunit and CPI-17 during Ca2+ sensitization in rabbit smooth muscle. J Physiol 546: 879–889, 2003.[Abstract/Free Full Text]
  23. Kitazawa T, Kobayashi S, Horiuti K, Somlyo AV, and Somlyo AP. Receptor-coupled, permeabilized smooth muscle. Role of the phosphatidylinositol cascade, G-proteins, and modulation of the contractile response to Ca2+. J Biol Chem 264: 5339–5342, 1989.[Abstract/Free Full Text]
  24. Kitazawa T, Masuo M, and Somlyo AP. G protein-mediated inhibition of myosin light-chain phosphatase in vascular smooth muscle. Proc Natl Acad Sci USA 88: 9307–9310, 1991.[Abstract/Free Full Text]
  25. Laher I and Zhang JH. Protein kinase C and cerebral vasospasm. J Cereb Blood Flow Metab 21: 887–906, 2001.[CrossRef][Web of Science][Medline]
  26. Magness RR, Rosenfeld CR, and Carr BR. Protein kinase C in uterine and systemic arteries during ovarian cycle and pregnancy. Am J Physiol Endocrinol Metab 260: E464–E470, 1991.[Abstract/Free Full Text]
  27. Magness RR, Shaw CE, Phernetton TM, Zheng J, and Bird IM. Endothelial vasodilator production by uterine and systemic arteries. II. Pregnancy effects on NO synthase expression. Am J Physiol Heart Circ Physiol 272: H1730–H1740, 1997.[Abstract/Free Full Text]
  28. Miura M, Iwanaga T, Ito KM, Seto M, Sasaki Y, and Ito K. The role of myosin light chain kinase-dependent phosphorylation of myosin light chain in phorbol ester-induced contraction of rabbit aorta. Pflügers Arch 434: 685–693, 1997.[CrossRef][Web of Science][Medline]
  29. Morgan KG and Gangopadhyay SS. Invited review: cross-bridge regulation by thin filament-associated proteins. J Appl Physiol 91: 953–962, 2001.[Abstract/Free Full Text]
  30. Nauli SM, Zhang L, and Pearce WJ. Maturation depresses cGMP-mediated decreases in [Ca2+]i and Ca2+ sensitivity in ovine cranial arteries. Am J Physiol Heart Circ Physiol 280: H1019–H1028, 2001.[Abstract/Free Full Text]
  31. Nelson SH, Steinsland OS, Wang Y, Yallampalli C, Dong YL, and Sanchez JM. Increased nitric oxide synthase activity and expression in the human uterine artery during pregnancy. Circ Res 87: 406–411, 2000.[Abstract/Free Full Text]
  32. Niiro N, Koga Y, and Ikebe M. Agonist-induced changes in the phosphorylation of the myosin- binding subunit of myosin light chain phosphatase and CPI17, two regulatory factors of myosin light chain phosphatase, in smooth muscle. Biochem J 369: 117–128, 2003.[CrossRef][Web of Science][Medline]
  33. Nishimura J, Moreland S, Moreland RS, and van Breemen C. Regulation of the Ca2+-force relationship in permeabilized arterial smooth muscle. Adv Exp Med Biol 304: 111–127, 1991.[Medline]
  34. Pfitzer G. Invited review: regulation of myosin phosphorylation in smooth muscle. J Appl Physiol 91: 497–503, 2001.[Abstract/Free Full Text]
  35. Rosenfeld CR. Mechanisms regulating angiotensin II responsiveness by the uteroplacental circulation. Am J Physiol Regul Integr Comp Physiol 281: R1025–R1040, 2001.[Abstract/Free Full Text]
  36. Sato K, Hori M, Ozaki H, Takano-Ohmuro H, Tsuchiya T, Sugi H, and Karaki H. Myosin phosphorylation-independent contraction induced by phorbol ester in vascular smooth muscle. J Pharmacol Exp Ther 261: 497–505, 1992.[Abstract/Free Full Text]
  37. Sobue K and Sellers JR. Caldesmon, a novel regulatory protein in smooth muscle and nonmuscle actomyosin systems. J Biol Chem 266: 12115–12118, 1991.[Free Full Text]
  38. Somlyo AP and Somlyo AV. Signal transduction and regulation in smooth muscle. Nature 372: 231–236, 1994.[CrossRef][Medline]
  39. Somlyo AP and Somlyo AV. Ca2+ sensitivity of smooth muscle and nonmuscle myosin II: modulated by G proteins, kinases, and myosin phosphatase. Physiol Rev 83: 1325–1358, 2003.[Abstract/Free Full Text]
  40. Uehata M, Ishizaki T, Satoh H, Ono T, Kawahara T, Morishita T, Tamakawa H, Yamagami K, Inui J, Maekawa M, and Narumiya S. Calcium sensitization of smooth muscle mediated by a Rho-associated protein kinase in hypertension. Nature 389: 990–994, 1997.[CrossRef][Medline]
  41. Veerareddy S, Cooke CL, Baker PN, and Davidge ST. Vascular adaptations to pregnancy in mice: effects on myogenic tone. Am J Physiol Heart Circ Physiol 283: H2226–H2233, 2002.[Abstract/Free Full Text]
  42. Walsh MP, Horowitz A, Clement-Chomienne O, Andrea JE, Allen BG, and Morgan KG. Protein kinase C mediation of Ca2+-independent contractions of vascular smooth muscle. Biochem Cell Biol 74: 485–502, 1996.[Web of Science][Medline]
  43. Weber LP, Seto M, Sasaki Y, Sward K, and Walsh MP. The involvement of protein kinase C in myosin phosphorylation and force development in rat tail arterial smooth muscle. Biochem J 352: 573–582, 2000.[Medline]
  44. Weiner CP, Martinez E, Chestnut DH, and Ghodsi A. Effect of pregnancy on uterine and carotid artery response to norepinephrine, epinephrine, and phenylephrine in vessels with documented functional endothelium. Am J Obstet Gynecol 161: 1605–1610, 1989.[Web of Science][Medline]
  45. Xiao D, Buchholz JN, and Zhang L. Pregnancy attenuates uterine artery pressure-dependent vascular tone: role of PKC/ERK pathway. Am J Physiol Heart Circ Physiol. 290: H2337–H2343, 2006.[Abstract/Free Full Text]
  46. Xiao D, Longo LD, and Zhang L. {alpha}1-Adrenoceptor-mediated phosphorylation of MYPT-1 and CPI-17 in the uterine artery: role of ERK/PKC. Am J Physiol Heart Circ Physiol 288: H2828–H2835, 2005.[Abstract/Free Full Text]
  47. Xiao D, Liu Y, Pearce WJ, and Zhang L. Endothelial nitric oxide release in isolated perfused ovine uterine arteries: effect of pregnancy. Eur J Pharmacol 367: 223–230, 1999.[CrossRef][Web of Science][Medline]
  48. Xiao D, Pearce WJ, Longo LD, and Zhang L. ERK-mediated uterine artery contraction: role of thick and thin filament regulatory pathways. Am J Physiol Heart Circ Physiol 286: H1615–H1622, 2004.[Abstract/Free Full Text]
  49. Xiao D, Pearce WJ, and Zhang L. Pregnancy enhances endothelium-dependent relaxation of ovine uterine artery: role of NO and intracellular Ca2+. Am J Physiol Heart Circ Physiol 281: H183–H190, 2001.[Abstract/Free Full Text]
  50. Xiao D and Zhang L. ERK MAP kinases regulate smooth muscle contraction in ovine uterine artery: effect of pregnancy. Am J Physiol Heart Circ Physiol 282: H292–H300, 2002.[Abstract/Free Full Text]
  51. Xiao D and Zhang L. Adaptation of uterine artery thick- and thin-filament regulatory pathways to pregnancy. Am J Physiol Heart Circ Physiol 288: H142–H148, 2005.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
K. Chang, D. Xiao, X. Huang, L. D. Longo, and L. Zhang
Chronic hypoxia increases pressure-dependent myogenic tone of the uterine artery in pregnant sheep: role of ERK/PKC pathway
Am J Physiol Heart Circ Physiol, June 1, 2009; 296(6): H1840 - H1849.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Regul. Integr. Comp. Physiol.Home page
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]

Home page
 Reproductive SciencesHome page
K. Chang and Lubo Zhang
Review Article: Steroid Hormones and Uterine Vascular Adaptation to Pregnancy
Reproductive Sciences, April 1, 2008; 15(4): 336 - 348.
[Abstract] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
291/1/H413    most recent
00103.2006v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Xiao, D.
Right arrow Articles by Zhang, L.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Xiao, D.
Right arrow Articles by Zhang, L.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2006 by the American Physiological Society.