Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta, Georgia 30912
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The signal transduction mechanisms defining the role of cyclic nucleotides in the regulation of pulmonary vascular tone is currently an area of great interest. Normally, signaling mechanisms that elevate cAMP and guanosine-3',5'-cyclic monophosphate (cGMP) maintain the pulmonary vasculature in a relaxed state. Modulation of the large-conductance, calcium- and voltage-activated potassium (BKCa) channel is important in the regulation of pulmonary arterial pressure, and inhibition (closing) of the BKCa channel has been implicated in the development of pulmonary hypertension. Accordingly, studies were done to determine the effect of cAMP-elevating agents on BKCa channel activity using patch-clamp studies in pulmonary arterial smooth muscle cells (PASMC) of the fawn-hooded rat (FHR), a recognized animal model of pulmonary hypertension. Forskolin (10 µM), a stimulator of adenylate cyclase and an activator of cAMP-dependent protein kinase (PKA), and 8-4-chlorophenylthio (CPT)-cAMP (100 µM), a membrane-permeable derivative of cAMP, opened BKCa channels in single FHR PASMC. Treatment of FHR PASMC with 300 nM KT5823, a selective inhibitor of cGMP-dependent protein kinase (PKG) activity inhibited the effect of both forskolin and CPT-cAMP. In contrast, blocking PKA activation with 300 nM KT5720 had no effect on forskolin or CPT-cAMP-stimulated BKCa channel activity. These results indicate that cAMP-dependent vasodilators activate BKCa channels in PASMC of FHR via PKG-dependent and PKA-independent signaling pathways, which suggests cross-activation between cyclic nucleotide-dependent protein kinases in pulmonary arterial smooth muscle and therefore, a unique signaling pathway for cAMP-induced pulmonary vasodilation.
high-conductance calcium-and voltage-activated potassium channel; cAMP-dependent protein kinase; cross-activation
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
SIGNAL TRANSDUCTION MECHANISMS defining the role of cyclic nucleotides in the modulation of vascular tone is currently an area of great interest. Agents that relax vascular smooth muscle via activation of adenylate cyclase, such as forskolin, are thought to elicit their biological effects by increasing the intracellular concentration of cAMP (16, 17, 34). Although the exact mechanism by which cAMP causes vascular relaxation is not known, it is believed that activation of PKA is involved (7, 17, 18). In addition to cAMP-mediated vascular relaxation, guanosine-3',5'-cyclic monophosphate (cGMP) has been reported to elicit vasodilatation by activation of cGMP-dependent PKG (6, 23). In the pulmonary circulation, cGMP and cAMP have been implicated as mediators of smooth muscle vasodilatation (13, 15, 16, 24), and in ovine pulmonary vascular smooth muscle, it has recently been reported that pulmonary arteries are more sensitive to relaxation induced by cGMP and that activation of PKG is involved in both cGMP- and cAMP-induced relaxation (10).
It is believed that cAMP and cGMP stimulate their associated protein kinases, PKA and PKG, respectively, but recent evidence suggests that the vasodilatory effects of cAMP-producing agents may involve "cross-activation" of PKG by cAMP (41, 43) and stimulation of PKA by cGMP (8). Lincoln and colleagues (24) originally showed that cross-activation of PKG by cAMP was important in cAMP-elevating agent vasodilatation, and it is thought that cAMP can activate both PKA and PKG. However, relaxation correlates most closely with PKG activation (13), and vasodilators that increase cGMP and cAMP seem to act synergistically on vasorelaxation (41). Thus, whereas studies have reported the existence of cross-activation of PKG by cAMP, few studies have investigated this signaling mechanism at the cellular and molecular levels in pulmonary arterial smooth muscle cells (PASMC).
Although multiple classes of K+ channels are expressed at varying densities in different vascular beds, the large-conductance, calcium- and voltage-activated potassium (BKCa) channel is the predominant potassium channel species in most arteries (27). Inhibition of BKCa channels produces membrane depolarization and subsequent vasoconstriction (29). Specific to cyclic nucleotide activity, evidence suggests that BKCa channels in arterial smooth muscle can be opened through activation of PKG, which phosphorylates the channel (31). It has been shown that PKG stimulates BKCa channel activity in coronary (43), cerebral (31), and pulmonary arteries (1), and BKCa channels are also opened by PKA (25) in a variety of tissues including in vascular smooth muscle to influence myogenic tone (37). In PASMC, it has been reported that nitric oxide causes vasodilation through PKG-mediated BKCa channel activation by channel phosphorylation (28, 35). Furthermore, Haynes et al. (17) observed that tetraethylammonium (TEA), a selective BKCa channel blocker at low concentrations (38), inhibited pulmonary vasodilation induced by the PKA activator 8-bromoadenosine 3',5'-cyclic monophosphate or forskolin, suggesting a vasodilatory mechanism involving cAMP and BKCa channel activation.
In light of these previous investigations, the present study was done to determine mechanisms of cAMP activation of BKCa channels in PASMC of the fawn-hooded rat (FHR), an animal model of idiopathic pulmonary hypertension. Because genetic factors are likely to contribute to the pulmonary hypertension that develops in the FHR, the pulmonary vasculature of this animal model is an excellent model to use to further understand potential mechanisms of pulmonary hypertension. Specifically, the effect of cAMP on PKA and PKG modulation of BKCa channel activity was investigated at the cellular and molecular levels via the patch-clamp technique in FHR PASMC.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Animals. All procedures and protocols were approved by the Animal Care and Use Committee at the Medical College of Georgia. FHRs were a gift from Dr. I. McMurtry (Cardiovascular Pulmonary Research Laboratory, Department of Medicine, University of Colorado Health Sciences Center, Denver, CO), and a breeding colony was established. Animals were maintained on standard rat chow, allowed free access to food and water, and exposed to a 12:12-h light-dark cycle. Male FHRs at 12-16 wk of age were used for this study.
Isolation of PASMC. After the rats were anesthetized with pentobarbital sodium (50 mg/kg ip), a midline incision was made, and the lungs were immediately excised and placed into ice-cold dissociation buffer consisting of (in mM): 110 NaCl, 5 KCl, 0.16 CaCl2, 2 MgCl2, 10 HEPES, 0.5 NaH2PO4, 10 NaHCO3, 0.5 KH2PO4, 10 glucose, 0.49 EDTA, and 10 taurine. Proximal conduit pulmonary arterial vessels were dissected from the lung lobes under a stereomicroscope. Endothelium was removed, and the adventitia was carefully teased away. The vascular tissue was then dissociated enzymatically at 35°C for 1 h in an incubation solution of dissociation buffer containing the following: papain (6.7 mg/5 ml), cysteine (3.5 mg/5 ml), and BSA (5.0 mg/ml). After incubation, 5.0 ml of dissociation buffer containing 2.0 mg/ml BSA was added to the cell solution, and the tissue was then triturated gently, which allowed individual smooth muscle cells to fall away from the larger pieces of undigested tissue with minimal damage to the cells. These cells exhibited morphology characteristics of vascular smooth muscle cells. The solution was then removed and centrifuged at 1,200 rpm for 10 min. The pellet was then resuspended in dissociation buffer. For patch-clamp experiments, several drops of cell suspension were placed in a microscope chamber containing recording solution, and all experiments were performed within 1-2 h after cell dissociation.
Ion channel recording. Whole cell currents were measured from metabolically intact PASMC using the amphotericin-perforated-patch technique (6, 11). In contrast to standard whole cell techniques, perforated-patch recordings provide accurate current measurement with only minimal current decay or loss of soluble cytoplasmic components due to cellular dialysis. Furthermore, endogenous calcium buffering is not inactivated by dialyzing cells with calcium chelators, as are required during whole cell recordings. In addition, identification and characterization of single ion channels were also assessed in these cells.
Perforated-patch experiments.
Cells were placed in a recording solution of the following composition
(in mM): 140 NaCl, 5 KCl, 2 MgCl2, 2 CaCl2, 20 HEPES, and 20 glucose (pH 7.2). Patch pipettes with a resistance of 3 M or less were fabricated from capillaries of Corning Glass 7052 or
other appropriate formulations. To measure potassium currents, the tip
of the patch pipette was filled with a solution containing (in mM) 90 KCH3SO3, 40 KCl, 5 MgCl2, and 20 HEPES to approximate normal cellular potassium concentration
([K+]) and chloride concentration ([Cl
])
(pH 7.2 with KOH). The remainder of the pipette was back filled with a
similar solution to which 200 mg/ml amphotericin B (diluted by dimethyl
sulfoxide) was added. Cells were studied only if the voltage drop
across the series resistance was reduced to
5 mV within 10-20
min after forming a gigaohm seal. Voltage-clamp and voltage-pulse
generation was controlled with an Axopatch 200A patch-clamp amplifier
(Axon Instruments), and data were analyzed with pCLAMP 6.0.3 (Axon
Instruments), which is a comprehensive software package for acquisition
and analysis of both whole cell and single-channel currents.
Voltage-activated currents were filtered at 2 kHz and digitized at 10 kHz, and capacitative and leakage currents were subtracted digitally.
All drugs were diluted into fresh bath solution and perfused into a
0.5-ml recording chamber (Warner Instruments).
Single-channel experiments.
Single potassium channels were measured in cell-attached patches by
filling the patch pipette (2-5 M) with the standard bath solution and making a gigaohm seal on an intact cell. The solution in
the recording chamber contained (in mM) 140 KCl, 10 MgCl2, 0.1 CaCl2, 10 HEPES, and 30 glucose (pH 7.2). This
manipulation allowed precise regulation of the patch membrane voltage
by the patch-clamp amplifier and yielded more accurate current
measurements. In experiments measuring potassium channel activity in
cell-free inside-out patches, the solution facing the cytoplasmic
surface of the membrane had the following composition (in mM): 115 KCH3SO3, 26 KCl, 2 MgCl2, 1 BAPTA,
0.47 CaCl2 (pCa 7), 5.0 Mg-ATP, 0.1 GTP, and 20 HEPES (pH
7.2). The solution facing the external surface was the standard
recording solution as described above. Currents were filtered at 2 kHz
and digitized at 10 kHz. Channel activity was quantified by calculating
the single, open-channel probability (NPo) as described previously
(11, 42).
Right ventricular hypertrophy measurements. At the time of death, hearts from adult FHR and control male Sprague-Dawley rats (SDR) were removed and dissected to isolate the free wall of the right ventricle from the left ventricle and septum. The ratio of right ventricle weight to left ventricle plus septum weight (RV/LV+S) was used as an index of right ventricular hypertrophy (14).
Drugs. TEA, iberiotoxin (IBTx), forskolin, 8-4-chlorophenylthio (CPT)-cAMP, CPT-cGMP, Rp-8-CPT-cGMPs, KT5823, and KT5720 were purchased from Calbiochem (San Diego, CA). All other agents were purchased from Sigma (St. Louis, MO).
Statistical analysis. All data were expressed as means ± SE. Statistical significance between two groups was evaluated by Student's t-test for paired data. Comparison among multiple groups was made by using one-way ANOVA for multiple comparisons. A P value <0.05 indicated a significant difference.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Identification of right ventricular hypertrophy. The RV/LV+S ratios in FHR (0.44 ± 0.05; n = 8) vs. control SDR (0.28 ± 0.03, P < 0.05; n = 8) were indicative of right ventricular hypertrophy in FHR as reported in other studies (22, 26, 36, 42).
Identification of BKCa channels in PASMC from FHR.
To date, we are aware of no previous electrophysiological studies that
have characterized potassium channels in PASMC from FHR. Therefore, our
initial experiments characterized outward currents in PASMC from these
animals, and these experiments were done in metabolically intact muscle
cells. We performed current-clamp experiments on single PASMC using the
perforated-patch method and measured an average resting membrane
potential (RMP) of 50.8 ± 2.8 mV (n = 4). After
treating cells with 100 nM IBTx for 15-20 min, RMP was depolarized
to an average of
25.8 ± 5.9 mV (n = 4), which
represented a highly significant (P = 0.006)
depolarization of RMP due to selective blockade of BKCa
channels indicating that these channels are a major regulator of RMP in
these cells.
|
Single BKCa channel properties.
Unitary current amplitude was plotted as a function of membrane
potentials (60 to +60 mV) as shown in Fig.
2. Recordings from excised (inside-out)
patches under conditions of symmetrical [K+] (140 mM)
demonstrated activity of a large-amplitude channel that reversed at
~0 mV, which is the calculated equilibrium potential for
K+ in these experiments (Fig. 2A).
Single-channel current-voltage relationships yielded an average
single-channel conductance of 142 ± 5 pS (n = 3;
Fig. 2B), which agrees with other studies reporting BKCa channel conductance in vascular smooth muscle
(6, 11). Furthermore, raising Ca2+
concentration at the cytoplasmic face of the patch membrane increased channel gating dramatically (Figs. 5B and 6A),
and channel activity was inhibited by 1 mM TEA (Fig.
3B), a selective inhibitor of BKCa channels at this concentration (37). Thus
the specific biophysical and pharmacological profile obtained from both
intact cells and isolated patches in PASMC from FHR clearly identify this protein as the BKCa channel, which is highly expressed
in vascular smooth muscle.
|
|
cAMP-dependent vasodilators stimulate BKCa channels by
a PKA-independent pathway.
Intracellular levels of cAMP were increased in PASMC with either
forskolin (stimulator of adenylyl cyclase) or a membrane-permeable derivative of cAMP (CPT-cAMP). As shown in Fig. 3, forskolin (10 µM;
10 min) stimulated BKCa channel activity (NPo from
~0.0000 ± 0.0000 to 0.2868 ± 0.0857; n = 5; P < 0.02; Fig. 3A). In contrast, 1 mM
TEA inhibited BKCa channel activity after excision into the inside-out configuration (Fig. 3B). In addition to
forskolin, treating cells with membrane-permeable 100 µM CPT-cAMP (20 min; Fig. 4B) dramatically
increased the activity of BKCa channels in cell-attached
patches (NPo ~0.0000 ± 0.0000 to 0.9832 ± 0.005; n = 5; P < 0.0001). Treatment with the
PKA inhibitor KT5720 (300 nM) did not attenuate the effect of forskolin
(n = 3; Fig. 4A) or CPT-cAMP
(n = 3; Fig. 4B) on BKCa channel
activity. These data indicate that cAMP activates BKCa
channels in FHR PASMC by a PKA-independent signaling pathway.
|
cAMP-dependent vasodilators stimulate BKCa channels by
a PKG-dependent pathway.
Fig. 5 shows the effect of KT5823 (300 nM), an inhibitor of PKG activity, on CPT-cAMP (Fig. 5A) and
forskolin-induced BKCa channel activation (Fig.
5B). As previously observed, 100 µM CPT-cAMP increased
BKCa channel NPo (0.000 ± 0.000 to 0.997 ± 0.0057; P < 0.0001; n = 3), but the
subsequent addition of 300 nM KT5823 completely reversed this effect
(0.000 ± 0.000; P < 0.0001; n = 3) (Fig. 5A), a phenomenon not observed in SDR PASMC
(control NPo: 0.000 ± 0.000; forskolin NPo: 0.3400 +
0.295; forskolin and KT5823 NPo: 0.5379 ± 0.071;
n = 2). KT5823 was also a potent inhibitor of 10 µM
forskolin-stimulated NPo, because pretreatment with 300 nM KT5823
prevented the response to 10 µM forskolin (n = 2) as
illustrated in the time-course histogram of BKCa channel activity (Fig. 5B). Subsequent incision of the patch
into an inside-out configuration and raising cytoplasmic
Ca2+ concentration produced an immediate increase in
BKCa channel activity in the presence of both KT5823 and
forskolin, indicating the continued BKCa channel viability
of these cells. Further experiments done with another inhibitor of PKG
activity, 100 µM Rp-8-CPT-cGMPS, also prevented the stimulatory
effect of CPT-cAMP on BKCa channels (n = 4)
(Fig. 6A). Once again,
excision into the inside-out configuration revealed the continued
BKCa channel viability of these cells in the presence of
Rp-8-CPT-cGMPS and CPT-cAMP (Fig. 6A, right).
|
|
cGMP mimics the effect of cAMP and forskolin on BKCa channels. If cAMP-dependent vasodilators activate BKCa channels through a PKG-dependent signaling pathway, then treating cells with a non-cAMP- dependent agent that increases levels of PKG via cGMP should also stimulate BKCa channel activity. To this end, subsequent experiments revealed that treating cells with CPT-cGMP (a membrane-permeable derivative of cGMP) stimulated BKCa channel openings in PASMC. Specifically, BKCa channel NPo was increased by >100-fold after a 20-min exposure to 100 µM CPT-cGMP (0.0003 ± 0.0002 to 0.0335 ± 0.0106; P < 0.03; n = 4) (Fig. 6B).
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Results of the present study indicate that the cAMP-dependent vasodilators forskolin (activator of adenylate cyclase) and CTP-cAMP (membrane-permeable derivative of cAMP) activate BKCa channels in PASMC of FHR via PKG-dependent and PKA-independent signaling pathways. These findings suggest cross-activation between cyclic nucleotide-dependent protein kinases in pulmonary arterial smooth muscle. Earlier studies have shown that agents such as forskolin can activate PKG by increasing cAMP levels in both vascular and nonvascular smooth muscle (13, 18, 24). Also, it has been observed that cAMP levels are up to fivefold higher in some vascular smooth muscle tissues (13), and autophosphorylation of PKG greatly improves its affinity for cAMP, making it more readily activated by cAMP than in the unphosphorylated state (12).
Specific to pulmonary vascular smooth muscle, few studies have shown that cAMP can activate PKG, although recent studies have shown that cGMP can elicit physiological effects by stimulating PKA activity in systemic vascular smooth muscle (8). In addition, cAMP and its analogs can activate both PKA and PKG, with vascular relaxation correlating more closely with PKG activation (13). In the pulmonary circulation, cGMP and cAMP have been implicated as mediators of smooth muscle vasodilation (10, 15-17), and in ovine pulmonary vascular smooth muscle, it has recently been reported (10) that pulmonary arteries are more sensitive to relaxation induced by cGMP and that both cGMP- and cAMP-induced relaxation involves activation of PKG. In addition, Gao and colleagues (15) observed PKG-mediated relaxation of newborn ovine pulmonary veins, and Haynes et al. (17) reported in rat pulmonary circulation that cAMP-mediated pulmonary vasodilatation was mediated via PKA.
Potassium channel activity is the main determinant of membrane potential, and associated K+ efflux causes hyperpolarization, which inhibits voltage-gated calcium channels and promotes vascular relaxation. Although multiple classes of potassium channels are expressed at varying densities in different vascular beds, the BKCa channel is the predominant potassium channel species in most arteries (27). BKCa channels are activated by submicromolar intracellular Ca2+ concentration and are blocked by external charybdotoxin, IBTx, and TEA ions (27, 32). The biophysical profile of the BKCa channel is that it is a large conducting channel (100-150 pS in physiological K+ gradients) that is both calcium and voltage dependent for activation (6, 27, 43). Because of their large conductance and high density, these channels influence RMP and provide an important repolarizing negative feedback mechanism. Inhibition of BKCa channels produces membrane depolarization and subsequent vasoconstriction (29). Specific to this study, the BKCa single-channel conductance was slightly lower (~142 pS) than is typically measured in vascular smooth muscle (>160 pS). The reason for this slightly lower value in this particular vascular smooth muscle type is not apparent. However, because our data are the first to report single BKCa channel conductance in PASMC from FHRs, the BKCa channels in pulmonary vascular smooth muscle in this species may indeed have a conductance of 140-150 pS, which is only ~10% lower than average pS values measured in other types of vascular smooth muscle. Regardless, the conductance values measured in this study are clearly indicative of average single-channel conductance of BKCa channels.
Evidence suggests that ion channels, including BKCa channels, may be modulated by cyclic nucleotide second messengers in various tissues including pulmonary vascular smooth muscle (4, 5, 9, 23, 30, 33). Barman (3) recently reported that dibutyryl cAMP, a cell membrane-permeable analog of cAMP, the cGMP/cAMP phosphodiesterase inhibitor IBMX, and the cAMP-dependent vasodilator isoproterenol blocked the pulmonary vasoconstrictor response to serotonin. In addition, it has been shown that the activity of BKCa channels is potentiated by the cGMP-dependent activation of G kinase (31), and Archer et al. (1) provided compelling evidence for the opening of BKCa channels by increased cGMP levels in rat pulmonary vascular smooth muscle cells. In their experimental model, Archer et al. (1) suggest that pulmonary vascular signal transduction is modulated by both cGMP and cAMP, which regulate potassium channel phosphorylation. Specifically, increased levels of these cyclic nucleotide second messengers promote opening of BKCa channels, which leads to membrane hyperpolarization, inhibition of voltage-gated calcium channels, and subsequent vasorelaxation. Torphy (40), in a recent review, documented that increases in cAMP and cGMP can simultaneously activate the PKA and PKG pathways to promote opening of BKCa channels and elicit membrane hyperpolarization.
Because vascular myocytes are a rich source of PKG, several studies suggest that cGMP-dependent phosphorylation is important in regulating vascular tone (6, 31). Evidence suggests that BKCa channels in arterial smooth muscle can be opened through activation of PKG, which phosphorylates the channel (31), and Carrier et al. (6) reported that BKCa channel activity via cGMP-dependent phosphorylation contributes to vasodilatation in mesenteric vascular smooth muscle. In PASMC, it has been reported that nitric oxide causes vasodilation through PKG-mediated BKCa channel activation by channel phosphorylation (28, 35). Finally, it has been shown that purified PKG stimulates BKCa channel activity in coronary (43), cerebral (31), and pulmonary arteries (1).
Cyclic nucleotide-elevating vasodilators have been used to treat a variety of cardiovascular diseases including heart failure relating to pulmonary hypertension, and vasodilator therapy plays an important role in the management of pulmonary hypertension. Agents that stimulate the cAMP or cGMP transduction pathway in the pulmonary vasculature are presently among the primary treatment measures for this pathophysiological state. For example, some of the most commonly used vasodilators, sodium nitroprusside and nitroglycerin, stimulate the cGMP signaling pathway, whereas isoproterenol, amrinone, and forskolin activate the cAMP transduction pathway. Most recently, the selective phosphodiesterase inhibitor zaprinast has been shown to induce pulmonary vasodilation through the cGMP pathway (19) and has been reported to be effective in the management of pulmonary hypertension. Although the vasodilatory effects of these agents are well documented, there is currently very little knowledge available about the cellular and molecular basis of cAMP- and cGMP-dependent vasodilator mechanisms of action in pulmonary vascular smooth muscle.
The study of pulmonary hypertension has also been problematic because of lack of relevant animal models. In addition, it is difficult in many experimental models to separate the vascular changes that may be pathogenic from the changes that occur secondary to the hypertension (2). Recently, the FHR has been found to be an excellent animal model to study pulmonary hypertension (22, 36). The FHR strain is characterized by platelet abnormalties and systemic hypertension and has been widely used to study genetic risk factors for the development of idiopathic pulmonary hypertension (20, 36). Studies show that the platelet storage disease is not involved in the pathogenesis of pulmonary hypertension (2, 20) and that the FHR strain develops significant pulmonary hypertension, which is not associated with differences in blood gas tension, polycythemia, or parenchymal lung disease (45). Unlike other rat strains, FHR develop severe pulmonary hypertension, which is age dependent (20), a phenomenon accelerated under mild hypoxic conditions (36). Evidence also suggests that the genetic locus for the hypertensive condition is PH1 on chromosome 1 (39).
In summary, the results of this study indicate that cAMP-dependent vasodilators activate BKCa channels in PASMC of FHR via PKG-dependent and PKA-independent signaling pathways. Thus this study suggests that cAMP increases levels of PKG in an animal model of pulmonary hypertension and demonstrates the phenomenon of cross-activation of cyclic-nucleotide-dependent protein kinases in pulmonary arterial smooth muscle under potential pathophysiologcial conditions.
![]() |
ACKNOWLEDGEMENTS |
---|
The authors thank Louise Meadows for excellent technical assistance.
![]() |
FOOTNOTES |
---|
This work was supported by National Heart, Lung, and Blood Institute Grants HL-68026 (to S. A. Barman) and HL-64779 (to R. E. White) and by the American Heart Association Grant 9950179N (to R. E. White).
Address for reprint requests and other correspondence: S. A. Barman, Dept. of Pharmacology and Toxicology, Medical College of Georgia, Augusta, GA 30912 (E-mail: sbarman{at}mail.mcg.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.
First published January 24, 2003;10.1152/ajplung.00295.2002
Received 27 August 2002; accepted in final form 13 January 2003.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Archer, SL,
Huang JMC,
Hampl V,
Nelson DP,
Schultz PJ,
and
Weir EK.
Nitric oxide and cGMP cause vasorelaxation by activation of a charybdotoxin-sensitive K channel by cGMP-dependent protein kinase.
Proc Natl Acad Sci USA
91:
7583-7587,
1994[Abstract].
2.
Ashmore, RC,
Rodman DM,
Sato K,
Webb SA,
O'Brien RF,
McMurtry IF,
and
Stelzner TJ.
Paradoxical constriction to platelets by arteries from rats with pulmonary hypertension.
Am J Physiol Heart Circ Physiol
260:
H1929-H1934,
1991
3.
Barman, SA.
Role of calcium-activated potassium channels and cyclic nucleotides on pulmonary vasoreactivity to serotonin.
Am J Physiol Lung Cell Mol Physiol
273:
L142-L147,
1997
4.
Cabell, F,
Weiss DS,
and
Price JM.
Inhibition of adenosine-induced coronary vasodilation by block of large-conductance Ca2+-activated K+ channels.
Am J Physiol Heart Circ Physiol
267:
H1455-H1460,
1994
5.
Cachelin, AB,
De Peyer JE,
Kokubun S,
and
Reuter H.
Ca2+ channel modulation by 8-bromocyclic-AMP in cultured heart cells.
Nature
304:
462-464,
1983[ISI][Medline].
6.
Carrier, GO,
Fuchs LC,
Winecoff AP,
Giulumar AD,
and
White RE.
Nitrovasodilators relax mesenteric microvessels by cGMP-induced stimulation of Ca-activated K channels.
Am J Physiol Heart Circ Physiol
273:
H76-H84,
1997
7.
Conti, MA,
and
Adelstein RS.
The relationship between calmodulin binding and phosphorylation of smooth muscle myosin kinase by the catalytic subunit of 3',5'-cAMP-dependent protein kinase.
J Biol Chem
256:
3178-3181,
1981
8.
Cornwell, TL,
Arnold E,
Boerth NJ,
and
Lincoln TM.
Inhibition of smooth muscle growth by nitric oxide and activation of cAMP-dependent protein kinase by cGMP.
Am J Physiol Cell Physiol
267:
C1405-C1413,
1994
9.
Demirel, E,
Rusko J,
Laskey RE,
Adams DJ,
and
van Breeman C.
TEA inhibits ACh-induced EDRF release: endothelial Ca2+-dependent K+ channels contribute to vascular tone.
Am J Physiol Heart Circ Physiol
267:
H1135-H1141,
1994
10.
Dhanakoti, SN,
Gao Y,
Nguyen MQ,
and
Raj JU.
Involvement of cGMP-dependent protein kinase in the relaxation of ovine pulmonary arteries to cGMP and cAMP.
J Appl Physiol
88:
1637-1642,
2000
11.
Dimitropoulou, C,
Han G,
Miller AW,
Molero M,
Fuchs LC,
White RE,
and
Carrier GO.
Potassium (BKCa) currents are reduced in microvascular smooth muscle cells from insulin-resistant rats.
Am J Physiol Heart Circ Physiol
282:
H908-H917,
2002
12.
Foster, JL,
Guttman J,
and
Rosen OM.
Autophosphorylation of cGMP-dependent protein kinase.
J Biol Chem
256:
5029-5036,
1981[ISI][Medline].
13.
Francis, SH,
Noblett BD,
Todd BW,
Wells JN,
and
Corbin JD.
Relaxation of vascular and tracheal smooth muscle by cyclic nucleotide analogs that preferentially activate purified cGMP-dependent protein kinase.
Mol Pharmacol
34:
506-517,
1988[Abstract].
14.
Fulton, RM,
Hutchinson EC,
and
Jones AM.
Ventricular weight in cardiac hypertrophy.
Br Heart J
14:
413-420,
1952.
15.
Gao, Y,
Dhanakoti S,
Tolsa JF,
and
Raj JU.
Role of protein kinase G in nitric oxide-and cGMP-induced relaxation of newborn ovine pulmonary veins.
J Appl Physiol
87:
993-998,
1999
16.
Haynes, J, Jr,
Kithas PA,
Taylor AE,
and
Strada SJ.
Selective inhibition of cGMP-inhibitable cAMP phosphodiesterase decreases pulmonary vasoreactivity.
Am J Physiol Heart Circ Physiol
261:
H487-H492,
1991
17.
Haynes, JH,
Robinson J,
Saunders L,
Taylor AE,
and
Strada SJ.
Role of cAMP-dependent protein kinase in cAMP-mediated vasodilation.
Am J Physiol Heart Circ Physiol
262:
H511-H516,
1992
18.
Hei, YJ,
MacDonell KL,
McNeill JH,
and
Diamond J.
Lack of correlation between activation of cyclic AMP-dependent protein kinase and inhibition of contraction of rat vas deferens by cyclic AMP analogs.
Mol Pharmacol
39:
233-238,
1991[Abstract].
19.
Ichinose, FI,
Adie C,
Hurford WE,
Bloch KD,
and
Zapol WE.
Selective pulmonary vasodilation induced by aerosolized zaprinast.
Anesthesiology
88:
410-416,
1998[ISI][Medline].
20.
Kentera, D,
Susic D,
Veljkovic V,
Tucakovic G,
and
Koko V.
Pulmonary artery pressure in rats with hereditary platelet function defect.
Respiration
54:
110-114,
1988[ISI][Medline].
21.
Le Cras, TD,
Dug-Ha K,
Markham NE,
and
Abman SH.
Early abnormalities of pulmonary vascular development in the fawn-hooded rat raised at Denver's altitude.
Am J Physiol Lung Cell Mol Physiol
279:
L283-L291,
2000
22.
Le Cras, TD,
Kim D,
Gebb S,
Markham NE,
Shannon JM,
Tuder RM,
and
Abman SH.
Abnormal lung growth and the development of pulmonary hypertension in the fawn-hooded rat.
Am J Physiol Lung Cell Mol Physiol
277:
L709-L718,
1999
23.
Lincoln, TM,
and
Cornwell TL.
Intracellular cyclic GMP receptor proteins.
FASEB J
7:
328-338,
1993
24.
Lincoln, TM,
Cornwell TL,
and
Taylor AE.
cGMP-dependent protein kinase mediates the reduction of Ca2+ by cAMP in vascular smooth muscle cells.
Am J Physiol Cell Physiol
258:
C399-C407,
1990
25.
Minami, K,
Fukuzawa K,
Nakaya Y,
Xeng XR,
and
Inoue I.
Mechanism of activation of the Ca-activated K+ channel by cyclic AMP in cultured porcine coronary artery smooth muscle cells.
Life Sci
53:
1129-1135,
1993[ISI][Medline].
26.
Nagaoka, T,
Muramatsu M,
Sato K,
McMurtry I,
Oka M,
and
Fukuchi Y.
Mild hypoxia causes severe pulmonary hypertension in fawn-hooded but not in Tester Moriyama rats.
Respir Physiol
127:
53-60,
2001[ISI][Medline].
27.
Nelson, MT,
and
Quayle JM.
Physiological roles and properties of potassium channels in arterial smooth muscle.
Am J Physiol Cell Physiol
268:
C799-C822,
1995
28.
Peng, W,
Hoidal JR,
and
Farrukh IS.
Regulation of Ca2+-activated K+ channels in pulmonary vascular smooth muscle cells: role of nitric oxide.
J Appl Physiol
81:
1264-1272,
1996
29.
Post, JM,
Hume JR,
Archer SL,
and
Weir EK.
Direct role for potassium channel inhibition in hypoxic pulmonary vasoconstriction.
Am J Physiol Cell Physiol
262:
C882-C890,
1992
30.
Reuter, H.
Calcium channel modulation by neurotransmitters, enzymes and drugs.
Nature
301:
569-574,
1983[ISI][Medline].
31.
Robertson, BE,
Schubert R,
Hescheler J,
and
Nelson MT.
cGMP-dependent protein kinase activates Ca-activated K channels in cerebral artery smooth muscle cells.
Am J Physiol Cell Physiol
265:
C299-C303,
1993
32.
Robertson, TP,
Aaronson PI,
and
Ward JPT
Hypoxic vasoconstriction and intracellular Ca2+ in pulmonary arteries: evidence for PKC-independent Ca2+ sensitization.
Am J Physiol Heart Circ Physiol
268:
H301-H307,
1995
33.
Sadoshima, J,
Akaike N,
Kanaide H,
and
Nakamura M.
Cyclic AMP modulates Ca-activated K+ channel in cultured smooth muscle cells of rat aortas.
Am J Physiol Heart Circ Physiol
255:
H754-H759,
1988
34.
Sakai, A,
and
Voelkel NF.
Dibutyryl cyclic adenosine monophosphate inhibits pulmonary vasoconstriction.
Lung
166:
223-231,
1988.
35.
Saqueton, CB,
Miller RB,
Porter VA,
Milla CE,
and
Cornfield DN.
NO causes perinatal pulmonary vasodilation through K+ channel activation and intracellular Ca2+ release.
Am J Physiol Lung Cell Mol Physiol
276:
L925-L932,
1999
36.
Sato, K,
Webb S,
Tucker A,
Rabinovitch M,
O'Brien RF,
McMurtry IF,
and
Stelzner TJ.
Factors influencing the idiopathic development of pulmonary hypertension in the fawn-hooded rat.
Am Rev Respir Dis
145:
793-797,
1992[ISI][Medline].
37.
Schubert, R,
Noack T,
and
Serebryakov VN.
Protein kinase C reduces the Kca current of rat tail artery smooth muscle cells.
Am J Physiol Cell Physiol
276:
C648-C658,
1999
38.
Soria, B.
The biophysical basis of K channel pharmacology.
In: Ion Channel Pharmacology, edited by Soria B,
and Cena V.. New York: Oxford University Press, 1988, p. 167-185.
39.
Stelzner, T,
Hofman TA,
Brown D,
Deng A,
and
Jacob HJ.
Genetic determinants of pulmonary hypertension in fawn-hooded rats.
Chest
6, Suppl 111:
96S,
1997.
40.
Torphy, TJ.
-adrenoceptors, cAMP, and airway smooth muscle relaxation: challenges to the dogma.
Trends Pharmacol Sci
15:
370-374,
1994[ISI][Medline].
41.
Toyoshima, H,
Nasa Y,
Hashizume Y,
Koseki Y,
Isayama Y,
Koshaka Y,
Yamada T,
and
Takeo S.
Modulation of cAMP-mediated vasorelaxation by endothelial nitric oxide and basal cGMP in vascular smooth muscle.
J Cardiovasc Pharmacol
32:
543-551,
1998[ISI][Medline].
42.
Tyler, RC,
Muramatsu M,
Abman SH,
Stelzner TJ,
Rodman DM,
Bloch KD,
and
McMurtry IF.
Variable expression of endothelial NO synthase in three forms of rat pulmonary hypertension.
Am J Physiol Lung Cell Mol Physiol
276:
L297-L303,
1999
43.
White, RE,
Darkow DJ,
and
Lang DL.
Estrogen relaxes coronary arteries by opening BKCa channels through a cGMP-dependent mechanism.
Circ Res
77:
936-942,
1995
44.
White, RE,
Kryman JP,
El-Mowafy AM,
Han G,
and
Carrier GO.
cAMP-dependent vasodilators cross-activate the cGMP-dependent protein kinase to stimulate BKCa channel activity in coronary artery smooth muscle cells.
Circ Res
86:
897-905,
2000
45.
Zamora, MR,
Stelzner TJ,
Webb S,
Panos RJ,
Ruff LJ,
and
Dempsey EC.
Overexpression of endothelin-1 and enhanced growth of pulmonary artery smooth muscle cells from fawn-hooded rats.
Am J Physiol Lung Cell Mol Physiol
270:
L101-L109,
1996