Department of Physiology, University of Massachusetts Medical School, Worcester, Massachusetts 01655
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In airway smooth muscle
cells (SMCs) from mouse lung slices, 10 µM ATP induced
Ca2+ oscillations that were accompanied by airway
contraction. After ~1 min, the Ca2+ oscillations subsided
and the airway relaxed. By contrast,
0.5 µM adenosine
5'-O-(3-thiotriphosphate) (nonhydrolyzable) induced Ca2+ oscillations in the SMCs and an associated airway
contraction that persisted for >2 min. Adenosine
5'-O-(3-thiotriphosphate)-induced Ca2+
oscillations occurred in the absence of external Ca2+ but
were abolished by the phospholipase C inhibitor U-73122 and the
inositol 1,4,5-trisphosphate receptor inhibitor xestospongin. Adenosine, AMP, and
,
-methylene ATP had no effect on airway caliber, and the magnitude of the contractile response induced by a
variety of nucleotides could be ranked in the following order: ATP = UTP > ADP. These results suggest that the SMC response to ATP
is impaired by ATP hydrolysis and mediated via P2Y2 or
P2Y4 receptors, activating phospholipase C to release
Ca2+ via the inositol 1,4,5-trisphosphate receptor. We
conclude that ATP can serve as a spasmogen of airway SMCs and that
Ca2+ oscillations in SMCs are required to sustain airway contraction.
calcium signaling; confocal microscopy; ATP hydrolysis
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
ASTHMA IS AN AIRWAY DISEASE characterized by a hyperreactive response of airway smooth muscle cells (SMCs) to spasmogens such as histamine, leukotrienes, and perhaps serotonin (2, 6, 25). A major source of histamine and other agonists is an exocytotic release from inflammatory cells (i.e., mast cells and basophils) within the submucosal tissue, in close proximity to the SMCs. However, mast cell degranulation can simultaneously release another potential spasmogen, ATP, which is packaged in the same exocytotic vesicles (23). In addition, airway epithelial cells have also been identified as a source of extracellular ATP. Although the active release mechanisms of ATP from epithelial cells are not understood (5, 7, 11, 36, 37), epithelial trauma associated with asthma (12, 18) can result in damaged cells that passively release ATP.
In many other tissues, ATP has been found to stimulate P2X and P2Y receptors, both of which have been identified in lung tissue (19, 29, 34, 35). P2X receptors are Ca2+ channels that, on binding of ATP, initiate an influx of extracellular Ca2+ into the cell. P2Y receptors are G protein-linked receptors that, on binding of ATP, activate phospholipase C (PLC), and this leads to the hydrolysis of phosphatidylinositol 4,5-bisphosphate to produce inositol 1,4,5-trisphosphate (IP3) and diacylglycerol. IP3 binds to specific receptors (IP3R) within the sarcoplasmic reticulum (SR) to release Ca2+ from the SR (for review see Ref. 4). Because increases in the intracellular free Ca2+ concentration ([Ca2+]i) of SMCs usually result in the phosphorylation of myosin light chain and the stimulation of contraction (15, 16), ATP is likely to be a potent spasmogen of airway SMCs.
Surprisingly, the effects of ATP on the [Ca2+]i of airway SMCs or on airway contraction have not been extensively studied, and the few reports that exist are inconsistent. For example, ATP was found to increase in vivo airway resistance in rats (9) or induce contraction in isolated guinea pig tracheae (8). On the other hand, ATP was reported to relax rabbit tracheal smooth muscle strips (1) and isolated mouse trachea (10, 17) that had been previously contracted with acetylcholine (ACh). These tissue preparations were derived from the large airways and consisted of tracheal rings or strips. As a result, these studies were not able to focus on the responses of individual SMCs. In addition, it is not clear whether results obtained from large airways (e.g., trachea) are applicable to the pathophysiology of smaller airways (e.g., lower bronchi and bronchioles). On the other hand, Michoud et al. (22) studied cultured rat tracheal SMCs and reported that ATP induced a Ca2+ transient followed by a lower, but sustained, Ca2+ elevation. However, the contractile responses of these cells were not investigated. By contrast, ATP-induced Ca2+ signaling is better characterized in vascular SMCs (e.g., from the pulmonary artery), where it was found that ATP could induce a series of Ca2+ oscillations (14, 21, 26).
Consequently, the objective of the present study was to characterize the effect of ATP on the Ca2+ signaling and the associated contractility of airway SMCs. To perform these studies, we have refined a technique to cut thin lung slices (~75 µm thick) and have used these slices to simultaneously measure airway contraction and Ca2+ signaling in single airway SMCs with confocal microscopy (3). The major advantages of lung slices are that the in situ organization of the lung and the contractility of the SMCs are maintained for several days.
With this approach, we found that ATP simultaneously induced Ca2+ oscillations in SMCs and contraction of mouse airways. This response is mediated by P2Y receptors acting via PLC and IP3Rs, but not via P2X receptors and Ca2+ influx. These results highlight the potential role for extracellular ATP in regulating airway caliber and are consistent with the hypothesis that sustained airway contraction is maintained by Ca2+ oscillations within SMCs.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Materials. Cell culture reagents were obtained from Invitrogen Life Technologies-GIBCO (Carlsbad, CA). Other reagents were obtained from Sigma (St. Louis, MO) unless otherwise stated. In most cases, the Hanks' balanced salt solution was supplemented (sHBSS) with HEPES buffer (25 mM) but lacked phenol red.
Lung slices.
Lung slices were cut as previously described (3). Briefly,
male BALB/C inbred mice (Charles River Breeding Labs, Needham, MA;
42-77 days old) were killed by intraperitoneal injection of pentobarbital sodium (Nembutal), and the chest wall was removed. The
trachea was cannulated using an intravenous catheter system (20G
Intima, Becton Dickinson, Sandy, UT), and the lungs were inflated with
2% agarose-sHBSS at 37°C (~1 ml). Subsequently, 0.1-0.2 ml of
air was injected to flush the agarose-sHBSS out of the airways. To
stiffen the soft lung tissue for sectioning, the agarose was gelled at
4°C. The lungs were removed, and ~75-µm-thick slices were cut
with a tissue slicer (model EMS-4000, Electron Microscopy Sciences,
Fort Washington, PA). The slices were maintained by flotation in DMEM
supplemented with 10% fetal bovine serum and antibiotics and
antimycotics at 37°C in 10% CO2 for up to 5 days.
Immunohistochemistry was performed using monoclonal mouse anti--actin antibodies and FITC-conjugated anti-mouse IgG antibodies (for details see Ref. 3).
Measurement of airway contraction. Experiments were performed on airways with a lumen that was free of agarose and lined by epithelial cells showing ciliary activity. For all experiments, the slices (~7,000-8,000 µm long × 3,000-4,000 µm wide × 75 µm thick) and were bathed in 150 µl of sHBSS. The cross-sectional area of the airways was 29,989 ± 1,885 (SE) µm2 (n = 104), and there were no significant differences between groups of airways used for different experiments. The slices were placed on cover glasses within a custom-made Plexiglas chamber and held in position by a piece of nylon mesh (CMN-300-B, Small Parts, Miami Lakes, FL). Phase-contrast images were recorded using a digital charge-coupled device camera (model TM-6710, Pulnix America, Sunnyvale, CA), a digital camera interface (Road Runner, BitFlow, Woburn, MA), and image acquisition software (Video Savant, IO Industries, London, ON, Canada). Frames were captured in time lapse (1 frame/s), and the cross-sectional area of the airway was measured with respect to time by pixel summing using the image analysis software Scion (Scion, Frederick, MD; download available at no cost at www.scioncorp.com).
Measurements of [Ca2+]i. Slices were loaded with the Ca2+ indicator dye Oregon green (20 µM; Molecular Probes, Eugene, OR) for 1 h (for details see Ref. 3), and confocal microscopy was performed using a custom-built microscope (31). Images were recorded in time lapse (2 frames/s). Regions of interest of 8.4 × 8.4 µm were defined in single SMCs, and the average fluorescence intensities of each region of interest were obtained. Bleach correction was calculated from the bleaching rate 15-30 s before the addition of drugs. Data were corrected with the bleach correction before a rolling average of three frames was calculated. Final fluorescence values are expressed as a fluorescence ratio (F/F0) normalized to the initial fluorescence (F0). Simultaneous airway contraction was analyzed by measuring the area of the part of the airway lumen that was visible in the confocal images.
Drug application.
ATP, adenosine 5'-O-(3-thiotriphosphate) (ATPS), UTP,
ADP, AMP,
,
-methylene ATP, and adenosine were dissolved in sHBSS. A 10× concentrated solution was added to the volume of the bath solution to be diluted to the final working concentration. Xestospongin (Calbiochem, La Jolla, CA), U-73122, U-73433, and indomethacin were
dissolved in dimethyl sulfoxide and diluted in sHBSS to the final
concentrations used. Final dimethyl sulfoxide concentrations were
0.1% for indomethacin, 0.05% for xestospongin, and 2.5% for U-73122
and U-73433.
Statistics. Statistical analysis was performed using the t-test and the Kruskal-Wallis one-way analysis of variance on ranks; for all pairwise multiple comparisons, the Student-Newman-Keuls method was used. Values are means ± SE. P < 0.05 was considered to be statistically significant.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
ATP-induced airway contraction.
The addition of 10-100 µM ATP initiated airway contraction
(Figs. 1 and
2). This airway contraction was followed
by relaxation of the airway in the continued presence of ATP
[relaxation began 61 ± 23 (SE) s after addition of 100 µM ATP,
n = 6; Fig. 2A]. To distinguish whether the
hydrolysis of ATP, presumably by ectonucleotidases, or receptor
accommodation was responsible for the brevity of the response to ATP,
we tested the effect of the nonhydrolyzable ATP analog ATPS on
airway contraction. At
500 nM, ATP
S induced contraction (Fig.
2B) that was maintained for a significantly longer time than
that induced by ATP (relaxation began 164 ± 5 s after
addition of 100 µM ATP
S, n = 6, P < 0.002). Airway contraction was dependent on ATP concentration in the
range 10-100 µM (Fig. 2C). A decrease in lumen size
of 10.9 ± 1.4% (n = 6) of the initial cross-sectional area occurred in response to 100 µM ATP. A similar concentration dependence of airway contraction was induced by 0.5-100 µM ATP
S (Fig. 2C). However, the airways
responded more strongly to ATP
S than to ATP, and a decrease of
~10% in the lumen area was induced by ~1 µM ATP
S compared
with 100 µM ATP. This greater sensitivity to ATP
S resulted in a
leftward shift of the concentration-dependence curve compared with ATP.
A near-maximal reduction in lumen area of 14.6 ± 5.1%
(n = 6) was induced by 100 µM ATP
S.
|
|
|
|
|
ATP-induced Ca2+ signaling in airway
SMCs.
The Ca2+ response of the SMCs to ATP consisted of a brief
burst of Ca2+ oscillations (Fig.
6A). Because of the small
number of oscillations that occurred, their frequency was difficult to
accurately calculate. However, the number of oscillations was dependent
on the ATP concentration: 1.25 ± 0.25 and 10.9 ± 1.4 oscillations in response to 10 and 100 µM ATP, respectively
(n = 6). The initial [Ca2+]i
increase associated with the ATP-induced oscillations, defined as the
percent change in the F/F0 measured immediately before and
at the peak of the initial Ca2+ oscillation, also increased
with concentration (Fig. 6D).
|
|
Correlation between ATP-induced airway contraction and Ca2+ signaling. The Ca2+ signaling in the SMCs was correlated with airway contraction by analysis of the changes that occurred in the cross-sectional area of the airway lumen that were visible in the confocal images simultaneously with the changes in SMC fluorescence. The change in lumen size is given in absolute units, rather than as a percentage of the starting lumen area. This data representation is necessary because, at the magnifications required to observe the changes in [Ca2+]i of single SMCs, only a part of the area of the airway lumen is visible in the confocal images. Furthermore, because the size of this area can vary depending on the tissue morphology and alignment, reporting the change in area as a percentage of the starting area may misrepresent the size of the airway contraction.
As described earlier, 100 µM ATP induced a small number of Ca2+ oscillations that terminated after ~1 min (Fig. 8A). The initial Ca2+ oscillation was associated with the initiation of airway contraction. The following two to three Ca2+ oscillations correlated with the maintenance of contraction. However, with the subsequent decline in magnitude and cessation of the ATP-induced Ca2+ oscillations, a simultaneous relaxation of the airway was observed. By contrast, ATP
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
We present a versatile airway preparation, the lung slice, which
we have used in conjunction with confocal microscopy to simultaneously record Ca2+ signaling in SMCs and the associated airway
contraction, a measure of the integrated activity of airway SMCs. We
have found that ATP and ATPS induced Ca2+ oscillations
in airway SMCs and that these Ca2+ oscillations correlated
with the initiation and maintenance of airway contraction. Conversely,
when the Ca2+ oscillations subsided, the airways relaxed.
ACh-induced Ca2+ oscillations in airway SMCs have been reported by several groups (20, 27, 28, 30, 32, 33), and Pabelick et al. (24) assumed that ACh-induced Ca2+ oscillations may function to regulate global [Ca2+]i in airway SMCs and, thereby, regulate airway contraction. In our previous study reporting ACh-induced Ca2+ oscillations in airway SMCs in lung slices (3), we demonstrated that the Ca2+ oscillations occurred during the maintenance of airway contraction and that whenever the Ca2+ oscillations were abolished by a variety of approaches, the airways relaxed. As a result, we hypothesized that ACh-induced Ca2+ oscillations maintain airway contraction. Our findings with ATP in the present study support and extend this hypothesis by demonstrating that this principle is not limited to a single agonist (e.g., ACh) but also applies to ATP. As a result, the maintenance of contraction by Ca2+ oscillations may be a general principle of airway SMCs in response to agonists.
The maximal frequencies of ACh-induced Ca2+ oscillations
(~20/min) (3) were higher than the frequencies of the
ATPS-induced Ca2+ oscillations (~8/min), and the
corresponding maximal contraction induced was also higher (~20 and
~14% of starting area for ACh and ATP
S, respectively). This
suggests that higher frequencies are required to maintain stronger
airway contraction. This relationship between Ca2+
oscillation frequency and airway contraction supports the idea that
airway contraction is generated or regulated in a frequency-modulated manner.
Although ATP induced airway contraction, the airways relaxed after ~1
min. Extracellular ATP is commonly subject to hydrolysis by
ectonucleotidases (for review see Ref. 38), and Gerwins
and Fredholm (13) found that, in a cultured SMC line,
hydrolysis could reduce ATP concentrations to <50% of starting value
within a few minutes. Lung slices contain a variety of cell types that are in close proximity to the SMCs, making it likely that
ectonucleotidase activity is present. Consistent with this idea is our
finding that lower concentrations of the nonhydrolyzable ATP analog
ATPS induced airway contraction that was sustained for longer
periods than that induced by ATP. Consequently, we believe that the
relaxation of the airway after the addition of ATP results primarily
from a degradation of ATP. This proposed hydrolysis of ATP would be expected to release adenosine, AMP, and ADP, but none of these had any
relaxing effect on airway size. This is consistent with the idea that
airway relaxation resulted from a reduction of ATP concentration.
Alternatively, it is possible that a complex mixture of ATP and ATP
metabolites might have a relaxant effect on airways and that this
contributed to the brevity of the ATP-induced contraction. Another
explanation for the airway relaxation is receptor desensitization. Although it was found that ATP
S induced contraction for a longer period than ATP, the contractile response to ATP
S subsided after ~10 min. If it is assumed that the ATP
S is still present,
relaxation could be explained by the fact that the P2Y receptors or
another intracellular site became desensitized (29). This
would also infer that the relaxation of airways after ATP exposure
resulted from ATP degradation and receptor accommodation.
The nucleotide receptor agonist ,
-methylene ATP is selective for
P2X over P2Y receptors (29) but had only marginally
contractile effects on airways. In addition, P2X receptors are
Ca2+ channels leading to Ca2+ influx on binding
of ATP (19, 29), but in Ca2+-free solution,
ATP
S still induced Ca2+ oscillations with a magnitude
and frequency comparable to that in the presence of external
Ca2+. The effects of ATP on airways are therefore likely to
be mediated by P2Y, rather than P2X, receptors. Given the order of
potency for inducing airway contraction (ATP = UTP > ADP),
the receptor subtypes P2Y2 and P2Y4 would be
predicted. In addition, because ADP induced a slight contraction,
P2Y1 receptors may also be present, but a small contractile
response could also be mediated via P2Y2 or
P2Y4 receptors. P2Y receptors are coupled to G proteins and increase intracellular IP3 levels via activation of PLC
(29, 35). This is consistent with our data, which indicate
a need for PLC activity and IP3Rs for the initiation of
Ca2+ oscillations. Therefore, we propose that ATP-induced
Ca2+ signaling requires an activation of PLC by G
protein-coupled P2Y2 or P2Y4 receptors, leading
to increased formation of IP3 and release of
Ca2+ from the SR via IP3Rs.
ATP, acting via prostaglandins released from epithelial cells, has been
previously reported to extensively relax perfused tracheae that were
precontracted with 10 µM ACh (1, 10, 17). However, in
this study, airways fully contracted with 10 µM ACh or airways partly
contracted with 100 nM ACh [~50% of their maximal contraction
(3)] showed no response or very little response to 100 µM ATP or ATPS. ATP probably did not induce further contraction, because ACh and ATP utilize similar signal transduction pathways, and
the stimulation by ACh exceeded the stimulation induced by ATP. The
small relaxing effect of ATP could be abolished with indomethacin, and
this is consistent with the hypothesis that cyclooxygenase products may
be released from epithelial cells. We also found that PGE2
had a relaxing effect on precontracted airways, but this effect was
reversible by ATP
S, a result indicating that the contraction induced
by ATP
S was sufficient to counter the relaxing effect of
PGE2. Because there was no difference in ATP-induced airway
contraction in the presence or absence of indomethacin, we believe that
neither ATP nor its metabolites release sufficient quantities of
prostaglandins to influence the ATP-induced airway contraction. We
therefore conclude that ATP primarily functions to contract airways in
mouse lung slices.
ATP-induced changes of [Ca2+]i have been investigated in several types of SMCs, and ATP-induced Ca2+ oscillations have been observed, for example, in vascular SMCs (14, 21, 26). In contrast, little is known about ATP-induced Ca2+ signaling in airway SMCs. Michoud et al. (22) reported a Ca2+ transient followed by a Ca2+ plateau in response to ATP in cultured rat tracheal SMCs. Besides differences in species and tissue preparation, the study averaged the fluorescence from ~20 cells, and this probably prevented the observation of Ca2+ oscillations.
In summary, we have demonstrated that ATP induces airway contraction and Ca2+ oscillations in mouse airway SMCs. We believe that these responses are mediated via P2Y receptors to stimulate Ca2+ oscillations that rely on PLC activity and IP3Rs, but not on Ca2+ influx. The strength of the response appears to be attenuated by hydrolysis of ATP. Also, in view of the fact that the airways remained contracted whenever Ca2+ oscillations were occurring in the SMCs, we provide further evidence that airway contraction is maintained by Ca2+ oscillations. These studies suggest that ATP is a potential spasmogen of airway SMCs and that it may play a role in the regulation of airway caliber and in the pathogenesis of airway disease.
![]() |
ACKNOWLEDGEMENTS |
---|
This work was supported by National Heart, Lung, and Blood Institute Grant HL-49288.
![]() |
FOOTNOTES |
---|
Address for reprint requests and other correspondence: M. J. Sanderson, Dept. of Physiology, University of Massachusetts Medical School, 55 Lake Ave. North, Worcester, MA 01655 (E-mail: michael.sanderson{at}umassmed.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.
August 2, 2002;10.1152/ajplung.00139.2002
Received 7 May 2002; accepted in final form 31 July 2002.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Aksoy, MO,
and
Kelsen SG.
Relaxation of rabbit tracheal smooth muscle by adenine nucleotides: mediation by P2 purinoceptors.
Am J Respir Cell Mol Biol
10:
230-236,
1994[Abstract].
2.
Barnes, PJ.
Histamine and serotonin.
Pulm Pharmacol Ther
14:
329-339,
2001[ISI][Medline].
3.
Bergner, A,
and
Sanderson MJ.
Acetylcholine-induced calcium signaling and contraction of airway smooth muscle cells in lung slices.
J Gen Physiol
119:
187-198,
2002
4.
Bootman, MD,
Collins TJ,
Peppiatt CM,
Prothero LS,
MacKenzie L,
De Smet P,
Travers M,
Tovey SC,
Seo JT,
Berridge MJ,
Ciccolini F,
and
Lipp P.
Calcium signallingan overview.
Semin Cell Dev Biol
12:
3-10,
2001[ISI][Medline].
5.
Braunstein, GM,
Roman RM,
Clancy JP,
Kudlow BA,
Taylor AL,
Shylonsky VG,
Jovov B,
Peter K,
Jilling T,
Ismailov I,
Benos DJ,
Schwiebert LM,
Fitz JG,
and
Schwiebert EM.
Cystic fibrosis transmembrane conductance regulator facilitates ATP release by stimulating a separate ATP release channel for autocrine control of cell volume regulation.
J Biol Chem
276:
6621-6630,
2001
6.
Cazzola, I,
and
Matera MG.
5-HT modifiers as a potential treatment of asthma.
Trends Pharmacol Sci
21:
13-16,
2000[ISI][Medline].
7.
Donaldson, SH,
Lazarowski ER,
Picher M,
Knowles MR,
Stutts MJ,
and
Boucher RC.
Basal nucleotide levels, release, and metabolism in normal and cystic fibrosis airways.
Mol Med
6:
969-982,
2000[ISI][Medline].
8.
Fedan, JS,
Belt JJ,
Yuan LX,
and
Frazer DG.
Contractile effects of nucleotides in guinea pig isolated, perfused trachea: involvement of respiratory epithelium, prostanoids and Na+ and Cl channels.
J Pharmacol Exp Ther
264:
210-216,
1993[Abstract].
9.
Flezar, M,
Olivenstein R,
Cantin A,
and
Heisler S.
Extracellular ATP stimulates elastase secretion from human neutrophils and increases lung resistance and secretion from normal rat airways after intratracheal instillation.
Can J Physiol Pharmacol
70:
1065-1068,
1992[ISI][Medline].
10.
Fortner, CN,
Breyer RM,
and
Paul RJ.
EP2 receptors mediate airway relaxation to substance P, ATP, and PGE2.
Am J Physiol Lung Cell Mol Physiol
281:
L469-L474,
2001
11.
Foskett, JK.
ClC and CFTR chloride channel gating.
Annu Rev Physiol
60:
689-717,
1998[ISI][Medline].
12.
Gaillard, D,
Jouet JB,
Egreteau L,
Plotkowski L,
Zahm JM,
Benali R,
Pierrot D,
and
Puchelle E.
Airway epithelial damage and inflammation in children with recurrent bronchitis.
Am J Respir Crit Care Med
150:
810-817,
1994[Abstract].
13.
Gerwins, P,
and
Fredholm BB.
ATP and its metabolite adenosine act synergistically to mobilize intracellular calcium via the formation of inositol 1,4,5-trisphosphate in a smooth muscle cell line.
J Biol Chem
267:
16081-16087,
1992
14.
Guibert, C,
Pacaud P,
Loirand G,
Marthan R,
and
Savineau JP.
Effect of extracellular ATP on cytosolic Ca2+ concentration in rat pulmonary artery myocytes.
Am J Physiol Lung Cell Mol Physiol
271:
L450-L458,
1996
15.
Gunst, SJ,
and
Tang DD.
The contractile apparatus and mechanical properties of airway smooth muscle.
Eur Respir J
15:
600-616,
2000
16.
Hall, IP.
Second messengers, ion channels and pharmacology of airway smooth muscle.
Eur Respir J
15:
1120-1127,
2000
17.
Kao, J,
Fortner CN,
Liu LH,
Shull GE,
and
Paul RJ.
Ablation of the SERCA3 gene alters epithelium-dependent relaxation in mouse tracheal smooth muscle.
Am J Physiol Lung Cell Mol Physiol
277:
L264-L270,
1999
18.
Laitinen, LA,
Heino M,
Laitinen A,
Kava T,
and
Haahtela T.
Damage of the airway epithelium and bronchial reactivity in patients with asthma.
Am Rev Respir Dis
131:
599-606,
1985[ISI][Medline].
19.
Lambrecht, G.
Agonists and antagonists acting at P2X receptors: selectivity profiles and functional implications.
Naunyn Schmiedebergs Arch Pharmacol
362:
340-350,
2000[ISI][Medline].
20.
Liu, X,
and
Farley JM.
Acetylcholine-induced chloride current oscillations in swine tracheal smooth muscle cells.
J Pharmacol Exp Ther
276:
178-186,
1996[Abstract].
21.
Mahoney, MG,
Randall CJ,
Linderman JJ,
Gross DJ,
and
Slakey LL.
Independent pathways regulate the cytosolic [Ca2+]i initial transient and subsequent oscillations in individual cultured arterial smooth muscle cells responding to extracellular ATP.
Mol Biol Cell
3:
493-505,
1992[Abstract].
22.
Michoud, MC,
Tolloczko B,
and
Martin JG.
Effects of purine nucleotides and nucleosides on cytosolic calcium levels in rat tracheal smooth muscle cells.
Am J Respir Cell Mol Biol
16:
199-205,
1997[Abstract].
23.
Osipchuk, Y,
and
Cahalan M.
Cell-to-cell spread of calcium signals mediated by ATP receptors in mast cells.
Nature
359:
241-244,
1992[ISI][Medline].
24.
Pabelick, CM,
Sieck GC,
and
Prakash YS.
Significance of spatial and temporal heterogeneity of calcium transients in smooth muscle.
J Appl Physiol
91:
488-496,
2001
25.
Page, S,
Ammit AJ,
Black JL,
and
Armour CL.
Human mast cell and airway smooth muscle cell interactions: implications for asthma.
Am J Physiol Lung Cell Mol Physiol
281:
L1313-L1323,
2001
26.
Pauvert, O,
Marthan R,
and
Savineau J.
NO-induced modulation of calcium oscillations in pulmonary vascular smooth muscle.
Cell Calcium
27:
329-338,
2000[ISI][Medline].
27.
Prakash, YS,
Kannan MS,
and
Sieck GC.
Regulation of intracellular calcium oscillations in porcine tracheal smooth muscle cells.
Am J Physiol Cell Physiol
272:
C966-C975,
1997
28.
Prakash, YS,
Pabelick CM,
Kannan MS,
and
Sieck GC.
Spatial and temporal aspects of ACh-induced [Ca2+]i oscillations in porcine tracheal smooth muscle.
Cell Calcium
27:
153-162,
2000[ISI][Medline].
29.
Ralevic, V,
and
Burnstock G.
Receptors for purines and pyrimidines.
Pharmacol Rev
50:
413-492,
1998
30.
Roux, E,
Guibert C,
Savineau JP,
and
Marthan R.
[Ca2+]i oscillations induced by muscarinic stimulation in airway smooth muscle cells: receptor subtypes and correlation with the mechanical activity.
Br J Pharmacol
120:
1294-1301,
1997[Abstract].
31.
Sanderson MJ and Parker I. Video-rate confocal microscopy. In:
Methods in Enzymology, edited by Marriott G and Parker
I. In press.
32.
Sieck, GC,
Kannan MS,
and
Prakash YS.
Heterogeneity in dynamic regulation of intracellular calcium in airway smooth muscle cells.
Can J Physiol Pharmacol
75:
878-888,
1997[ISI][Medline].
33.
Sims, SM,
Jiao Y,
and
Zheng ZG.
Intracellular calcium stores in isolated tracheal smooth muscle cells.
Am J Physiol Lung Cell Mol Physiol
271:
L300-L309,
1996
34.
Suarez-Huerta, N,
Pouillon V,
Boeynaems J,
and
Robaye B.
Molecular cloning and characterization of the mouse P2Y4 nucleotide receptor.
Eur J Pharmacol
416:
197-202,
2001[ISI][Medline].
35.
Von Kugelgen, I,
and
Wetter A.
Molecular pharmacology of P2Y receptors.
Naunyn Schmiedebergs Arch Pharmacol
362:
310-323,
2000[ISI][Medline].
36.
Walsh, DE,
Harvey BJ,
and
Urbach V.
CFTR regulation of intracellular calcium in normal and cystic fibrosis human airway epithelia.
J Membr Biol
177:
209-219,
2000[ISI][Medline].
37.
Watt, WC,
Lazarowski ER,
and
Boucher RC.
Cystic fibrosis transmembrane regulator-independent release of ATP: its implications for the regulation of P2Y2 receptors in airway epithelia.
J Biol Chem
273:
14053-14058,
1998
38.
Zimmermann, H.
Extracellular metabolism of ATP and other nucleotides.
Naunyn Schmiedebergs Arch Pharmacol
362:
299-309,
2000[ISI][Medline].