| pA2
online © Copyright 2003 The British Pharmacological Society |
033P
University
of Manchester Autumn Meeting September 2003 |
|
Evidence
for the involvement of the two-pore domain k+
channel task-1 in setting resting membrane potential and resting
tension in rat small pulmonary arteries |
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The pulmonary circulation is a low pressure high flow system that responds to acidic challenge and hypoxia with constriction. The resting membrane potential (EM) of isolated pulmonary artery myocytes is approximately -55mV. However, although evidence implicates voltage-gated potassium (K+) channels in setting pulmonary artery myocyte EM and in the response to hypoxia (Coppock et al., 2001), channel kinetics suggest they would be inactive at rest; thus their contribution to EM maintenance remains doubtful. However, the TASK family of two-pore domain (2P) K+ channels whose members are sensitive to extracellular pH and may be involved in setting EM are more likely mediators of this response(Lesage & Lazdunski, 2000). In the present study we demonstrate functional evidence for the presence of 2P K+ channels in rat pulmonary artery myocytes.
Male
Sprague-Dawley rats (175-225g) were killed by stunning and cervical dislocation.
Rat small pulmonary arteries were dissected on ice. Microelectrode:
Intact small pulmonary arteries were superfused (10ml.min-1)
with HEPES-buffered Tyrode at pH 7.4, 6.4 or 8.4 (37°C - adjusted
with NaOH or HCl). Myocytes were impaled from the adventitial side, using
microelectrodes with a resistance of 50-80M
and filled with 3M KCl. Myography: Small pulmonary arteries were
mounted on a wire myograph and tensioned to an equivalent transmural pressure
of 30mm Hg. Vessels bathed in Krebs (37°C) were gassed with 21% O2/5%
CO2. Application of anandamide (0.3-30µM),
bupivacaine (0.3-300µM) or clofilium (0.3-100µM) were used
to assess the role of 2P K+ channels
in determining resting tension.
Switching from control (pH 7.4) HEPES-buffered Tyrode to alkaline Tyrode significantly hyperpolarised EM (-51.2±0.4 to -60.4±1.3mV at pH 8.4, P <0.01). Exchange of control to acidic (pH 6.4) HEPES-buffered n=4; E=Eese Tyrode resulted in a significant depolarisation of EM (-43.6±1.3Mv; P < 0.01). Anandamide, an inhibitor of TASK-1 (Maingret et al., 2001), completely and reversibly increased EM in small pulmonary arteries maintained in alkaline HEPES-buffered Tyrode equivalent to that seen in pulmonary arteries in acidic Tyrode (pH 8.4 plus anandamide -41.9±5.1mV). Anandamide and bupivacaine significantly increased resting isometric tension in isolated small pulmonary arteries (P < 0.05), whilst clofilium an inhibitor of TASK-2 (Niemeyer et al., 2001) was without effect.
TASK-1, TASK-2 and TASK-3 are modulated by changes in pH (range 6.4-8.4) with acidic pH resulting in channel closure (O'Connell et al., 2002).We have previously demonstrated the presence of 2P K+ channels including TASK-1 and TASK-2 in rat pulmonary arteries, while TASK-3 channels were absent (Johnson et al., 2002). The increase in EM by anandamide and acidic pH, coupled with the increase in resting tension elicited by anandamide and bupivacaine suggests that TASK-1 has a functional role in rat pulmonary arteries. The lack of effect of clofilium on resting tension, coupled with apparent low levels of expression of TASK-2 suggest this channel contributes little to resting tension.
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E.A., et al. (2001). Am J Physiol Lung Cell Mol Physiol,
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