Reverse-mode NCX current in mouse airway smooth muscle: Na(+)  and voltage dependence, contributions to Ca(2+)  influx and contraction, and altered expression in a model of allergen-induced hyperresponsiveness.
OBJECTIVE
We examined the electrophysiological properties of reverse-mode Na(+) /Ca(2+)  exchange (NCX) in mouse airway smooth muscle (ASM), assessing its contributions to regulation of [Ca(2+) ], and its expression in acute and chronic airway hyperresponsiveness (AHR).


METHODS
Membrane currents were studied in single murine ASM cells under voltage clamp at -60 mV using ramp depolarizing commands to +80 mV. Confocal fluorimetric and RT-PCR techniques were used to monitor changes in cytosolic [Ca(2+) ] and NCX expression, respectively.


RESULTS
With standard KCl-containing electrode, 30 μm KB-R7943 (an inhibitor of reverse-mode NCX activity) exhibited variable effects on membrane current, indicating modulation of more than one conductance. KB-R7943 activated outwardly rectifying current that was inhibited by 100 μm iberiotoxin (blocker of large-conductance Ca(2+) -dependent K(+)  channels), indicating a direct enhancing effect of KB-R7943 on those K(+)  channels. After obviating K(+)  currents, we found that a current sensitive to 4-4'-diisothiocyanostilbene-2,2'-disulfonic acid (blocker of Ca(2+) -dependent Cl- channels) was markedly increased by elevating [Na(+) ] in the electrode solution to 13, 15.5 and 18 mm and suppressed by KB-R7943, indicating Ca(2+)  influx via reverse-mode NCX activity. With conditions preventing Ca(2+)  influx through voltage-dependent Ca(2+)  channels but promoting that through NCX, we found that introduction of Ca(2+)  led to marked but transient KB-R7943-sensitive elevation of [Ca(2+) ]. Additionally, KB-R7943 suppressed cholinergically evoked Ca(2+)  waves. Finally, NCX1 expression was not significantly changed in allergen-induced AHR acute model but increased approx. 2.5-fold in a chronic model.


CONCLUSIONS
Reverse-mode NCX activity leads to a physiologically relevant increase in [Ca(2+) ] even under control conditions, and this may be exaggerated in allergen-induced AHR and asthma.
Firestone Institute for Respiratory Health, St. Joseph's Hospital, Department of Medicine, McMaster University, Hamilton, ON, Canada. mrahman@mcmaster.ca
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Rahman
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Inman
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Kiss
L J
LJ
Janssen
D000485:Allergens; D000818:Animals; D039741:Antigens, Dermatophagoides; D001980:Bronchi / Q000502:physiology; D016535:Bronchial Hyperreactivity / Q000276:immunology / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D005260:Female; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D010455:Peptides / Q000494:pharmacology; D024681:Potassium Channels, Calcium-Activated / Q000037:antagonists & inhibitors; D019831:Sodium-Calcium Exchanger / Q000378:metabolism*; D013890:Thiourea / Q000031:analogs & derivatives / Q000494:pharmacology; D014132:Trachea / Q000502:physiology
C101670:2-(2-(4-(4-nitrobenzyloxy)phenyl)ethyl)isothiourea methanesulfonate; D000485:Allergens; D039741:Antigens, Dermatophagoides; D010455:Peptides; D024681:Potassium Channels, Calcium-Activated; D019831:Sodium-Calcium Exchanger; C064719:iberiotoxin; D013890:Thiourea; D002118:Calcium