Two distinct membrane currents activated by cyclopiazonic acid‐induced calcium store depletion in single smooth muscle cells of the mouse anococcygeus

• Published in: British journal of pharmacology 1996-02, Vol.117 (3), p.566-572
• Main Authors: Wayman, Christopher P., McFadzean, Ian, Gibson, Alan, Tucker, John F.
• Format: Article
• Language: English
• Subjects: Animals; Anococcygeus (mouse); Biological and medical sciences; caffeine; Caffeine - pharmacology; Calcium - metabolism; Calcium - physiology; Calcium Channel Blockers - pharmacology; calcium store depletion; Calcium-Transporting ATPases - antagonists & inhibitors; calcium‐dependent chloride current; Cell membranes. Ionic channels. Membrane pores; Cell structures and functions; Chloride Channels - drug effects; Chloride Channels - metabolism; cyclopiazonic acid; Electrophysiology; Enzyme Inhibitors - pharmacology; Fundamental and applied biological sciences. Psychology; In Vitro Techniques; Indoles - pharmacology; Male; Membrane Potentials - drug effects; Mice; Molecular and cellular biology; Muscle Contraction - drug effects; Muscle, Smooth - drug effects; Muscle, Smooth - enzymology; Muscle, Smooth - metabolism; non‐selective cation current; Patch-Clamp Techniques; Phosphodiesterase Inhibitors - pharmacology; whole‐cell patch‐clamp
• ISSN: 0007-1188; 1476-5381
• DOI: 10.1111/j.1476-5381.1996.tb15228.x

1 By use of the whole‐cell configuration of the patch‐clamp technique, membrane currents induced by cyclopiazonic acid (CPA; an inhibitor of the sarcoplasmic reticulum (SR) calcium‐ATPase) were investigated in single smooth muscle cells freshly dispersed from the mouse anococcygeus. Voltage‐dependent calcium currents were blocked with extracellular nifedipine and caesium and tetraethylammonium chloride were used to block voltage‐dependent potassium currents. 2 At a holding potential of −40 mV, CPA (10 μm) activated an inward current that consisted of two distinct components. The first was an initial transient current with an amplitude of 19.6±1.9 pA while the second was sustained and had an amplitude of 3.5±0.3 pA. 3 The current‐voltage (I‐V) relationship for the transient current showed marked outward rectification. The current had a reversal potential of 9.1±1.1 mV which was shifted to 29.0±4.2 mV when the extracellular chloride concentration was lowered from 148.4 to 58.4 mM. The sustained current had a near‐linear I‐V relationship and a reversal potential of 31.0±2.7 mV. Removal of extracellular calcium had no effect on the transient current, but shifted the reversal potential of the sustained current to 18.2±5.7 mV. 3 The initial transient current was abolished in cells bathed in extracellular solutions containing the chloride channel blockers, 4,4′ diisothiocyanato‐stilbene‐2,2′‐disulphonic acid (DIDS; 1 mM) or anthracene‐9‐carboxylic acid (A‐9‐C; 1 mM), and was absent in cells containing the calcium buffers EGTA (1 to 5 mM) or BAPTA (10 mM). The second sustained current was unaffected by either the chloride channel blockers or the intracellular calcium buffers. 4 Treatment of the cells with caffeine (10 mM) produced similar inward currents to those produced by CPA. In the presence of caffeine, CPA (10 μm) induced no further inward current. 5 In organ bath studies, CPA (10 μm)‐induced contractions of the mouse anococcygeus were inhibited by cadmium and nickel (both 50–400 μm) and the general calcium entry blocker, SKF 96365 (10 μm); lanthanum and gadolinium had no effect at concentrations up to 400 μm. The pharmacology of the CPA‐induced non‐selective cation current mirrored that of the CPA‐induced whole muscle contraction being reversed by cadmium (100 μm) and SKF 96365 (10 μm), but unaffected by lanthanum (400 μm). The initial chloride conductance was unaffected by cadmium, SKF 96365 or lanthanum. 6 It is concluded that CPA activates a transient ca