title: Temporal and spatial dynamics underlying capacitative calcium entry in human colonic smooth muscle.
abstract: Following smooth muscle excitation and contraction, depletion of intracellular Ca(2+) stores activates capacitative Ca(2+) entry (CCE) to replenish stores and sustain cytoplasmic Ca(2+) (Ca(2+)(i)) elevations. The objectives of the present study were to characterize CCE and the Ca(2+)(i) dynamics underlying human colonic smooth muscle contraction by using tension recordings, fluorescent Ca(2+)-indicator dyes, and patch-clamp electrophysiology. The neurotransmitter acetylcholine (ACh) contracted tissue strips and, in freshly isolated colonic smooth muscle cells (SMCs), caused elevation of Ca(2+)(i) as well as activation of nonselective cation currents. To deplete Ca(2+)(i) stores, the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) inhibitors thapsigargin and cyclopiazonic acid were added to a Ca(2+)-free bathing solution. Under these conditions, addition of extracellular Ca(2+) (3 mM) elicited increased tension that was inhibited by the cation channel blockers SKF-96365 (10 microM) and lanthanum (100 microM), suggestive of CCE. In a separate series of experiments on isolated SMCs, SERCA inhibition generated a gradual and sustained inward current. When combined with high-speed Ca(2+)-imaging techniques, the CCE-evoked rise of Ca(2+)(i) was associated with inward currents carrying Ca(2+) that were inhibited by SKF-96365. Regional specializations in Ca(2+) influx and handling during CCE were observed. Distinct "hotspot" regions of Ca(2+) rise and plateau were evident in 70% of cells, a feature not previously recognized in smooth muscle. We propose that store-operated Ca(2+) entry occurs in hotspots contributing to localized Ca(2+) elevations in human colonic smooth muscle.
authors.affiliation: Department of Physiology and Pharmacology, Schulich School of Medicine and Dentistry, The University of Western Ontario, London, ON N6A 5C1, Canada.
authors.forename: Jason R
authors.initials: JR
authors.lastname: Kovac
authors.forename: Tom
authors.initials: T
authors.lastname: Chrones
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Sims
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000368:Aged; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D018678:Cholinergic Agents / Q000494:pharmacology; D003106:Colon / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007811:Lanthanum / Q000494:pharmacology; D008297:Male; D008564:Membrane Potentials; D008856:Microscopy, Fluorescence; D009119:Muscle Contraction / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors
chemical_list: D002121:Calcium Channel Blockers; D018678:Cholinergic Agents; D004791:Enzyme Inhibitors; D007093:Imidazoles; D007211:Indoles; D019284:Thapsigargin; D007811:Lanthanum; D000252:Calcium-Transporting ATPases; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; D000109:Acetylcholine; D002118:Calcium; C000543:cyclopiazonic acid