title: nNOS in canine lower esophageal sphincter: colocalized with Cav-1 and Ca2+-handling proteins?
abstract: Immunochemical studies with light microscopy, confocal microscopy, and electron microscopy were used to examine proteins associated with caveolin (Cav) in canine lower esophageal sphincter. The main Cav was Cav-1. It appeared to be colocalized at the cell periphery, in punctate sites, with immunoreactivity to antibodies against different COOH- and NH2-terminal epitopes of neuronal nitric oxide (NO) synthase (nNOS). One COOH-terminal-directed antibody, made in guinea pig, was used to colocalize other immunoreactivities. Those that apparently colocalized with nNOS were L-Ca2+ channels, the PM Ca2+ pump, and, in part, calreticulin and calsequestrin. The large-conductance Ca2+-activated K+ (BK(Ca)) channels were located in discrete peripheral sites, some with Cav. Immunoreactivities not fully colocalized with nNOS were to the sarcoplasmic reticulum Ca2+ pump, connexins 43, 40, and 45, and vinculin. In patch-clamp studies, NO-driven outward currents, mainly through BK(Ca) channels, were inhibited by antibodies to Cav-1 and not by calmodulin and were restored by an NO donor. Several Ca2+-handling molecules are localized at the PM with and/or near Cav. This may allow intracellular calcium concentration levels to be controlled differently than those in the cytosol near caveolae.
authors.affiliation: Department of Medicine, Faculty of Health Sciences, McMaster University, Hamilton, Ontario L8N 3Z5, Canada. edaniel@ualberta.ca
authors.forename: E E
authors.initials: EE
authors.lastname: Daniel
authors.forename: J
authors.initials: J
authors.lastname: Jury
authors.forename: Y F
authors.initials: YF
authors.lastname: Wang
mesh_terms: D000818:Animals; D002135:Calcium-Binding Proteins / Q000032:analysis*; D021941:Caveolae / Q000737:chemistry; D051242:Caveolin 1; D022461:Caveolins / Q000032:analysis*; D018031:Connexin 43 / Q000032:analysis; D004285:Dogs; D004943:Esophagogastric Junction / Q000737:chemistry* / Q000166:cytology; D007150:Immunohistochemistry; D009130:Muscle, Smooth / Q000737:chemistry / Q000166:cytology; D019001:Nitric Oxide Synthase / Q000032:analysis*; D052248:Nitric Oxide Synthase Type I; D018408:Patch-Clamp Techniques; D016596:Vinculin / Q000032:analysis
chemical_list: D002135:Calcium-Binding Proteins; D051242:Caveolin 1; D022461:Caveolins; D018031:Connexin 43; D016596:Vinculin; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I