title: Elementary purinergic Ca2+ transients evoked by nerve stimulation in rat urinary bladder smooth muscle.
abstract: The translation of nerve transmission to Ca2+ signals in urinary bladder smooth muscle (UBSM) is incompletely understood. Thus, we sought to characterize Ca2+ signals in strips of UBSM loaded with the Ca2+-sensitive fluorescent dye, fluo-4, using laser scanning confocal microscopy. Two types of Ca2+ signals occurred spontaneously and could be evoked with field stimulation: large, rapid, global Ca2+ transients termed 'global Ca2+ flashes', and much smaller, localized Ca2+ transients. Global Ca2+ flashes were inhibited by the L-type voltage-dependent Ca2+ channel (VDCC) inhibitor, diltiazem and with P2X receptor blockade. Simultaneous intracellular recordings and Ca2+ measurements indicated that these events are caused by Ca2+ influx through VDCCs during action potentials. Small, local Ca2+ transients occurred spontaneously, and their frequency could be elevated with field stimulation. Atropine, an inhibitor of muscarinic receptors, did not affect these local Ca2+ transients. However, the desensitizing P2X receptor agonist alpha,beta-methylene ATP, and the purinergic antagonist, suramin, effectively inhibited the local Ca2+ transients. The frequency of these 'purinergic Ca2+ transients' was increased about 7-fold by a 10 s stimulus train (1 Hz). The amplitude, duration at one-half amplitude and the spatial spread of the evoked purinergic Ca2+ transients were F/F(o) = 2.4 +/- 0.13, 111.7 +/- 9.3 ms and 14.0 +/- 1.0 microm2, respectively. Tetrodotoxin inhibited evoked purinergic Ca2+ transients, indicating that they were dependent on nerve fibre activation. Purinergic Ca2+ transients were not dependent on VDCC activity. Neither 2-APB, an inhibitor of inositol 1,4,5-triphosphate (Ins(1,4,5)P3) (IP3)-induced Ca2+ release, nor ryanodine inhibited the purinergic Ca2+ transients. We have identified two novel Ca2+ signals in rat UBSM. Large, rapid, global Ca2+ flashes that represent Ca2+ influx through VDCCs during action potentials, and local, purinergic Ca2+ transients that represent Ca2+ entry through P2X receptors. Our results indicate that purinergic Ca2+ transients evoked by release of ATP from nerve varicosities are elementary signals in the process of nerve-smooth muscle communication.
authors.affiliation: Department of Pharmacology, University of Vermont, Given Building, Rm C-315, 89 Beaumont Avenue, Burlington, VT 05405-0068, USA. thomas.heppner@uvm.edu
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Heppner
authors.forename: Adrian D
authors.initials: AD
authors.lastname: Bonev
authors.forename: Mark T
authors.initials: MT
authors.lastname: Nelson
mesh_terms: D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D004110:Diltiazem / Q000494:pharmacology; D004558:Electric Stimulation / Q000379:methods; D066298:In Vitro Techniques; D008297:Male; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology*; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000502:physiology*; D009435:Synaptic Transmission / Q000187:drug effects / Q000502:physiology*; D001743:Urinary Bladder / Q000187:drug effects / Q000502:physiology*
chemical_list: D018048:Receptors, Purinergic P2; D004110:Diltiazem