Nitric Oxide Increases Calcium Oscillations in Response to Muscarinic Receptor Stimulation
Department
Biological Sciences
Major
Chemical Engineering /Biochemical Emphasis
Research Advisor
Aronstam, Robert
Advisor's Department
Biological Sciences
Funding Source
Missouri S&T cDNA Resource Center
Abstract
The effect of NO on calcium oscillations elicited by low concentrations of a muscarinic agonist in CHO cells stably transfected with the gene for the human M3 receptor. Nitric oxide’s targets include several synaptic receptor-mediated signaling processes. Agonist stimulation of M3 muscarinic acetylcholine receptors leads to release Ca2+ into the cytosol; this depletion of calcium stores stimulates calcium influx. At near threshold agonist concentrations, a cyclical release and re-uptake of calcium from intracellular stores (calcium oscillations) is the predominant response. Calcium influx was eliminated by the use of calcium-free medium; fura-2 fluorescence was used to quantify cytosolic calcium concentration; carbamylcholine was used to activate muscarinic receptors; nitric oxide was provided by varying concentrations of S-nitrosylglutathione (SNG) before measuring muscarinic responses. Calcium release from intracellular stores was characterized by carbamylcholine sensitivity, maximum initial response, and the presence, amplitude and decay of calcium oscillations. Pretreatment with nitric oxide donor decreased the amplitude of the initial Ca2+ peak by ≈15%, but increased the proportion of cells displaying oscillatory behavior from < 10% to > 90%. The threshold for the elicitation of Ca2+ responses by carbamylcholine increased, consistent with a NOmediated subsensitivity. The average delay in the manifestation of the Ca2+ response increased markedly from 5.3 sec to 8.0 and 12.2 sec in the presence of 4.3 and 8.5 mM SNG, respectively. Nitric oxide modulates the ability of M3 muscarinic receptors to activate calcium signaling pathways as indicated by an enhanced development of calcium oscillations. These changes may reflect changes in the potency of the muscarinic ligand.
Biography
Barbara Wheelden plans on attending graduate school in BioMedical Engineering. She is active in Engineers Without Borders and the S&T Chapters of ISPE and Omega Chi Epsilon.
Research Category
Sciences
Presentation Type
Poster Presentation
Document Type
Poster
Award
Sciences poster session, Third place
Location
Upper Atrium/Hallway
Presentation Date
08 Apr 2009, 9:00 am - 11:45 am
Nitric Oxide Increases Calcium Oscillations in Response to Muscarinic Receptor Stimulation
Upper Atrium/Hallway
The effect of NO on calcium oscillations elicited by low concentrations of a muscarinic agonist in CHO cells stably transfected with the gene for the human M3 receptor. Nitric oxide’s targets include several synaptic receptor-mediated signaling processes. Agonist stimulation of M3 muscarinic acetylcholine receptors leads to release Ca2+ into the cytosol; this depletion of calcium stores stimulates calcium influx. At near threshold agonist concentrations, a cyclical release and re-uptake of calcium from intracellular stores (calcium oscillations) is the predominant response. Calcium influx was eliminated by the use of calcium-free medium; fura-2 fluorescence was used to quantify cytosolic calcium concentration; carbamylcholine was used to activate muscarinic receptors; nitric oxide was provided by varying concentrations of S-nitrosylglutathione (SNG) before measuring muscarinic responses. Calcium release from intracellular stores was characterized by carbamylcholine sensitivity, maximum initial response, and the presence, amplitude and decay of calcium oscillations. Pretreatment with nitric oxide donor decreased the amplitude of the initial Ca2+ peak by ≈15%, but increased the proportion of cells displaying oscillatory behavior from < 10% to > 90%. The threshold for the elicitation of Ca2+ responses by carbamylcholine increased, consistent with a NOmediated subsensitivity. The average delay in the manifestation of the Ca2+ response increased markedly from 5.3 sec to 8.0 and 12.2 sec in the presence of 4.3 and 8.5 mM SNG, respectively. Nitric oxide modulates the ability of M3 muscarinic receptors to activate calcium signaling pathways as indicated by an enhanced development of calcium oscillations. These changes may reflect changes in the potency of the muscarinic ligand.
Comments
Joint project with C. Anna Growcock