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Activity and subcellular trafficking of the sodium-coupled choline transporter CHT is regulated acutely by peroxynitrite
- Source :
- Molecular pharmacology. 73(3)
- Publication Year :
- 2007
-
Abstract
- Excess formation of nitric oxide and superoxide by-products (peroxynitrite, reactive oxygen, and reactive nitrogen species) attenuates cholinergic transmission potentially having a role in Alzheimer disease pathogenesis. In this study, we investigated mechanisms by which acute exposure to peroxynitrite impairs function of the sodium-dependent hemicholinium-3 (HC-3)-sensitive choline transporter (CHT) that provides substrate for acetylcholine synthesis. The peroxynitrite generator 3-morpholinosydnonimine (SIN-1) acutely inhibited choline uptake in cells stably expressing FLAG-tagged rat CHT in a dose- and time-dependent manner, with an IC(50) = 0.9 +/- 0.14 mM and t((1/2)) = 4 min. SIN-1 significantly reduced V(max) of choline uptake without altering the K(m). This correlated with a SIN-1-induced decrease in cell surface CHT protein, observed as lowered levels of HC-3 binding and biotinylated CHT at the plasma membrane. It is noteworthy that short-term exposure of cells to SIN-1 accelerated the rate of internalization of CHT from the plasma membrane, but it did not alter return of CHT back to the cell surface. SIN-1 did not disrupt cell membrane integrity or cause cell death. Thus, the inhibitory effect of SIN-1 on choline uptake activity and HC-3 binding was related to enhanced internalization of CHT proteins from the plasma membrane to subcellular organelles.
- Subjects :
- Luminescence
Time Factors
Cell
Cell Culture Techniques
Cholinergic Agents
Kidney
Choline
Membrane Potentials
chemistry.chemical_compound
Neuroblastoma
Enzyme Inhibitors
Internalization
media_common
Superoxide
Cell biology
Protein Transport
medicine.anatomical_structure
Biochemistry
Data Interpretation, Statistical
Molecular Medicine
Peroxynitrite
Subcellular Fractions
Nitrogen
media_common.quotation_subject
Transfection
Nitric oxide
Cell Line
Inhibitory Concentration 50
Cell Line, Tumor
Peroxynitrous Acid
medicine
Animals
Humans
Biotinylation
Reactive nitrogen species
Pharmacology
Dose-Response Relationship, Drug
L-Lactate Dehydrogenase
Cell Membrane
Sodium
Membrane Transport Proteins
Hemicholinium 3
Culture Media
Rats
Choline transporter
Kinetics
Oxidative Stress
chemistry
Gene Expression Regulation
Molsidomine
Cholinergic
Tyrosine
Subjects
Details
- ISSN :
- 15210111
- Volume :
- 73
- Issue :
- 3
- Database :
- OpenAIRE
- Journal :
- Molecular pharmacology
- Accession number :
- edsair.doi.dedup.....de156898f0da209008c9c98a542597d0