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Cloning and functional characterization of a high-affinity Na(+)/dicarboxylate cotransporter from mouse brain
1Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston, Texas 77555, USA. ampajor@utmb.edu
American Journal of Physiology. Cell Physiology
|April 5, 2001
Summary
Researchers identified a novel high-affinity sodium/dicarboxylate cotransporter (mNaDC-3) in the mouse brain. This transporter is crucial for absorbing neurotransmitter precursor substrates, impacting neurotransmitter levels like glutamate.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- Neurons utilize high-affinity Na+/dicarboxylate cotransporters to absorb neurotransmitter precursor substrates.
- Substrates like alpha-ketoglutarate and malate are metabolized to replenish neurotransmitter pools, including glutamate.
Purpose of the Study:
- To isolate and characterize a high-affinity Na+/dicarboxylate cotransporter from the mouse brain.
- To understand the substrate specificity, kinetic properties, and electrical characteristics of the identified transporter.
Main Methods:
- cDNA isolation from mouse brain.
- mRNA expression analysis in various tissues (brain, choroid plexus, kidney, liver).
- Transport assays to determine substrate specificity and kinetic parameters (Michaelis-Menten constant).
- Two-electrode voltage clamp assays to study electrical properties.
Main Results:
- Isolated cDNA coding for mNaDC-3, a high-affinity Na+/dicarboxylate cotransporter.
- Detected mNaDC-3 mRNA in brain, choroid plexus, kidney, and liver.
- Demonstrated broad substrate specificity for dicarboxylates (succinate, alpha-ketoglutarate, fumarate, malate, dimethylsuccinate).
- Observed pH sensitivity for succinate transport and partial substitution of sodium by lithium.
- Determined Michaelis-Menten constants for succinate.
Conclusions:
- mNaDC-3 likely represents the high-affinity Na+/dicarboxylate cotransporter in the brain.
- mNaDC-3 exhibits unique electrical properties compared to other family members.
- This transporter plays a significant role in neurotransmitter precursor uptake and metabolism.