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Related Experiment Videos

Synaptic vesicular glutamate uptake: modulation by a synaptosomal cytosolic factor.

A T Lobur1, P E Kish, T Ueda

  • 1Department of Pharmacology, University of Michigan, Ann Arbor 48109.

Journal of Neurochemistry
|May 1, 1990
PubMed
Summary

A novel protein factor in nerve terminals inhibits ATP-dependent glutamate uptake into synaptic vesicles. This discovery suggests endogenous regulation of neurotransmitter transport in the brain.

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Area of Science:

  • Neuroscience
  • Cell Biology
  • Biochemistry

Background:

  • L-glutamate functions as a neurotransmitter, and its uptake into synaptic vesicles is ATP-dependent.
  • Understanding the regulation of neurotransmitter uptake is crucial for comprehending synaptic function.

Purpose of the Study:

  • To identify and characterize factors that regulate ATP-dependent glutamate uptake into synaptic vesicles.
  • To investigate the nature of an endogenous cytosolic factor inhibiting glutamate vesicular transport.

Main Methods:

  • Size exclusion chromatography was used to estimate the molecular weight of the inhibitory factor.
  • Ammonium sulfate precipitation was employed to isolate the protein.
  • Thermal inactivation and proteolytic digestion (trypsin, chymotrypsin) were performed to assess factor stability and activity.

Related Experiment Videos

  • Glutamate uptake assays were conducted using isolated synaptic vesicles from rat and bovine brains.

    • Main Results:

    • A nerve terminal cytosolic factor was identified that dose-dependently inhibits ATP-dependent glutamate uptake into synaptic vesicles.
    • The inhibitory factor is a protein >100,000 molecular weight, precipitated by 40% ammonium sulfate, and inactivated by heating to 100°C.
    • Proteolytic digestion by trypsin or chymotrypsin slightly increased inhibitory activity and retained activity after heating, suggesting active fragment generation.
    • The inhibitory effect was not species-specific, with rat and bovine factors inhibiting uptake in vesicles from both species.

    Conclusions:

    • An endogenous protein factor regulates the ATP-dependent vesicular uptake of glutamate.
    • Proteolytic digestion may yield active fragments of the inhibitory factor.
    • This regulation mechanism may be conserved across species, suggesting in vivo relevance for controlling glutamate neurotransmission.