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Related Concept Videos

Reporter Genes02:11

Reporter Genes

Reporter genes are a type of protein-coding gene that are often tagged to a gene of interest. Once inside a target cell, reporter genes usually produce visually identifiable characteristics like fluorescence and luminescence when expressed along with the gene of interest. Thus, reporter genes “report” the presence or absence of genes of interest in an organism, determine the gene expression pattern, or track the physical location of a DNA segment or protein in the cell.
Commonly used reporter...

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Development of pH-responsive fluorescent false neurotransmitters.

Minhee Lee1, Niko G Gubernator, David Sulzer

  • 1Department of Chemistry, Columbia University, 3000 Broadway, New York, New York 10027, USA.

Journal of the American Chemical Society
|June 15, 2010
PubMed
Summary

Researchers developed pH-responsive fluorescent false neurotransmitters (FFNs) that act as vesicular monoamine transporter (VMAT) substrates and pH sensors. These novel probes allow measurement of vesicular pH and drug-induced changes in neuroscience research.

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Area of Science:

  • Neuroscience
  • Biochemistry
  • Chemical Biology

Background:

  • Vesicular monoamine transporter (VMAT) plays a critical role in neurotransmitter storage and release.
  • Accurate measurement of vesicular pH is essential for understanding neurotransmitter dynamics and drug mechanisms.
  • Existing probes lack the specificity and functionality for simultaneous VMAT substrate activity and ratiometric pH sensing.

Purpose of the Study:

  • To develop novel pH-responsive fluorescent false neurotransmitters (pH-FFNs) capable of acting as VMAT substrates and ratiometric pH sensors.
  • To characterize the optical and biological properties of synthesized pH-FFNs for neuroscience applications.
  • To utilize a lead pH-FFN probe to investigate methamphetamine's effect on vesicular pH and content release.

Main Methods:

  • Systematic molecular design integrating VMAT recognition, ratiometric pH sensing, and lipophilicity modulation.
  • Synthesis and screening of 14 pH-FFN compounds.
  • Evaluation of probe uptake in VMAT2-transfected HEK cells.
  • Two-photon fluorescence microscopy to measure vesicular pH in PC-12 cells.
  • Assessment of methamphetamine's impact on vesicular pH and probe localization.

Main Results:

  • Successfully synthesized and characterized novel pH-FFNs.
  • Identified Mini202 as a lead probe with high VMAT2 uptake and desirable optical properties.
  • Measured basal catecholamine secretory vesicle pH in PC-12 cells at approximately 5.9.
  • Observed an increase in vesicular pH to approximately 6.4 upon methamphetamine incubation.
  • Demonstrated methamphetamine-induced redistribution of vesicular content from vesicles to cytoplasm.

Conclusions:

  • pH-responsive FFNs, exemplified by Mini202, are effective tools for simultaneous VMAT substrate activity and ratiometric pH sensing.
  • Mini202 enables precise measurement of vesicular pH and real-time monitoring of drug effects on vesicular function.
  • This novel probe facilitates fundamental neuroscience research, neuroendocrine studies, and drug screening efforts.