Gene discovery in genetically labeled single dopaminergic neurons of the retina

Proc Natl Acad Sci U S A. 2004 Apr 6;101(14):5069-74. doi: 10.1073/pnas.0400913101. Epub 2004 Mar 26.


In the retina, dopamine plays a central role in neural adaptation to light. Progress in the study of dopaminergic amacrine (DA) cells has been limited because they are very few (450 in each mouse retina, 0.005% of retinal neurons). Here, we applied transgenic technology, single-cell global mRNA amplification, and cDNA microarray screening to identify transcripts present in DA cells. To profile gene expression in single neurons, we developed a method (SMART7) that combines a PCR-based initial step (switching mechanism at the 5' end of the RNA transcript or SMART) with T7 RNA polymerase amplification. Single-cell targets were synthesized from genetically labeled DA cells to screen the RIKEN 19k mouse cDNA microarrays. Seven hundred ninety-five transcripts were identified in DA cells at a high level of confidence, and expression of the most interesting genes was confirmed by immunocytochemistry. Twenty-one previously undescribed proteins were found in DA cells, including a chloride channel, receptors and other membrane glycoproteins, kinases, transcription factors, and secreted neuroactive molecules. Thirty-eight percent of transcripts were ESTs or coding for hypothetical proteins, suggesting that a large portion of the DA cell proteome is still uncharacterized. Because cryptochrome-1 mRNA was found in DA cells, immunocytochemistry was extended to other components of the circadian clock machinery. This analysis showed that DA cells contain the most common clock-related proteins.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Base Sequence
  • DNA Primers
  • DNA, Complementary
  • Dopamine / metabolism*
  • Immunohistochemistry
  • Mice
  • Mice, Transgenic
  • Neurons / cytology
  • Neurons / metabolism*
  • Nucleic Acid Hybridization
  • Oligonucleotide Array Sequence Analysis
  • Retina / cytology
  • Retina / metabolism*


  • DNA Primers
  • DNA, Complementary
  • Dopamine