Matching neural morphology to molecular expression: single cell injection following immunostaining

J Neurocytol. 2003 Mar;32(3):245-51. doi: 10.1023/B:NEUR.0000010083.03446.78.

Abstract

To match a neuron's morphology with its expression of a particular protein, it is useful to first identify the cell by immunostaining and then inject it with fluorescent dye. Such targeted injection cannot be performed with a hydrophilic dye (such as Lucifer yellow) because the neuron, once rendered porous to antibodies, does not retain it. But a lipophilic dye (such as DiI) injected iontophoretically into the soma forms a crystal and is thereby trapped. From this intracellular depot dye diffuses into the cell membrane to reveal the detailed morphology. We have used this strategy to identify the morphology of a GABAergic retinal bipolar cell and several types of GABAergic amacrine cell. In addition, we demonstrate probable connections from a narrow-field, GABAergic amacrine cell to the OFF brisk-transient ganglion cell. Finally, we show that the strategy works in the cortical slice, showing a layer IV cell immunostained for parvalbumin to be a "nest basket cell".

Publication types

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

MeSH terms

  • Amacrine Cells / cytology
  • Amacrine Cells / metabolism
  • Animals
  • Carbocyanines
  • Cats
  • Cell Size / physiology
  • Cerebral Cortex / cytology
  • Cerebral Cortex / metabolism
  • Coloring Agents
  • Dendrites / metabolism
  • Dendrites / ultrastructure
  • Fluorescent Dyes*
  • Glutamate Decarboxylase / metabolism
  • Guinea Pigs
  • Immunohistochemistry
  • In Vitro Techniques
  • Isoenzymes / metabolism
  • Microinjections / methods*
  • Neural Inhibition / physiology
  • Neural Pathways / cytology*
  • Neural Pathways / metabolism
  • Neurons / cytology*
  • Neurons / metabolism
  • Organic Chemicals
  • Parvalbumins / metabolism
  • Rats
  • Retina / cytology*
  • Retina / metabolism
  • Retinal Ganglion Cells / cytology
  • Retinal Ganglion Cells / metabolism
  • Synaptic Transmission / physiology
  • gamma-Aminobutyric Acid / metabolism

Substances

  • 3,3'-dihexadecylindocarbocyanine
  • Carbocyanines
  • Coloring Agents
  • Fluorescent Dyes
  • Isoenzymes
  • Organic Chemicals
  • Parvalbumins
  • SYTO 13
  • gamma-Aminobutyric Acid
  • Glutamate Decarboxylase
  • glutamate decarboxylase 2