Combining laser scanning confocal microscopy and electron microscopy to determine sites of synaptic contact between two identified neurons

Marie Jeanne Cabirol-Pol, Adi Mizrahi, John Simmers, Pierre Meyrand

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

Abstract

Here we report a double labelling method for correlative confocal and electron microscopy (EM) which allows selective characterisation of structural relationships between two single identified neurons in the same preparation. Using the lobster stomatogastric nervous system, we labelled pairs of identified, synaptically-connected neurons by intracellular injection of Lucifer Yellow (LY) in one neuron and a mixture of Rhodamine (Rdh) and Horseradish Peroxidase (HRP) in its partner. First, whole-mounts of LY- and Rdh-stained neurons were visualized using laser scanning confocal microscopy (LSCM) in order to isolate neuropilar regions of possible synaptic contact. Second, after conventional treatment for electron microscopy (LY was revealed with immunogold and HRP with DAB), areas of close appositions were viewed in EM. This technique allowed us to determine all the regions of close contact between two cells, and then to use electron microscopy to determine the presence or absence of synaptic contact within each of these restricted areas. These techniques enabled us to show that there were few areas of apposition and that only an extremely small proportion of these areas was in fact regions of synaptic contact between the two labelled neurons.

Original languageAmerican English
Pages (from-to)175-181
Number of pages7
JournalJournal of Neuroscience Methods
Volume97
Issue number2
DOIs
StatePublished - 15 Apr 2000
Externally publishedYes

Bibliographical note

Funding Information:
This work was supported by a doctoral studentship from the Ministère de l’Education Nationale de la Recherche et de la Technologie to M.J. Cabirol-Pol, a Chateaubriand grant to A. Mizhari, by the Conseil Régional d’Aquitaine and by the Human Science Program. We thank Drs Valérie Fénelon and Patsy Dickinson for comments on the manuscript.

Keywords

  • Confocal microscopy
  • Double labelling
  • Electron microscopy
  • Horseradish peroxidase
  • Lucifer Yellow
  • Rhodamine

Fingerprint

Dive into the research topics of 'Combining laser scanning confocal microscopy and electron microscopy to determine sites of synaptic contact between two identified neurons'. Together they form a unique fingerprint.

Cite this