A systematic random sampling scheme optimized to detect the proportion of rare synapses in the neuropil.
Synapses can only be morphologically identified by electron microscopy and this is often a very labor-intensive and time-consuming task. When quantitative estimates are required for pathways that contribute a small proportion of synapses to the neuropil, the problems of accurate sampling are particularly severe and the total time required may become prohibitive. Here we present a sampling method devised to count the percentage of rarely occurring synapses in the neuropil using a large sample (approximately 1000 sampling sites), with the strong constraint of doing it in reasonable time. The strategy, which uses the unbiased physical disector technique, resembles that used in particle physics to detect rare events. We validated our method in the primary visual cortex of the cat, where we used biotinylated dextran amine to label thalamic afferents and measured the density of their synapses using the physical disector method. Our results show that we could obtain accurate counts of the labeled synapses, even when they represented only 0.2% of all the synapses in the neuropil.
da Costa
Nuno Maçarico
NM
Institute for Neuroinformatics, University of Zürich and ETH Zürich, Winterthurerstrasse 190, 8057 Zürich, Switzerland. ndacosta@ini.phys.ethz.ch
Hepp
Klaus
K
Martin
Kevan A C
KA
D000818:Animals; D001710:Biotin / Q000031:analogs & derivatives; D002415:Cats; D002452:Cell Count / Q000379:methods*; D003911:Dextrans; D019044:Image Cytometry / Q000379:methods*; D008854:Microscopy, Electron / Q000379:methods*; D009445:Neuroanatomy / Q000379:methods*; D019581:Neuropil / Q000502:physiology / Q000648:ultrastructure*; D017729:Presynaptic Terminals / Q000502:physiology / Q000648:ultrastructure; D012984:Software; D013194:Staining and Labeling / Q000379:methods; D013569:Synapses / Q000502:physiology / Q000648:ultrastructure*; D013788:Thalamus / Q000502:physiology / Q000648:ultrastructure; D014793:Visual Cortex / Q000502:physiology / Q000648:ultrastructure; D014795:Visual Pathways / Q000502:physiology / Q000648:ultrastructure
D003911:Dextrans; C076397:biotinylated dextran amine; D001710:Biotin