iScience (Sep 2020)

Systematic Quantification of Synapses in Primary Neuronal Culture

  • Peter Verstraelen,
  • Gerardo Garcia-Diaz Barriga,
  • Marlies Verschuuren,
  • Bob Asselbergh,
  • Rony Nuydens,
  • Peter H. Larsen,
  • Jean-Pierre Timmermans,
  • Winnok H. De Vos

Journal volume & issue
Vol. 23, no. 9
p. 101542

Abstract

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Summary: Most neurological disorders display impaired synaptic connectivity. Hence, modulation of synapse formation may have therapeutic relevance. However, the high density and small size of synapses complicate their quantification. To improve synapse-oriented screens, we analyzed the labeling performance of synapse-targeting antibodies on neuronal cell cultures using segmentation-independent image analysis based on sliding window correlation. When assessing pairwise colocalization, a common readout for mature synapses, overlap was incomplete and confounded by spurious signals. To circumvent this, we implemented a proximity ligation-based approach that only leads to a signal when two markers are sufficiently close. We applied this approach to different marker combinations and demonstrate its utility for detecting synapse density changes in healthy and compromised cultures. Thus, segmentation-independent analysis and exploitation of resident protein proximity increases the sensitivity of synapse quantifications in neuronal cultures and represents a valuable extension to the analytical toolset for in vitro synapse screens.

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