Locus Coeruleus and Dopamine-Dependent Memory Consolidation

Figure 4

Identification of key PRPs (plasticity-related proteins) by using optogenetics and translational profiling. (a) The critical test session would include (i) a behavioural condition that enhances memory (novelty), (ii) optogenetic activation of LC neurons (LC on), and (iii) LC activation with D₁/D₅ receptor blocker (LC on with D₁/D₅-R blocker) that might block the relevant synthesis of PRPs mediated by DAergic signaling in key target neurons. These conditions are compared to a home cage condition. (b) The TRAP technology, involving cell type-specific expression of green fluorescent protein- (GFP-) tagged ribosomal protein and GFP immunoprecipitation, enables the selective isolation of “translated mRNAs” in genetically defined neurons. (c) BONCAT (bioorthogonal noncanonical amino acid tagging) technology, involving labelling of newly synthesized proteins by AHA (azidohomoalanine), which can be later tagged for isolation and identification by mass spectrometry. (d) Candidate PRPs would be identified through the Venn diagram overlap of experimental conditions. (e) Optogenetic inhibition of a candidate PRP using “miniSOG,” a genetically encoded singlet oxygen generator [168]. After light illumination, singlet oxygen (¹O₂) is generated by miniSOG leading to the inactivation of fusion protein of interest.

(a) Critical test session

(b) TRAP

(c) BONCAT

(d) Translational profiling

(e) Functional assay