Summary Lattice-like structures known as perineuronal nets (PNNs) are key components of the extracellular matrix (ECM). Once fully crystallized by adulthood, they are largely stable throughout life. Contrary to previous reports that PNNs inhibit processes involving plasticity, here we report that the dynamic regulation of PNN expression in the adult auditory cortex is vital for fear learning and consolidation in response to pure tones. Specifically, after first confirming the necessity of auditory cortical activity for fear learning and consolidation, we observed that mRNA levels of key proteoglycan components of PNNs were enhanced 4 hours after fear conditioning but were no longer different from the control groups 24 hours later. A similar pattern of regulation was observed in numbers of cells surrounded by PNNs and area occupied by them in the auditory cortex. Finally, the removal of auditory cortex PNNs resulted in a deficit in fear learning and consolidation.

Metabotropic glutamate receptors (mGluRs) are class C, synaptic G-protein-coupled receptors (GPCRs) that contain large extracellular ligand binding domains (LBDs) and form constitutive dimers. Despite the existence of a detailed picture of inter-LBD conformational dynamics and structural snapshots of both isolated domains and full-length receptors, it remains unclear how mGluR activation proceeds at the level of the transmembrane domains (TMDs) and how TMD-targeting allosteric drugs exert their effects. Here, we use time-resolved functional and conformational assays to dissect the mechanisms by which allosteric drugs activate and modulate mGluR2. Single-molecule subunit counting and inter-TMD fluorescence resonance energy transfer measurements in living cells reveal LBD-independent conformational rearrangements between TMD dimers during receptor modulation. Using these assays along with functional readouts, we uncover heterogeneity in the magnitude, direction, and the timing of the action of both positive and negative allosteric drugs. Together our experiments lead to a three-state model of TMD activation, which provides a framework for understanding how inter-subunit rearrangements drive class C GPCR activation.

Highlights d Branched photoswitchable tethered ligands enable nearcomplete optical GPCR agonism d Efficient optical control of mGluR2, mGluR3, and mGluR5 across labeling and spectral modalities d mGluR5 activation in astrocytic processes leads to local calcium oscillations d mGluR2 activation in prefrontal cortex rapidly and reversibly modulates working memory

Highlights d Single-cell RNA sequencing shows overlapping mGluR expression throughout cortex d Fluorescence complementation assays enable quantification of mGluR dimerization d Many heterodimers form with higher efficiency than parent homodimers d mGluR2/3 heterodimers are found natively in mouse frontal cortex

Employing self-labelling protein tags for the attachment of fluorescent dyes has become a routine and powerful technique in optical microscopy to visualize and track fused proteins. However, membrane permeability of...