Rapid 3D Enhanced Resolution Microscopy Reveals Diversity in Dendritic Spinule Dynamics, Regulation, and Function

• Published in: Neuron (Cambridge, Mass.) Mass.), 2020-08, Vol.107 (3), p.522-537.e6
• Main Authors: Zaccard, Colleen R., Shapiro, Lauren, Martin-de-Saavedra, Maria D., Pratt, Christopher, Myczek, Kristoffer, Song, Amy, Forrest, Marc P., Penzes, Peter
• Format: Article
• Language: English
• Subjects: Animals; Brain slice preparation; Calcium; Calcium - metabolism; calcium transients; Cerebral Cortex - cytology; Confocal microscopy; dendritic spine dynamics; Dendritic spines; Dendritic Spines - metabolism; Dendritic Spines - physiology; Dendritic Spines - ultrastructure; Glutamate receptors; Guanine Nucleotide Exchange Factors - metabolism; Imaging, Three-Dimensional; intrabodies; kalirin-7; Light microscopy; live imaging; Mice; Microscopy; Microscopy, Confocal; multi-synaptic spines; N-Methyl-D-aspartic acid receptors; Neural networks; Post-Synaptic Density - physiology; Post-Synaptic Density - ultrastructure; postsynaptic density; Pyramidal cells; Pyramidal Cells - physiology; Pyramidal Cells - ultrastructure; Rac1 protein; Receptors, N-Methyl-D-Aspartate - agonists; Regulation; Spatio-Temporal Analysis; Spinules; structured illumination microscopy; Studies; Synapses; Synapses - physiology; Synapses - ultrastructure; synaptic plasticity; synaptic spinules
• ISSN: 0896-6273; 1097-4199
• DOI: 10.1016/j.neuron.2020.04.025

Dendritic spinules are thin protrusions, formed by neuronal spines, not adequately resolved by diffraction-limited light microscopy, which has limited our understanding of their behavior. Here we performed rapid structured illumination microscopy and enhanced resolution confocal microscopy to study spatiotemporal spinule dynamics in cortical pyramidal neurons. Spinules recurred at the same locations on mushroom spine heads. Most were short-lived, dynamic, exploratory, and originated near simple PSDs, whereas a subset was long-lived, elongated, and associated with complex PSDs. These subtypes were differentially regulated by Ca2+ transients. Furthermore, the postsynaptic Rac1-GEF kalirin-7 regulated spinule formation, elongation, and recurrence. Long-lived spinules often contained PSD fragments, contacted distal presynaptic terminals, and formed secondary synapses. NMDAR activation increased spinule number, length, and contact with distal presynaptic elements. Spinule subsets, dynamics, and recurrence were validated in cortical neurons of acute brain slices. Thus, we identified unique properties, regulatory mechanisms, and functions of spinule subtypes, supporting roles in neuronal connectivity. [Display omitted] •Most spinules are short-lived, dynamic, recurrent, and originate near simple PSDs•An elongated stable subset traffics PSD fragments and can form secondary synapses•Spinule subsets are positively regulated by local Ca2+ transients and kalirin-7•Neuronal activity induces spinule elongation and contact with distal synapses Enhanced resolution 3D microscopy reveals the dynamics of thin dendritic spine protrusions, termed spinules, which are uniquely regulated by local Ca2+ and the Rac1-GEF kalirin-7. The majority are small, short-lived, dynamic, and recurrent, whereas an elongated subset is activity-induced, stabilized by distal presynaptic terminals, and can form connectivity-altering secondary synapses.