Authors:

Hareem Tariq

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About Paper:

Alzheimer's disease (AD) is a neurodegenerative disorder associated with memory loss and cognitive decline, primarily attributed to calcium signalling dysfunction. Canonical Ca2+ signalling activates Calcium-Calmodulin Kinase II (CaMKII), the most abundant protein in excitatory synapses and central to synaptic plasticity (SP), which further modulates actin-driven morphological changes in dendritic spines at excitatory synapses. The underlying mechanisms of CaMKII-dependent actin regulation are complex given that the extent to which spine membranes, and in particular the spine neck, expand due to these protein signals remain unresolved. The aim of this study is to explore the interactions of CaMKII and actin in rat hippocampal neurons, utilizing a proximity ligation assay (PLA). PLA is a powerful tool that allows in situ detection of protein interactions with high specificity and sensitivity. The proteins CaMKII and actin are detected with primary antibodies, after which a pair of secondary antibodies (PLA probes) bind. Hybridization of connector DNA oligonucleotides join the probes within proximity and a ligase closes the circular DNA template rolling-circle amplification (RCA). The incorporation of fluorescently labelled nucleotides within the template generates fluorescent dots, indicative of interaction, which can be detected using confocal microscopy fluorescence imaging techniques. Our results show detection and visualization of the close proximity of these proteins and shed light on CaMKII and actin colocalization throughout the dendritic spine. Thus, the interplay between Ca2+ signalling, CaMKII and actin offers insights into dendritic spines organization, determining synaptic strength, and influencing SP, with potential therapeutic implications for synaptic dysfunction in neurodegenerative disorders like AD.

Source:

Purdue University / 2023

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Co-authors:

Hareem Tariq