Imaging ERK and PKA Activation in Single Dendritic Spines during Structural Plasticity

Area of Science:

• Neuroscience
• Molecular Biology
• Biochemistry

Background:

• Extracellular signal-regulated kinase (ERK) and protein kinase A (PKA) are crucial for synaptic plasticity and dendritic spine structural changes.
• Existing Förster resonance energy transfer (FRET)-based kinase sensors lack the sensitivity for imaging activity in small neuronal compartments like single dendritic spines.

Purpose of the Study:

• To develop highly sensitive FRET-based biosensors for monitoring ERK and PKA activity in real-time within individual dendritic spines.
• To visualize the spatial dynamics of ERK and PKA activation during structural long-term potentiation (sLTP) in hippocampal neurons.

Main Methods:

• Improvement of FRET-based biosensors to enhance sensitivity for kinase activity detection.
• Utilization of two-photon fluorescence lifetime imaging microscopy (2pFLIM) for high-resolution imaging.
• Application of enhanced sensors in hippocampal CA1 pyramidal neurons to study sLTP.

Main Results:

• Successfully imaged ERK and PKA activation in single dendritic spines during sLTP.
• Demonstrated that kinase activation during sLTP spreads widely, with length constants exceeding 10 μm.
• Validated the enhanced sensitivity of the improved FRET biosensors.

Conclusions:

• The developed FRET biosensors provide unprecedented sensitivity for imaging kinase activity in small neuronal structures.
• Kinase activation during sLTP exhibits widespread propagation within dendritic compartments.
• The sensor improvement strategy is adaptable for creating sensitive biosensors for other protein kinases.