# Development of kinase biosensors for multiplex neuronal imaging of signaling pathways in behaving mice

> **NIH NIH RF1** · JOHNS HOPKINS UNIVERSITY · 2022 · $2,409,828

## Abstract

Project Summary
Cell signaling pathways in the brain are an essential part of a complex system regulating the activity and
coordination of neuronal networks. During learning and memory these neuronal networks can be modified
through neuronal and synaptic plasticity processes in which information is stored in the synaptic network.
Intracellular signaling pathways play critical roles in regulating neuronal excitability and synaptic strength,
thereby comprising an important part of the cellular and molecular mechanisms underlying learning and memory.
Disruptions in the proper regulation of synaptic plasticity are involved in a number of neurological and psychiatric
disorders including autism, schizophrenia, and intellectual disability. Revealing the dynamic activities and
interactions of different signaling pathways is therefore crucial for understanding the mechanisms controlling
neuronal networks both in health and disease. However, direct interrogation of signaling pathway activity in live
animals has been challenging due to a lack of appropriate tools. Monitoring of multiple signaling pathways in
such a setting has not been achieved. The goal of this research proposal is to develop novel tools to
simultaneously monitor the activity of several signaling pathways and to use rapid, sensitive in vivo imaging
techniques to visualize dynamic activity of these signaling pathways in live animals during physiologically
relevant sensory experience and learning. Most existing biosensors for signaling activities are based on
fluorescence resonance energy transfer (FRET) and the use of two different fluorescent proteins, which limits
their use for monitoring of multiple signaling pathways in parallel. In this proposal, new single-color fluorescent
protein-based kinase biosensors with high sensitivity and optimized two-photon excitation properties will be
developed for imaging signaling pathways involved in synaptic plasticity, especially PKA, CaMKII, ERK, and
PKC. The ultimate goal of this research proposal is to establish the use of these new biosensors in the mouse
brain and to monitor both rapid dynamics of signaling pathways on the order of seconds to minutes and the long-
term stability of signaling pathways on the order of weeks to months using two-photon microscopy in awake
behaving animals. This proposed project will be the first investigation of multiple neuronal activities beyond
calcium and voltage changes in live animals. These studies will allow us to examine the regulation of kinase
pathways in vivo and will help elucidate the complexity of signaling pathways during synaptic plasticity in the
brain.

## Key facts

- **NIH application ID:** 10505852
- **Project number:** 1RF1MH126707-01A1
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Richard L Huganir
- **Activity code:** RF1 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $2,409,828
- **Award type:** 1
- **Project period:** 2022-08-01 → 2025-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10505852

## Citation

> US National Institutes of Health, RePORTER application 10505852, Development of kinase biosensors for multiplex neuronal imaging of signaling pathways in behaving mice (1RF1MH126707-01A1). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10505852. Licensed CC0.

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