# Screening using split fluorescent protein tags for neurotransmitter receptors that define a synaptic balance in neuralcircuits

> **NIH NIH R21** · UNIVERSITY OF GEORGIA · 2023 · $404,340

## Abstract

The function of neural circuits relies on the number and distribution of synapses composed of neurotransmitter
receptors, which establish and maintain a synaptic balance between excitation and inhibition. Mutations in
these receptors or pharmacologic perturbations of their function can disrupt neural circuit function, leading to
abnormal development and function of the brain. However, our knowledge of spatial patterning of the synaptic
balance by neurotransmitter receptors is incomplete. The missing pieces of this puzzle include molecular
characterizations that define excitatory and inhibitory (E and I) synapses in the developing brain. In the
proposed project, we will develop a method to detect the presence of known neurotransmitter receptor
subunits in Drosophila. We will generate a fly library in which endogenously expressed neurotransmitter
receptor subunits are tagged by split fluorescent proteins (split FPs). A short region of the split FP coding
sequence will be inserted into an array of neurotransmitter receptor subunit genes. The remainder of the FP
will be expressed in particular cell types using available Drosophila expression lines. This will allow us to
examine neurotransmitter receptor localization relative to individual dendritic branches. In Aim 1, we will
optimize the split FP tags for multicolor imaging of endogenous proteins in Drosophila. We will also develop a
cloning-free approach to insert the split FP tags into Drosophila genomic loci via CRISPR-mediated homology-
directed DNA repair, which is cost-effective and scalable for library generation. In Aim 2, we will apply the
knock-in approach to generate a fly library of 15 neurotransmitter receptor subunits tagged with split FPs. We
will employ this library to delineate a map of neurotransmitter receptors along dendrites in the cells of neural
circuits and ultimately study the molecular basis of E and I synapse formation in establishing circuit function.
The proposed work will provide crucial new information on the distribution of neurotransmitter receptors relative
to the establishment and maintenance of E and I synaptic balance as well as valuable resources for the
scientific community. Moreover, the project will develop a transferrable novel approach for generating large-
scale libraries of split FP-tagged molecules, giving researchers a means to localize proteins of interest in a
variety of cell types in Drosophila.

## Key facts

- **NIH application ID:** 10805112
- **Project number:** 1R21NS128750-01A1
- **Recipient organization:** UNIVERSITY OF GEORGIA
- **Principal Investigator:** Daichi Kamiyama
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2023
- **Award amount:** $404,340
- **Award type:** 1
- **Project period:** 2023-09-15 → 2026-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10805112

## Citation

> US National Institutes of Health, RePORTER application 10805112, Screening using split fluorescent protein tags for neurotransmitter receptors that define a synaptic balance in neuralcircuits (1R21NS128750-01A1). Retrieved via AI Analytics 2026-05-27 from https://api.ai-analytics.org/grant/nih/10805112. Licensed CC0.

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