# Covalent labeling endogenous G-protein coupled receptors in living cells

> **NIH NIH R21** · OREGON HEALTH & SCIENCE UNIVERSITY · 2020 · $192,500

## Abstract

Opioid receptors are widely distributed in the central and peripheral nervous
system and upon activation have manifold actions including; depression of
respiration, activation of the reward pathway, disruption of normal
gastrointestinal motility and analgesia. It is currently difficult to examine this
widespread distribution in wild type animals. Antibodies against the opioid
receptors are often very unreliable and the epitopes are most often intracellular
such that detection in living tissue is not possible. The use of transgenic or
knockin animals to identify receptors requires extensive characterization, can be
limited by varied expression levels and has been limited to studies in mice. The
goal of this proposal is to use a ligand directed approach to covalently label
endogenous opioid receptors in living tissue. The guide compound, naltrexamine,
is a non-selective opioid antagonist that has high affinity for all subtype opioid
receptors. Naltrexamine will be coupled through a series of linkers, a reactive acyl
imidazole and fluorescent ligand. Once bound to the receptor, the reactive acyl
imidazole reacts with a lysine on the receptor and simultaneously releases the
naltrexamine. Thus natrexamine is free to dissociate from the receptor, however
the fluorescent ligand is left covalently bound to the receptor. Preliminary results
indicate that this compound interacts very effectively with the mu-opioid
receptor. Thus endogenous receptor are labeled and the labeling is not dependent
on species.
This compound and future refinements will be used areas of the CNS where
opioid receptors are expressed in only a small population of neurons. This
proposal will center on the VTA and Substantia Nigra in mouse and rat. Opioid
receptor expressing neurons in living brain slices will be identified and whole cell
recordings will be made. By targeting these cells a complete understanding of
opioid action in this key area of the reward system will be possible. The results of
this cell based study will address a long-standing question surrounding the
mechanisms that underlie one aspect of the addictive properties of opioids.

## Key facts

- **NIH application ID:** 9891997
- **Project number:** 5R21DA048136-02
- **Recipient organization:** OREGON HEALTH & SCIENCE UNIVERSITY
- **Principal Investigator:** John T Williams
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $192,500
- **Award type:** 5
- **Project period:** 2019-04-01 → 2021-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9891997

## Citation

> US National Institutes of Health, RePORTER application 9891997, Covalent labeling endogenous G-protein coupled receptors in living cells (5R21DA048136-02). Retrieved via AI Analytics 2026-05-21 from https://api.ai-analytics.org/grant/nih/9891997. Licensed CC0.

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