Mu-Opioid Receptor Agonist Induces Kir3 Currents in Mouse Peripheral Sensory Neurons - Effects of Nerve Injury.

Neuropathic pain often arises from damage to peripheral nerves and is difficult to treat. Activation of opioid receptors in peripheral sensory neurons is devoid of respiratory depression, sedation, nausea, and addiction mediated in the brain, and ameliorates neuropathic pain in animal models. Mechanisms of peripheral opioid analgesia have therefore gained interest, but the role of G protein-coupled inwardly rectifying potassium (Kir3) channels, important regulators of neuronal excitability, remains unclear.

Analgesic effects of a novel pH-dependent μ-opioid receptor agonist in models of neuropathic and abdominal pain.

Recently, (±)-N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide (NFEPP), a newly designed μ-opioid receptor (MOR) agonist with a low pKa, has been shown to produce injury-restricted analgesia in models of inflammatory and postoperative pain, without exhibiting typical opioid side effects. Here, we investigated MOR binding of NFEPP in brain and dorsal root ganglia, pH in injured tissues, and the analgesic efficacy of NFEPP compared with fentanyl in a chronic constriction injury model of neuropathic pain, and in the acetic acid-induced abdominal writhing assay in rats. Binding experiments revealed significantly lower affinity of NFEPP compared with fentanyl at pH 7.

Opioid receptor signaling, analgesic and side effects induced by a computationally designed pH-dependent agonist.

Novel pain killers without adverse effects are urgently needed. Opioids induce central and intestinal side effects such as respiratory depression, sedation, addiction, and constipation. We have recently shown that a newly designed agonist with a reduced acid dissociation constant (pK) abolished pain by selectively activating peripheral μ-opioid receptors (MOR) in inflamed (acidic) tissues without eliciting side effects.

Polyglycerol-opioid conjugate produces analgesia devoid of side effects.

Novel painkillers are urgently needed. The activation of opioid receptors in peripheral inflamed tissue can reduce pain without central adverse effects such as sedation, apnoea, or addiction. Here, we use an unprecedented strategy and report the synthesis and analgesic efficacy of the standard opioid morphine covalently attached to hyperbranched polyglycerol (PG-M) by a cleavable linker.

Novel Opioid Analgesics and Side Effects.

Conventional opioids mediate analgesia as well as severe adverse effects via G-protein coupled opioid receptors (OR) in both inflamed (peripheral injured tissue) and healthy (brain, intestinal wall) environments. To exclude side effects, OR activation can be selectively achieved in damaged tissue by lowering the pK of an opioid ligand to the acidic pH of inflammation. As a result, protonation of the ligand and consequent OR binding and activation of G-proteins is pH- and injury-specific.

Production of G protein-coupled receptors in an insect-based cell-free system.

The biochemical analysis of human cell membrane proteins remains a challenging task due to the difficulties in producing sufficient quantities of functional protein. G protein-coupled receptors (GPCRs) represent a main class of membrane proteins and drug targets, which are responsible for a huge number of signaling processes regulating various physiological functions in living cells. To circumvent the current bottlenecks in GPCR studies, we propose the synthesis of GPCRs in eukaryotic cell-free systems based on extracts generated from insect (Sf21) cells.

Targeting delta opioid receptors for pain treatment: drugs in phase I and II clinical development.

Opioids are widely used to treat severe pain. Most clinically used opioids activate µ-opioid receptors (MOR). Their ligands induce potent analgesia but also adverse effects.

Opioids and TRPV1 in the peripheral control of neuropathic pain--Defining a target site in the injured nerve.

Targeting peripheral neuropathic pain at its origin may prevent the development of hypersensitivity. Recently we showed this can be mediated by opioid receptors at the injured nerve trunk. Here, we searched for the most relevant peripheral site to block transient receptor potential vanilloid 1 (TRPV1), and investigated analgesic interactions between TRPV1 and opioids in neuropathy.

Electrophysiological patch clamp assay to monitor the action of opioid receptors.

The patch clamp is a valuable electrophysiological technique, which allows the study of single or multiple ion channels in cells, and it is particularly useful in testing the excitable cells such as neurons. Activation of neuronal opioid receptors results in the modulation of various ion channels, which enables to examine the receptors' action with the patch clamp. In this chapter, we analyze the activation of the G-protein-coupled inwardly rectifying potassium channel 2 by opioids, and the capsaicin-induced transient receptor potential vanilloid 1 channel currents during opioid withdrawal, using the whole cell patch clamp in transfected human embryonic kidney 293 cells as well as in mouse and rat primary dorsal root ganglion neurons.

Analysis of potassium and calcium imaging to assay the function of opioid receptors.

As the activation of opioid receptors leads to the modulation of potassium and calcium channels, the ion imaging represents an attractive method to analyze the function of the receptors. Here, we describe the imaging of potassium using the FluxOR™ potassium ion channel assay, and of calcium using Fura-2 acetoxymethyl ester. Specifically, we (1) characterize the activation of the G-protein-coupled inwardly rectifying potassium 2 channel by agonists of μ- and δ-opioid receptors with the aid of the FluxOR™ assay in cultured mouse dorsal root ganglion neurons, and (2) describe calcium imaging protocols to measure capsaicin-induced transient receptor potential vanilloid 1 channel activity during opioid withdrawal in transfected human embryonic kidney 293 cells.

Modulation of transient receptor vanilloid 1 activity by transient receptor potential ankyrin 1.

Transient receptor potential vanilloid 1 (TRPV1) is a nonselective ligand-gated cation channel responding to noxious heat, protons, and chemicals such as capsaicin. TRPV1 is expressed in sensory neurons and plays a critical role in pain associated with tissue injury, inflammation, and nerve lesions. Transient receptor potential ankyrin 1 (TRPA1) is coexpressed with TRPV1.

Opioid withdrawal increases transient receptor potential vanilloid 1 activity in a protein kinase A-dependent manner.

Hyperalgesia is a cardinal symptom of opioid withdrawal. The transient receptor potential vanilloid 1 (TRPV1) is a ligand-gated ion channel expressed on sensory neurons responding to noxious heat, protons, and chemical stimuli such as capsaicin. TRPV1 can be inhibited via μ-opioid receptor (MOR)-mediated reduced activity of adenylyl cyclases (ACs) and decreased cyclic adenosine monophosphate (cAMP) levels.