Voltage modulates the effect of μ‐receptor activation in a ligand‐dependent manner

Abstract

Background and Purpose: Various GPCRs have been described as being modulated in a voltage-dependent manner. Opioid analgesics act via activation of μ receptors in various neurons. As neurons are exposed to large changes in membrane potential, we were interested in studying the effects of depolarization on μ receptor signalling. Experimental Approach: We investigated potential voltage sensitivity of μ receptors in heterologous expression systems (HEK293T cells) using electrophysiology in combination with Förster resonance energy transfer-based assays. Depolarizationinduced changes in signalling were also tested in physiological rat tissue containing locus coeruleus neurons. We applied depolarization steps across the physiological range of membrane potentials. Key Results: Studying μ receptor function and signalling in cells, we discovered that morphine-induced signalling was strongly dependent on the membrane potential (V M). This became apparent at the level of G-protein activation, G-protein coupled inwardly rectifying potassium channel (K ir 3.X) currents and binding of GPCR kinases and arrestin3 to μ receptors by a robust increase in signalling upon membrane depolarization. The pronounced voltage sensitivity of morphine-induced μ receptor activation was also observed at the level of K ir 3.X currents in rat locus coeruleus neurons. The efficacy of peptide ligands to activate μ receptors was not (Met-enkephalin) or only moderately ([D-Ala 2 , N-Me-Phe 4 , Gly 5-ol]-enkephalin) enhanced upon depolarization. In contrast, depolarization reduced the ability of the analgesic fentanyl to activate μ receptors. Conclusion and Implications: Our results indicate a strong ligand-dependent modulation of μ receptor activity by the membrane potential, suggesting preferential activity of morphine in neurons with high neuronal activity.

Depolarization-Dependent C-Raf Signaling Promotes Hyperexcitability and Reduces Opioid Sensitivity of Isolated Nociceptors after Spinal Cord Injury

“…3F,G), and is similar to the voltage-dependent reduction in efficacy observed with other GPCRs (Vickery et al, 2016). Moreover, regulation of MOR by voltage was recently reported (Ruland et al, 2020). The rapid onset of [K 1 ] e -induced effects (within 5 min) indicates that, under these testing conditions, the observed effects are independent of transcription and/or translation and point toward direct signaling events triggered by depolarization.…”

“…In contrast, depolarization effects on opioid efficacy were C-Raf independent, agonist-specific (morphine vs DAMGO), and were observed in neurons and HEK cells, suggesting a receptor-mediated mechanism. Voltage can modulate the efficacy of MOR coupling to G proteins and b -arrestin in an agonist-specific manner (Ruland et al, 2020). Altered balance of voltage-regulated G-protein/ b -arrestin activation may explain our E max effects at high DAMGO concentrations.…”

“…6C). Thus, treatment with [K 1 ] e requires C-Raf activation to alter DAMGO potency, while the effects of depolarization on DAMGO E max may indeed indicate a voltage-sensitive mechanism acting directly on the MOR (Vickery et al, 2016;Ruland et al, 2020).…”

Differential recognition of opioid analgesics by µ opioid receptors: Predicted interaction patterns correlate with ligand-specific voltage sensitivity

“…S1A, B-G). Ligand dependence of the voltage sensitivity, mainly based on a change in efficacy in receptor activation, was additionally reported previously for morphine, Met-enkephalin, DAMGO and fentanyl (Ruland et al, 2020). Therefore, the MOR shows a strong ligand-specific voltage sensitivity.…”

“…However, based on the above-mentioned differences between the different opioids, it is important to understand details of ligand-receptor interactions. We recently showed that ligand-induced MOR activity was modulated by the membrane potential, and the effect and extent of this voltage sensitivity was ligand specific (Ruland et al, 2020). Since the first report of voltage sensitivity of the muscarinic M2 receptor (Ben-Chaim et al, 2003), several other GPCRs have been observed to be modulated in their activity depending on the membrane potential.…”