Internalized GPCRs as Potential Therapeutic Targets for the Management of Pain

Abstract

Peripheral and central neurons in the pain pathway are well equipped to detect and respond to extracellular stimuli such as pro-inflammatory mediators and neurotransmitters through the cell surface expression of receptors that can mediate rapid intracellular signaling. Following injury or infection, activation of cell surface G protein-coupled receptors (GPCRs) initiates cell signaling processes that lead to the generation of action potentials in neurons or inflammatory responses such as cytokine secretion by immune cells. However, it is now appreciated that cell surface events alone may not be sufficient for all receptors to generate their complete signaling repertoire. Following an initial wave of signaling at the cell surface, active GPCRs can engage with endocytic proteins such as the adaptor protein β-arrestin (βArr) to promote clathrin-mediated internalization. Classically, βArr-mediated internalization of GPCRs was hypothesized to terminate signaling, yet for multiple GPCRs known to contribute to pain, it has been demonstrated that endocytosis can also promote a unique "second wave" of signaling from intracellular membranes, including those of endosomes and the Golgi, that is spatiotemporally distinct from initial cell-surface events. In the context of pain, understanding the cellular and molecular mechanisms that drive spatiotemporal signaling of GPCRs is invaluable for understanding how pain occurs and persists, and how current analgesics achieve efficacy or promote side-effects. This review article discusses the importance of receptor localization for signaling outcomes of pro- and anti-nociceptive GPCRs, and new analgesic opportunities emerging through the development of "location-biased" ligands that favor binding with intracellular GPCR populations.

Keywords: GPCR; analgesia; drug delivery; endosome; pain; signal transduction; trafficking.

Figure 1

Role of G protein-coupled receptors (GPCRs) in pain and neurogenic inflammation. Injury or damaged tissues and infiltrating immune cells stimulate GPCRs on the peripheral sensory nerve terminals through release of painful and inflammatory mediators. Activated peptidergic and non-peptidergic Aδ and C fibers contribute to the response via the release of glutamate, Substance P (SP) and Calcitonin Gene-Related Peptide (CGRP) at the injury site and central terminals. The presence of endogenous mediators in the spinal cord (neuro- and glio-transmitters) can promote activation and recycling of GPCRs including pre-synaptic CB_1/2 cannabinoid and Mu-opioid receptor (MOR)/Delta-opioid receptor (DOR) and the exocytic trafficking of the γ-aminobutyric acid_A receptor (GABAR) which is an inhibitory receptor that can normalize neuronal excitability where excitatory neurotransmitters are released. Stimulation and endocytosis of receptors such as Neurokinin 1 Receptor (NK₁R) and Calcitonin Receptor-Like Receptor/Receptor Activity-Modifying Protein 1 (CLR/RAMP1) in post-synaptic neurons are known to modify firing frequency and the duration of pain responses (Jensen et al., ; Yarwood et al., ; Stoeber et al., 2018).

Figure 2

GPCR localization influences compartmentalized signaling and neuronal hyper-excitability. Activation of GPCRs on central neurons by extracellular neuropeptides (e.g., NK₁R or CL/RAMP1) initiates cell surface-delimited G protein-dependent signaling events. This is followed by GPCR kinase (GRK) phosphorylation, arrestin-binding, and clathrin-mediated endocytosis into endosomes to promote the recruitment of unique signaling complexes and drive spatiotemporally distinct signaling that is associated with sustained excitability of neurons in spinal cord slice preparations (Jensen et al., ; Yarwood et al., 2017). Lipid conjugation can influence membrane partitioning of antagonists. Palmitoylated pepducins are proposed to inhibit G protein-mediated inflammatory processes on the cytoplasmic interface of the plasma membrane (PM); whereas the sterol moiety cholestanol increases drug accumulation in endosomes to enhance inhibition of sustained endosomal-delimited signaling.