Abstract
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. Whereas functional Kir3 channels have been detected in dorsal root ganglion (DRG) neurons in rats, some studies question their contribution to opioid analgesia in inflammatory pain models in mice. However, neuropathic pain can be diminished by activation of peripheral opioid receptors in mouse models. Therefore, here we investigated effects of the selective μ-opioid receptor (MOR) agonist [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO) on potassium conductance in DRG neurons upon a chronic constriction injury (CCI) of the sciatic nerve in mice. For verification, we also tested human embryonic kidney (HEK) 293 cells transfected with MOR and Kir3.2. Using patch clamp, we recorded currents at -80 mV and applied voltage ramps in high extracellular potassium concentrations, which are a highly sensitive measures of Kir3 channel activity. We found a significantly higher rate of HEK cells responding with potassium channel blocker barium-sensitive inward current (233 ± 51 pA) to DAMGO application in transfected than in untransfected group, which confirms successful recordings of inward currents through Kir3.2 channels. Interestingly, DAMGO induced similar inward currents (178 ± 36–207 ± 56 pA) in 15–20% of recorded DRG neurons from naïve mice and in 4–27% of DRG neurons from mice exposed to CCI, measured in voltage clamp or voltage ramp modes. DAMGO-induced currents in naïve and CCI groups were reversed by barium and a more selective Kir3 channel blocker tertiapin-Q. These data indicate the coupling of Kir3 channels with MOR in mouse peripheral sensory neuron cell bodies, which was unchanged after CCI. A comparative analysis of opioid-induced potassium conductance at the axonal injury site and peripheral terminals of DRG neurons could clarify the role of Kir3 channel–MOR interactions in peripheral nerve injury and opioid analgesia.
Highlights
In European countries, 12–30% of adults suffer from chronic pain and many of them experience severe limitations in managing daily life activities (Breivik et al, 2006)
To establish the protocol for patch clamp recordings of inward potassium currents, we first used human embryonic kidney (HEK) 293 cells transfected with mu-opioid receptor (MOR) and Kir3.2 and untransfected HEK 293 cells
Application of the potassium channel blocker barium (3 mM BaCl2) reversed DAMGOmediated currents in one untransfected cell responding to DAMGO (Figure 1G) and in all DAMGO-responding transfected cells (Figure 1H), indicating that inward currents were mediated by potassium channels
Summary
In European countries, 12–30% of adults suffer from chronic pain and many of them experience severe limitations in managing daily life activities (Breivik et al, 2006). Among these are opioids such as morphine and fentanyl, which are the most powerful analgesics available They induce constipation which predominantly results from activation of intestinal opioid receptors (Imam et al, 2018), as well as respiratory depression, sedation, dizziness, and nausea mediated in the central nervous system (CNS) (Li and van den Pol, 2008; Imam et al, 2018). Their prolonged use leads to the development of addiction, which has resulted in a worldwide opioid epidemic (Volkow et al, 2018). Interactions between opioid receptors and potassium channels pose a promising basis for the development of novel therapies with an improved side effect profile
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