Inhibition of the phosphoinositide 3-kinase-AKT-cyclic GMP-c-Jun N-terminal kinase signaling pathway attenuates the development of morphine tolerance in a mouse model of neuropathic pain.

Research presented here sought to determine if opioid induced tolerance is linked to activity changes within the PI3Kγ-AKT-cGMP-JNK intracellular signaling pathway in spinal cord or peripheral nervous systems. Morphine or saline injections were given subcutaneously twice a day for five days (15 mg/kg) to male C57Bl/6 mice. A separate cohort of mice received spinal nerve ligation (SNL) one week prior to the start of morphine tolerance. Afterwards, spinal cord, dorsal root ganglia, and sciatic nerves were isolated for quantifying total and phosphorylated- JNK levels, cGMP, and gene expression analysis of Pik3cg, Akt1, Pten, and nNos1. This pathway was downregulated in the spinal cord with increased expression in the sciatic nerve of morphine tolerant and morphine tolerant mice after SNL. We also observed a significant increase in phosphorylated- JNK levels in the sciatic nerve of morphine tolerant mice with SNL. Pharmacological inhibition of PI3K or JNK, using thalidomide, quercetin, or SP600125, attenuated the development of morphine tolerance in mice with SNL as measured by thermal paw withdrawal. Overall, the PI3K/AKT intracellular signaling pathway is a potential target for reducing the development of morphine tolerance in the peripheral nervous system. Continued research into this pathway will contribute to the development of new analgesic drug therapies.

Modulation of SUR1 KATP Channel Subunit Activity in the Peripheral Nervous System Reduces Mechanical Hyperalgesia after Nerve Injury in Mice.

The ATP-sensitive K+ channel (KATP) is involved in hypersensitivity during chronic pain and is presumed to be a downstream target of mu opioid receptors. Multiple subtypes of KATP channels exist in the peripheral and central nervous system and their activity may be inversely correlated to chronic pain phenotypes in rodents. In this study, we investigated the different KATP channel subunits that could be involved in neuropathic pain in mice. In chronic pain models utilizing spinal nerve ligation, SUR1 and Kir6.2 subunits were found to be significantly downregulated in dorsal root ganglia and the spinal cord. Local or intrathecal administration of SUR1-KATP channel subtype agonists resulted in analgesia after spinal nerve ligation but not SUR2 agonists. In ex-vivo nerve recordings, administration of the SUR1 agonist diazoxide to peripheral nerve terminals decreased mechanically evoked potentials. Genetic knockdown of SUR1 through an associated adenoviral strategy resulted in mechanical hyperalgesia but not thermal hyperalgesia compared to control mice. Behavioral data from neuropathic mice indicate that local reductions in SUR1-subtype KATP channel activity can exacerbate neuropathic pain symptoms. Since neuropathic pain is of major clinical relevance, potassium channels present a target for analgesic therapies, especially since they are expressed in nociceptors and could play an essential role in regulating the excitability of neurons involved in pain-transmission.

Morphine Efficacy, Tolerance, and Hypersensitivity Are Altered After Modulation of SUR1 Subtype KATP Channel Activity in Mice.

ATP-sensitive potassium (KATP) channels are found in the nervous system and are downstream targets of opioid receptors. KATP channel activity can effect morphine efficacy and may beneficial for relieving chronic pain in the peripheral and central nervous system. Unfortunately, the KATP channels exists as a heterooctomers, and the exact subtypes responsible for the contribution to chronic pain and opioid signaling in either dorsal root ganglia (DRG) or the spinal cord are yet unknown. Chronic opioid exposure (15 mg/kg morphine, s.c., twice daily) over 5 days produces significant downregulation of Kir6.2 and SUR1 in the spinal cord and DRG of mice. In vitro studies also conclude potassium flux after KATP channel agonist stimulation is decreased in neuroblastoma cells treated with morphine for several days. Mice lacking the KATP channel SUR1 subunit have reduced opioid efficacy in mechanical paw withdrawal behavioral responses compared to wild-type and heterozygous littermates (5 and 15 mg/kg, s.c., morphine). Using either short hairpin RNA (shRNA) or SUR1 cre-lox strategies, downregulation of SUR1 subtype KATP channels in the spinal cord and DRG of mice potentiated the development of morphine tolerance and withdrawal. Opioid tolerance was attenuated with intraplantar injection of SUR1 agonists, such as diazoxide and NN-414 (100 µM, 10 µL) compared to vehicle treated animals. These studies are an important first step in determining the role of KATP channel subunits in antinociception, opioid signaling, and the development of opioid tolerance, and shed light on the potential translational ability of KATP channel targeting pharmaceuticals and their possible future clinical utilization. These data suggest that increasing neuronal KATP channel activity in the peripheral nervous system may be a viable option to alleviate opioid tolerance and withdrawal.