Microglia Disrupt Mesolimbic Reward Circuitry in Chronic Pain
Anna M. W. Taylor, Annie Castonguay, Alison J. Taylor, Niall P. Murphy, Atefeh Ghogha, Christopher Cook, Lihua Xue, Mary C. Olmstead, Yves De Koninck, Christopher J. Evans and Catherine M. Cahill
Journal of Neuroscience 3 June 2015, 35 (22) 8442-8450; DOI: https://doi.org/10.1523/JNEUROSCI.4036-14.2015
Anna M. W. Taylor
1Department of Anesthesiology and Perioperative Care, University of California Irvine, Irvine, California 92697, 2Hatos Center for Neuropharmacology, Semel Institute for Neuroscience and Human Behavior, University of California Los Angeles, Los Angeles, California 90095,
1Department of Anesthesiology and Perioperative Care, University of California Irvine, Irvine, California 92697, 5Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada, and
Loss of opioid- and cocaine-evoked DA release in chronic pain. Morphine (10 mg/kg) failed to stimulate extracellular DA in the PNI group, but treatment with minocycline (PNI+MC; 30 mg/kg) restored morphine-evoked extracellular DA levels. Similarly, cocaine-evoked (10 mg/kg) DA levels were lower in the PNI group compared with the sham group and cocaine-evoked extracellular DA could be recovered by pretreatment with minocycline (PNI+MC). Amphetamine-evoked (1 mg/kg) extracellular DA was not significantly different between PNI and sham groups and was not altered by MC treatment. ***p < 0.001. Error bars indicate SEM. Insets depict area under the curve (AUC) for the entire measurement period of DA dialysis for each group. *p < 0.05. Error bars indicate SEM.
Systemic opioid- and cocaine-induced place preference in chronic pain. a, Systemic morphine (10 mg/kg) produced a place preference in control and chronic pain (PNI) groups. When animals received limited conditioning trials (one exposure to 10 mg/kg morphine and one to saline), only the sham group displayed a robust place preference. Systemic low dose of morphine (1 mg/kg) produced a trend toward a place preference, but was not significant in either the sham or the PNI group. *p < 0.05, **p < 0.01. Error bars indicate SEM. b, Systemic cocaine (10 mg/kg) produced a place preference in both sham and PNI groups. **p < 0.01, ***p < 0.001. Error bars indicate SEM. c, Morphine (10 mg/kg) and cocaine (10 mg/kg) antinociception was measured using the tail withdrawal assay. Neither morphine nor cocaine antinociception was significantly different between sham or PNI groups. Further, inhibition of microglia in chronic pain animals (PNI+MC) did not affect morphine antinociception. Error bars indicate SEM. n.s., Not significant; %MPE, percent maximum possible effect.
Loss of mesolimbic-specific opioid- and cocaine-induced place preference in chronic pain. a, Repeated conditioning to intra-VTA DAMGO (1 ng) failed to produce a robust place preference in the PNI groups, whereas a high dose of DAMGO (25 ng) produced a robust place preference in both the sham and PNI groups. Pretreatment with the microglial inhibitor minocycline (30 mg/kg) recovered intra-VTA DAMGO (1 ng) place preference in the PNI group. Top left pictograph depicts intra-VTA injection sites. *p < 0.05, **p < 0.01, ***p < 0.001. Error bars indicate SEM. b, Cocaine (100 μg) administered directly into the NAc produced a place preference in the sham group, but not the PNI group. Left pictograph depicts intra-NAc injection sites. *p < 0.05. Error bars indicate SEM.
Chronic pain activates microglia in the VTA. a, Microglia were significantly activated in the VTA of the chronic pain (PNI) group and displayed a robust activated phenotypes. Pretreatment with the microglial inhibitor minocycline (MC; 30 mg/kg) blocked microglial activation in the PNI+MC group. **p < 0.01, ***p < 0.001. Error bars indicate SEM. Scale bar, 75 μm. b, Mechanical withdrawal thresholds of the ipsilateral hindpaw were measured with von Frey filaments. The PNI group showed significantly lowered mechanical thresholds 2 weeks after nerve injury. Systemic treatment with the microglial inhibitor MC did not affect evoked pain thresholds. **p < 0.01, ***p < 0.001. Error bars indicate SEM. n.s., Not significant.
Chronic pain disrupts Cl− transport in VTA GABAergic through a microglia-BDNF signaling pathway. a, KCC2 labeling in the VTA was primarily localized in processes not labeled for TH. Immunolabeling of KCC2 (green) was identified in non-TH (red)-labeled neurons and the labeling was punctate, which is indicative of terminal arborization. Scale bar, 75 μm. The right micrographs represent a single image of a TH+ neuron (middle, arrowhead) surrounded by KCC2-immunoreactive arbors (bottom). The top image is a merge of the bottom two images. b, The chronic pain (PNI) group exhibited decreased KCC2 expression in the VTA that could be recovered by systemic treatment with minocycline (PNI+MC; 30 mg/kg). Left black and white micrographs are representative images of KCC2 labeling in the VTA of sham and PNI groups. The right histogram shows KCC2 quantification. Error bars indicate SEM. *p < 0.05, **p < 0.01. AU, Arbitrary units. c, Top left, KCC2 function was assayed in VTA slices using MQAE FLIM. The VTA was localized with brain atlas coordinates and confirmed with TH labeling (red) after Cl− imaging assay. Bottom left, Representative micrographs showing MQAE loading in GAD65-GFP cells (white arrows) in the VTA (2-photon optical section ∼1 μm). Top right, Pseudocolor images showing lifetime maps from control VTA slices in the presence of 2.5 or 15 mm KCl. d, Representative lifetime plots from VTA slices. e, Graph plotted by averaging the slope in the lifetime plots immediately after administration of 15 mm KCl. The PNI group had significantly slower fluorescence lifetime change rate indicative of a slower transport of Cl− into the cell. Normal transport rate was restored in animals pretreated with minocycline (PNI+MC) or in slices treated with a TrkB antagonist (PNI+anti-trkB). (*p < 0.05; n = 7 animals with at least 2 slices per animal, 14–20 slices/condition, and 70–100 cells, on average 5/slice). Error bars indicate SEM. f, VTA punches from the PNI group showed significantly elevated BDNF protein, which was blocked with pretreatment with MC. *p < 0.05 compared with sham, #p < 0.05 compared with PNI+saline. Error bars indicate SEM.