Acute inflammation sensitizes knee-innervating sensory neurons and decreases mouse digging behavior in a TRPV1-dependent manner

Abstract

threshold, as well as increased GABA and capsaicin sensitivity, but have unaltered acid sensitivity. 10The inflammation-induced sensitization of knee neurons persisted for 24-hours in culture, but was

Intraarticular Adeno‐Associated Virus Serotype AAV‐PHP.S–Mediated Chemogenetic Targeting of Knee‐Innervating Dorsal Root Ganglion Neurons Alleviates Inflammatory Pain in Mice

“…Consistent with our hypothesis, when AAV‐PHP.S‐CAG‐dTomato and the commonly used retrograde tracer, fast blue, were co‐injected unilaterally into 1 knee in mice (n = 3, all female), we observed both fast blue and virus labeling (Figure A). Comparable to findings of previous studies that used fast blue and other retrograde tracers , we observed a similar proportion of labeling in the lumbar DRG neurons with fast blue and AAV‐PHP.S‐CAG‐dTomato (Figure B). Across L2–L5 DRG, there was ~40% co‐labeling of neurons with fast blue and AAV‐PHP.S‐CAG‐dTomato, which suggests that neither retrograde tracer is able to label the entire knee neuron population (Figure C and Supplementary Figure 1, http://onlinelibrary.wiley.com/doi/10.1002/art.41314/abstract).…”

“…Based on previous studies , we hypothesized that the increased excitability of knee‐innervating neurons in vitro via the G q ‐DREADD system would cause pain‐like behavior in mice in vivo, which was tested by assessing digging behavior (a measure of spontaneous pain as previously described ), dynamic weight bearing, and rotarod behavior (a measure of motor coordination ) (Figure D). Three weeks after virus injection into both knee joints, mice injected with vehicle or C21 (n = 4 male and 4 female mice in each group) did not show significant changes in digging behavior (mean ± SEM digging duration pre‐vehicle 31.6 ± 3.7 seconds versus post‐vehicle 21.3 ± 4.4 seconds; mean ± SEM digging duration pre‐C21 25.2 ± 4.9 seconds versus post‐C21 32.7 ± 4.7 seconds; mean ± SEM number of burrows pre‐vehicle 4.6 ± 0.5 versus post‐vehicle 3.5 ± 0.4; mean ± SEM number of burrows pre‐C21 3.9 ± 0.7 versus post‐C21 3.6 ± 0.5) (Figures E and F) or weight bearing (mean ± SEM front paw to rear paw weight ratio pre‐vehicle 0.5 ± 0.02 versus post‐vehicle 0.5 ± 0.04; mean ± SEM front paw to rear paw weight ratio pre‐C21 0.4 ± 0.06 versus post‐C21 0.4 ± 0.03) (Figures G and H).…”

“…The titer values were chosen based on a pilot experiment conducted with AAV‐PHP.S‐CAG‐YFP, in which injection of 2 μl (~3 × 10 11 vg) and 10 μl (~10 12 vg) labeled a similar number of neurons as assessed by acute culture. For experiments involving inflammation, 7.5 μl Freund's complete adjuvant (CFA) (10 mg/ml; Chondrex) was injected into the left knee, and Vernier calipers were used to measure knee width before and 24 hours after CFA injection .…”

“…Digging behavior was measured as an assessment of spontaneous pain, as previously described , for 3 minutes in a standard cage with a wire lid and filled with Aspen midi 8/20 wood chip bedding (LBS Biotechnology). For training, mice were habituated in the test room in their home cages for 30 minutes, and then they were allowed to dig twice for 3 minutes with a 30‐minute break in between.…”

“…The variation in DRG neurons offers a unique opportunity to selectively tune the excitability of a distinct subset of DRG neurons in order to provide pain relief with reduced side effects. For example, we have recently shown that the excitability of knee‐innervating DRG neurons (identified by retrograde tracing) is increased in a mouse model of inflammatory joint pain and after incubation with human osteoarthritis (OA) synovial fluid samples . These findings suggest that modulation of the knee‐innervating DRG neuron subset (knee neurons) could help control arthritic pain.…”