Research ArticlePain

CT-guided injection of a TRPV1 agonist around dorsal root ganglia decreases pain transmission in swine

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Science Translational Medicine  16 Sep 2015:
Vol. 7, Issue 305, pp. 305ra145
DOI: 10.1126/scitranslmed.aac6589
  • Fig. 1. CT-guided injections in a pig model.

    (A) Three-dimensional (3D) surface rendering of posterior skin surface with animal in prone position on CT table. Bilateral spinal needles are in place at the L2/3, L3/4, L4/5, and L5/6 levels (blue arrowheads). (B) In the same animal, 3D volume rendering of pig lumbosacral spine CT with spinal needles in place immediately adjacent to DRGs at the neuroforamina. (C) Axial CT image with pig in prone position. Scan obtained after injection of 2000 ng of RTX at the L4/L5 level. Periganglionic distribution of drug/contrast mixture is seen (black arrowheads) surrounding the exiting L4 DRG (black arrow) on the right (needle tip is out of plane of scan). Needle is in place to deliver vehicle solution (not yet injected) to left DRG. Black appearance distal to needle tip is a common CT artifact due to beam hardening along the needle path.

  • Fig. 2. Withdrawal latency to C-fiber activation with laser stimulus.

    (A) Diagram depicts the injected levels in pigs at L2/3, L3/4, L4/5, and L5/6, and the sensory dermatomes innervated by these ganglia (orange shading). C-fiber stimulations were confined to this shaded area. (B) Withdrawal latency at week 2 after injection with a 10.5-s cutoff threshold shows symmetric baseline withdrawal latency in uninjected animals. n = 2 (P = 0.257; left, 6.2 ± 0.4 s; right, 6.5 ± 0.4 s; 68 total stimuli). Significantly delayed or absent response in 500 ng (9.2 ± 0.3 s, n = 2) and 2000 ng (9.5 ± 0.3, n = 3) RTX groups relative to contralateral dermatomes injected with vehicle only (contralateral to 500 ng, 6.8 ± 0.4; contralateral to 2000 ng, 7.8 ± 0.3). Data points reaching 10.5-s cutoff (orange shading) have been staggered slightly to minimize overlap. Individual stimuli, circle or square; mean response time, gray box. (C) Withdrawal latency at week 4 after injection with a 15.5-s cutoff. There is a significantly delayed or absent response in RTX-treated hindlimb (500 ng, 13.7 ± 0.5, n = 3; 2000 ng, 12.1 ± 0.6, n = 4) compared to the contralateral limb injected with vehicle only (500 ng, 9.3 ± 0.6; 2000 ng, 8.5 ± 0.4). (D) RTX-treated dermatomes have an increase in number of trials reaching cutoff (combined 2- and 4-week data): uninjected, 4.4% (3 of 68); 500 ng, 60.7% (37 of 61) versus vehicle control 15.7% (11 of 70); and 2000 ng, 59.6% (53 of 89) versus vehicle control 17.4% (19 of 109). Nonparametric Kruskal-Wallis test with Dunn’s posttest to correct for multiple comparisons.

  • Fig. 3. Relative TRPV1 mRNA expression in treated DRG compared with control.

    Relative TRPV1 expression compared to control at the same lumbar level injected with 500 ng of RTX: 63.9 ± 7.3%, n = 7 DRG (P = 0.02), and with 2000 ng of RTX: 33.7 ± 4.1%, n = 12 DRG (P = 0.0005). Wilcoxon matched pairs test.

  • Fig. 4. Immunofluorescence in L4/5 DRG and nerve terminals in the spinal cord (L5).

    (A to C) TRPV1 IR cell bodies from vehicle- (A) and RTX-injected (B) ganglia. (C) Bar graph depicts a 66% relative decrease in TRPV1 IR cells in RTX-treated (11.1 ± 1.6%, 140 of 1259) versus contralateral ganglia (32.7 ± 3.4%, 476 of 1457; P = 0.002). (D to F) NF200-IR cell bodies were not significantly affected by RTX injection (69.3 ± 3.9%, 567 of 818) (D); vehicle-injected DRG (71.6 ± 4.7%, 985 of 1375; P = 0.27) (E). (G to I) SP-IR cell bodies in RTX-treated ganglia (10.7 ± 2.8%, 99 of 923) (G) compared with contralateral ganglia (25.8 ± 2.7%, 418 of 1621) (H) were decreased by 59% (P < 0.0001). (J to L) CGRP-IR cell bodies from vehicle-injected (27 ± 1.4%, 290 of 1055) (J) and RTX-injected (42 ± 0.7%, 440 of 1044; P = 0.005) (K) ganglia. (L) Bar graph depicts a 35.6% relative decrease. (M to O) CGRP-IR nerve terminals in the dorsal horn (DH) from vehicle- (M) and RTX-injected (N) sides of the same animal with a 40.8 ± 8.5% reduction (O) in CGRP nociceptive terminals. Cell counts are a summation of DRG sections from three animals and are expressed as % IR cells per total cells. Scale bars, 100 μm (A to K) and 300 μm (M and N).

  • Fig. 5. Comparative histology of DRG injected with either vehicle or RTX.

    Hematoxylin and eosin (H&E) preparation of DRG parenchyma from lumbar DRG at ×40 magnification. (A and C) DRG injected with vehicle only demonstrating multiple large-, medium-, and small-diameter cell bodies with a generally thinly dispersed and uniform distribution of satellite cells, typically arrayed linearly along axons and in single cell layers around cell bodies. (B and D) In contrast, DRG injected with RTX demonstrate an overall reduction in the number of ganglion cells and an increase in the number of intervening, multilayered satellite cells (black arrows). Additionally, in some foci, the satellite cells are seen forming small aggregates resembling the structures that have been previously described as nodules of Nageotte (black arrowheads). Scale bar, 50 μm.

Supplementary Materials

  • www.sciencetranslationalmedicine.org/cgi/content/full/7/305/305ra145/DC1

    Fig. S1. Axial CT at the L5 level showing drug/contrast mixture in the ventral epidural space from bilateral periganglionic injections.

    Table S1. Criteria for disability scoring before and after CT-guided injections.

    Table S2. Clinical gait scoring before and after RTX injection.

    Table S3. Withdrawal latency to C-fiber activation with laser stimulus—all animals.

    Movie S1. Withdrawal behavior from infrared laser heat stimulus.

    Movie S2. Withdrawal behavior from noxious mechanical (pinprick) stimulus.

  • Supplementary Material for:

    CT-guided injection of a TRPV1 agonist around dorsal root ganglia decreases pain transmission in swine

    Jacob D. Brown, Maythem Saeed, Loi Do, Joao Braz, Allan I. Basbaum, Michael J. Iadarola, David M. Wilson, William P. Dillon*

    *Corresponding author. E-mail: william.dillon{at}ucsf.edu

    Published 16 September 2015, Sci. Transl. Med. 7, 305ra145 (2015)
    DOI: 10.1126/scitranslmed.aac6589

    This PDF file includes:

    • Fig. S1. Axial CT at the L5 level showing drug/contrast mixture in the ventral epidural space from bilateral periganglionic injections.
    • Table S1. Criteria for disability scoring before and after CT-guided injections.
    • Table S2. Clinical gait scoring before and after RTX injection.
    • Table S3. Withdrawal latency to C-fiber activation with laser stimulus—all animals.
    • Legends for movies S1 and S2

    [Download PDF]

    Other Supplementary Material for this manuscript includes the following:

    • Movie S1 (.mp4 format). Withdrawal behavior from infrared laser heat stimulus.
    • Movie S2 (.mov format). Withdrawal behavior from noxious mechanical (pinprick) stimulus.

    [Download Movie S1]

    [Download Movie S2]

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