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note
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125I]α-bgt to membranes of T. californica electroplax was done according to methods previously described [see (9)].
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24
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0029146901
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To assess nociceptive responses to an acute thermal stimulus, a commercially available paw thermal stimulator was used (Anesthesiology Research Laboratory, Department of Anesthesiology, University of California at San Diego, La Jolla, CA). This device has been previously described [D. M. Dirig and T. L. Yaksh, Pain 62, 321 (1995)] and is based on the initial work of Hargreaves et al. [K. Hargreaves, R. Dubner, F. Brown, C. Flores, J. Joris, ibid. 32, 77 (1988)]. Briefly, rats were placed in Plexiglas cubicles that were located on a glass, surface of the apparatus. The surface of the glass was maintained at 30°C. A thermal stimulus was applied to the bottom of the rear foot of the rat by means of a movable focused projection bulb. The stimulus current was maintained at 4.8 A. The latency until the animal moved its foot from the stimulus was recorded automatically by photodiode motion sensors. In the current studies, a 20-s cutoff was employed to limit possible tissue damage after exposure to the stimulus. For measurements, the following protocol was used. Six animals were used in each run. For each measure (that is, each time point), one foot of each of the six animals was tested and then the process was repeated for the opposite foot. Mean values for the response were then computed based on the two scores.
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Pain
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Dirig, D.M.1
Yaksh, T.L.2
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0023924046
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To assess nociceptive responses to an acute thermal stimulus, a commercially available paw thermal stimulator was used (Anesthesiology Research Laboratory, Department of Anesthesiology, University of California at San Diego, La Jolla, CA). This device has been previously described [D. M. Dirig and T. L. Yaksh, Pain 62, 321 (1995)] and is based on the initial work of Hargreaves et al. [K. Hargreaves, R. Dubner, F. Brown, C. Flores, J. Joris, ibid. 32, 77 (1988)]. Briefly, rats were placed in Plexiglas cubicles that were located on a glass, surface of the apparatus. The surface of the glass was maintained at 30°C. A thermal stimulus was applied to the bottom of the rear foot of the rat by means of a movable focused projection bulb. The stimulus current was maintained at 4.8 A. The latency until the animal moved its foot from the stimulus was recorded automatically by photodiode motion sensors. In the current studies, a 20-s cutoff was employed to limit possible tissue damage after exposure to the stimulus. For measurements, the following protocol was used. Six animals were used in each run. For each measure (that is, each time point), one foot of each of the six animals was tested and then the process was repeated for the opposite foot. Mean values for the response were then computed based on the two scores.
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Pain
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Hargreaves, K.1
Dubner, R.2
Brown, F.3
Flores, C.4
Joris, J.5
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0017643233
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The method used for the formalin test was based on a modified version of a previously published method [D. Dubuisson and S. G. Dennis, ibid. 4, 161 (1977)]. After a 20-min period of acclimation to individual cages, 50 μl of a 5% formalin solution was injected sc into the dorsal aspect of one of the rear paws, and the rats were then returned to the clear observation cages suspended above mirror panels. Only phase 2 of the formalin test was scored, and phase 2 was defined as the 20-min period of time from 30 to 50 min after formalin injection. The investigator recorded nocifensive behaviors in the injected paw of four animals during the session by observing each animal for one 15-s observation period during each 1-min interval. Nocifensive behaviors recorded included flinching, licking, or biting the injected paw. In dose-response studies, ABT-594 (or saline) was administered 5 min before the injection of formalin. In antagonist studies, the antagonists or saline were administered 10 min before ABT-594 treatment.
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Pain
, vol.4
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Dubuisson, D.1
Dennis, S.G.2
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F. V. Abbott, R. Melzack, B. F. Leber, Pharmacol. Biochem. Behav. 17, 1213 (1982); H. L. Tripathi, B. R. Martin, M. D. Aceto, J. Pharmacol. Exp. Ther. 221, 91 (1982).
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F. V. Abbott, R. Melzack, B. F. Leber, Pharmacol. Biochem. Behav. 17, 1213 (1982); H. L. Tripathi, B. R. Martin, M. D. Aceto, J. Pharmacol. Exp. Ther. 221, 91 (1982).
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Tripathi, H.L.1
Martin, B.R.2
Aceto, M.D.3
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15144360055
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in preparation
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A. W. Bannon et al., in preparation.
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Bannon, A.W.1
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0027477465
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T. J. Coderre, J. Katz, A. L. Vaccarino, R. Melzack, Pain 52, 259 (1993); A. Tjolsen, O. G. Berge, S. Hunskaar, J. H. Rosland, K. Hole, ibid. 51, 5 (1992).
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Pain
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Coderre, T.J.1
Katz, J.2
Vaccarino, A.L.3
Melzack, R.4
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31
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T. J. Coderre, J. Katz, A. L. Vaccarino, R. Melzack, Pain 52, 259 (1993); A. Tjolsen, O. G. Berge, S. Hunskaar, J. H. Rosland, K. Hole, ibid. 51, 5 (1992).
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Pain
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Tjolsen, A.1
Berge, O.G.2
Hunskaar, S.3
Rosland, J.H.4
Hole, K.5
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S. H. Kim and J. M. Chung, ibid. 50, 355 (1992). A within-subjects design in which all animals received all treatments was used for dose-response studies in the Chung model. Before the start of drug studies, baseline allodynia scores were determined for all animals. Only rats with threshold scores ≤4 g were considered allodynic and were used in further testing. Drug studies (separate studies for each compound) began approximately 2 weeks after the nerve ligation surgery. For dose-response experiments, animals were tested over a 2-week period. Test days were separated by 2-to 3-day intervals during which no testing was conducted and no treatment was given. On test days, animals were placed in the individual chambers and allowed to acclimate for 15 to 20 min. After acclimation, baseline scores were determined. Next, animals were treated and then scores were determined 15, 30, 60, and 120 min after treatment. This procedure was repeated on test days until each animal had received all treatments for any given drug. The treatment order was counterbalanced across animals. For statistical analysis, the time point of peak effect was compared. This was 15 min for ABT-594 and (-)-nicotine and 30 min for morphine.
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Pain
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Kim, S.H.1
Chung, J.M.2
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Z. Wiensenfeld-Hallin et al., Neurosci. Lett. 52, 199 (1984); S. De Biasi and A. Rustioni, Proc. Natl. Acad. Sci. U.S.A. 85, 7820 (1988); R. A. Cridland and J. L. Henry, Brain Res. 462, 15 (1988).
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Cridland, R.A.1
Henry, J.L.2
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note
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Spinal cord slices (250 μm thick) of the dorsal half of the lumbar enlargement were continuously supertused with oxygenated buffer, and fractions were collected every 3 min. After a 36-min equilibrium period, the tissue was superfused with 1 μM capsaicin in the absence and presence of 30 μM ABT-594 (n = 4 per group) or 30 μM ABT-594 and 100 μM mecamylamine (n = 4 per group) for 6 min. SP-like immunoreactivity (SP-LI) was assayed in both perfusates and lysates by radioimmunoassay (Peninsula Laboratories, Belmont, CA). Release was expressed as a percentage of the peptide content per 3-min fraction [(peptide content in perfusate)/(peptide content in perfusale + peptide content in tissue lysate)]. Release detected both during (6 min) and 3 min after capsaicin treatment was significantly higher than the baseline value. For this reason, data are expressed as the sum of SP-LI content measured in the three fractions (9 min) immediately following treatment. The value for the capsaicin treatment alone was 0.2 ± 0.03%. The value for the capsaicin treatment + 30 μM ABT-594 was 0.07 ± 0.02%; which is significantly different from capsaicin treatment alone (P < 0.05). In the capsaicin + 30 μM ABT-594 + mecamylamine (100 μM) group, the percent of SP-LI measure was 0.14 ± 0.3, which indicates blockade of the effect of ABT-594 by mecamylamine.
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0031043244
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Electrophysiological studies were conducted in anesthetized Sprague-Dawley rats as previously described [A. Diaz and A. H. Dickenson, Pain 69, 93 (1997)]. Briefly, for extracellular recordings, a tungsten electrode was lowered into the dorsal horn of the spinal cord, and recordings were made from convergent neurons (n = 5) within the dorsal horn that responded to both noxious thermal (4°C) and mechanical (50g) stimuli, as well as to non-noxious thermal (40°C) and mechanical (5g) stimuli. The depth of the cells from the spinal cord surface was established with the use of a SCAT microdriver (Digitimer, Hertfordshire, UK), and the cells used in this experiment were located deep in the dorsal horn with a mean depth ranging from 741 to 817 μm. During recording, animals were spontaneously breathing, and the level of halothane was kept at 2 to 2.5% to maintain complete areflexia.
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(1997)
Pain
, vol.69
, pp. 93
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Diaz, A.1
Dickenson, A.H.2
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note
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ABT-594 treatment inhibited responses to noxious mechanical and thermal stimuli by 70 and 61%, repectively. ABT-594 treatment had no effect on responses of these neurons to non-noxious mechanical and thermal stimuli.
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0002158209
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P. Wall and R. Melzack, Eds. Churchill Livingstone, Edinburgh
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T. L. Yaksh and A. B. Malmberg, in Textbook of Pain, P. Wall and R. Melzack, Eds. (Churchill Livingstone, Edinburgh, 1994), pp. 165-200; in Progress in Pain Research, H. L. Fields and J. C. Liebeskind, Eds. (IASP Press, Seattle, WA, 1994), pp. 151-171.
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(1994)
Textbook of Pain
, pp. 165-200
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Yaksh, T.L.1
Malmberg, A.B.2
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IASP Press, Seattle, WA
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T. L. Yaksh and A. B. Malmberg, in Textbook of Pain, P. Wall and R. Melzack, Eds. (Churchill Livingstone, Edinburgh, 1994), pp. 165-200; in Progress in Pain Research, H. L. Fields and J. C. Liebeskind, Eds. (IASP Press, Seattle, WA, 1994), pp. 151-171.
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(1994)
Progress in Pain Research
, pp. 151-171
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Fields, H.L.1
Liebeskind, J.C.2
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note
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Immunohistochemistry of free-floating sections (anti-Fos staining) was as follows. After overnight incubation in 20% sucrose-phosphate-buffered saline (PBS), brains from saline-and ABT-594-treated rats were cut on a cryostat (in 40-μm coronal sections). Free-floating sections were immunostained for Fos protein with a three-step ABC-peroxidase technique, beginning with a 30-min incubation with blocking serum. Sections were next incubated with antibody (Ab) to Fos (sheep polyclonal immunoglobulin G, 1:2000; Genosys, The Woodlands, TX) for 48 hours at 4°C, washed with PBS, and incubated for 1 hour with a biotinylated secondary sheep Ab solution (1:200). Finally, sections were washed in PBS, incubated with ABC reagent (Vector), and then developed in a peroxidase substrate solution. The sections were mounted and cover-slipped, then examined and photographed with a light microscope (Leica, DMRB). Fos-like immunoreactivity (FLI) was quantitated with an image analysis system (Leica, Quantimet 500).
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Rats were anesthetized with sodium pentobarbital (55 mg/kg, ip). Animals were placed in a David Kopf student stereotaxic instrument (Tujunga, CA) with the skull on an even horizontal plane. For NRM cannula, the coordinates from infra-aural zero were AP -2.5 mm, lateral 0.0 mm, and -0.5 mm deep. Animals were implanted with a C315G (26-gauge) guide cannula (Plastics One, Roanoke, VA) cut to 7 mm. The guide cannula was implanted so that the injector cannula (33 gauge) extended 4 mm beyond the tip of the guide. For NRM injections, 0.3 μl was delivered over 60 s. and the cannula was left in place for another 40 s.
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S. R. Chaplan, F. W. Bach, J. W. Pogrel, J. M. Chung, T. L. Yaksh, J. Neurosci. Methods 53, 55 (1994).
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J. Neurosci. Methods
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Chaplan, S.R.1
Bach, F.W.2
Pogrel, J.W.3
Chung, J.M.4
Yaksh, T.L.5
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unpublished observations
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R. Porsolt, unpublished observations.
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Porsolt, R.1
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note
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All experiments involving animals were conducted under protocols approved by the Institutional Animal Care and Use Committee of Abbott Laboratories. The authors acknowledge the synthetic contributions of J. Lynch, J. Wasicak, and H. Bai; the biological contributions of D. Anderson and D. J. B. Kim; and J. Daly for his pioneering work in the field and his thoughtful input to this manuscript.
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