Anesth Analg 1999;89:403
© 1999 International Anesthesia Research Society
REGIONAL ANESTHESIA AND PAIN MANAGEMENT
Experimental Arthritis in the Rat Does Not Alter the Analgesic Potency of Intrathecal or Intraarticular Morphine
Hartmut Buerkle, MD,
Esther Pogatzki, MD,
Matthias Pauser, MS,
Carsten Bantel, MD,
Gerd Brodner, MD, PhD,
Thomas Möllhoff, MD, MSc, and
Hugo Van Aken, MD, PhD
Klinik und Poliklinik für Anästhesiologie und operative Intensivmedizin, Westfälische Wilhelms-Universität Münster, Münster, Germany
Address correspondence to Hartmut Buerkle, MD, Klinik und Poliklinik für Anästhesiologie und operative Intensivmedizin, Westfälische Wilhelms-Universität Münster, Albert-Schweitzer-Strasse 33, 48129 Münster, Germany. Address e-mail to buerkle{at}uni-muenster.de
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Abstract
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To explore further the role of inflammatory processing on peripheral opioid pharmacology, we examined whether the potency of intraarticular (IA) or intrathecal (IT) morphine in tests of thermal and mechanical nociception changed during the induction of experimental arthritis in the rat. Thermal nociception by IT morphine (3, 10, and 50 µg) or IA morphine (100, 1000, and 3000 µg) was assessed by means of a modified Hargreaves box ever) 28 h. Mechanical antinociception was determined for the largest applied doses of morphine using von Frey hairs. Morphine produced dose-dependent thermal antinociception after IT or IA administration: a 50% increase in maximum antinociceptive thermal response (50% effective dose) was produced by IT doses of 9.7 µg at the start and 9.1 µg at the end of this 28-h observational interval, whereas after IA administration, 50% effective dose values were 553 µg at the start and 660 µg at the end. The largest applied dose of either IT or IA morphine produced mechanical antinociception. On Day 1, the antinociceptive effect for mechanical nociception (expressed as the area under the curve of the percentage of maximal possible effect values at 0.5, 1, 2, and 4 h) was 68% for IT morphine 50 µg and 53% for IA morphine 3000 µg. Neither result differed from the corresponding area under the curve values on Day 2. Naloxone administered either IT or IA abolished the antinociceptive action of morphine given at the same site. We conclude that, although morphine has a peripheral analgesic site of action in a rat arthritis model, its potency for both IA and IT routes of administration does not change during the onset of arthritis.
Implications: In this animal study, we showed that the administration of morphine modulates thermal and mechanical antinociception at central and peripheral sites in inflammatory pain.
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Introduction
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Several studies have indicated that exogenous and endogenous opioids exert antinociceptive effects at peripheral receptor sites, in addition to the well–established central (supraspinal and spinal) site of action (1–4). However, the pharmacologic action of the peripheral analgesic effect of opioids is not yet well characterized. Although there is general agreement regarding the antinociceptive action of opioids in certain inflammatory conditions, there are still equivocal clinical data regarding its effectiveness (5,6). Inflammation clearly activates opioid receptor systems through the breakdown of the perineurium by inflammatory processes, and this results in a facilitated accessibility of opioid receptors (2,3). In addition, "silent" opioid receptors are activated by inflammation, and new synthesis and axonal migration of opioid receptors from the dorsal horn to the peripheral nociceptor terminal are induced through inflammatory modulation (3). These changes eventually result in an enhanced analgesic response to opioids. It has been shown that mainly µ and 
receptors can be activated at the peripheral afferent neuron, which has µ, 
, and receptors (7,8). However, with regard to the relative efficacy of central versus peripheral opioid analgesics, few data are available on the antinociceptive effect in blocking autonomic responses of centrally and peripherally applied opioids (8). In addition, it has not yet been elucidated whether peripheral or central opioid antinociception can be further enhanced during continually inflammation. Thus, to assess further the central and peripheral antinociceptive pharmacology of morphine in a standardized model, we performed the present animal study with the repeated administration of intrathecal (IT) and intraarticular (IA) injections of different morphine doses in a well-defined model of persistent, tonic inflammatory pain (9,10).
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Methods
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Animal surgery and testing protocols were approved by our institutional animal care committee. All procedures were performed according to the guidelines issued by the International Association for the Study of Pain. Male rats (Sprague–Dawley; Harlan Industries, Borchen, Germany; 300–325 g) were kept in individual cages on a 12-h light/dark cycle, with water and food provided ad libitum. Animals were randomly assigned to treatment groups and were only used once. The animals were killed with pentobarbital after the experiment.
For the spinal delivery of study drugs, animals were prepared with IT catheters in a modified version of techniques described previously (11). Immediately after recovery from anesthesia on the day of surgery, animals received 20 µL of IT lidocaine 1% to ensure that the IT catheter was properly located by demonstrating bilateral similar, reversible motor blockade for both hindlimbs. Only animals with normal motor function revealing no gross abnormalities (normal stepping, normal paw withdrawal, and normal righting reflex) after the IT placement were used in subsequent experiments. A postsurgical recovery period of 5 days was allowed for all animals.
Testing for thermal and mechanical nociceptive effects was performed as described previously (10). Briefly, after assessing baseline values for thermal and mechanical nociception, animals were anesthetized with halothane 2–3%. A mixture of 3% kaolin and 3% carrageenan (KC) 0.1 mL was injected into the right knee joint (9,10). Recovery from anesthesia occurred within 5–10 min. For all animals, the degree of the inflammatory response was measured by assessing the circumference of the right knee joint before KC injection (30 min) and 24 h after KC IA injection.
As a measurement of thermal nociception, the thermal withdrawal response (TWR), was assessed by an observer blinded to the compound injected, using a modified technique described previously (12). Briefly, each animal was placed in a clear plastic cage (9 x 22 x 25 cm) on top of a glass plate, with the surface temperature maintained at 30°C using a feedback-controlled heater fan. Before any stimulation, the animal was left on this surface for at least 15 min to allow proper adaptation to the environment. The thermal stimulus, a halogen bulb, was positioned under the glass and focused on either the plantar surface of the ipsilateral paw of the inflamed knee joint or the contralateral paw of the noninflamed knee joint. The stimulus was automatically terminated when paw elevation was sensed by photodiodes or when an interval of 20 s (cutoff time) had passed. The light beam intensity was monitored by a measurement of bulb current, and the stimulus intensity was calibrated daily by assessing the temperature change after 10 s sensed by an underglass thermocouple (t1/2 0.2 s). The intensity of the light was adjusted and maintained at mean (±SD) baseline latencies of 8.2 ± 1.2 s. Withdrawal latencies to the nearest 0.1 s were measured for both paws.
To test mechanical nociception, the mechanical withdrawal response (MWR) was assessed as described previously (13). Rats were placed on an elevated wire mesh bottom in clear plastic cages. Calibrated von Frey hair filaments (0.4, 0.69, 1.20, 2.04, 3.63, 5.49, 8.51, 15.13, 28.84, 75.88 g) (Semmes-Weinstein Monofilaments; Stoelting, Wood Dale, IL) were applied to the ipsilateral paw of the inflamed knee joint from below the mesh floor until a positive sign of pain behavior (withdrawal, licking) was elicited. The Dixon up-and-down method, described by Chaplan et al. (13), was used to assess the threshold. The minimal stimulation period was 5 s, with a maximum of six stimuli per trial and paw.
All experiments were performed in a randomized manner, and each measurement was obtained by an observer blinded to the treatment. Dose-response curves were derived from thermal antinociceptive effects.
Experiment 1: IT Morphine
To assess the effect of IT morphine on thermal nociception in rats with knee joint inflammation, three groups of rats (n = 5) were assigned to receive, according to preliminary pilot studies, single doses of morphine IT (3, 10, 50 µg) in volumes of 10 µL, followed by 10 µL of saline to flush the catheter. Mechanical antinociception was assessed for the IT dose of 50 µg of morphine. After an adaptation period in the plastic cage, each animal was tested on two consecutive days (Day 1: 0–4 h; Day 2: 24–28 h) after receiving the KC injection on Day 1. Baseline values were obtained 30 min before the KC injection. IT morphine was injected 30 min after the induction of knee joint inflammation (KC). Testing of nociception was subsequently performed 30, 60, 120, and 240 min after IT morphine delivery on Day 1 and 24 h later on Day 2 (after 30 min, another IT injection of morphine of the same dose used on Day 1).
Experiment 2: IA Morphine
To assess the effect of IA morphine on thermal nociception in rats with knee joint inflammation, three groups of rats (n = 5) were assigned to receive, according to preliminary pilot studies, single doses of morphine IA (100, 1000, 3000 µg) in volumes of 100 µL. Mechanical antinociception was assessed for the IA dose of 3000 µg of morphine. After an adaptation period, each animal was tested on two consecutive days (Day 1: 0–4 h; Day 2: 24–28 h) after receiving the KC injection on Day 1. Baseline values were obtained 30 min before the KC injection. IA morphine was injected 30 min after the induction of knee joint inflammation (KC). Testing of nociception was subsequently performed 30, 60, 120, and 240 min after IA morphine delivery on Day 1 and 24 h later on Day 2 (after another IA injection of morphine of the same dose used on Day 1). Naloxone IA in 100 µL) was used 10 min before the IA delivery of the largest IA dose of morphine used, in a separate group (n = 5 rats) (150 µL).
The percentage of maximal possible effect (%MPE) of antinociception in animals with inflammation was calculated as follows: 
The area under the curve (AUC) of the %MPE values at 0.5, 1, 2, and 4 h was calculated for inhibition of MWRs (AUC of %MPE). Dose-response curves are presented as the %MPE of thermal analgesia. Statistical analysis of the data was performed by using nonparametric tests. The Mann-Whitney rank-sum test and the Kruskal-Wallis test were used. Multiple comparisons after the Kruskal-Wallis test were performed using the Scheffé test. Statistical significance was set at P < 0.05. For all drugs, the dose-response analysis was conducted as described by Tallarida and Murray (14). The mean 50% effective dose (ED50) and the 95% CIs were calculated using the least-squares linear regression model, with the log dose values being used (14).
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Results
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There was no difference in the baseline values of thermal nociception for all groups and in the nociceptive thresholds for mechanical nociception (Table 1). No significant difference was observed regarding the baseline knee joint circumferences for all groups. After KC injection, no significant difference in the circumferences was shown between the different treatment groups (IT or IA morphine) (Table 2).
Experiment 1: IT Morphine
There was a fast onset of thermal antinociception, with a peak effect at 30 and 60 min, respectively, for the largest dose of IT morphine (50 µg) on Days 1 and 2 (Fig. 1, top). This dose-dependent thermal antinociception did not differ from the ED50 values required on both days (Day 1: 9.7 µg [CI 6.37–14.83]; Day 2: 9.1 µg [CI: 7.25–11.34]) (Fig. 2, top). The MWR was also suppressed by IT morphine at the largest dose (50 µg), with a corresponding time-effect curve to thermal antinociception. The AUC for percent inhibition of the MWR for IT morphine (50 µg) did not differ between the 2 days (P > 0.05) (Fig. 3). The IT injection of the antagonist naloxone before the IT delivery of the largest dose of morphine resulted in a significant suppression of the thermal antinociception (P < 0.05) (Fig. 4).
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Figure 1. Top, Thermal antinociception time-effect curves for the largest applied dose of intrathecal (IT) morphine (50 µg). Each point represents the mean ± SD for five animals. Bottom, Thermal antinociception time-effect curves for the largest applied dose of intraarticular (IA) morphine (3000 µg). Each time point represents the mean ± SD for five animals. Testing was conducted on Days 1 and 2. The percent maximal possible effect (%MPE), where 100%MPE = no withdrawal after 20 s of thermal stimulation.
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Figure 2. The 50% effective dose of intrathecal (IT) or intraarticular (IA) morphine for thermal antinociception. The analgesic response to the thermal stimulus was tested on Days 1 and 2 after IT or IA delivery of morphine. The percent maximal possible effect (%MPE), where 100%MPE = no withdrawal after 20 s of thermal stimulation. Top, The 50% effective dose of IT morphine. There was no significant difference for the contralateral paw of the noninflamed knee joint versus the right ipsilateral paw of the inflamed knee joint for all applied doses of IT morphine (P > 0.05). Bottom, The 50% effective dose of IA morphine. There was a significant difference for the contralateral paw of the noninflamed knee joint versus the right ipsilateral paw of the injected and inflamed knee joint for the two largest applied doses of IA morphine (*P < 0.05).
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Figure 3. Mechanical antinociception displayed for the largest applied dose of intrathecal (IT) or intraarticular (IA) morphine as area under the curve (AUC) (summed percent maximal possible effect [%MPE] values 0.5, 1, 2, and 4 h postdrug). The %MPE, where 100%MPE = no response to 76-g von Frey hair stimulation. Testing was conducted on Days 1 and 2. Each bar presents the mean ± SD for five animals. *Significant difference between the paws after IA injection of morphine on Days 1 and 2 (P < 0.05).
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Figure 4. Suppression of thermal antinociception using the antagonist naloxone at the site of the largest dose of either intrathecal (IT) or intraarticular (IA) morphine delivery. Each bar presents the mean ± SD for five animals. The percent maximal possible effect (%MPE), where 100%MPE = no withdrawal after 20 s of thermal stimulation. *Significant difference between agonist and agonist plus antagonist (P < 0.05).
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Experiment 2: IA Morphine
IA morphine resulted in dose-dependent inhibition of the TWR, with the ED50 values on the 2 days being 553 µg (CI 365.78–836.67) and 660 µg (CI 444.89–979.04) (Fig. 2, bottom). With an increase of thermal antinociception in the ipsilateral paw, there was a concomitant increase observed for the contralateral paw of the noninflamed, noninjected knee joint. However, the latency increase for the ipsilateral paw of the treated, inflamed knee joint was statistically significantly different after the injection of 1000 µg of morphine IA on Days 1 and 2 (P < 0.05). The IA injection of 3000 µg of morphine resulted in an increased thermal antinociception for both the ipsilateral and the contralateral paws. As with IT delivery, there was a fast onset of action observed after 60 min on Days 1 and 2 (Fig. 1, bottom) for suppression of the TWR.
The MWR was also suppressed by IA morphine at the largest dose (3000 µg), with a time-effect curve corresponding to that for thermal antinociception. The %AUC for inhibition of the MWR after IA morphine (3000 µg) differed between the 2 days (53% ± 5% vs 38% ± 8%; P < 0.05) (Fig. 3).
The IA injection of the antagonist naloxone before IA delivery of the largest dose of morphine resulted in significant suppression of thermal antinociception (P < 0.05) (Fig. 4).
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Discussion
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The peripheral analgesic activity of morphine has been demonstrated in preclinical and clinical studies (1,5,6). These showed that inflammation is mandatory for activation of peripherally distributed opioid receptors (2,3), inducing synthesis and migration of these receptors from central to peripheral sites and facilitating their accessibility to exogenously delivered opioids. However, it has not been determined whether continuing inflammation further enhances opioid antinociception and results in a dose-response shift in the antinociception. In the present study, using a well defined model of persistent tonic inflammatory pain (9,10,16), we did not find this type of leftward shift for the inhibition of thermal hyperalgesia, nor was there a more profound suppression of mechanical hyperalgesia for the largest applied dose of morphine on the second day of inflammation. These findings were not dependent on the route of administration (i.e., IT or IA injection).
As demonstrated in previous studies, the IA injection of antinociceptive drugs provides a reliable model for the identification of peripherally mediated analgesia (15,16). The model used in this study—IA injection of KC to induce inflammatory pain—has several distinct advantages compared with other models, such as the intraplantar injection of Freund adjuvants and subsequent analgesic injections. The IA injection of opioids and other drugs into the closed compartment of the knee-joint results in a reduced probability of systemic drug reabsorption that might be higher after intraplantar injection (15). In addition, this model has been characterized for the time course of the thermal and mechanical nociception, including the presumed underlying mechanisms, the release of excitatory neurotransmitters at the spinal cord level (9,10). The IA opioid injection effect on induced autonomic responses in rats has also been characterized (8). In the present study, peripherally applied morphine resulted in thermal and mechanical antinociception. In agreement with the findings of the study by Nagasaka et al. (8), this peripheral analgesic action of morphine is further supported by the observation that local naloxone attenuated the effects of IA morphine. The peripheral delivery of opioids has been advocated because it might reduce typical µ opioid-related side effects (pruritus, respiratory depression, nausea, etc.) (1). However, although no decrease in knee joint circumferences was observed in the limited time period of our experimental design, morphine may be beneficial by affecting the inflammatory cascade after peripheral administration (1). Clinical data on the analgesic effect of peripherally active opioids (i.e., IA morphine) are equivocal (5). Both routes of morphine administration result in similar antinociception, but the IA doses required were approximately 60 times larger, which suggests a higher potency of centrally delivered morphine. This observed difference in potency between the two routes might be due to a higher level occupancy being required to block the transduction at the peripheral nociceptor (8). Although the efficacy of peripheral opioid analgesia is equivalent to that of spinal injection, a difference in occupancy would support the controversial clinical data obtained with locally administered opioids. However, we cannot exclude the onset of acute opioid tolerance by the single IA injection of morphine and its impact on the dose-response curves obtained; yet, data only support acute opioid tolerance with a single IT morphine dose (17). In conclusion, we contend that peripheral antinociception mediated by morphine is not further enhanced during continuing inflammatory pain. Both routes (IT and IA opioid administration) result in a dose-dependent antinociceptive effect, for which the observed difference in potency may be due to different pharmacokinetics and pharmacodynamics associated with the two routes of morphine administration.
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Acknowledgments
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This study was funded by a grant from the Klinik und Poliklinik für Anästhesiologie und operative Intensivemedizin.
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References
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-
Stein C, Schafer M, Hassan AH. Peripheral opioid receptors. Ann Med 1995;27:219–21.[Web of Science][Medline]
-
Antonijevic I, Mousa SA, Schafer M, Stein C. Perineurial defect and peripheral opioid analgesia in inflammation. Neurosci 1995;15:165–72.[Abstract]
-
Hassan AH, Ableitner A, Stein C, Herz A. Inflammation of the rat paw enhances axonal transport of opioid receptors. Neuroscience 1993;55:185–95.[Web of Science][Medline]
-
Schafer M, Imai Y, Uhl GR, Stein C. Inflammation enhances peripheral mu-opioid receptor-mediated analgesia, but not mu-opioid receptor transcription in dorsal root ganglia. Pharmacol 1995;279:165–9.
-
Kalso E, Tramer MR, Carroll D, et al. Pain relief from intra-articular morphine after knee surgery: a qualitative systemic review. Pain 1997;71:127–34.[Web of Science][Medline]
-
Picard PR, Tramer MR, McQuay HJ, Moore RA. Analgesic efficacy of peripheral opioids (all except intra-articular): a meta-analysis. Pain 1997;72:309–18.[Web of Science][Medline]
-
Barber A. Mu- and kappa-opioid receptor agonists produce peripheral inhibition of neurogenic plasma extravasation in rat skin. Eur J Pharmacol 1993;236:113–20.[Web of Science][Medline]
-
Nagasaka H, Awad H, Yaksh TL. Peripheral and spinal actions of opioids in the blockade of the autonomic response evoked by compression of the inflamed knee joint. Anesthesiology 1996;85:808–16.[Web of Science][Medline]
-
Yang LC, Marsala M, Yaksh TL. Characterization of time course of spinal amino acids, citrulline and PGE2 release after carrageenan/kaolin-induced knee joint inflammation: a chronic microdialysis study. Pain 1996;67:345–54.[Web of Science][Medline]
-
Buerkle H, Boschin M, Marcus M, et al. Central and peripheral analgesia mediated by the acetylcholinesterase inhibitor neostigmine in the rat inflamed knee joint model. Anesth Analg 1998;86:1027–32.[Abstract]
-
Yaksh TL, Rudy TA. Analgesia mediated by a direct spinal action of narcotics. Science 1976;192:1357–8.[Abstract/Free Full Text]
-
Hargreaves K, Dubner R, Brown F, et al. A new and sensitive method for measuring thermal nociception in cutaneous hyperalgesia. Pain 1988;32:77–88.[Web of Science][Medline]
-
Chaplan SR, Bach FW, Pogrel JW, et al. Quantitative assessment of tactile allodynia in the rat paw. J Neurosci Methods 1994;53:55–63.[Web of Science][Medline]
-
Tallarida RJ, Murray RB. Manual of pharmacologic calculations with computer programs. New York:Springer, 1986.
-
Gentili M, Juhel A, Bonnet F. Peripheral analgesic effect of intra-articular clonidine. Pain 1996;64:593–6.[Web of Science][Medline]
-
Tonussi CR, Ferreira SH. Rat knee-joint carrageenin incapacitation test: an objective screen for central and peripheral analgesics. Pain 1992;48:421–7.[Web of Science][Medline]
-
Fairbanks CA, Wilcox GL. Acute tolerance to spinally administered morphine compares with chronically induced morphine tolerance. J Pharmacol Exp Ther 1997;282:1408–17.[Abstract/Free Full Text]
Accepted for publication April 4, 1999.
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Abstract
Introduction
