The Role of Central and Peripheral Opioid Receptors in Inflammatory Pain and Edema: A Study Using Morphine and DiPOA ([8-(3,3-Diphenyl-propyl)-4-oxo-1-phenyl-1,3,8-triaza- spiro[4.5]dec-3-yl]-acetic Acid)

The role of opioid receptors located in the central nervous system (CNS) and peripheral nervous system in inflammatory pain is well established. In contrast, although it is has been shown that agonists can reduce other manifestations of inflammation, such as edema, the mechanism of action remains unclear. In this study, we have activated receptors located centrally, those located peripherally, and those located both centrally and peripherally and compared the effects on pain and edema using the rat carrageenan model of acute inflammation. Activation of receptors located only in the periphery, by administration of the peripheralized agonist [8-(3,3-diphenylpropyl)-4-oxo-1-phenyl-1,3,8-triaza-spiro[4.5]dec-3-yl]-acetic acid (DiPOA) or local administration of morphine, resulted in antihyperalgesia (30 mg/kg DiPOA, 83% inhibition; 100 g/rat morphine, 75% inhibition) without affecting edema. In contrast, activation of both central and peripheral receptors using systemically administered morphine resulted in antihyperalgesia (1 mg/kg, 80% inhibition) and inhibition of edema (10 mg/kg, 54% inhibition). Finally, activation of only receptors located in the CNS, by central administration of DiPOA or systemic administration of morphine after block of only the peripheral receptors using q-naltrexone, resulted in a significant reduction in edema. Our findings confirm the role of peripheral receptors in the pathology of pain associated with acute inflammation and argue against the involvement of these receptors in edema formation. Furthermore, our data demonstrate that activation of receptors in the brain inhibits carrageenan-induced edema and suggest that the antiedematous effect of morphine is due to action at central receptors alone. Opioid receptors are expressed throughout the central nervous system (CNS), and their activation results in potent analgesia via inhibition of ascending excitatory nociceptive transmissions and activation of descending inhibitory systems (Fields and Basbaum, 1999; Yaksh 1999). In situ hybridization and immunohistochemistry has localized mRNA and protein for the opioid receptor to the peripheral nervous system (PNS), specifically to the cell bodies of primary afferent sensory neurons located in the dorsal root ganglia (Wang and Wessendorf, 2001). Inflammation can modulate peripheral opioid receptor function (Selley et al., 1993; Ingram and Williams, 1994; Antonijevic et al., 1995; Mousa, 2003; Zöllner et al., 2003). Indeed, local administration of exogenous opioids at the site of inflammation has shown therapeutic utility in animals and humans (for review, see Stein et al., 2003). In humans, intra-articular administration of morphine reverses the hyperalgesia associated with osteoarthritis (Stein et al., 1999) or resulting from arthroscopic knee surgery (Kalso et al., 1997). Preclinically the hyperalgesia caused by acute chemical injury to the rat cornea was reversed by direct application of morphine to the surface of the eye (Wenk et al., 2003). In addition, local administration of the agonists loperamide, morphine, or fentanyl is antihyperalgesic in rodent models of inflammatory pain (Stein et al., 1988; Zhou et al., 1998; DeHaven-Hudkins et al., 2002). In addition to producing potent analgesia, morphine can also reduce inflammation. Several studies have demonstrated that systemically administered morphine produces a substantial reduction of inflammagen-induced extravasation (Hargreaves et al., 1988; Joris et al., 1990) and edema (Hargreaves et al., 1988; Joris et al., 1990; Sacerdote et al., 1996; Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. doi:10.1124/jpet.105.088351. ABBREVIATIONS: CNS, central nervous system; PNS, peripheral nervous system; DiPOA, [8-(3,3-diphenyl-propyl)-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]dec-3-yl]-acetic acid; FCA, Freund’s complete adjuvant; i.pl., intraplantar; PWT, paw withdrawal threshold; PLSD, protected least significant difference. 0022-3565/05/3143-1234–1240$20.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 314, No. 3 Copyright © 2005 by The American Society for Pharmacology and Experimental Therapeutics 88351/3046762 JPET 314:1234–1240, 2005 Printed in U.S.A. 1234 at A PE T Jornals on A uust 5, 2017 jpet.asjournals.org D ow nladed from Walker et al., 1996; Alebouyeh et al., 2002; Amann et al., 2002). Furthermore, Sacerdote et al. (1996) and Planas et al. (1995) observed a proinflammatory effect of the opioid antagonists naltrexone and naloxone, respectively. The mechanisms of the antiedematous effect of systemically administered opioids are not clearly understood (Joris et al., 1990; Perrot et al., 1999). Proposed hypotheses include activation of neuronal receptors present in either the CNS or the PNS. Morphine, and other centrally active opioids, reduce muscular tone in conjunction with sedation. This reduction in tone could have effects on local blood flow in peripheral tissues. Alternatively, opioids could reduce edema via interaction with receptors present on cells of the immune system such as leukocytes and macrophages either, circulating or resident at the site of inflammation. In contrast to the well documented anti-inflammatory effect of systemically administered morphine, the effect of locally administered agonists on inflammation-induced edema has received much less attention. [8-(3,3-Diphenyl-propyl)-4-oxo-1-phenyl-1,3,8-triaza-spiro[4.5]dec-3-yl]-acetic acid (DiPOA) is a recently described, highaffinity, high-efficacy, systemically available, and peripherally restricted selective receptor agonist (Valenzano et al., 2004). Systemically administered DiPOA is antihyperalgesic in the Freund’s complete adjuvant (FCA) model of inflammatory pain in a naltrexone-sensitive manner, at doses that do not produce analgesia or sedation (Whiteside et al., 2004). The role of peripheral receptors in pain associated with inflammation is therefore well established. It is not known, however, whether antihyperalgesia due to agonism of peripheral receptors is secondary to a reduction of edema; in fact, the role of peripheral receptors in edema formation is not well understood. We sought to investigate the role of centrally and peripherally located receptors in acute inflammation. The carrageenan model of acute inflammation was chosen because it allows both preemptive and curative actions of compounds to be determined. We have compared the effects of DiPOA administered either systemically or intracerebroventricularly (i.c.v.) with morphine administered either systemically or locally. In addition, we antagonized peripheral receptors by administration of the peripheral antagonist, naltrexone methiodide (qnaltrexone), before systemic administration of morphine. Comparison of these different conditions allowed us to examine the relative contributions of central and peripheral opioid receptors against the carrageenan-induced inflammatory response. Materials and Methods Compounds and Administration Procedures. All reagents are from Sigma-Aldrich (St. Louis, MO) unless otherwise stated. The synthetic route for DiPOA (free base) has been recently disclosed in patent application WO 2003101953 (Victory and Chen, 2003). DiPOA was used in all experiments as its free base (molecular weight 483.6; Ki at 0.76 0.15 nM; solubility in 100 mM K2HPO4 at pH 7.4 50 M). DiPOA was administered either intraperitoneally (i.p.) or i.c.v. in 25% -cyclodextrin dissolved in distilled water in a dose volume of 2 ml/kg or 50 l/rat, respectively. The opioid agonist morphine sulfate was administered either subcutaneously (s.c.) in 0.9% saline in a dose volume of 2 ml/kg or intraplantar (i.pl.) in 0.9% saline in a dose volume of 50 l/rat. The peripheral opioid antagonist q-naltrexone was administered s.c. in 0.9% saline in a dose volume of 2 ml/kg. The nonsteroidal anti-inflammatory drug naproxen hydrochloride served as a positive control and was administered i.p. in 0.9% saline in a dose volume of 2 ml/kg. Animals. The Purdue Institutional Animal Care and Use Committee approved all animal procedures according to the guidelines of the Office of Laboratory Animal Welfare. Male Sprague-Dawley rats (Taconic Farms, Germantown, NY), weighing 180 to 200 g at the start of experiments, were used. Animals were group-housed and had free access to food and water at all times. For comparison with compound-treated groups, animals treated with appropriate drug vehicle were included in each experiment. The volume of administration and all other experimental procedures and conditions for vehicle and compound-treated rats were identical. Carrageenan Model: Inflammatory Hyperalgesia. For this assay, hind paw withdrawal thresholds (PWTs) to a noxious mechanical stimulus were determined using an analygesymeter (model 7200; Ugo Basile, Varese, Italy). Cut-off was set at 250 g, and the endpoint was taken as complete paw withdrawal. PWT was determined once for each rat at each time point. Baseline PWT was determined (predose PWT), the rats were anesthetized with isofluorane (2% in oxygen) and received an intraplantar injection of 2% carrageenan (50 l, diluted in 0.9% saline) to the left hind paw. Compounds were administered either 30 min before carrageenan injection (to investigate the effect on the development of inflammatory hyperalgesia) or 210 min after carrageenan injection (to investigate the effect on an established hyperalgesia). Rats received either a single dose of 3, 10, or 30 mg/kg DiPOA i.p.; a single dose of 1, 3, or 10 mg/kg morphine s.c.; or a single dose of 30, 100, or 200 g/rat morphine i.pl. (animals were briefly anesthetized as described above for i.pl. injections). In each experiment, a positive control (naproxen i.p., 30 mg/kg, 30 min before carrageenan) was included. Four hours after carrageenan injection, PWTs were again measured as described above (postdose PWT). Carrageenan Model: Inflammatory Edema. For this assay, hind paw volume was determined using a plethysmometer (model 7140; Ugo Basile). Paw volume was determined once for each rat at each time point. Baseline paw volume was determined (predose paw volume), the rats were anesthetized with isofluorane (2% in oxygen) and received an intraplantar injection of 2% carrageenan (50 l, diluted in 0.9% saline) to the left hind paw. Compounds were administered either 30 min before carrageenan injection (to investigate the effect on the development of inflammatory edema) or 210 min after carrageenan injection (to investigate the effect on an established edema). Rats received a single dose of either 3, 10, or 30 mg/kg DiPOA i.p.; a single dose of 10 g/rat DiPOA i.c.v.; a single dose of 1, 3, or 10 mg/kg morphine s.c.; or a single dose of 30, 100, or 200 g/rat morphine i.pl. (animals were anesthetized as described above for i.pl. and i.c.v. injections). q-Naltrexone (10 mg/kg s.c.) was administered 10 min before administration of 10 mg/kg morphine s.c. In each experiment, a positive control (naproxen i.p., 30 mg/kg, 30 min before carrageenan) was included. Four hours after carrageenan injection, paw volume was again measured as described above (postdose paw