Synergistic antinociceptive effects of ketamine and morphine in the orofacial capsaicin test in the rat

BACKGROUND: The clinical efficacy of the noncompetitive N-methyl-d-aspartate receptor antagonist ketamine for treating orofacial pain has already been reported. Side effects related to psychotomimetic disturbances, however, limit ketamine use as an analgesic. Theoretically, this limitation could be minimized by using low doses of ketamine in combination with other analgesics. In the present study, the potential synergistic antinociceptive interaction between ketamine and morphine in the orofacial capsaicin test in rats was investigated. METHODS: Male Sprague-Dawley rats were subcutaneously injected with solvent, ketamine, morphine, or combination of both drugs. Thirty minutes later, the orofacial capsaicin test was performed by injecting of 1.5 microg/25 microl of a capsaicin solution into the vibrissa pad. Animal behavior was recorded on videotape and analyzed off-line. The total time spent on rubbing-scratching nociceptive behavior during a period of 42 min was measured. RESULTS: Subcutaneously administered ketamine (0.4, 1.25, 4, 12.5 mg/kg), morphine (0.5, 1, 2, 4 mg/kg) and ketamine + morphine (0.20 + 0.12, 0.40 + 0.24, 0.80 + 0.49, 1.61 + 0.97, 3.21 + 1.94 mg/kg) reduced the rat facial rubbing-scratching behavior in a dose-dependent manner. Isobolographic analysis showed that the ketamine + morphine association inhibited the studied behavior in a superadditive manner. CONCLUSIONS: These results indicate that ketamine and morphine have antinociceptive effects on the orofacial capsaicin test. Furthermore, their combination produces synergistic antinociception. It is therefore suggested that, used together, ketamine and morphine might be clinically efficient at lower doses than those currently used when administered separately. This could provide a useful strategy for the clinical management of orofacial pain.     

Cervicomedullary intrathecal injection of morphine produces antinociception in the orofacial formalin test in the rat.

BACKGROUND: High cervical and medullary drug delivery has been advocated for the treatment of refractory head and neck pain in humans. Currently, parallel models in animals have not been developed to support this methodology. We combined an accepted animal model of pain of cranial origin with a novel technique of neuraxial drug delivery to address this issue. METHODS: Male Wistar rats were implanted with intrathecal catheters that were advanced cephalad through a lumbar guide cannula to the high cervical spinal cord. The orofacial formalin test was used to assess antinociception. Vehicle or morphine (1, 3, 6, 10, 30 microg) was injected intrathecally followed 10 minutes later by injection of formalin solution, 2.5%, into the vibrissal pad. Motor assessment and hemodynamic and respiratory blood gas measurements were evaluated in a separate group of animals. RESULTS: Intrathecal morphine produced a dose-dependent decrease in the first and second phases of the behavioral response (P < 0.05). The ED50 (95% confidence limits) values for the first and second phases were 6.65 microg (3.52-14.9 microg) and 3.40 microg (2.37-4.61 microg), respectively. Ten micrograms intrathecal naloxone antagonized the morphine effect (P < 0.05). Significant cardiovascular and respiratory depression was observed. No significant motor dysfunction was observed. CONCLUSIONS: Cervicomedullary injection of morphine produced antinociception in the orofacial formalin test in the rat. This animal model may be useful to assess analgesics designed for parallel clinical application in humans.     

NOS inhibitors exhibit antinociceptive properties in the rat formalin test

PURPOSE: To assess the systemic and nociceptive effects of nitric oxide synthase (NOS) inhibitors in the modulation of acute pain in rats subjected to the formalin test. METHODS: Formalin 5% was injected in the hind paw in the presence and absence of NOS inhibitors (e.g., 7-nitro indazole, N-nitro-L-arginine and aminoguanidine). Catheters were chronically implanted to continuously record mean arterial blood pressure (MAP) and heart rate (HR). MAP, HR and paw lifting time were recorded at control and every five minutes for 35 min following formalin and NOS inhibitors. RESULTS: Formalin injected into the rat hind paw induced a biphasic nociceptive behaviour: an initial acute phase (phase 1: during zero to five minutes after the formalin injection) followed by a prolonged tonic response (phase 2: beginning about ten minutes after the formalin injection). Aminoguanidine, an inhibitor of the inducible NOS and 7-nitro indazole, an inhibitor of the neuronal NOS, did not affect phase 1, whereas N-nitro-L-arginine, a non-selective NOS inhibitor decreased it (49%). All three NOS inhibitors diminished nociceptive behaviours during phase 2. L-arginine reversed antinociceptive effects of N-nitro-L-arginine in phase 1 and in phase 2. Pressor effects induced by formalin in phase 1 were abolished following all three NOS inhibitors. During phase 2, formalin-induced pressor effects remained unaffected by N-nitro-L-arginine and aminoguanidine but were inhibited by 7-nitro indazole. CONCLUSION: Our data demonstrate that NO is predominantly generated by vascular endothelial NOS in phase 1 and phase 2, whereas the neuronal NOS and the inducible NOS exhibit antinociceptive effects through a non-NO related pathway in phases 1 and 2 in rats subjected to the formalin test.     

Gabapentin produces dose-dependent antinociception in the orofacial formalin test in the rat

BACKGROUND AND OBJECTIVES: High neuraxial drug infusion has been advocated for the treatment of intractable cranial and facial pain in humans. Currently, parallel animal models have not been characterized to support this methodology. We combined an accepted animal model of pain of cranial origin with a novel technique of cervico-medullary drug delivery to determine the antinociceptive potential of gabapentin. Gabapentin was chosen because of its reported efficacy in a wide array of complex cranial pain syndromes. METHODS: Male Wistar rats were implanted with intrathecal catheters that were advanced cephalad through a lumbar guide cannula to terminate in the high cervical spinal cord (C1-C4). Antinociception was assessed by the orofacial formalin test. Vehicle or gabapentin (3, 10, 30, 100 microg) was injected intrathecally followed 10 minutes later by injection of 2.5% formalin solution into the vibrissal pad. Motor assessment was evaluated in a separate group of animals. RESULTS: Intrathecal gabapentin (10, 30, 100 microg) produced a dose-dependent decrease in the second phase of the behavioral response to formalin (P <.05). First-phase responses were unaffected by all doses of gabapentin. The ED50 (95% confidence limit) value for the second phase was 8.27 microg (3.50 to 14.5). No overt motor dysfunction or behavioral impairment was observed. CONCLUSIONS: Gabapentin produced dose-dependent antinociception in the second phase of the orofacial formalin test in the rat after injection into the cervico-medullary cerebrospinal fluid. This animal model may be useful to assess analgesics designed for parallel clinical application in humans for the treatment of intractable head and neck pain that is refractory to conventional modalities.     

The interaction between gabapentin and amitriptyline in the rat formalin test after systemic administration

We examined the effects of systemically administered gabapentin on flinching and biting/licking behaviors produced by 2.5% formalin in the rat, compared these with those of amitriptyline, and determined the effects of combinations of gabapentin with amitriptyline. Gabapentin produced a dose-related inhibition of Phase 2, but not Phase 1, flinching and biting/licking behaviors. In contrast, amitriptyline produced an increase in Phase 2 flinching behaviors while simultaneously decreasing biting/licking behaviors. Fifty percent effective dose (ED50) values against biting/licking behaviors were 22.9 +/- 1.3 mg/kg and 8.5 +/- 1.3 mg/kg for gabapentin and amitriptyline, respectively. Combinations of increasing fractional increments of ED50 doses of gabapentin and amitriptyline produced an additive effect against biting/licking behaviors, as revealed by isobolographic analysis. These increments had no effect on flinching behaviors except at the ED25 + ED25 doses, at which flinching was increased, again revealing additivity between the two drugs. Flinching behaviors in rats do not reflect the analgesic properties of systemically administered amitriptyline observed in humans and may not be useful for predicting an effect of combinations of drugs with amitriptyline. Biting/licking behaviors do reflect analgesic properties for both drugs and may be more useful in this regard. IMPLICATIONS: By use of the rat formalin test, a model of persistent pain, we examined the effect of a combination of amitriptyline and gabapentin, which are used to treat chronic pain in humans. The drug combination produced additive analgesia against one outcome, but another outcome was more ambiguous.     

Synergistic antinociception between zaprinast and morphine in the spinal cord of rats on the formalin test

BACKGROUND AND OBJECTIVE: The cyclic guanosine monophosphate level, which causes an antinociception, is increased in cells as a direct result of phosphodiesterase inhibition. This study used a nociceptive test to examine the nature of the pharmacological interaction between intrathecal zaprinast, a phosphodiesterase inhibitor, and morphine. METHODS: Catheters were inserted into the intrathecal space through an incision in the atlantooccipital membrane of male Sprague-Dawley rats. As a nociceptive model, 50 microL of a 5% formalin solution was injected into the hind paw. After observing the effect of zaprinast (37, 111, 369 nmol) and morphine (1, 4, 10, 40 nmol) alone, the interactions of their combination were examined by an isobolographic analysis. RESULTS: Intrathecal zaprinast (P < 0.05) and morphine (P < 0.05) dose-dependently suppressed the flinching observed during phase 1 and phase 2 in the formalin test. The ED50 values (95% confidence intervals) of zaprinast and morphine in phase 1 were 161.9 (87.9-298.3) and 11.6 nmol (4.8-27.9 nmol), respectively. The phase 2 ED50 values (95% confidence intervals) of zaprinast and morphine were 229.9 (142.5-370.9) and 3.9 nmol (1.9-7.6 nmol), respectively. Isobolographic analysis revealed a synergistic interaction after intrathecal delivery a zaprinast-morphine mixture in both phases. The ED50 values of (95% confidence intervals) zaprinast in the combination of zaprinast with morphine in phase 1 and phase 2 were 14.2 (4.9-40.6) and 10.4 nmol (3-35.9 nmol), respectively. CONCLUSIONS: Intrathecal zaprinast and morphine are effective against acute pain and facilitated pain state. Zaprinast interacts synergistically with morphine.     

Comparison of the antinociceptive effects of pre- and posttreatment with intrathecal morphine and MK801, an NMDA antagonist, on the formalin test in the rat.

Current thinking emphasizes that protracted small afferent input can evoke mechanisms that mediate a significant potentiation of spinal nociceptive processing and that this facilitory component has a unique pharmacology. To investigate the behavioral parallels of this spinal facilitation, we evaluated the effects of pre- and post-treatment of intrathecal morphine (mu agonist) and MK801 (N-methyl-D-aspartate [NMDA] antagonist) on the formalin test. Intraplantar formalin resulted in a biphasic appearance of flinching behavior (phase 1 = 0-5 min; phase 2 = 10-60 min). Morphine and MK801 were administered intrathecally 15 min before formalin injection in the pretreatment study and 9 min after formalin injection in the posttreatment study. Pretreatment with intrathecal morphine produced comparable dose-dependent suppressions of the phase 1 and phase 2 behaviors (ED50 = 0.5 micrograms [95% CI = 0.3-0.9] and 0.3 micrograms [95% CI = 0.1-0.7], respectively). Posttreatment with morphine also resulted in comparable suppression of the phase 2 response (ED50 = 0.2 micrograms [95% CI = 0.1-0.3]). At the highest dose of intrathecal morphine (10 micrograms), an almost complete suppression of formalin-evoked behavior was observed. Pretreatment with MK801 inhibited the second-phase response more strongly than the first-phase response (ED50 = 1.6 micrograms [95% CI = 0.5-5.7] vs. 0.1 microgram [95% CI = 0.3 - 0.4], respectively). In contrast, posttreatment with the highest dose of MK801 had no effect on the phase 2 response.(ABSTRACT TRUNCATED AT 250 WORDS)     

Orally administered paracetamol does not act locally in the rat formalin test: evidence for a supraspinal, serotonin-dependent antinociceptive mechanism

BACKGROUND: The mechanism of action of paracetamol (acetaminophen) remains elusive because it is still under discussion as to whether it acts locally and/or centrally. The primary aim of this study was to clarify its site(s) of action (central and/or local) using the rat formalin test. METHODS: Spontaneous biting and licking of the injected paw following intraplantar injection of formalin 2.5% was monitored during the two phases of nociceptive behavior (0-5 and 20-40 min after injection), and the authors examined the antinociceptive activity of paracetamol following oral, intravenous, intraplantar, and intrathecal administrations as well as the reversion of this effect by an intrathecal injection of WAY 100,635, a selective 5-HT1A receptor antagonist. RESULTS: The oral administration of paracetamol (300, 400 mg/kg) reduced nociceptive behavior in both phases (400 mg/kg: 36.9 +/- 4.6% and 61.5 +/- 5.2% of inhibition in phases I and II, respectively, P <0.05), whereas lower doses reduced primarily the score of the second phase of the test. Only high doses of 10 to 20 mg/kg intraplantarly administered paracetamol, which were ineffective when administered subcutaneously, produced a significant but limited reduction in the early phase of the test and had no effect on the second phase or any antiinflammatory activity. Thus, this local effect did not seem to participate in the antinociceptive action of 400 mg/kg orally given paracetamol, which was totally blocked in both phases by an intrathecal injection of 40 microg WAY 100,635 per rat. Such an inhibition was not observed when paracetamol (200 microg per rat) was intrathecally coinjected with WAY 100,635, whereas the antinociceptive action of 5-HT (50 microg per rat, intrathecally) during both phases of pain was inhibited by WAY 100,635 (intrathecally). CONCLUSIONS: Orally administered paracetamol does not seem to exert any relevant local action in the formalin model of tonic pain in rats, but it might activate the serotonergic bulbospinal pathways via a supraspinal site of action that remains to be elucidated.     

The antinociceptive effect of the combination of spinal morphine with systemic morphine or buprenorphine.

We sought to analyze the mode of interaction of spinal morphine with systemic morphine or buprenorphine, administered in a wide range of antinociceptive doses. The study was performed on Sprague-Dawley rats by using a plantar stimulation test and isobolographic and fractional analyses of drug interaction. The isobolographic and fractional analyses demonstrated that intrathecal morphine interacted with subcutaneous morphine in a synergistic manner while producing a 50% or 75% antinociceptive effect. The sum of D(75) fractions was more than that for 50% antinociception, suggesting a less dramatic interaction. The combination with a maximal relative dose of systemic morphine (0.66:1) showed a maximal degree of supraadditivity. The interaction between spinal morphine and systemic buprenorphine was similar to that of morphine/morphine, although the supra-additivity was not as pronounced. For the doses that produced a 50% antinociceptive effect, a synergistic interaction was observed only for the combination with a morphine/buprenorphine ratio of 1.33:1. When the relative amount of intrathecal morphine was decreased or increased, the effect became additive. At the doses that produced 75% antinociception, both combinations of morphine and buprenorphine demonstrated supraadditive interaction. Implications: Spinal morphine interacts with systemic morphine or buprenorphine in asupraadditive manner. This mode of interaction most probably results from the simultaneous activation of spinal and supraspinal antinociceptive systems.Supraspinal structures played a more important role in the antinociceptive effect of experimental combinations than structures of the spinal cord.     

The effect of intrathecal gabapentin on pain behavior and hemodynamics on the formalin test in the rat.

In this study, we examined the effect of intrathecal (i.t.) gabapentin, administered before and after the injection of formalin into the rat hindpaw, on pain behavior and hemodynamics. Formalin evoked a biphasic flinching behavior and hypertension. I.t. gabapentin administered 10 min before formalin produced a dose-dependent reduction of the Phase 2, but not Phase 1, flinching and cardiovascular response. In contrast, i.t. gabapentin administered 9 min after formalin had no effect on either phase of flinching. I.t. D-serine (100 micrograms) administered 10 min before i.t. galapentin reversed the Phase 2 effect of gabapentin. I.t. gabapentin did not affect the thermal escape latency or the baseline cardiovascular measures even at the largest dose (300 micrograms). These results indicate that the spinal effect of gabapentin reduces the somatosympathetic reflex and somatosensory response to tissue injury without an accompanying effect on acute nociception or resting sympathetic outflow. Implications: After tissue injury, there is an enhanced pain behavior and cardiovascular response, representing a facilitated state of spinal processing. Spinally delivered gabapentin had no evident effect on resting heart rate or blood pressure, but it attenuated the enhanced pain behavior and cardiovascular response otherwise produced by injury.     

The role of cyclooxygenase-1 and -2 in the rat formalin test

Prostaglandins are thought to play an important role in nociceptive transmission at peripheral sites and in the spinal cord. Prostaglandins are produced by cyclooxygenase (COX), which catalyzes the conversion of arachidonic acid. Two forms of COX have been identified: COX-1, which is constitutively expressed, and COX-2, which is an inducible enzyme. To define the role of COX-1 in nociceptive transmission, we examined the effect of oral and intrathecal administration of 5-(4-chlorophenyl)-1-(4-methoxyphenyl)-3-trifluoromethylpyrazole (SC-560), a selective COX-1 inhibitor, on the rat formalin test and compared the effect of SC-560 with that of celecoxib, a COX-2 selective inhibitor, and indomethacin, a nonselective COX-1 and COX-2 inhibitor, on the rat formalin test. Oral and intrathecal administration of SC-560 had no effect on the agitation behavior in the rat formalin test. Oral and intrathecal administration of celecoxib and indomethacin depressed agitation behavior during the rat formalin test. These data suggest that prostaglandins synthesized by COX-1 are not involved in nociceptive transmission during the rat formalin test but that COX-2 does play an important role in the rat formalin test. IMPLICATIONS: Our data suggest that a COX-2 selective inhibitor, but not a COX-1 selective inhibitor, may produce a good analgesic effect on the inflammatory pain state in a clinical situation.     

Inhibitory effect of low-dose pentazocine on the development of antinociceptive tolerance to morphine

Purpose  The development of antinociceptive tolerance to morphine is one of the major problems in its clinical use. Therefore, exploring effective measures to prevent morphine tolerance is of great clinical relevance. We evaluated whether pentazocine could prevent morphine tolerance in mice. Methods  Five groups of male ICR mice received repeated subcutaneous (s.c.) injections of morphine at a high dose (10 mg·kg⁻¹) or saline, concomitantly with s.c. injections of pentazocine at low, subanalgesic doses (0.1, 0.3, or 1.0 mg·kg⁻¹) or saline, respectively, once daily for 14 days. On day 15, mice received co-injections of morphine and pentazocine 120 min after pretreatment with nor-binaltorphimine (5 mg·kg⁻¹), a selective κ-opioid receptor antagonist. The tail pressure threshold was measured before and 60 min after the daily drug co-injections. Results  Repeated s.c. co-injections of morphine and saline resulted in a progressive decrease in morphine-induced antinociception, due to the development of morphine tolerance. Co-injections of pentazocine (0.1, 0.3, and 1.0 mg·kg⁻¹) with morphine potentiated the morphine-induced antinociception dose-dependently by preventing the development of morphine tolerance. Nor-binaltorphimine completely inhibited the chronic antinociception maintained by co-injections of morphine and pentazocine. Conclusion  When chronically co-administered with morphine, pentazocine at low, subanalgesic doses dose-dependently potentiated morphine-induced antinociception in morphine-tolerant mice, through its κ-opioid-receptor-mediated tolerance-preventing activity. Because pentazocine is the only agonist-antagonist analgesic that has an effective oral formulation suitable for chronic administration, the results of the present study warrant clinical trials of pentazocine to assess its tolerance-preventing activity in patients with cancer pain.     

Perineural antinociceptive effect of opioids in a rat model

BACKGROUND: The research on conductive analgesia induced by perineural opioids generated a large body of conflicting data. In this study we reassessed the antinociceptive response to perineural administration of morphine, fentanyl or meperidine in a rat model. METHODS: Analgesia was assessed using the hind paw withdrawal latency (HPWL) response to radiant heat. The opioid dose producing 20% of maximal possible effect (20%MPE) for systemic analgesia was calculated for each drug. Then sciatic blockade was performed with the dose corresponding to 20%MPE. The injected hind paw was used to measure direct perineural effect and the contralateral hind paw was used as an indicator of systemic effect. RESULTS: The response latency produced by morphine or fentanyl was not significantly different for ipsilateral (perineural effect) or contralateral (systemic effect) paw (27+/-11 vs. 28+/-16 and 3l+/-16 vs. 23+/-16 s, respectively). However, the meperidine group showed significantly higher %MPE for the ipsilateral paw (79+/-32 s) than for the contralateral paw (27+/-22 s). CONCLUSIONS: The results indicate that perineural fentanyl or morphine do not produce analgesia. Perineural block produced by meperidine was attributed to local anesthetic-like effect, rather than to drug interaction with opioid receptor.     

Antinociception of intrathecal adenosine receptor subtype agonists in rat formalin test

Adenosine has shown antinociceptive action via spinal adenosine receptors. There are four types of adenosine receptors: A1, A2A, A2B, and A3. We characterized the nature of types of adenosine receptors for the control of nociception at the spinal level. For nociception, formalin solution (5%, 50 microL) was injected into the hindpaw of male Sprague-Dawley rats. The effects of intrathecal adenosine A1 (CPA), A2A (DPMA), and A3 (IB-MECA) receptor agonists were examined. CPA and IB-MECA produced limited or no effect on the early phase response of the formalin test, respectively, but the two drugs depressed the late phase response. DPMA suppressed both phase responses. CPA was the most potent drug among the three in the late phase. These results suggest that spinal adenosine A1 and A2A receptors may be involved in the modulation of the early and the late phase responses of the formalin test, whereas adenosine A3 receptor may be involved in the regulation of the late phase response.     

Analgesic effectiveness of ketorolac compared to meperidine in the rat formalin test.

The rat formalin test is an analgesic behavioral observation assessment method that demonstrates two phases of nociceptive behavior. The test consists of injecting the right hind paw with a 5% formalin solution and then observing the animal for specific nociceptive behavior. The phases represent two different types of pain. Phase 1 is pain produced by direct nerve stimulation and phase 2 is an inflammation-induced pain. The nociceptive behavior measured in this experiment was licking and biting the injected paw. A comparison of nociceptive behavior was made when ketorolac and meperidine were injected (i.p.) 10 min prior to formalin injection. As expected, a biphasic pattern of licking and biting the injected paw ensued. It was found that ketorolac had no significant reduction in licking and biting, while meperidine dramatically reduced the nociceptive response in phase 1. In phase 2, both ketorolac and meperidine caused a reduction in licking and biting; however, meperidine reduced the nociceptive response to a greater extent. This experiment demonstrates that ketorolac, when compared to meperidine, is less effective in treating pain from inflammatory origin and is not effective in treating pain from direct nerve stimulation.