n-methylnaloxone and Disease-Models--Animal

Bone cancer metastasis is extremely painful and decreases the quality of life of the affected patients. Available pharmacological treatments are not able to sufficiently ameliorate the pain, and as patients with cancer are living longer, new treatments for pain management are needed. Decitabine (5-aza-2'-deoxycytidine), a DNA methyltransferases inhibitor, has analgesic properties in preclinical models of postsurgical and soft-tissue oral cancer pain by inducing an upregulation of endogenous opioids. In this study, we report that daily treatment with decitabine (2 µg/g, intraperitoneally) attenuated nociceptive behavior in the 4T1-luc2 mouse model of bone cancer pain. We hypothesized that the analgesic mechanism of decitabine involved activation of the endogenous opioid system through demethylation and reexpression of the transcriptionally silenced endothelin B receptor gene, Ednrb. Indeed, Ednrb was hypermethylated and transcriptionally silenced in the mouse model of bone cancer pain. We demonstrated that expression of Ednrb in the cancer cells lead to release of β-endorphin in the cell supernatant, which reduced the number of responsive dorsal root ganglia neurons in an opioid-dependent manner. Our study supports a role of demethylating drugs, such as decitabine, as unique pharmacological agents targeting the pain in the cancer microenvironment.

Topics: Animals; Antimetabolites, Antineoplastic; beta-Endorphin; Bone Density; Bone Neoplasms; Cancer Pain; Cell Line, Tumor; Culture Media, Conditioned; Decitabine; Disease Models, Animal; Endothelin-1; Female; Ganglia, Spinal; Locomotion; Mice; Mice, Inbred BALB C; Naloxone; Neurons; Quaternary Ammonium Compounds; Receptor, Endothelin B; Weight-Bearing

Pain and inflammation are inherently linked responses to injury, infection, or chronic diseases. Given that acute inflammation in humans or mice enhances the analgesic properties of opioids, there is much interest in determining the inflammatory transducers that prime opioid receptor signaling in primary afferent nociceptors. Here, we found that activation of the transient receptor potential vanilloid type 1 (TRPV1) channel stimulated a mitogen-activated protein kinase (MAPK) signaling pathway that was accompanied by the shuttling of the scaffold protein β-arrestin2 to the nucleus. The nuclear translocation of β-arrestin2 in turn prevented its recruitment to the μ-opioid receptor (MOR), the subsequent internalization of agonist-bound MOR, and the suppression of MOR activity that occurs upon receptor desensitization. Using the complete Freund's adjuvant (CFA) inflammatory pain model to examine the role of TRPV1 in regulating endogenous opioid analgesia in mice, we found that naloxone methiodide (Nal-M), a peripherally restricted, nonselective, and competitive opioid receptor antagonist, slowed the recovery from CFA-induced hypersensitivity in wild-type, but not TRPV1-deficient, mice. Furthermore, we showed that inflammation prolonged morphine-induced antinociception in a mouse model of opioid receptor desensitization, a process that depended on TRPV1. Together, our data reveal a TRPV1-mediated signaling pathway that serves as an endogenous pain-resolution mechanism by promoting the nuclear translocation of β-arrestin2 to minimize MOR desensitization. This previously uncharacterized mechanism may underlie the peripheral opioid control of inflammatory pain. Dysregulation of the TRPV1-β-arrestin2 axis may thus contribute to the transition from acute to chronic pain.

Topics: Acute Pain; Analgesia; Analgesics, Opioid; Animals; beta-Arrestin 2; Chronic Pain; Disease Models, Animal; Freund's Adjuvant; Humans; Inflammation; Mice; Mice, Knockout; Naloxone; Narcotic Antagonists; Quaternary Ammonium Compounds; Signal Transduction; TRPV Cation Channels

Fentanyl is an opioid commonly prescribed for cancer pain. Using melanoma-bearing mice, we investigated whether peripheral action would contribute to fentanyl analgesia in cancer pain. Intravenous injection of fentanyl inhibited mechanical nociception in healthy mice, which was markedly inhibited by the opioid antagonist naloxone, but not naloxone methiodide, a peripherally acting opioid antagonist. Melanoma-bearing mice showed mechanical allodynia and spontaneous licking, a pain-related behavior, which were suppressed by intravenous and local injections of fentanyl. Both naloxone and naloxone methiodide inhibited the analgesic effect of intravenous fentanyl to the same degree. Electrophysiological analysis showed that melanoma growth increased the spontaneous and mechanical stimuli-evoked activity of the tibial nerve, which were inhibited by intravenous fentanyl. There was a greater expression of µ- opioid receptors in skin with a melanoma mass than in the contralateral normal skin. In addition, we found µ-opioid receptors in cultured melanoma cells. There was no difference between the number of µ-opioid receptors in the dorsal root ganglia and spinal cord of the melanoma-bearing and contralateral skin side. These results suggest that the analgesic effect of systemic fentanyl is produced via central and peripheral µ- opioid receptors in cancer pain, and cancer cells are a key site of peripheral action.

Topics: Action Potentials; Analgesics, Opioid; Animals; Cancer Pain; Cell Line, Tumor; Disease Models, Animal; Fentanyl; Humans; Hyperalgesia; Injections, Intramuscular; Injections, Intravenous; Male; Melanoma; Mice; Mice, Inbred C57BL; Naloxone; Narcotic Antagonists; Quaternary Ammonium Compounds; Receptors, Opioid; Receptors, Opioid, mu; Skin; Skin Neoplasms; Spinal Cord; Tibial Nerve; Xenograft Model Antitumor Assays

The search for new drugs for cancer pain management has been a long-standing goal in basic and clinical research. Classical opioid drugs exert their primary antinociceptive effect upon activating opioid receptors located in the central nervous system. A substantial body of evidence points to the relevance of peripheral opioid receptors as potential targets for cancer pain treatment. Peptides showing limited blood-brain-barrier permeability promote peripheral analgesia in many pain models. In the present study we examined the peripheral and central analgesic effect of intravenously administered biphalin - a dimeric opioid peptide in a mouse skin cancer pain model, developed by an intraplantar inoculation of B16F0 melanoma cells. The effect of biphalin was compared with morphine - a golden standard in cancer pain management. Biphalin produced profound, dose-dependent and naloxone sensitive spinal analgesia. Additionally, the effect in the tumor-bearing paw was largely mediated by peripheral opioid receptors, as it was readily attenuated by the blood-brain-barrier-restricted opioid receptor antagonist - naloxone methiodide. On the contrary, morphine facilitated its analgesic effect primarily by activating spinal opioid receptors. Both drugs induced tolerance in B16F0 - implanted paws after chronic treatment, however biphalin as opposed to morphine, showed little decrease in its activity at the spinal level. Our results indicate that biphalin may be considered a future alternative drug in cancer pain treatment due to an enhanced local analgesic activity as well as lower tolerance liability compared with morphine.

Topics: Analgesia; Analgesics, Opioid; Animals; Blood-Brain Barrier; Cancer Pain; Cell Line, Tumor; Disease Models, Animal; Drug Tolerance; Enkephalins; Male; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Morphine; Naloxone; Opioid Peptides; Permeability; Quaternary Ammonium Compounds; Receptors, Opioid; Skin Neoplasms

Clinical studies have tested the use of an engineered herpes virus to treat pain. We hypothesized that subcutaneous injections of an engineered herpes virus that expresses enkephalin would attenuate orofacial nociception and hypersensitivity in male and female rats by a central mechanism.. Herpes virus was injected subcutaneously around the mouth of male and female rats seventy-two hours before ligatures were placed on the masseter tendon, control treatment groups received either no virus or no ligature. Enkephalin expression was measured and von Frey filament testing and meal duration were utilized to measure mechanical hypersensitivity and the nociceptive response, respectively. Naloxone or naloxone methiodide was administered to rats injected with the enkephalin expressing virus to test if enkephalin was acting peripherally or centrally.. Ligature significantly lengthened meal duration and reduced the threshold to von Frey filaments for 18 days. Infection with the enkephalin transgene significantly decreased this response for at least 11 days but only in male rats. Virus injection significantly increased expression of enkephalin in the mental nerve that innervates the mouth region, the trigeminal ganglia and the trigeminal nucleus caudalis but no increase was observed in the masseter nerve after virus injection. Naloxone but not naloxone methiodide reversed the response to the enkephaline expressing virus.. The data suggests that sex should be a considered when using this virus and that viral transfection of the mental nerve with an enkephalin transgene can reduce nociception and hypersensitivity through a central mechanism.

Topics: Animals; Disease Models, Animal; Enkephalins; Female; Herpesviridae; Herpesviridae Infections; Hyperalgesia; Male; Masseter Muscle; Naloxone; Narcotic Antagonists; Nociception; Quaternary Ammonium Compounds; Rats; Rats, Sprague-Dawley; Trigeminal Ganglion; Trigeminal Nerve; Trigeminal Nuclei

Endomorphins are endogenous opioid peptides in mammals and display a strong antinociceptive effect after central administration. However, the clinical usage of these peptides is limited because of their diminished analgesic effect following systemic injection and their inability to cross the blood-brain barrier. In this study, we characterized the in vivo effects of four novel endomorphin-1 analogues (termed MELs), which previously showed potential as highly potent analgesics with a good pharmacological profile in vitro. The analogues were administered intravenously to several rodent pain models to examine their antinociception and blood-brain barrier permeability. The tested peptides, especially MEL1214, showed good analgesic activity and blood-brain barrier permeability. Behavioral studies showed dose-dependent analgesic effect after systematic administration of MEL1214 in the tested pain models. Pre-treatment of subcutaneous administration of naloxone methiodide did not affect the antinociception of these peptides. As compared to morphine, MEL1214 was less prone to induce tolerance after consecutive intravenous administration for 5 days. Gastrointestinal transit was evaluated by the isolated colon response and bead expulsion to determine the potential constipation effect. In contrast to morphine, MEL1214 produced no significant constipation effect at an equivalent dose. MEL1214 shows promise as a suitable compound to treat pain with reduced side effects, and exhibits good potential to be further developed as a novel opioid analgesic in pain treatment.

Topics: Analgesics, Opioid; Animals; Blood-Brain Barrier; Capillary Permeability; Colon; Constipation; Disease Models, Animal; Drug Tolerance; Male; Mice; Morphine; Naloxone; Narcotic Antagonists; Nociceptive Pain; Oligopeptides; Quaternary Ammonium Compounds

This study describes the antinociceptive effects of μ-opioid agonists, d-Ala(2),N-Me-Phe(4),Gly(5)-ol-enkephalin (DAMGO) and morphine in a model of rat visceral pain in which nociceptive responses were triggered by 2% acetic acid intraperitoneal (i.p.) injections. DAMGO and morphine were administered i.p., to the same site where acetic acid was delivered or intracerebroventricularly (i.c.v.). The antinociceptive actions of i.p. versus i.c.v. administered DAMGO or morphine were evaluated in the late phase of permanent visceral nociceptive responses. Both compounds inhibited the nociceptive responses in a dose-dependent manner and exhibited more potent agonist activity after i.c.v. than i.p. administration. DAMGO and morphine showed comparable ED(50) values after i.p. injections. However, DAMGO was much stronger than morphine after central administration. Co-administration of the peripherally restricted opioid antagonist, naloxone methiodide (NAL-M), significantly attenuated the antinociceptive effects of i.p. DAMGO or morphine. On the other hand, i.c.v. injections of NAL-M partially antagonized the antinociceptive effect of i.p. morphine and failed to affect the antinociceptive action of i.p. DAMGO indicating the partial and pure peripheral antinociceptive effects of morphine and DAMGO, respectively. These results suggest the role of either central or peripheral μ-opioid receptors (MOR) in mediating antinociceptive effects of i.p. μ-opioid agonists in the rat late permanent visceral pain model which closely resembles the clinical situation.

Topics: Acetic Acid; Analgesics, Opioid; Analysis of Variance; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Routes; Drug Interactions; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Morphine; Naloxone; Narcotic Antagonists; Quaternary Ammonium Compounds; Rats; Visceral Pain

We previously reported the synthesis of three new opioid agonists as well as their in vitro and in vivo activity [Girón, R., Abalo, R., Goicoechea, C., Martín, M.I., Callado, L.F., Cano, C., Goya, P., Jagerovic, N. 2002. Synthesis and opioid activity of new fentanyl analogs. Life Sci. 71, 1023-1034]. One of them, N-[1-phenylpyrazol-3-yl]-N-[1-(2-phenethyl)-4-piperidyl)] propenamide (IQMF-4), showed an interesting antinociceptive activity. Intraperitoneally (i.p.) administered, it was as effective as fentanyl or morphine, being less potent than fentanyl but more so than morphine. The aim of the present work was to evaluate its antinociceptive effect by different routes of administration, using the hot plate test, and to investigate possible side effects, such as tolerance and withdrawal, in vitro, using the myenteric plexus-longitudinal muscle strip preparation from guinea pig ileum, and in vivo, using the hot plate test. IQMF-4 was more potent than morphine when administered per os (p.o.), but less potent when administered intracerebroventricularly (i.c.v.). By both routes, fentanyl is more potent that IQMF-4. When IQMF-4 was administered i.p., naloxone methiodide, a peripherally acting antagonist, was able to completely block its antinociceptive effect, whereas, after i.c.v. administration, the blockade was only partial. An interesting feature of the new compound is that it induces tolerance in vitro but not in vivo. Moreover, though in vitro withdrawal was not different from fentanyl or morphine, in vivo withdrawal symptoms were significantly less frequent in mice treated with IQMF-4 than in those treated with morphine or fentanyl. Although more assays are required, these results show that IQMF-4 appears to be a potent analgesic compound with an interesting peripheral component, and reduced ability to induce dependence.

Topics: Administration, Oral; Analgesics, Opioid; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Tolerance; Fentanyl; Guinea Pigs; Hot Temperature; Ileum; Injections, Intraperitoneal; Injections, Intraventricular; Male; Mice; Morphine; Muscle Contraction; Muscle, Smooth; Naloxone; Narcotic Antagonists; Opioid-Related Disorders; Pain; Pain Measurement; Pain Threshold; Piperidines; Propane; Quaternary Ammonium Compounds; Reaction Time; Substance Withdrawal Syndrome; Time Factors

A visceral pain model incorporating use of cyclophosphamide (CP) to induce bladder inflammation has been described. CP treatment in rats produces changes in behavior (abnormal postures and eye closure) and respiration rate indicative of visceral pain. We characterized the dose-dependency and progression of CP-induced cystitis pain after intraperitoneal (i.p.) CP. The behavioral and respiration rate changes were ameliorated by systemic morphine and ABT-594 [(R)-5-(2-azetidinylmethoxy)-2-chloropyridine], a neuronal nicotinic acetylcholine receptor agonist, in a manner reversible by naloxone and mecamylamine, respectively. Sites of antinociceptive actions of morphine and ABT-594 were investigated using systemic, intrathecal (i.t.), or intracerebroventricular (i.c.v.) administration of blood-brain barrier impenetrant antagonists. Naloxone methiodide produced a complete antagonism of morphine antinociception after i.c.v. but not i.p. or i.t. administration. Chlorisondamine blocked ABT-594 antinociception after i.c.v. but not i.p. administration. Further pharmacological characterization of behavioral and respiration changes in CP-cystitis was performed using standard analgesics. The alpha(2)-adrenoceptor agonist clonidine produced a weak attenuation of CP-pain behavior. NSAIDs (ibuprofen, acetaminophen, and celecoxib) and anticonvulsants (gabapentin and lamotrigine) were without effect. These results demonstrate that morphine and ABT-594 produce antinociception in CP-cystitis by a predominantly supraspinal site of action, and that mechanisms producing robust centrally-mediated antinociception could be beneficial in cystitis pain.. In this article, potential antinociceptive effects of a variety of pharmacological agents were evaluated in a rat cystitis pain model. Morphine and a nicotinic acetylcholine receptor agonist ABT-594 were found to exert potent antinociception in this model. Findings presented here aid identification of agents to treat cystitis pain in the clinic.

Topics: Analgesics; Animals; Azetidines; Behavior, Animal; Brain; Chlorisondamine; Cyclophosphamide; Cystitis; Disease Models, Animal; Male; Mecamylamine; Morphine; Naloxone; Narcotic Antagonists; Nicotinic Antagonists; Pain; Pyridines; Quaternary Ammonium Compounds; Rats; Rats, Sprague-Dawley; Receptors, Nicotinic; Viscera

Opioid overdose, which is commonly associated with opioid induced respiratory depression, is a problem with both therapeutic and illicit opioid use. While the central mechanisms involved in the effects of opioids are well described, it has also been suggested that a peripheral component may contribute to the effects observed. This study aimed to further characterise the effects of the peripherally acting naloxone methiodide on the respiratory, analgesic and withdrawal effects produced by various opioid agonists. A comparison of the respiratory and analgesic effects of morphine, methadone and heroin in male Swiss-Albino mice was conducted and respiratory depressive ED(80) doses of each opioid determined. These doses (morphine 9 mg/kg i.p., methadone 7 mg/kg i.p., and heroin 17 mg/kg i.p.) were then used to show that both naloxone (3 mg/kg i.p.) and naloxone methiodide (30-100 mg/kg i.p.) could reverse the respiratory and analgesic effects of these opioid agonists, but only naloxone precipitated withdrawal. Further investigation in female C57BL/6J mice using barometric plethysmography found that both opioid antagonists could reverse methadone induced decreases in respiratory rate and increases in tidal volume. Its effects do not appear to be strain or sex dependent. It was concluded that naloxone methiodide can reverse the respiratory and analgesic actions of a variety of opioid agonists, without inducing opioid withdrawal.

Topics: Analgesics, Opioid; Animals; Disease Models, Animal; Drug Interactions; Female; Heroin; Male; Methadone; Mice; Mice, Inbred C57BL; Morphine; Naloxone; Plethysmography, Whole Body; Quaternary Ammonium Compounds; Respiration; Respiratory Insufficiency; Substance Withdrawal Syndrome

Trigeminal neuropathic pain represents a real challenge to therapy because commonly used drugs are devoid of real beneficial effect or patients frequently become intolerant or refractory to some of these compounds. In a rat model of trigeminal neuropathic pain, which shares numerous similarities with human trigeminal neuralgia and trigeminal neuropathic pain, we used a genomic herpes simplex virus-derived vector (HSVLatEnk) to examine the possible effect of a local overproduction of proenkephalin A (PA) targeted to the trigeminal primary sensory neurons. Unilateral peripheral inoculation of recombinant vectors on the vibrissal pad territory resulted in an about ninefold increase in proenkephalin A mRNA levels in trigeminal ganglion ipsilateral to the infected side. Transgene-derived met-enkephalin accumulated in numerous nerve cell bodies of trigeminal ganglion and was transported through the sensory nerve fibers located in the infraorbital nerve. Bilateral mechanical hyperresponsiveness, which developed 2 weeks after chronic constrictive injury of the left infraorbital nerve, was significantly attenuated in animals overproducing PA in the trigeminal ganglion ipsilateral to the lesioned infraorbital nerve. This antiallodynic effect was reversed by both the opioid receptor antagonist naloxone and the peripherally acting antagonist naloxone methiodide. Our data demonstrate that the local overproduction of PA-derived peptides in trigeminal ganglion sensory neurons evoked a potent antiallodynic effect through the stimulation of mainly peripherally located opioid receptors and suggest that targeted delivery of endogenous opioids may be of interest for the treatment of some severe forms of neuropathic pain.

Topics: Animals; Disease Models, Animal; Enkephalins; Genetic Therapy; Male; Naloxone; Narcotic Antagonists; Neurons; Pain Threshold; Protein Precursors; Quaternary Ammonium Compounds; Rats; Rats, Sprague-Dawley; RNA, Messenger; Simplexvirus; Transcriptional Activation; Trigeminal Ganglion; Trigeminal Neuralgia

The effects of the selective delta-1 (delta(1)) opioid receptor agonist, DPDPE, and the selective delta(2) opioid receptor agonist, DSLET, have been studied on the ventricular fibrillation threshold (VFT) in rats with an experimental post-infarction cardiosclerosis (CS). It has been found that CS induced a significant decrease in VFT. This CS-induced decrease in VFT was significantly reversed by intravenous administration of DPDPE (0.1 mg/kg) 10 min before VFT measurement. On the contrary, intravenous injection of DSLET (0.5 mg/kg) exacerbated the CS-induced cardiac electrical instability. Pretreatment with the selective delta opioid receptor antagonist, ICI 174,864 (0.5 mg/kg), completely abolished the changes in VFT produced by both DPDPE and DSLET. Previous administration of a nonselective peripherally acting opioid receptor antagonist, naloxone methiodide (5 mg/kg) also completely reversed the antifibrillatory action of DPDPE. Naloxone methiodide and ICI 174,864 alone had no effect on VFT. Pretreatment with the nonselective K(ATP) channel blocker, glibenclamide (0.3 mg/kg), or with the mitochondrial selective K(ATP) channel blocker, 5-hydroxydecanoic acid (5-HD, 5 mg/kg), completely abolished the DPDPE-induced increase in cardiac electrical stability. Glibenclamide and 5-HD alone had no effect on VFT. These results demonstrate that the delta opioid receptor plays an important role in the regulation of electrical stability in rats with post-infarction cardiosclerosis. We propose that peripheral delta(1) opioid receptor stimulation reverses CS-induced electrical instability via mitochondrial K(ATP) channels. On the contrary, delta(2) opioid receptor stimulation may exacerbate the CS-induced decrease in VFT. Further studies are necessary to determine the delta opioid receptor subtype which mediates the antifibrillatory effect of DPDPE and pro-fibrillatory effect of DSLET.

Topics: Adenosine Triphosphate; Analgesics, Opioid; Animals; Decanoic Acids; Disease Models, Animal; Drug Antagonism; Enkephalin, D-Penicillamine (2,5)-; Enkephalin, Leucine; Glyburide; Hydroxy Acids; Male; Mitochondria, Heart; Myocardial Infarction; Myocardium; Naloxone; Potassium Channels; Quaternary Ammonium Compounds; Rats; Rats, Wistar; Receptors, Opioid, delta; Sclerosis; Ventricular Fibrillation

Opiate withdrawal behaviors in the infant differ from those of the adult. The neural circuitry underlying opioid withdrawal in the adult rat is well defined and includes the locus coeruleus (LC) and periaqueductal gray (PAG), with a minor role of the amygdala. Because the different behaviors that constitute the infant syndrome may be mediated by different neural circuits, we tested the hypothesis that these three sites are involved in opiate withdrawal. Pups were injected with morphine from day 1-6 after birth (b.i.d.) and on the morning of the seventh day. Withdrawal was then elicited by local injection of the opioid antagonist methylnaloxonium into the LC, PAG, or amygdala. Withdrawal signs were precipitated in a dose-dependent manner following injection into the LC or PAG, but not the amygdala. The withdrawal behaviors elicited from the LC and PAG included both the same and different behaviors. The results support the hypothesis that the neural circuitry mediating opiate withdrawal behaviors is similar in infant and adult animals, but the behaviors expressed are age-specific.

Topics: Age Factors; Amygdala; Animals; Animals, Newborn; Behavior, Animal; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Locus Coeruleus; Male; Morphine; Naloxone; Naltrexone; Narcotic Antagonists; Neural Pathways; Opioid-Related Disorders; Periaqueductal Gray; Quaternary Ammonium Compounds; Rats; Rats, Long-Evans; Substance Withdrawal Syndrome

In a rat model of peripheral mononeuropathy produced by moderate constriction of the sciatic nerve, we have shown that various i.v. doses of morphine and selective opioid agonists produce potent and long-lasting antinociceptive effects on the vocalization threshold to paw pressure. For all the opioids, the antinociceptive effects were more marked for the paw on the nerve-injured side (nerve-injured paw) than for the sham-operated paw. One contributory mechanism could be a peripheral action of the opioid agonists in the nerve-injured paw. This hypothesis was tested in the present study, using systemic morphine and low doses of local naloxone or its quaternary salt naloxone methiodide, exhibiting peripherally acting antagonist properties. The effects of escalating doses of naloxone (0.5-2 microgram injected i.v. or intraplantar into the nerve-injured paw) or naloxone methiodide (5-30 micrograms into the nerve-injured paw) on the antinociceptive effect of morphine (1 mg/kg i.v.) were evaluated using the vocalization threshold to paw pressure in neuropathic rats at two weeks after placing ligatures, a time when the behavioural pain-related disorders have reached a maximum.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Analgesics; Animals; Constriction; Disease Models, Animal; Dose-Response Relationship, Drug; Male; Morphine; Naloxone; Peripheral Nervous System Diseases; Quaternary Ammonium Compounds; Rats; Rats, Sprague-Dawley; Sciatic Nerve; Vocalization, Animal