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Meeting Abstracts  |   September 1996
Role Central Mu, Delta-1, and Kappa-1 Opioid Receptors in Opioid-induced Muscle Rigidity in the Rat
Author Notes
  • (Vankova) Postgraduate Researcher, Department of Anesthesiology, University of California, San Diego.
  • (Weinger) Associate Professor of Anesthesiology, University of California, San Diego; Staff Physician, San Diego Veterans Affairs Medical Center; Adjunct Associate Member, Department of Neuropharmacology, The Scripps Research Institute.
  • (Chen) Postdoctoral Fellow, Department of Anesthesiology, University of California, San Diego.
  • (Bronson, Motis) Research Student, Department of Anesthesiology, University of California, San Diego.
  • (Koob) Member, Department of Neuropharmacology, The Scripps Research Institute.
  • Received from the Department of Anesthesiology, University of California, San Diego School of Medicine; the Anesthesia Research Service of the San Diego Veterans Affairs Medical Center; and Department of Neuropharmacology, The Scripps Research Institute. Submitted for publication November 7, 1995. Accepted for publication May 5, 1996. Supported by grant DA06616 to M.B.W. from the National Institute on Drug Abuse. Preliminary results of this work were presented at the annual meeting of the American Society of Anesthesiologists, New Orleans, Louisiana, October 18, 1989, and the annual meeting of the International Anesthesia Research Society, San Antonio, Texas, March 9, 1991.
  • Address reprint requests to Dr. Weinger: Veterans Affairs Medical Center (125), 3350 La Jolla Village Drive, San Diego, California 92161. Address electronic mail to: mweinger@uscd.edu.
Article Information
Meeting Abstracts   |   September 1996
Role Central Mu, Delta-1, and Kappa-1 Opioid Receptors in Opioid-induced Muscle Rigidity in the Rat
Anesthesiology 9 1996, Vol.85, 574-583.. doi:
Anesthesiology 9 1996, Vol.85, 574-583.. doi:
Key words: Anesthetics, intravenous: alfentanil. Complications: muscle rigidity. Analgesics, opioid: receptors. Antagonists: narcotic. Brain: drug injections. Measurement techniques: electromyography. Animals: rat.
HIGH-DOSE opiate administration produces intense analgesia and decreased sympathetic response to painful stimulation. Because of these advantageous clinical properties, opiates have increasingly enjoyed widespread use in anesthesia. Unfortunately, the profound analgesia of acute high-dose opiate* administration may be accompanied by prolonged respiratory depression and intense generalized muscle rigidity. Laboratory efforts have been undertaken to understand the mechanism of opiate-induced muscle rigidity.** The development of opiate agonists that do not produce muscle rigidity or other undesirable side effects would be a major advance in anesthesia and clinical pain management.
The cloning, localization, and functional validation of three similar but distinct gene products [1] now provide convincing evidence for the existence of at least three opioid receptor types (i.e., mu [micro], delta [delta], and kappa [kappa]) with different distributions throughout the central nervous system (CNS). [2-4] These three receptors are also differentially implicated in the mediation of various physiologic and behavioral effects of opiates. [1,4-7] *** In vitro and in vivo pharmacology strongly substantiates a further subdivision of each of the receptor types into subtypes (or isoreceptors). [4,6,7,8] 
With respect to the role of different receptors in opiate-induced muscle rigidity, initial studies focused on the micro receptor and used relatively poorly selective opioid receptor agonists. [9] The elucidation of the in vivo receptor selectivity of opioid agonists, such as D-Ala2,N-Me-Phe4-Gly5-ol-enkephalin (DAMGO; micro selective), [10,11] D-Pen2,D-Pen5-enkephalin (DPDPE; delta-1selective), [12-14] as well as the non-peptide agonist trans-(plus/minus)-3,4-dichloro-N-methyl-N-2-(1-pyrrolidinyl)cyclohexyl) -benzene-acetamide methane sulfonate (U50, 488H; kappa-1), [15,16] now allows a more detailed study of the central opioid receptors mediating systemic opiate-induced muscle rigidity.
The purpose of the current study was to determine whether opioid-induced muscle rigidity, as measured by extremity electromyographic activity, is the result solely of activation of central micro opioid receptors or whether central delta and/or kappa receptors might also play a role. To pursue the hypothesis that central delta or kappa opioid receptors do mediate systemic opiate-induced muscle rigidity, the effect of intracerebroventricular administration of opioid receptor-selective agonists was investigated in intact, healthy spontaneously ventilating rats. In addition, the interaction between central opioid receptor activation and the rigidity induced by systemic administration of the potent micro-preferring agonist alfentanil [17,18] was examined. The receptor specificity of the effects observed was confirmed with the use of opioid receptor-selective antagonists.
Methods
Animals
All procedures were approved by our institution's Animal Care Committee. The subjects were 114 male albino Wistar rats (Harlan Laboratories, Indianapolis, IN) that weighed approximately 250-320 g at the time of the experiments. Animals were housed 2-3 per cage in a temperature-controlled room with a 12-h light/dark cycle.
Drugs
The drugs administered intracerebroventricularly were DAMGO (courtesy of NIDA, Bethesda, MD), DPDPE (courtesy of NIDA), U50,488H (gift of Upjohn Laboratories, Kalamazoo, MI), D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2(CTAP; Peninsula Laboratories, Belmont, CA), naltrindole hydrochloride (NTI, Searle Research and Development, Skokie, IL), and nor-binaltorphimine (Research Biochemicals International, Natick, MA). All drugs were freshly dissolved in sterile physiologic saline (0.9%) and administered in a volume of 5 micro liter. The doses are expressed as free base.
In some of the experiments, systemic opiate-induced muscle rigidity was induced with alfentanil (Alfenta; Janssen Pharmaceutica, Piscataway, NJ) dissolved in saline and administered subcutaneously at a dose of 500 micro gram/kg in a volume of 1 ml/kg. This dose of alfentanil was chosen because it was shown, in previous studies, to consistently produce profound muscle rigidity. [17-19] 
Surgical Procedure and Habituation
All rats were anesthetized with halothane and secured in a stereotaxic apparatus (Kopf Instruments, Tujunga, CA). Under aseptic conditions, a 7-mm, 23-gauge, stainless-steel guide cannula was implanted in the lateral ventricle. The coordinates used were: -0.6 mm to bregma, 2.0 mm lateral to the midline, and -3.2 mm to the skull surface at the point of entry. [20] The cannula was anchored to the skull with stainless-steel screws and dental cement and kept patent with a 30-gauge stainless-steel dummy stylet. The rats were allowed to recuperate for at least 5 days and to acclimate to the experimental cylindrical restraining apparatus during three separate daily 2-h sessions during this postsurgical period.
Experimental Design
The first series of experiments was undertaken to define agonist dose-response relations. Rats were pretreated with a single intracerebroventricular dose of either the delta1receptor agonist DPDPE, the kappa1receptor agonist U50,488H, or the micro-receptor agonist DAMGO, and then randomly assigned to one of two groups. In one group, alfentanil was injected subcutaneously 10 min after agonist administration, whereas the other received the same volume of physiologic saline. The animals pretreated with DAMGO intracerebroventricularly received only saline subcutaneous injection. The doses tested were: DPDPE at 0, 50, or 150 nmol; U50,488H at 0, 22, or 107 nmol; and DAMGO at 0, 4, or 8 nmol.
In the second series of experiments, the same agonists were used at doses that were demonstrated in the first experiment to have an effect alone or in combination with alfentanil on muscle rigidity. In these experiments, the rats first received an intracerebroventricular injection of either an opioid receptor-selective antagonist or saline. Then, the rats received a second intracerebroventricular injection of either the corresponding agonist or saline. The receptor-selective antagonists used in these experiments were as follows: (1) for delta opioid receptors, NTI (10 nmol); (2) for kappa1receptors, nor-binaltorphimine (10 nmol); and (3) for micro receptors, CTAP (3 nmol). Ten minutes later, alfentanil was injected (except for the DAMGO group, which received only the intracerebroventricular treatments).
In all tests, animals were assigned randomly to treatment groups, and the observer was blinded to the experimental conditions. Each rat was studied only once. The drug doses and the time interval between treatments were chosen based on preliminary experiments**** and were consistent with those found, in this laboratory, to produce other relevant physiologic effects, including antinociception,***** respiratory effects, [21,22] and reinforcement. [23,24] 
Experimental Protocol
One to two hours before experiments, animals were transported from the animal care facility; food and water was withheld. Rats were placed individually into the cylindrical holding apparatus inside a sound-attenuating box (Coulbourn Instruments, Lehigh Valley, PA). Muscle rigidity was assessed by measuring hindlimb electromyographic activity, as described previously. [17-19,25,26] Briefly, two monopolar electrodes were inserted percutaneously into the left gastrocnemius muscle of each rat, and a third (ground) electrode was inserted into the right hind limb. Leads were held in place with cellophane tape. Limbs were secured with surgical tape in a manner that allowed unimpeded joint movement. After lead placement, the door of the box was closed, and baseline electromyographic activity was recorded for 15 min. Then, one or two (according to the experimental design) intracerebroventricular injections were performed, using an 8-mm, 30-gauge, stainless-steel injector cannula attached to polyethylene tubing. Five microliters of drug were injected by pressure using a Hamilton pump for 2 min. Electromyographic activity was then recorded at 5-min intervals for 60 min after the alfentanil (or saline) injection.
Raw muscle potentials were differentially amplified 200 times and band-pass filtered (10 Hz-3 KHz). The resulting signals, viewed on an oscilloscope, were converted with a root-mean-squared voltage rectifier (t 1/2 = 3 s) to produce time-varying analog deflections on Triplet 200 mV meters. [17-19,26] Full-scale deflection of the meter corresponded to 100 micro Volt of electromyographic activity. During data collection, care was taken to exclude the effects of transient movement artifacts, thereby permitting assessment of tonic rather than phasic muscle activity. At the conclusion of the experiment, the position of each cannula was verified with an intracerebroventricular infusion of india ink followed by necropsy.
Data Analysis
Data from the two experimental series were analyzed separately. In the first series, the mean area under the electromyographic curve was calculated and, for each agonist, a two-way analysis of variance was performed to evaluate drug effect (alfentanil or saline) and the effect of drug dose. In the second experimental series, statistical differences between agonist, antagonist, antagonist-agonist, and saline (control) groups were determined using two-way analysis of variance with repeated measures. [27] Then, Student-Newman-Keuls a posteriori tests were used to assess differences between treatment groups at individual time points, as well as differences over time within each treatment group. Mean animal weights between different treatment groups were analyzed in the same manner. Data were expressed as mean plus/minus SEM (P < 0.05 considered statistically significant).
Results
In all experimental groups, the baseline electromyographic activity, recorded during the 15-min pretreatment period, ranged from 2 to 5 micro Volt root-mean-squared. No significant differences in body weight were found among the different treatment groups.
When intracerebroventricular administration of either DPDPE (50 or 150 nmol) or U50,488H (43 or 107 nmol) was followed by a saline subcutaneous injection, the electromyographic activity was not significantly different from its own preinjection values or from the values in the saline-saline control groups (Figure 1(a) and Figure 2(a)). In contrast, the intracerebroventricular administration of DAMGO resulted in a dose-dependent increase in muscle tone (Figure 3).
Figure 1. Effect of different doses of DPDPE on electromyographic activity. Y axis gives the area under the electromyographic data versus time curve (AUC): (a) saline-treated groups; and (b) alfentanil-treated groups (500 micro gram/kg subcutaneously). N = 6 for each dose examined. Significant decrease in alfentanil-induced muscle rigidity (*P < 0.05) after intracerebroventricular pretreatment with DPDPE (150 nmol), compared with saline intracerebroventricularly (zero dose group).
Figure 1. Effect of different doses of DPDPE on electromyographic activity. Y axis gives the area under the electromyographic data versus time curve (AUC): (a) saline-treated groups; and (b) alfentanil-treated groups (500 micro gram/kg subcutaneously). N = 6 for each dose examined. Significant decrease in alfentanil-induced muscle rigidity (*P < 0.05) after intracerebroventricular pretreatment with DPDPE (150 nmol), compared with saline intracerebroventricularly (zero dose group).
Figure 1. Effect of different doses of DPDPE on electromyographic activity. Y axis gives the area under the electromyographic data versus time curve (AUC): (a) saline-treated groups; and (b) alfentanil-treated groups (500 micro gram/kg subcutaneously). N = 6 for each dose examined. Significant decrease in alfentanil-induced muscle rigidity (*P < 0.05) after intracerebroventricular pretreatment with DPDPE (150 nmol), compared with saline intracerebroventricularly (zero dose group).
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Figure 2. Effect of different doses of U50,488H on the electromyographic activity (electromyography-time area under the curve): (a) saline-treated groups; and (b) alfentanil-treated groups. N = 6 for each dose. U50,488H (107 nmol) pretreatment significantly decreased alfentanil-induced muscle rigidity (*P < 0.05) when compared with saline intracerebroventricularly (zero dose group).
Figure 2. Effect of different doses of U50,488H on the electromyographic activity (electromyography-time area under the curve): (a) saline-treated groups; and (b) alfentanil-treated groups. N = 6 for each dose. U50,488H (107 nmol) pretreatment significantly decreased alfentanil-induced muscle rigidity (*P < 0.05) when compared with saline intracerebroventricularly (zero dose group).
Figure 2. Effect of different doses of U50,488H on the electromyographic activity (electromyography-time area under the curve): (a) saline-treated groups; and (b) alfentanil-treated groups. N = 6 for each dose. U50,488H (107 nmol) pretreatment significantly decreased alfentanil-induced muscle rigidity (*P < 0.05) when compared with saline intracerebroventricularly (zero dose group).
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Figure 3. Effect of different doses of the micro receptor agonist DAMGO on electromyographic activity (electromyographytime area under the curve). N = 6 for the saline group, n = 7 for the 4 nmol, and n = 8 for the 8 nmol group. *P < 0.005 compared with saline group.
Figure 3. Effect of different doses of the micro receptor agonist DAMGO on electromyographic activity (electromyographytime area under the curve). N = 6 for the saline group, n = 7 for the 4 nmol, and n = 8 for the 8 nmol group. *P < 0.005 compared with saline group.
Figure 3. Effect of different doses of the micro receptor agonist DAMGO on electromyographic activity (electromyographytime area under the curve). N = 6 for the saline group, n = 7 for the 4 nmol, and n = 8 for the 8 nmol group. *P < 0.005 compared with saline group.
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The pretreatment with 50 nmol DPDPE failed to alter the alfentanil-induced muscle rigidity. However, 150 nmol DPDPE significantly attenuated the alfentanil-induced muscle rigidity (Figure 1(b)). Similarly, pretreatment with U50,488H dose-dependently decreased electromyographic activity after alfentanil injection, although this effect only attained statistical significance with the 107-nmol dose (Figure 2(b)).
In the second experimental series, DPDPE 150 nmol or U50,488H 107 nmol was administered 10 min before alfentanil. DPDPE preceded by saline intracerebroventricularly decreased the alfentanil-induced muscle rigidity at all time points when compared with the control group (two saline intracerebroventricular treatments followed by alfentanil). In addition, when the delta antagonist NTI was injected before DPDPE, the electromyographic activity after alfentanil administration was not significantly different from control (alfentanil produced its usual full effect; Figure 4). Intracerebroventricular administration of NTI followed by saline failed to alter the alfentanil-induced muscle rigidity.
Figure 4. Effect of pretreatment with 150 nmol DPDPE (Sal/DPDPE group,); 10 nmol NTI (NTI/Sal group); 10 nmol NTI and 150 nmol DPDPE (NTI/DPDPE group); or saline (Sal/Sal group) on the alfentanil-induced muscle rigidity. The first intracerebroventricular injection (saline or NTI) was performed at 0 min, the second intracerebroventricular injection (DPDPE or saline) was at 15 min. All animals received 500 micro gram/kg alfentanil subcutaneously at 25 min. N = 7 for all experimental groups. The administration of DPDPE (Sal/DPDPE group) significantly decreased alfentanil-induced muscle rigidity compared with Sal/Sal group (*P < 0.05; +P < 0.001). There was no significant difference in electromyographic activity between Sal/NTI; NTI/DPDPE, and Sal/Sal groups.
Figure 4. Effect of pretreatment with 150 nmol DPDPE (Sal/DPDPE group,); 10 nmol NTI (NTI/Sal group); 10 nmol NTI and 150 nmol DPDPE (NTI/DPDPE group); or saline (Sal/Sal group) on the alfentanil-induced muscle rigidity. The first intracerebroventricular injection (saline or NTI) was performed at 0 min, the second intracerebroventricular injection (DPDPE or saline) was at 15 min. All animals received 500 micro gram/kg alfentanil subcutaneously at 25 min. N = 7 for all experimental groups. The administration of DPDPE (Sal/DPDPE group) significantly decreased alfentanil-induced muscle rigidity compared with Sal/Sal group (*P < 0.05; +P < 0.001). There was no significant difference in electromyographic activity between Sal/NTI; NTI/DPDPE, and Sal/Sal groups.
Figure 4. Effect of pretreatment with 150 nmol DPDPE (Sal/DPDPE group,); 10 nmol NTI (NTI/Sal group); 10 nmol NTI and 150 nmol DPDPE (NTI/DPDPE group); or saline (Sal/Sal group) on the alfentanil-induced muscle rigidity. The first intracerebroventricular injection (saline or NTI) was performed at 0 min, the second intracerebroventricular injection (DPDPE or saline) was at 15 min. All animals received 500 micro gram/kg alfentanil subcutaneously at 25 min. N = 7 for all experimental groups. The administration of DPDPE (Sal/DPDPE group) significantly decreased alfentanil-induced muscle rigidity compared with Sal/Sal group (*P < 0.05; +P < 0.001). There was no significant difference in electromyographic activity between Sal/NTI; NTI/DPDPE, and Sal/Sal groups.
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The administration of U50,488H preceded by saline decreased alfentanil-induced rigidity at most time points compared with the control group (Figure 5). This effect of U50,488H was abolished by pretreatment with nor-binaltorphimine. The intracerebroventricular injection of nor-binaltorphimine followed by saline intracerebroventricularly appeared to increase muscle tone before and after alfentanil compared with the control group; however, this effect just failed to attain statistical significance (P < 0.10) because of appreciable variability between animals.
Figure 5. Effect of pretreatment with 107 nmol U50,488H (Sal/U50,488H group, n = 7); 10 nmol nor-binaltorphimine (nor-binaltorphimine/Sal group, n = 6); 10 nmol nor-binaltorphimine and 107 nmol U50,488H (nor-binaltorphimine/U50,488H group, n = 6); or saline (Sal/Sal group, n = 7) on alfentanil-induced muscle rigidity. The intracerebroventricular injections were performed at time points 0 (saline or nor-binaltorphimine) and 60 (saline or U50,488H); alfentanil was administered subcutaneously at 70 min. The pretreatment with U50,488H (Sal/U50,488H group) significantly decreased the alfentanil-induced muscle rigidity compared with the control group (Sal/Sal) (*P < 0.05; +P < 0.005). No significant differences between Sal/Sal, nor-binaltorphimine/Sal, and nor-binaltorphimine/U50,488H groups were observed.
Figure 5. Effect of pretreatment with 107 nmol U50,488H (Sal/U50,488H group, n = 7); 10 nmol nor-binaltorphimine (nor-binaltorphimine/Sal group, n = 6); 10 nmol nor-binaltorphimine and 107 nmol U50,488H (nor-binaltorphimine/U50,488H group, n = 6); or saline (Sal/Sal group, n = 7) on alfentanil-induced muscle rigidity. The intracerebroventricular injections were performed at time points 0 (saline or nor-binaltorphimine) and 60 (saline or U50,488H); alfentanil was administered subcutaneously at 70 min. The pretreatment with U50,488H (Sal/U50,488H group) significantly decreased the alfentanil-induced muscle rigidity compared with the control group (Sal/Sal) (*P < 0.05; +P < 0.005). No significant differences between Sal/Sal, nor-binaltorphimine/Sal, and nor-binaltorphimine/U50,488H groups were observed.
Figure 5. Effect of pretreatment with 107 nmol U50,488H (Sal/U50,488H group, n = 7); 10 nmol nor-binaltorphimine (nor-binaltorphimine/Sal group, n = 6); 10 nmol nor-binaltorphimine and 107 nmol U50,488H (nor-binaltorphimine/U50,488H group, n = 6); or saline (Sal/Sal group, n = 7) on alfentanil-induced muscle rigidity. The intracerebroventricular injections were performed at time points 0 (saline or nor-binaltorphimine) and 60 (saline or U50,488H); alfentanil was administered subcutaneously at 70 min. The pretreatment with U50,488H (Sal/U50,488H group) significantly decreased the alfentanil-induced muscle rigidity compared with the control group (Sal/Sal) (*P < 0.05; +P < 0.005). No significant differences between Sal/Sal, nor-binaltorphimine/Sal, and nor-binaltorphimine/U50,488H groups were observed.
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Finally, the effect of the micro agonist DAMGO was studied. When preceded by saline intracerebroventricularly, DAMGO increased muscle tone within 5 min of intracerebroventricular administration, and this effect lasted for at least 80 min (Figure 6). The DAMGO time-effect curve appeared biphasic in some rats, although the group mean differences in electromyographic values over time were not statistically significant. Pretreatment with the micro antagonist CTAP significantly attenuated the muscle rigidity caused by DAMGO, although the CTAP/DAMGO group still had a significant increase in electromyographic activity for the first 30 min after DAMGO, compared with the pretreatment baseline. Thereafter, electromyographic activity was not different from baseline values.
Figure 6. Effect of 8 nmol DAMGO on muscle tone. Two intracerebroventricular injections were performed: one at time 0 and one at 10 min. Experimental group Sal/DAMGO (n = 9) received first saline and then DAMGO; the CTAP/DAMGO group first received 3 nmol CTAP and then DAMGO (n = 10). The administration of DAMGO significantly increased the muscle tone ((section)P < 0.001) compared with the baseline values. This effect was attenuated by pretreatment with CTAP (P < 0.05 for all time points). The electromyographic activity at some time points (indicated by *) within CTAP/DAMGO group was no different from the baseline values.
Figure 6. Effect of 8 nmol DAMGO on muscle tone. Two intracerebroventricular injections were performed: one at time 0 and one at 10 min. Experimental group Sal/DAMGO (n = 9) received first saline and then DAMGO; the CTAP/DAMGO group first received 3 nmol CTAP and then DAMGO (n = 10). The administration of DAMGO significantly increased the muscle tone ((section)P < 0.001) compared with the baseline values. This effect was attenuated by pretreatment with CTAP (P < 0.05 for all time points). The electromyographic activity at some time points (indicated by *) within CTAP/DAMGO group was no different from the baseline values.
Figure 6. Effect of 8 nmol DAMGO on muscle tone. Two intracerebroventricular injections were performed: one at time 0 and one at 10 min. Experimental group Sal/DAMGO (n = 9) received first saline and then DAMGO; the CTAP/DAMGO group first received 3 nmol CTAP and then DAMGO (n = 10). The administration of DAMGO significantly increased the muscle tone ((section)P < 0.001) compared with the baseline values. This effect was attenuated by pretreatment with CTAP (P < 0.05 for all time points). The electromyographic activity at some time points (indicated by *) within CTAP/DAMGO group was no different from the baseline values.
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Discussion
The current study examined the role of three major opioid receptor types in muscle rigidity by means of in vivo pharmacologic manipulation. The dose of alfentanil chosen for this study (500 micro gram/kg) produced a sustained and near maximal level of muscle rigidity alter systemic administration. [18] The selective delta-1 agonist DPDPE alone had no effect on hindlimb muscle tone; however, at an antinociceptive dose (150 nmol), DPDPE significantly decreased the alfentanil-induced muscle rigidity. Similarly, the kappa-1 agonist U50, 488H, although having no effect on electromyographic activity when given alone, produced a dose-related decrease in alfentanil-induced rigidity. Both of the non-micro opioid agonists failed to alter baseline muscle tone during the first 10 min after intracerebroventricular administration. In contrast, intracerebroventricular injection of the micro agonist DAMGO alone produced muscle rigidity in a dose-dependent manner. This effect had a rapid onset (within 5 min after intracerebroventricular injection), and the electromyographic values attained were comparable with those observed after systemic alfentanil administration. [18,19,25,26] 
Opiate-induced muscle rigidity is eliminated after spinal cord transection [28] or with systemic administration of an opiate antagonist. [25] Similarly, intracerebroventricular administration of the opiate antagonist methylnaloxonium blocks systemic opiate rigidity. [26,29] In contrast, intrathecal naloxone or methylnaloxonium will not attenuate systemic opiate rigidity (unpublished data), thereby supporting the assertion that this opiate effect is mediated solely by supraspinal opioid receptors. [17,26,29] ** An intracerebroventricular route for study drug administration was chosen in the current experiments to assure a CNS site of action of the effects under study, and also because DPDPE, DAMGO, and CTAP can be metabolized in plasma and, therefore, are not very active systemically.
There is substantial evidence for the existence of different opioid isoreceptors. [6-8] The delta isoreceptors (delta1and delta2) have been the most fully described. [6] DPDPE is a relatively delta1-selective agonist, [6,30] whereas the delta antagonist naltrindole is either not particularly isoreceptor selective or is, at best, weakly delta1-selective. [6,30,31] The role of delta2receptors in opiate rigidity remains to be elucidated.
The micro1receptor is postulated to mediate analgesia [5] and muscle rigidity, [25] whereas the micro2receptor is hypothesized to play a role in respiratory depression. [5] As appears to be the case for most micro-selective agonists, DAMGO may have a slightly higher affinity for the micro1(vs. micro2) isoreceptor. [32] Research on micro isoreceptor pharmacology is currently hampered by the absence of more isoreceptor-selective agonists. Kappa isoreceptors have been described and classified. [7] Both the agonist U50,488H and the antagonist norbinaltorphimine appear to be kappa1selective. [1,7,33-35] Unfortunately, currently available kappa2and kappa3isoreceptor ligands are insufficiently selective to permit straightforward interpretation of the results of their use in vivo.
As was observed in previous studies, [19] there was some variability in electromyographic values between study groups. Although drug doses, drug combinations, and control groups within each experiment (DAMGO, DPDPE, or U50,488H) were studied concurrently, the different opioid receptor agonists were investigated at different times throughout a 2-year period. Although all of the rats used in the experiments were of the same species (Wistar) and from the same vendor (Harlan), it is possible that minor genetic differences existed between batches. During this time period, other factors that could have contributed to the variability observed, including technical (e.g., location of electrodes) or environmental (e.g., season of the year) factors. Nevertheless, significant dose-related effects on muscle tone caused by intracerebroventricular injection of different opiate agonists (compared with parallel controls) were easily demonstrated.
The delta1agonist DPDPE, at a dose of 150 nmol, significantly attenuated systemic alfentanil-induced muscle rigidity, suggesting a modulatory role for CNS delta1receptors in opiate rigidity. This finding appears consistent with other studies, demonstrating negative or positive modulatory effects of delta agonists on micro opioid receptor agonist-mediated antinociception. [36,37] The finding that DPDPE-induced attenuation of alfentanil rigidity was blocked by pretreatment with the delta-selective antagonist naltrindole provides additional evidence for a delta receptor-mediated action. The absence of a significant effect of naltrindole alone on alfentanil-induced muscle rigidity suggests that alfentanil itself may not act at CNS delta receptors. In addition, naltrindole alone did not affect muscle tone. Further study with other selective delta ligands such as TIPP [38] and deltorphin II, a delta2-selective agonist, [13,14] will help to refine this hypothesis.
The results of earlier studies on the role of kappa receptors in muscle tone were inconclusive. Chaillet et al. [9] reported that intracerebroventricular injection of micro agonists (morphine and fentanyl) produced muscle rigidity, whereas the kappa1-preferring agonist ketocyclazocine produced muscle flaccidity. However, ketocyclazocine is not particularly kappa-selective. [8] In contrast, in the current study, intracerebroventricular administration of a highly kappa1-selective agonist, U50,488H, [7,15] failed to decrease muscle tone. The current results demonstrate, for the first time, that central injections of U50,488H antagonize alfentanil-induced muscle rigidity. The antagonism of micro receptor-mediated muscle rigidity by U50,488H is consistent with previous data on negative modulatory effects of the kappa system in other micro opioid effects. Using the kappa antagonist nor-binaltorphimine, Spanagel and Shippenberg [39] demonstrated that the endogenous kappa opioids play a role in morphine-induced behavioral sensitization. Kappa agonists can antagonize micro receptor-mediated decreased bladder motility, [40] dopamine release in locomotor systems, [41] respiratory depression, [42] alteration of seizure threshold, [43] and antinociception. [44] However, it is still unclear whether these kappa-mediated effects are due to a direct effect of kappa agonists on micro receptors or a functional interaction between the two receptor populations.
In the current study, the role of kappa1receptors in U50,488H's effects was confirmed by pretreatment with nor-binaltorphimine, a specific kappa1-opioid receptor antagonist. [45,46] Because nor-binaltorphimine is an irreversible antagonist with a slow onset, rats were pretreated intracerebroventricularly with nor-binaltorphimine 60 min before U50,488H. [34] Nor-binaltorphimine pretreatment effectively blocked U50,488H's effect on alfentanil-induced rigidity. The group that received nor-binaltorphimine alone appeared to have slightly higher electromyographic values after alfentanil than the saline group, although this effect failed to meet the criteria for statistical significance because of an increased variability of response between individual animals. Additional experiments will be required to determine the role of endogenous CNS kappa systems in the control of muscle tone.
Intracerebroventricular administration of DAMGO increased electromyographic activity in a dose-dependent manner. In addition, the pattern of the time-effect curve after 8 nmol of DAMGO appeared biphasic in many individual animals (although this pattern was obscured in the group mean), with an initial peak in the electromyographic activity at 5 min postintracerebroventricular injection and a secondary, less distinct, peak at 40-60 min. One hypothesis to explain this apparent biphasic pattern may be that, after intracerebroventricular administration, DAMGO diffuses to several active brain sites at variable distances away from the site of injection. This hypothesis is bolstered by several lines of evidence. First, a number of different brain structures have been implicated in opioid-induced muscle rigidity. These putative brain structures include the basal ganglia, [47] nucleus raphe pontis, [17,29] periaqueductal gray, [17,29,48] and locus coeruleus. [49,50] Microinjection of DAMGO into the periaqueductal gray [48] or of fentanyl into the locus coeruleus [51] both produce significant muscle rigidity. Similarly, microinjection of the antagonist methylnaloxonium into periventricular or pontine sites blocked systemic alfentanil-induced rigidity. [17] A significant decrease in alfentanil rigidity also was observed after microinjections of very small doses (0.125 micro gram in 0.1 micro liter) of methylnaloxonium into the periaqueductal gray (unpublished data). Second, the distribution after intracerebroventricular administration of a hydrophilic peptide like DAMGO has been shown to initially be to periventricular structures (e.g., periaqueductal gray) and then, after a time delay (to allow for tissue diffusion), to sites more distant from the ventricular system (e.g., substantia nigra, raphe nuclei, and globus pallidus). [51] This hypothesis must be tested with experiments specifically designed to examine the intracranial pharmacokinetics of intracerebroventricular drug administration.
Pasternak postulated the existence of two subtypes of the micro-opioid receptor: micro1and micro2. [5,8] Using naloxonazine, a putative micro1-selective irreversible antagonist, they demonstrated that catalepsy and analgesia [52] were mediated by the putative micro1receptor, whereas respiratory depression [53] was mediated by the micro2receptor. Subsequently, Negus et al. [25] showed that both naloxonazine and beta-funaltrexamine, a nonsubtype selective irreversible micro antagonist, produced similar rightward shifts in the alfentanil antinociception and muscle rigidity dose-effect curves. These data suggested that opiate-induced muscle rigidity may not be pharmacologically separable from opiate-induced antinociception on the basis of micro isoreceptor specificity. Consequently, development of even more potent micro-selective agonists may not represent an advance in pharmacotherapeutics, because opiates with greater efficacy at the micro receptor may be more likely to induce muscle rigidity. Therefore, the finding of antagonist effects of central delta and kappa agonists on mu agonist-induced rigidity potentially take on increased practical significance. In fact, significant pharmacotherapeutic benefit may accrue from the development of delta-and kappa-selective analgesics.
In conclusion, the current study supports the hypothesis that opiate-induced muscle rigidity is the result primarily of activation of central mu opioid receptors. A negative modulatory role of central delta sub 1 - and kappa1-opioid receptors on systemic micro-agonist-mediated muscle rigidity also was demonstrated. The site and mechanism of the interaction between these central opioid receptor populations remain to be elucidated. These findings have implications for the development of new opiate drugs for anesthesia and analgesia.
The authors thank Dr. Negus for his insightful suggestions with regard to the design of the experiments. They also thank Cory Campbell, David Wood, and Jeff Stuart, for technical assistance.
*In this paper, the term "opiate" is used to refer to exogenously administered drugs with morphine-like properties (e.g., alfentanil is an opiate, and alfentanil produces opiate-induced muscle rigidity). In contrast, the term "opioid" is used to refer to endogenous peptides, their derivatives, and their receptors (e.g., D-Ala2,N-Me-Ph4-Gly5-ol-enkephalin [DAMGO] is an opioid agonist that primarily acts at mu opioid receptors).
**Weinger MB: Opiate-induced muscle rigidity: Clinical implications and pathophysiology. Progress in Anesthesiology 1993; 7:198-212.
***Weinger MB: Opiates: Basic pharmacology. Progress in Anesthesiology 1995; 9:151-65.
****Bronson JB, Weinger MB: Opiate-induced muscle rigidity is mediated by mu, and not delta or kappa receptors, in the rat (abstract). ANESTHESIOLOGY 1989; 71:A600.
*****Weinger MB, Tang R, Wood DL: The effects of opioid receptor selective agonists on three anesthetic endpoints in the rat (abstract). ANESTHESIOLOGY 1993; 79:A777.
REFERENCES
Reisine T: Opiate receptors. Neuropharmacology 1995; 34:463-72.
George SR, Zastawny RL, Biones-Urbina R, Cheng R, Nguyen T, Heiber M, Kouvelas A, Chan AS, O'Dowd BF: Distinct distributions of mu, delta, and kappa opioid receptor mRNA in rat brain. Biochem Biophys Res Commun 1994; 205:1438-44.
Mansour A, Fox CA, Burke S, Meng F, Thompson RC, Akil H, Watson SJ: Mu, delta, and kappa opioid receptor mRNA expression in the rat CNS: In situ hybridization study. J Comp Neurol 1994; 350:412-38.
Uhl GR, Childers S, Pasternak G: An opiate-receptor gene family reunion. Trends Neurosci 1994; 17:89-93.
Pasternak GW: Pharmacological mechanisms of opioid analgesics. Clin Neuropharmacol 1993; 1:1-18.
Traynor JR, Elliott J: Delta-opioid receptor subtypes and crosstalk with micro-receptors. Trends Pharmacol Sci 1993; 14:84-6.
Wollemann M, Benye S, Simon J: The kappa-opioid receptor: Evidence for the different subtypes. Life Sci 1993; 52:599-611.
Pasternak GW: Multiple mu opiate receptors. ISI Atlas Sci: Pharmacol 1988; 890:148-54.
Chaillet P, Marcais-Collado H, Costentin J: Catatonic or hypotonic immobility induced in mice by intracerebroventricular injection of mu or kappa opioid receptor agonists as well as enkephalins or inhibitors of their degradation. Life Sci 1983; 33:2105-11.
Devine DP, Wise RA: Self-administration of morphine, DAMGO, and DPDPE into the ventral tegmental area of rats. J Neurosci 1994; 14:1978-84.
Frank LS, Miaskowsi C, Putris J, Levine JD: Dissociation of antinociceptive and motor effects of supraspinal opioid agonists in the rat. Brain Res 1991; 563:123-6.
Cotton R, Kosterlitz HW, Paterson SJ, Rance MJ, Traynor JR: The use of [sup 3 Hydrogen]-[D-Pen sup 2,D-Pen sup 5]enkaphalin as a highly selective ligand for the delta-binding site. Br J Pharmacol 1985; 84:927-32.
Mattia A, Vanderah T, Mosberg HI, Porreca F: Lack of antinociceptive cross-tolerance between [D-Pen sup 2,D-Pen sup 5]enkephalin and [D-Ala sup 2]deltorphin II in mice: Evidence for delta receptor subtypes. J Pharmacol Exp Ther 1991; 258:583-7.
Vanderah T, Takemori AE, Sultana M, Portoghese PS, Mosberg HI, Hruby VJ, Haaseth RC, Matsunaga TO, Porreca F: Interaction of [D-Pen sup 2,D-Pen sup 5]enkephalin and [D-Ala sup 2,Glu sup 4]deltorphin with delta-opioid receptor subtypes in vivo. Eur J Pharmacol 1994; 252:133-7.
Von Voightlander PF, Lewis RA: U50,488H: A selective and structurally novel non-mu (kappa) opioid kappa agonist. J Pharmacol Exp Ther 1983; 224:7-12.
Chien C-C, Brown G, Pan Y-X, Pasternak GW: Blockade of U50488H analgesia by antisense oligodeoxynucleotides to a kappa-opioid receptor. Eur J Pharmacol 1994; 253:R7-8.
Weinger MB, Smith NT, Blasco TA, Koob GF: Brain sites mediating opiate-induced muscle rigidity in the rat: Methylnaloxonium mapping study. Brain Res 1991; 544:181-90.
Yang P, Weinger MB, Negus SS: Elucidation of dose-effect relationships for different opiate effects using alfentanil in the spontaneously ventilating rat. ANESTHESIOLOGY 1992; 77:153-61.
Weinger MB, Bednarzyck J: Atipamezole, an alpha-2 antagonist, augments opiate-induced muscle rigidity in the rat. Pharmacol Biochem Behav 1994; 49:523-9.
Pellegrino L, Pellegrino A, Cushman A: A Stereotaxic Atlas of the Rat Brain. New York, Plenum Press, 1979.
Wood DL, Weinger MB, Furst SR: Antagonism of the respiratory effects of mu and delta opioid agonists (abstract). ANESTHESIOLOGY 1994; 81:A1408.
Wood DL, Weinger MB, Furst SR, Coleman MJ: Biphasic respiratory effects of a central delta opioid receptor agonist (abstract). ANESTHESIOLOGY 1994; 81:A1409.
Negus SS, Weinger MB, Hendricksen SJ, Koob GF: Effect of mu, delta, and kappa opioid agonists on heroin-maintained responding in the rat. Problems of Drug Dependence 1991. NIDA Res Monogr 1992; 119:410.
Negus SS, Henriksen SJ, Mattox A, Pasternak GW, Portoghese PS, Takemori AE, Weinger MB, Koob GF: Effect of antagonists selective for mu-opioid, delta-opioid, and kappa-opioid receptors on the reinforcing effects of heroin in rats. J Pharmacol Exp Ther 1993; 265:1245-52.
Negus SS, Pasternak GW, Koob GF, Weinger MB: Antagonist effects of betafunaltrexamine and naloxonazine on alfentanil-induced antinociception and muscle rigidity in the rat. J Pharmacol Exp Ther 1993; 264:739-45.
Weinger MB, Chen D-Y, Lin T, Lau C, Koob GF, Smith NT: A role for CNS alpha-2 adrenergic receptors in opiate-induced muscle rigidity in the rat. Brain Res 1995; 669:10-8.
Winer BJ: Statistical Principles in Experimental Design. 2nd edition. New York, McGraw-Hill, 1971.
Seeber U, Kuschinsky K, Sontag KH: Inhibition by opiate narcotics of rat flexor alpha-motoneurons. Naunyn Schmeidberg's Arch Pharmacol 1978; 301:181-5.
Blasco TA, Lee DE, Almaric M, Swerdlow NR, Smith NT, Koob GF: The role of the nucleus raphe pontis and the caudate nucleus in alfentanil rigidity in the rat. Brain Res 1986; 386:280-6.
Jiang QI, Takemori AE, Sultana M, Portoghese PS, Bowen WD, Mosberg HI, Porreca F: Differential antagonism of opioid delta antinociception by [D-Ala sup 2,Leu sup 5,Cys sup 6]enkephalin and naltrindole 5'-isothiocyanate: Evidence for delta receptor subtypes. J Pharmacol Exp Ther 1991; 257:1069-75.
Portoghese PS, Sultana M, Takemori AE: Naltrindole, a highly selective and potent non-peptide delta opioid receptor antagonist. Eur J Pharmacol 1988; 146:185-6.
Clark JA, Houghton R, Pasternak GW: Opiate binding in calf thalamic membranes: A selective micro sub 1 binding assay. Mol Pharmacol 1988; 34:308-17.
Clark JA, Liu L, Price M, Hersh B, Edelson M, Pasternak GW: Kappa opiate receptor multiplicity: Evidence for two U50488-sensitive kappa sub 1 subtypes and a novel kappa sub 3 subtype. J Pharmacol Exp Ther 1989; 251:461-8.
Horan P, Taylor J, Yamamura HI, Porreca F: Extremely long-lasting antagonistic actions of nor-binaltorphimine (nor-BNI) in the mouse tail flick test. J Pharmacol Exp Ther 1992; 260:1237-43.
Mansour A, Fox CA, Meng F, Akil H, Watson SJ: kappa sub 1 receptor mRNA distribution in the rat CNS: Comparison to kappa receptor binding and prodynorphin mRNA. Mol Cell Neurosci 1994; 5:124-44.
Horan P, Tallarida RJ, Haaseth R, Matsunaga T, Hruby VJ, Porreca F: Antinociceptive interactions of opioid delta receptor agonists with morphine in mice: Supra- and subadditivity. Life Sci 1992; 50:1535-41.
Porreca F, Takemori AE, Portoghese PS, Sultana M, Bowen WD, Mosberg HI: Modulation of mu-mediated antinociception by a subtype of opioid delta receptor in the mouse. J Pharmacol Exp Ther 1992; 263:147-52.
Lee PH, Nguyen TM, Chung NN, Schiller PW, Chang KJ: Tyrosine-iodination converts the delta-opioid peptide antagonist TIPP to an agonist. Eur J Pharmacol 1995; 280:211-4.
Spanagel R, Shippenberg T: Modulation of morphine-induced sensitization by endogenous kappa opioid system in the rat. Neurosci Lett 1993; 153:232-6.
Sheldon RJ, Nunan L, Porreca F: Mu antagonist properties of kappa agonists in a model of rat urinary bladder motility in vivo. J Pharmacol Exp Ther 1987; 243:234-40.
Kim HS, Iyengar S, Wood PL: Reversal of the actions of morphine on mesocortical dopamine metabolism in the rat by the kappa agonist MR-2034: Tentative mu-2 opioid control of mesocortical dopaminergic projections. Life Sci 1987; 41:1711-5.
Dosaka-Akita K, Tortella FC, Holaday JW, Long JB: The kappa opioid agonist U-50,488H antagonizes respiratory effects of mu opioid receptor agonists in conscious rats. J Pharmacol Exp Ther 1993; 264:631-7.
Porreca F, Tortella FC: Differential antagonism of mu agonists by U50488H in the rat. Life Sci 1987; 41:2511-6.
Bhargava HN, Ramarao P, Gulati A: Effects of morphine in rats treated chronically with U-50,488H, a kappa opioid receptor agonist. Eur J Pharmacol 1989: 162:257-64.
Endoh T, Matsuura H, Tanaka C, Nagase H: Nor-binaltorphimine:a potent and selective kappa-opioid receptor antagonist with long-lasting activity in vivo. Arch Int Pharmacodyn Ther 1992; 316:30-42.
Portoghese PS, Lipkowski AW, Takemori AE: Binaltorphimine and nor-binaltorphimine, potent and selective kappa-opioid receptor antagonists. Life Sci 1987; 40:1287-92.
Slater P, Starkie DA: Changes in limb tone produced by regional injections of opiates into rat brain. Naunyn-Schmiedbergs Arch Pharmacol 1987; 335:54-8.
Widdowson PS, Griffiths EC, Slater P: The effects of opioids in the periaqueductal gray region of rat brain on hind-limb muscle tone. Neuropeptides 1986; 7:251-8.
Lui PW, Lee TY, Chan SHH: Involvement of locus coeruleus and noradrenergic neurotransmission in fentanyl-induced muscular rigidivity in the rat. Neurosci Lett 1989; 96:114-9.
Lui P, Lee T, Chan S: Involvement of coerulospinal noradrenergic pathway in fentanyl-induced muscular rigidity in rats. Neurosci Lett 1990; 108:183-8.
Dauge V, Petit F, Rossignol P, Roques BP: Use of micro and delta opioid peptides of various selectivity gives evidence of specific involvement of micro opioid receptors in supraspinal analgesia (tail-flick test). Eur J Pharmacol 1987; 141:171-8.
Ling GSF, Pasternak GW: Morphine catalepsy in the rat: Involvement of mu1 (high affinity) opioid binding sites. Prog Neurobiol 1982; 32:193-6.
Ling GSF, Spiegel K, Lockhart SH, Pasternak GW: Separation of opioid analgesia from respiratory depression: Evidence for different receptor mechanisms. J Pharmacol Exp Ther 1985; 232:149-55.
Figure 1. Effect of different doses of DPDPE on electromyographic activity. Y axis gives the area under the electromyographic data versus time curve (AUC): (a) saline-treated groups; and (b) alfentanil-treated groups (500 micro gram/kg subcutaneously). N = 6 for each dose examined. Significant decrease in alfentanil-induced muscle rigidity (*P < 0.05) after intracerebroventricular pretreatment with DPDPE (150 nmol), compared with saline intracerebroventricularly (zero dose group).
Figure 1. Effect of different doses of DPDPE on electromyographic activity. Y axis gives the area under the electromyographic data versus time curve (AUC): (a) saline-treated groups; and (b) alfentanil-treated groups (500 micro gram/kg subcutaneously). N = 6 for each dose examined. Significant decrease in alfentanil-induced muscle rigidity (*P < 0.05) after intracerebroventricular pretreatment with DPDPE (150 nmol), compared with saline intracerebroventricularly (zero dose group).
Figure 1. Effect of different doses of DPDPE on electromyographic activity. Y axis gives the area under the electromyographic data versus time curve (AUC): (a) saline-treated groups; and (b) alfentanil-treated groups (500 micro gram/kg subcutaneously). N = 6 for each dose examined. Significant decrease in alfentanil-induced muscle rigidity (*P < 0.05) after intracerebroventricular pretreatment with DPDPE (150 nmol), compared with saline intracerebroventricularly (zero dose group).
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Figure 2. Effect of different doses of U50,488H on the electromyographic activity (electromyography-time area under the curve): (a) saline-treated groups; and (b) alfentanil-treated groups. N = 6 for each dose. U50,488H (107 nmol) pretreatment significantly decreased alfentanil-induced muscle rigidity (*P < 0.05) when compared with saline intracerebroventricularly (zero dose group).
Figure 2. Effect of different doses of U50,488H on the electromyographic activity (electromyography-time area under the curve): (a) saline-treated groups; and (b) alfentanil-treated groups. N = 6 for each dose. U50,488H (107 nmol) pretreatment significantly decreased alfentanil-induced muscle rigidity (*P < 0.05) when compared with saline intracerebroventricularly (zero dose group).
Figure 2. Effect of different doses of U50,488H on the electromyographic activity (electromyography-time area under the curve): (a) saline-treated groups; and (b) alfentanil-treated groups. N = 6 for each dose. U50,488H (107 nmol) pretreatment significantly decreased alfentanil-induced muscle rigidity (*P < 0.05) when compared with saline intracerebroventricularly (zero dose group).
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Figure 3. Effect of different doses of the micro receptor agonist DAMGO on electromyographic activity (electromyographytime area under the curve). N = 6 for the saline group, n = 7 for the 4 nmol, and n = 8 for the 8 nmol group. *P < 0.005 compared with saline group.
Figure 3. Effect of different doses of the micro receptor agonist DAMGO on electromyographic activity (electromyographytime area under the curve). N = 6 for the saline group, n = 7 for the 4 nmol, and n = 8 for the 8 nmol group. *P < 0.005 compared with saline group.
Figure 3. Effect of different doses of the micro receptor agonist DAMGO on electromyographic activity (electromyographytime area under the curve). N = 6 for the saline group, n = 7 for the 4 nmol, and n = 8 for the 8 nmol group. *P < 0.005 compared with saline group.
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Figure 4. Effect of pretreatment with 150 nmol DPDPE (Sal/DPDPE group,); 10 nmol NTI (NTI/Sal group); 10 nmol NTI and 150 nmol DPDPE (NTI/DPDPE group); or saline (Sal/Sal group) on the alfentanil-induced muscle rigidity. The first intracerebroventricular injection (saline or NTI) was performed at 0 min, the second intracerebroventricular injection (DPDPE or saline) was at 15 min. All animals received 500 micro gram/kg alfentanil subcutaneously at 25 min. N = 7 for all experimental groups. The administration of DPDPE (Sal/DPDPE group) significantly decreased alfentanil-induced muscle rigidity compared with Sal/Sal group (*P < 0.05; +P < 0.001). There was no significant difference in electromyographic activity between Sal/NTI; NTI/DPDPE, and Sal/Sal groups.
Figure 4. Effect of pretreatment with 150 nmol DPDPE (Sal/DPDPE group,); 10 nmol NTI (NTI/Sal group); 10 nmol NTI and 150 nmol DPDPE (NTI/DPDPE group); or saline (Sal/Sal group) on the alfentanil-induced muscle rigidity. The first intracerebroventricular injection (saline or NTI) was performed at 0 min, the second intracerebroventricular injection (DPDPE or saline) was at 15 min. All animals received 500 micro gram/kg alfentanil subcutaneously at 25 min. N = 7 for all experimental groups. The administration of DPDPE (Sal/DPDPE group) significantly decreased alfentanil-induced muscle rigidity compared with Sal/Sal group (*P < 0.05; +P < 0.001). There was no significant difference in electromyographic activity between Sal/NTI; NTI/DPDPE, and Sal/Sal groups.
Figure 4. Effect of pretreatment with 150 nmol DPDPE (Sal/DPDPE group,); 10 nmol NTI (NTI/Sal group); 10 nmol NTI and 150 nmol DPDPE (NTI/DPDPE group); or saline (Sal/Sal group) on the alfentanil-induced muscle rigidity. The first intracerebroventricular injection (saline or NTI) was performed at 0 min, the second intracerebroventricular injection (DPDPE or saline) was at 15 min. All animals received 500 micro gram/kg alfentanil subcutaneously at 25 min. N = 7 for all experimental groups. The administration of DPDPE (Sal/DPDPE group) significantly decreased alfentanil-induced muscle rigidity compared with Sal/Sal group (*P < 0.05; +P < 0.001). There was no significant difference in electromyographic activity between Sal/NTI; NTI/DPDPE, and Sal/Sal groups.
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Figure 5. Effect of pretreatment with 107 nmol U50,488H (Sal/U50,488H group, n = 7); 10 nmol nor-binaltorphimine (nor-binaltorphimine/Sal group, n = 6); 10 nmol nor-binaltorphimine and 107 nmol U50,488H (nor-binaltorphimine/U50,488H group, n = 6); or saline (Sal/Sal group, n = 7) on alfentanil-induced muscle rigidity. The intracerebroventricular injections were performed at time points 0 (saline or nor-binaltorphimine) and 60 (saline or U50,488H); alfentanil was administered subcutaneously at 70 min. The pretreatment with U50,488H (Sal/U50,488H group) significantly decreased the alfentanil-induced muscle rigidity compared with the control group (Sal/Sal) (*P < 0.05; +P < 0.005). No significant differences between Sal/Sal, nor-binaltorphimine/Sal, and nor-binaltorphimine/U50,488H groups were observed.
Figure 5. Effect of pretreatment with 107 nmol U50,488H (Sal/U50,488H group, n = 7); 10 nmol nor-binaltorphimine (nor-binaltorphimine/Sal group, n = 6); 10 nmol nor-binaltorphimine and 107 nmol U50,488H (nor-binaltorphimine/U50,488H group, n = 6); or saline (Sal/Sal group, n = 7) on alfentanil-induced muscle rigidity. The intracerebroventricular injections were performed at time points 0 (saline or nor-binaltorphimine) and 60 (saline or U50,488H); alfentanil was administered subcutaneously at 70 min. The pretreatment with U50,488H (Sal/U50,488H group) significantly decreased the alfentanil-induced muscle rigidity compared with the control group (Sal/Sal) (*P < 0.05; +P < 0.005). No significant differences between Sal/Sal, nor-binaltorphimine/Sal, and nor-binaltorphimine/U50,488H groups were observed.
Figure 5. Effect of pretreatment with 107 nmol U50,488H (Sal/U50,488H group, n = 7); 10 nmol nor-binaltorphimine (nor-binaltorphimine/Sal group, n = 6); 10 nmol nor-binaltorphimine and 107 nmol U50,488H (nor-binaltorphimine/U50,488H group, n = 6); or saline (Sal/Sal group, n = 7) on alfentanil-induced muscle rigidity. The intracerebroventricular injections were performed at time points 0 (saline or nor-binaltorphimine) and 60 (saline or U50,488H); alfentanil was administered subcutaneously at 70 min. The pretreatment with U50,488H (Sal/U50,488H group) significantly decreased the alfentanil-induced muscle rigidity compared with the control group (Sal/Sal) (*P < 0.05; +P < 0.005). No significant differences between Sal/Sal, nor-binaltorphimine/Sal, and nor-binaltorphimine/U50,488H groups were observed.
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Figure 6. Effect of 8 nmol DAMGO on muscle tone. Two intracerebroventricular injections were performed: one at time 0 and one at 10 min. Experimental group Sal/DAMGO (n = 9) received first saline and then DAMGO; the CTAP/DAMGO group first received 3 nmol CTAP and then DAMGO (n = 10). The administration of DAMGO significantly increased the muscle tone ((section)P < 0.001) compared with the baseline values. This effect was attenuated by pretreatment with CTAP (P < 0.05 for all time points). The electromyographic activity at some time points (indicated by *) within CTAP/DAMGO group was no different from the baseline values.
Figure 6. Effect of 8 nmol DAMGO on muscle tone. Two intracerebroventricular injections were performed: one at time 0 and one at 10 min. Experimental group Sal/DAMGO (n = 9) received first saline and then DAMGO; the CTAP/DAMGO group first received 3 nmol CTAP and then DAMGO (n = 10). The administration of DAMGO significantly increased the muscle tone ((section)P < 0.001) compared with the baseline values. This effect was attenuated by pretreatment with CTAP (P < 0.05 for all time points). The electromyographic activity at some time points (indicated by *) within CTAP/DAMGO group was no different from the baseline values.
Figure 6. Effect of 8 nmol DAMGO on muscle tone. Two intracerebroventricular injections were performed: one at time 0 and one at 10 min. Experimental group Sal/DAMGO (n = 9) received first saline and then DAMGO; the CTAP/DAMGO group first received 3 nmol CTAP and then DAMGO (n = 10). The administration of DAMGO significantly increased the muscle tone ((section)P < 0.001) compared with the baseline values. This effect was attenuated by pretreatment with CTAP (P < 0.05 for all time points). The electromyographic activity at some time points (indicated by *) within CTAP/DAMGO group was no different from the baseline values.
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