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The local monoaminergic dependency of spinal ketamine
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European Journal of Pharmacology, 194 (1991) 167-172 © 1991 Elsevier Science Publishers B.V. 0014-2999/91/$03.50 ADONIS 001429999100201G EJP 51743
The local monoaminergic dependency of spinal ketamine Terriann Crisp, Jeannette M. Perrotti, Deborah L. Smith, Janet L. Stafinsky and David J. Smith Department o/Anesthesiology, West Virginia University Health Sciences Center, Morgantown, WV 26506, U.S.A. and Department o/Pharmacology, Northeastern Ohio Universities College of Medicine, Rootstown, OH 44272, U.S.A. Received 28 June 1990, revised MS received 31 October 1990, accepted 4 December 1990
The effects of s.c. doses of naloxone, methysergide and phentolamine on ketamine-induced spinal analgesia were assessed to determine the involvement of opiate, serotonergic and noradrenergic components mediating ketamine's antinociceptive action. Ketamine administered intrathecally (i.t.) produced a significant elevation in tail-flick latency in rats. The spinal antinociceptive effects of ketamine were dose dependently reversed by methysergide (IDs0 = 0.008 mg/kg s.c.), phentolamine (IDs0 = 0.88 mg/kg s.c.) and naloxone (IDs0 = 3.0 mg/kg s.c.). Unlike morphine, which remains analgesic and dependent on opiate interactions following bilateral lesions of the dorsolateral funiculus (DLF), ketamine analgesia was absent following DLF lesions. Thus, ketamine appears to produce an antinociceptive response which is dependent upon the neuronal activity of the descending pain-inhibitory pathways. The monoaminergic components comprising the descending pathways appear to be more prominent in the action of ketamine than they are in the spinal action of morphine. Furthermore, the spinal opioid receptors involved in ketamine's effect may be different from the/~ subtype preferred by morphine. Ketamine; Analgesia; Opioid receptors; 5-HT (5-hydroxytryptamine,serotonin); Norepinephrine; Tail-flick test; (Intrathecal)
1. Introduction
Racemic ketamine HC1 is a dissociative anesthetic agent which exhibits analgesic properties in rodents (Ryder et al., 1978) and humans (White et al., 1980). Although the mechanisms underlying the analgesic action of the drug are not clearly understood, it has been shown that ketamine stereoselectively binds with a weak affinity to opiate receptors (Smith et al., 1980; 1987). Additionally, the antinociceptive action of systemically administered ketamine is naloxone-reversible (Pekoe and Smith, 1982; Ryder et al., 1978; Smith et al., 1985). The fact that ketamine-induced analgesia is reversed by naloxone does not imply direct opiate receptor activation as the primary mechanism by which the opiate produces its analgesic action. For instance, ketamine has repeatedly been shown to interact with monoaminergic (serotonergic, noradrenergic and dopaminergic) neuronal systems (Tung and Yaksh, 1981; Martin and Smith, 1982). Moreover, the analgesic properties of ketamine may be mediated via the ability of the agent to block high affinity mohoaminergic uptake sites (Pekoe and Smith, 1982; Smith et al., 1981; Azzaro and Smith, 1977). Correspondence to: T. Crisp, Department of Pharmacology, Northeastern Ohio Universities, College of Medicine, Rootstown, OH 44272, U.S.A.
Recent evidence suggests the involvement of both monoaminergic (Tung and Yaksh, 1981; Pekoe and Smith, 1982) and opiate (Smith et al., 1985) components mediating ketamine-induced analgesia. However, the analgesic efficacy of ketamine may change as a consequence of the route of administration and/or the site of injection of the drug (Smith et al., 1985). For instance, ketamine injected intracerebroventricularly (i.c.v.) or into the periaqueductal gray region of the rat mesencephalon, an area rich in opiate receptor sites (Bennet and Mayer, 1979), fails to elevate tail-flick latency (TFL) in rats (Smith et al., 1985; 1990). On the other hand, Tung and Yaksh (1981) reported that ketamine injected directly into the subarachnoid space of the rat spinal cord produced a weak analgesic action that was reversed by the serotonin (5-HT) receptor antagonist methysergide. That 5-HT was involved in mediating the local spinal action of ketamine was further substantiated by reports demonstrating that intraperitoneal (i.p.) injections of ketamine produced methysergide-reversible elevations in TFL 1-2 days following spinal transection in rats (Pekoe and Smith, 1982). However, neither naloxone nor the a-adrenoceptor antagonist phentolamine effectively reversed the antinociceptive effects of i.p. ketamine in spinally transected rats. Thus, a prominent 5-HT component may mediate the spinal analgesic effects of ketamine (Pekoe and Smith, 1982; Crisp et al., 1987).
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The purpose of the present study was to more thoroughly identify the mediatory role of opioid, serotonergic and noradrenergic neuronal systems in spinal ketamine-induced analgesia. This was done by testing the ability of various subcutaneous (s.c.) doses of naloxone, methysergide and phentolamine to alter the antinociceptive effects of ketamine following intrathecal (i.t.) injections of the anesthetic.
2. Materials and methods
2.1. Animal model and surgical preparations Male Sprague-Dawley rats (350 _+ 50 g) were chronically implanted with i.t. polyethylene (PE-10) catheters (LoPachin et al., 1981) to allow for the direct spinal administration of ketamine. Under ketamine anesthesia, each rat was fitted with an indwelling spinal catheter which was inserted through a slit in the atlanto-occipital membrane and passed distally approximately 8.5 cm to the rostral edge of the lumbar enlargement. Rats were allowed at least 7 days to recuperate from surgery, and only those animals which displayed unimpaired motor function after i.t. catheterization were used in the experiments. An interval of at least 2 weeks was maintained between repeated injections, and rats were not injected more than twice. After the second injection, rats were killed and cannula placement was checked using methylene blue dye. Data obtained from animals with malpositioned catheters (e.g. tip location in the epidural space) were excluded from the study. Bilateral dorsolateral funiculus (DLF) lesions were made in rats previously cannulated with i.t. catheters using a lesioning technique described by Hayes et al. (1978). In short, a laminectomy was done to expose the cord at the level of T1-T4. The dura was removed and a pair of dissecting forceps were inserted into the cord and closed for 15-20 s bilaterally to lesion the DLF. After the lesions were made, the exposed cord was covered with Gelfoam, soaked with 0.9% saline, and the wound was then sutured. Analgesic tests were conducted in DLF-lesioned rats 24-48 h after the lesions were made, at a time when monoaminergic nerve terminals below the site of the lesion were still intact (Magnusson, 1973). The analgesic efficacy of ketamine administered i.t. was evaluated in rats with intact spinal cords, and in rats with bilateral D L F lesions.
2.2. Analgesiometric assay The spinal antinociceptive efficacy of different doses of ketamine was measured using a modified version of the tail-flick test (D'Amour and Smith, 1941). A Model 33 Analgesia Meter was used, and the rat's tail (blackened with India Ink) was placed in a depression over
the photocell in the tail-flick meter. The time (in s) required for the rat to remove its tail from the light source was automatically determined and was expressed as TFL. Routinely, four baseline values were obtained, and the mean of the last three was used as the predrug TFL. Preliminary experiments from this laboratory and others (Tung and Yaksh, 1981) have demonstrated that the analgesic action of spinal ketamine is only minimally detectable when using 'standard' T F L values (i.e. baseline T F L between 2-4 s, cut-off T F L values of 10 s). By decreasing the intensity of the light source on the tail-flick meter, we were able to increase the sensitivity of the tail-flick analgesiometric assay (Tung and Yaksh, 1981). Therefore, for these experiments, the intensity of the bulb on the tail-flick meter was adjusted to obtain predrug baseline values of 5-8 s, and a 15 s maximal exposure to the heat source was employed as the cut-off value to preclude tissue damage to the tail.
2.3. The effects of ketamine administered i.t. in rats Different doses of ketamine (0.3, 1 and 3 ~tmol/10 /~1) were administered i.t. in rats using a Harvard Model 975 Infusion Pump that was pre-set to deliver 10/zl of drug or saline per min. Each drug injection was followed by a 5/~1 saline flush to clear the spinal catheter of the drug. Drug-induced changes in T F L were obtained 5, 10, 15, 30 and 60 min post-ketamine injection, and dose-response curves were generated using individual groups of rats for each dose of ketamine. Ketamine-induced alterations in T F L were evaluated in rats with intact spinal cords and in rats with bilateral D L F lesions.
2.4. The relative participation of opiate, serotonergic and noradrenergic components mediating the spinal antinociceptive action of ketamine To determine if ketamine administered i.t. interacted with opioid a n d / o r monoaminergic neuronal systems to produce spinal analgesia, varying doses of naloxone (0.3-10 mg/kg), methysergide (0.001-0.1 mg/kg) or phentolamine (0.01-10 m g / k g ) were administered s.c. and tested for an ability to alter ketamine-induced spinal analgesia. In these experiments, the 3/~mol dose of ketamine was tested against the various antagonists because it produced consistent elevations in T F L without taking animals to cut-0ff. The range of inhibitory doses for the antagonists used in this study was obtained from previous experiments demonstrating the inhibitory efficacy of s.c. naloxone, methysergide and phentolamine on the spinal analgesic effects of morphine, 5-HT and norepinephrine (NE), respectively (Crisp and Smith, 1989). Naloxone was injected s.c. immediately prior to i.t. ketamine injections, whereas
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methysergide and phentolamine were administered 15 min prior to spinal ketamine injections. In this manner, the maximal inhibitory effect of the antagonists would overlap with the time of ketamine's peak antinociceptive effect (established in the initial dose-response and duration experiments). Doses of naloxone, methysergide and phentolamine that were effective in reversing ketamine-induced spinal analgesia were injected alone in other experiments and tested for an ability to significantly alter T F L values by themselves. These values were compared to s.c. saline control T F L values.
2.5 Data analysis In order to analyze the ability of naloxone, methysergide and phentolamine to alter the spinal antinociceptive effects of ketamine, comparative assessments were made using apparent IDs0 values (defined as the dose of antagonist producing 50% inhibition of ketamine-induced analgesia; Tallarida and Murray, 1981). The probit analysis required that graded responses, obtained with a fixed dose of ketamine in the presence of various doses of each antagonist, be transformed to a quantal form. For this purpose, analgesia was defined as a post-drug T F L value that was greater than, or equal to, two S.D.s above the mean pre-drug latency of the study group (Crisp and Smith, 1989). All data were derived from experiments having an n of at least six animals per treatment group. Tail-flick latency values in figs. 1-5 are expressed as means + S.E.M. Statistical analyses of dose-response and duration data were conducted using a repeated measures analysis of variance (ANOVA) and a Dunnett's test as applicable to calculate significant differences between the means (P < 0.05). To statistically analyze the effects of different doses of naloxone, m e t h y s e r g i d e and p h e n t o l a m i n e on ketamine-induced analgesia, an ANOVA with no replications was used.
15-
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Fig. 1. Dose-responseand duration of i.t. ketamine((o) saline; (-) 0.3 #mol; (It) 1 #mol; (zx)3/xmol). Values represent the means+ S.E.M. of at least six animals/dose. * P < 0.05 versuspre-drug TFL values. with bilateral D L F lesions, the 3 /~mol i.t. dose of ketamine was no longer analgesic, and did not significantly elevate T F L values above pre-drug control values (fig. 2). Overt motor changes resulting from ketamine injections were assessed by placing rats on a 60 ° vertical wire mesh (Tung and Yaksh, 1981). In none of the rats tested did 0.3 or 1 /~mol dose of ketamine produce an inability to negotiate the mesh. The 3 /~mol dose produced a reversible hind limb paralysis within 3-4 min post-injection, but this effect lasted no longer than 8-10 min.
3.2. The effects of naloxone, methysergide and phentolamine administered s.c. on the spinal analgesic effects of ketamine In experiments designed to assess the involvement of opiate a n d / o r monoaminergic components mediating ketamine's spinal action, the 3 /~mol dose of ketamine was used. This dose produced a consistent analgesic 15'
3.
Results
,~
3.1. The effects of ketamine administered i.t. in rats In rats with intact spinal cords, ketamine injected i.t. produced a dose-dependent elevation in T F L that was significantly greater than pre-drug control values for 15 min post-injection (fig. 1). The onset of drug action occurred within 10 min, and the peak analgesic effect for the 1 and 3/~mol doses of ketamine was observed at 15 .m in post-injection. The T F L response to the 0.3 /~mol dose of the anesthetic was indistinguishable from pre-drug control values for the 60 min duration of the dose-response experiments. Ketamine-induced elevations in T F L were not significantly greater than controls after 15 min post-injection (fig. 1). Interestingly, in rats
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Fig. 2. Lack of response of DLF-lesioned rats to 3 /~mol ketamine. Values represent the means+S.E.M, of at least six animals. Fifteen minutes post-drug TFL values are compared to pre-drug controls.
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Fig. 3. High doses of naloxone administered s.c. reverse the analgesic action of i.t. ketamine. Values are the means + S.E.M. of at least six animals/treatment group. Time =15 min post-ketamine and 15 min post-naloxone. * P < 0.05 versus saline control, * * P < 0.05 versus 3 t~mol ketamine alone. response o n the tail-flick test w i t h o u t elevating T F L values a b o v e the 15 s cut-off point. T h e o p i a t e r e c e p t o r a n t a g o n i s t n a l o x o n e p r o d u c e d a d o s e - d e p e n d e n t reversal of k e t a m i n e ' s spinal a c t i o n (fig. 3), a n d h a d an a p p a r e n t IDs0 of 3.0 (2.11-4.28) m g / k g . V a r y i n g doses of m e t h y s e r g i d e were also tested against k e t a m i n e in a t t e m p t s to assess the relative i n v o l v e m e n t o f 5 - H T n e u r o n a l processes in k e t a m i n e - i n d u c e d spinal analgesia. M e t h y s e r g i d e dose d e p e n d e n t l y reversed k e t a m i n e - i n d u c e d elevations in T F L (fig. 4), having an a p p a r e n t IDs0 value of 0.008 (0.003-0.025) m g / k g . P h e n t o l a m i n e also decreased the spinal analgesic effects of k e t a m i n e in a d o s e - d e p e n d e n t m a n n e r (fig. 5). T h e a - a d r e n o c e p tor a n t a g o n i s t h a d an a p p a r e n t IDs0 of 0.88 (0.23-3.38) m g / k g s.c. D o s e s of naloxone, m e t h y s e r g i d e a n d p h e n 15'i --,
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Fig. 4. Methysergide administered s.c. dose dependently reversed the spinal effects of ketamine. Values are the means + S.E.M. of at least six animals/treatment group. Time = 15 min post-ketamine, 30 min post-methysergide. * P < 0.05 versus i.t. saline controls, * * P < 0.05 (one-sided) versus 3 t~mol ketamine alone.
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Fig. 5. Phentolamine administered s.c. reversed the effects of ketamine in a dose-dependent manner. Values represent the means_+S.E.M. of at least six animals/treatment group. Time =15 min post-ketamine and 30 min post-phentolamine. * P < 0.05 versus i.t. saline controls, • * P < 0.05 versus 3 #mol ketamine alone.
t o l a m i n e t h a t b l o c k e d the spinal a n t i n o c i c e p t i v e effects of the 3 /~mol d o s e of k e t a m i n e also reversed the analgesic effects o f the 1 ~ m o l d o s e of k e t a m i n e a d m i n istered i.t. A d d i t i o n a l l y , w h e n s.c. doses of n a l o x o n e (10 m g / k g ) , m e t h y s e r g i d e (1 m g / k g ) a n d p h e n t o l a m i n e (10 m g / k g ) were a d m i n i s t e r e d alone in a n o t h e r g r o u p of animals, no significant changes in T F L were o b s e r v e d for 30 m i n p o s t - i n j e c t i o n when c o m p a r e d to i.t. saline values.
4. D i s c u s s i o n
T h e p r e s e n t d a t a suggest that the analgesic effects of k e t a m i n e a d m i n i s t e r e d i.t. are spinally m e d i a t e d . W h e n k e t a m i n e was injected i.c.v, in rats, even high doses failed to p r o d u c e a n a l g e s i a ( S m i t h et al., 1990), suggestm g that the r o s t r a l diffusion of i.t. k e t a m i n e to s u p r a s p i n a l sites was n o t requisite for an analgesic r e s p o n s e to b e evoked. F u r t h e r m o r e , the r a p i d onset of a n a l g e s i a (5-10 min) also suggests that local spinal processes are involved. T h e c u r r e n t s t u d y also c o n f i r m s the i n v o l v e m e n t of o p i o i d a n d m o n o a m i n e r g i c processes in the spinal action of k e t a m i n e . H o w e v e r , the m e c h a n i s m ( s ) u n d e r l y ing the i n t e r a c t i o n o f k e t a m i n e with these systems rem a i n unclear. T h e fact that b i l a t e r a l lesions of the D L F d i m i n i s h e d the a n t i n o c i c e p t i v e a c t i o n of i.t. k e t a m i n e does imply, however, that n e u r o n a l activity in descending p a i n i n h i b i t o r y nerves ( R i v o t et al., 1987; Crisp a n d Smith, 1989) is necessary for k e t a m i n e - i n d u c e d spinal a n a l g e s i a to b e expressed. I n this regard, it has been e s t a b l i s h e d that k e t a m i n e has the p o t e n t i a l to e n h a n c e m o n o a m i n e r g i c (5-HT, N E
171 and DA) neuronal transmission by inhibiting the high affinity monoaminergic uptake carriers (Azzaro and Smith, 1977; Smith et al., 1981). Perhaps the monoaminergic dependency of ketamine-induced analgesia in intact animals is related to a blockade of monoamine uptake and the enhancement of the spinal synaptic activity of 5-HT (Yaksh and Wilson, 1979) and NE (Reddy et al., 1980). An action such as this would require neuronal activity in the descending monoaminergic pathways to initiate the release of neurotransmitters, and would be interrupted by DLF lesions as observed in this study. This mechanism is also consistent with the suggestion by Hwang and Wilcox (1987) that the analgesic activity of various heterocyclic antidepressants (e.g. desipramine, protriptyline, fluoxetine and citalopram) is related to their ability to block monoaminergic uptake. Furthermore, preliminary experiments in this laboratory have demonstrated that the 5-HT uptake blocker fluoxetine produces antinociceptive effects similar to ketamine when administered i.t. using the extended baseline (Crisp, studies in progress). On the other hand, since ketamine also has the potential to interact at opiate receptors as an agonist (Smith et al., 1982), the inability to observe an antinociceptive action in the absence of tonic neuronal activity is more difficult to explain. Morphine-like opiate agonists produce analgesia in DLF-lesioned animals, presumably as a result of their interaction with intrinsic /x opiate receptors in the dorsal horn of the spinal cord (Crisp and Smith, 1989). In contrast however, it has been suggested that the # opiate receptor subtype preferred by morphine is not the same opiate receptor responsible for ketamine's opiate agonistic action (Smith et al., 1982; 1987; 1990). Data from the present study seem to confirm this suggestion since higher doses of naloxone were required to block the spinal effects of ketamine than were needed to reverse the antinociceprive effect of i.t. morphine (cf. Crisp and Smith, 1989). Thus, an opiate receptor subtype less sensitive to naloxone (e.g. K or putative phencyclidine (PCP)/o sites) may be involved (Zukin and Zukin, 1981; Contreras et al., 1986; Smith et al., 1987; Anis et al., 1983; Thompson et al., 1985). The inactivity of ketamine in DLF-lesioned rats may suggest that ketamine-sensitive opiate receptors lie on interneurons that modulate activity in tonically active descending antinociceptive neuronal pathways, as has been suggested by Monroe et al. (1986). On the other hand, the reliance of ketamine on opiate mechanisms may be secondary to actions on other systems. With regard to the latter case, Basbaum and Fields (1978) proposed the existence of enkephalinergic interneurons between descending serotonergic nerve terminals and primary afferent nerve terminals in the spinal cord. Such neuronal circuitry could provide a basis for a naloxone-reversible action of ketamine secondary to its
ability to block 5-HT uptake. Alternatively, the inactivity of ketamine in DLF-lesioned rats may suggest that it lacks meaningful interactions with opiate systems in the spinal cord as has been suggested by Tung and Yaksh (1981). Accordingly, the requirement for high doses of naloxone to reverse ketamine-induced analgesia in intact animals may be related to non-opiate neuronal effects of the antagonist. The spinal or epidural administration of ketamlne in man (Guinto-Enriquez et al., 1990; Schulman et al., 1990) and animals (Tung and Yaksh, 1981) is attended by sensory blockade as well as a shorter-rived reversible motor blockade at doses only slightly higher than those that are selectively antinociceptive. A similar pattern was also observed in the present study where a 1/~mol dose of ketamine produced only analgesia while most of the animals receiving a 3 ttmol dose experienced a short period of motor blockade. Although the mechanism underlying the motor effect is not clearly understood, most investigators suggest that it is similar to a reversible local anesthetic effect (Altura et al., 1990; Tung and Yaksh, 1981). There is no histological evidence of cellular neurotoxicity (Guinto-Enriquez et al., 1990). Because of the uncertainty regarding the neurochemical basis of this effect, in the present study analgesia was assessed in animals receiving the 3/~mol dose at a point and time when the motor effects had subsided and only sensory blockade was apparent. Thus, the monoaminergic dependence of ketamine analgesia is not likely to be artifactual as a consequence of altered neurochemical changes associated with the appearance of motor alterations. In support of this, the relative effectiveness of adrenoceptor, 5-HT and opioid receptor antagonists was similar at the lower ketamine dose (1 #mol) where the action was exclusively antinociceptive, but less consistent, and of such a short duration as to make extensive studies of that dose level difficult. Ketamine is being evaluated extensively as an epidural and i.t. analgesic in man (E1-Khateeb et al., 1990; Guinto-Enriquez et al., 1990; Schulman et al., 1990). The impetus for these clinical studies included the need to find an alternative to the classical narcotic analgesics that produce deleterious side effects such as delayed respiratory depression, urinary retention, nausea, vomiting and pruritis following administration at the spinal level. Furthermore, an alternative compound with a mechanism of action different from prototypical narcotics like morphine would be useful in patients who have developed tolerance. Therefore, it seems possible that ketamine may provide such an alternative since its spinal action seems to rely heavily on monoaminergically mediated components (present study; Tung and Yaksh, 1981; Pekoe and Smith, 1982) while morphineinduced spinal antinociception primarily involves an opiate (naloxone-reversible) component (Crisp and Smith, 1989).
172 Acknowledgements Supported by NIH Grants GM 30002 and 2 S07 RR 05433-26, West Virginia University Health Sciences Center (D.J.S.) and a Postdoctoral Fellowship from West Virginia University Health Sciences Center (T.C.).
References Altura, B.M., B.T. Altura and A. Corella, 1980, Effects of ketamine on vascular smooth muscle function, Br. J. Pharmacol. 70, 257. Anis, N.A., S.C. Berry, N.R. Burton and D. Lodge, 1983, The dissociative anesthetics, ketamine and phencyclidine, selectively reduce excitation of central mammalian neurons by N-methylaspartate, Br. J. Pharmacol. 79, 565. Azzaro, A.J. and D.J. Smith, 1977, The inhibitory action of ketamine HC1 on 3H-5-hydroxytryptamine accumulation in rat brain synaptosomal-rich fractions: Comparison with [3H] catecholamine and [gamma-3H) aminobutyric acid uptake, Neuropharmacology 16, 349. Basbaum, A.I. and H.L. Fields, 1978, Endogenous pain control mechanisms: review and hypothesis. Ann, Neurol. 4, 451. Bennett, G.J. and D.J. Mayer, 1979, Inhibition of spinal cord interneurons by narcotic microinjection and focal electrical stimulation in the periaqueductal central gray matter, Brain Res. 172, 243. Contreras, P.C., K.C. Rice, A.E. Jackobson and T.L. O'Donohue, 1986, Stereotyped behavior correlates better than ataxia with phencyclidine-receptor interactions, European J. Pharmacol. 121. Crisp, T., J.M. Perrotti and D.J. Smith, 1987, Spinal antinociceptive properties of morphine, ketamine, serotonin and norepinephrine, Abstr. Soc. Neurosci. 13, 1592. Crisp, T. and D.J. Smith, 1989, A local serotonergic component involved in the spinal antinociceptive action of morphine, Neuropharmacology 28, 1047. D'Amour, F.E. and D. Smith, 1941, A method for determining loss of pain sensation, J. Pharmacol. Exp. Ther. 72, 74. E1-Khateeb, O.E., A. Ragab, M. Metwalli and H.A. Hassan, 1990, Assessment of epidural ketamine for relief of pain following vaginal and lower abdominal surgery, in: Status of Ketamine in Anesthesiology, ed. E.F. Domino (NPP Books, Ann Arbor, MI), p. 403. Guinto-Enriquez, G., R.Y. Enriquez and L. Reyes de Castro, 1990, Epidural injection of ketamine hydrochloride: an experimental study in rats, in: Status of Ketamine in Anesthesiology, ed. E.F. Domino (NPP Books, Ann Arbor, MI) p. 381. Hayes, R.L., D.D. Price, G.J. Bennett, G.L. Wilcox and D.J. Mayer, 1978, Differential effects of spinal cord lesions on narcotic and non-narcotic suppression of nociceptive reflexes: Further evidence for the physiologic multiplicity of pain modulation, Brain Res. 155, 91. Hwang, A.S. and G.L. Wilcox, 1987, Analgesic properties of intrathecally administrered heterocyclic antidepressants, Pain 28, 343. LoPachin, R.M., T.A. Rudy and T.L. Yaksh, 1981, An improved method for chronic catheterization of the rat spinal subarachnoid space, Physiol. Behav. 27, 559. Magnusson, T., 1973, Effect of chronic transection on dopamine, noradrenaline and 5-hydroxytryptamine in the rat spinal cord, Naunyn-Schmiedeb. Arch. Pharmacol. 278, 13. Martin, L.L. and D.J. Smith, 1982, Ketamine inhibits serotonin synthesis and metabolism in vivo, Neuropharmacology 21, 119.
Monroe, P.J., K. Michaux and D.J. Smith, 1986, Evaluation of the direct actions of drugs with a serotonergic link in spinal analgesia on the release of [3H]serotonin from spinal cord synaptosomes, Neuropharmacology, 25, 261. Pekoe, G.M. and D.J. Smith, 1982, The involvement of opiate and monoaminergic neuronal systems in the analgesic effect of ketamine, Pain 12, 57. Reddy, S.V.R., J.L. Maderdrut and T.L. Yaksh, 1980, Spinal cord pharmacology of adrenergic agonist-mediated antinociception, J. Pharmacol. Exp. Ther. 213, 525. Rivot, J.P., B. Calvino and J.M. Besson, 1987, Is there a tonic descending inhibition on the response of dorsal horn convergent neurons on C-fibre inputs?, Brain Res. 403, 142. Ryder, S., W.L. Way and A.J. Trevor, 1978, Comparative pharmacology of the optional isomers of ketamine in mice, European. J. Pharmacol. 49, 15. Schulman, S.M., A.T.C. Perg, L.S. Blancato, F. Cutrone and K. Nyunt, 1990, Studies with epidural ketamine and local anesthetic combinations for obstetrical analgesia and anesthesia, in: Status of Ketamine in Anesthesiology, ed. E.F. Domino (NPP Books, Ann Arbor, MI) p. 395. Smith, D.J., A.J. Azzaro, S.B. Zaldivar, S. Palmer and H.S. Lee, 1981, Properties of the optical isomers and metabolites of ketamine on the high affinity transport and catabolism of monoamines, Neuropharmacology 20, 391. Smith, D.J., R.L. Bouchal, C.A. DeSanctis, P.J. Monroe, J.B. Amedro, J.M. Perrotti and T. Crisp, 1987, Properties of the interaction between ketamine and opiate binding sites in vivo and in vitro, Neuropharmacology 26, 1253. Smith, D.J., G.M. Pekoe, L.L. Martin and B. Coalgate, 1980, The interaction of ketamine with the opiate receptor, Life Sci. 26, 789. Smith, D.J., G.M. Pekoe, P.J. Monroe, L.L. Martin, M.E.Y. Cabral and T. Crisp, 1990, Ketamine analgesia in rats may be mediated by an interaction with opiate receptors, in: Status of Ketamine in Anesthesiology, ed. E.F. Domino (NPP Books, Ann Arbor, MI) p. 199. Smith, D.J., J.M. Perrotti, A.L. Mansell and P.J. Monroe, 1985, Ketamine analgesia is not related to an opiate action in the periaqueductal gray region of the rat brain, Pain 21, 253. Smith, D.J., D.P. Westfall and J.D. Adams, 1982, Assessment of the potential agonistic and antagonistic properties of ketamine at opiate receptors in the guinea-pig ileum, Neuropharmacology 21, 605. Tallarida, R.J. and R.B. Murray (eds.), 1981, Method of Litchfield and Wilcoxon: Confidence limits of EDs0, in: Manual of Pharmacological Calculations with Computer Programs (Springer-Verlag, New York, Heidelberg, Berlin) p. 153. Thompson, A.M., D.C. West and D. Lodge, 1985, An N-methylaspartate receptor-mediated synapse in rat cerebral cortex: A site of action of ketamine?, Nature (London) 313, 479. Tung, A.S. and T.L. Yaksh, 1981, Analgesic effect of intrathecal ketamine in the rat, Reg. Anesth. 6, 91. White, P.F., J. Ham, W.L. Way and A.J. Trevor, 1980, Pharmacology of ketamine isomers in surgical patients, Anesthesiology 52, 231. Yaksh, T.L. and P.R. Wilson, 1979, Spinal serotonin terminal system mediates antinociception, J. Pharmacol. Exp. Ther. 208, 446. Zukin, S.R. and R.S. Zukin, 1981, Identification and characterization of 3H-phencylidine binding to specific brain receptor sites, in: PCP (Phencyclidine): Historical and Current Perspectives, ed. E.F. Domino, (NPP books, Ann Arbor, MI) p. 105.
European Journal of Pharmacology, 194 (1991) 167-172 © 1991 Elsevier Science Publishers B.V. 0014-2999/91/$03.50 ADONIS 001429999100201G EJP 51743
The local monoaminergic dependency of spinal ketamine Terriann Crisp, Jeannette M. Perrotti, Deborah L. Smith, Janet L. Stafinsky and David J. Smith Department o/Anesthesiology, West Virginia University Health Sciences Center, Morgantown, WV 26506, U.S.A. and Department o/Pharmacology, Northeastern Ohio Universities College of Medicine, Rootstown, OH 44272, U.S.A. Received 28 June 1990, revised MS received 31 October 1990, accepted 4 December 1990
The effects of s.c. doses of naloxone, methysergide and phentolamine on ketamine-induced spinal analgesia were assessed to determine the involvement of opiate, serotonergic and noradrenergic components mediating ketamine's antinociceptive action. Ketamine administered intrathecally (i.t.) produced a significant elevation in tail-flick latency in rats. The spinal antinociceptive effects of ketamine were dose dependently reversed by methysergide (IDs0 = 0.008 mg/kg s.c.), phentolamine (IDs0 = 0.88 mg/kg s.c.) and naloxone (IDs0 = 3.0 mg/kg s.c.). Unlike morphine, which remains analgesic and dependent on opiate interactions following bilateral lesions of the dorsolateral funiculus (DLF), ketamine analgesia was absent following DLF lesions. Thus, ketamine appears to produce an antinociceptive response which is dependent upon the neuronal activity of the descending pain-inhibitory pathways. The monoaminergic components comprising the descending pathways appear to be more prominent in the action of ketamine than they are in the spinal action of morphine. Furthermore, the spinal opioid receptors involved in ketamine's effect may be different from the/~ subtype preferred by morphine. Ketamine; Analgesia; Opioid receptors; 5-HT (5-hydroxytryptamine,serotonin); Norepinephrine; Tail-flick test; (Intrathecal)
1. Introduction
Racemic ketamine HC1 is a dissociative anesthetic agent which exhibits analgesic properties in rodents (Ryder et al., 1978) and humans (White et al., 1980). Although the mechanisms underlying the analgesic action of the drug are not clearly understood, it has been shown that ketamine stereoselectively binds with a weak affinity to opiate receptors (Smith et al., 1980; 1987). Additionally, the antinociceptive action of systemically administered ketamine is naloxone-reversible (Pekoe and Smith, 1982; Ryder et al., 1978; Smith et al., 1985). The fact that ketamine-induced analgesia is reversed by naloxone does not imply direct opiate receptor activation as the primary mechanism by which the opiate produces its analgesic action. For instance, ketamine has repeatedly been shown to interact with monoaminergic (serotonergic, noradrenergic and dopaminergic) neuronal systems (Tung and Yaksh, 1981; Martin and Smith, 1982). Moreover, the analgesic properties of ketamine may be mediated via the ability of the agent to block high affinity mohoaminergic uptake sites (Pekoe and Smith, 1982; Smith et al., 1981; Azzaro and Smith, 1977). Correspondence to: T. Crisp, Department of Pharmacology, Northeastern Ohio Universities, College of Medicine, Rootstown, OH 44272, U.S.A.
Recent evidence suggests the involvement of both monoaminergic (Tung and Yaksh, 1981; Pekoe and Smith, 1982) and opiate (Smith et al., 1985) components mediating ketamine-induced analgesia. However, the analgesic efficacy of ketamine may change as a consequence of the route of administration and/or the site of injection of the drug (Smith et al., 1985). For instance, ketamine injected intracerebroventricularly (i.c.v.) or into the periaqueductal gray region of the rat mesencephalon, an area rich in opiate receptor sites (Bennet and Mayer, 1979), fails to elevate tail-flick latency (TFL) in rats (Smith et al., 1985; 1990). On the other hand, Tung and Yaksh (1981) reported that ketamine injected directly into the subarachnoid space of the rat spinal cord produced a weak analgesic action that was reversed by the serotonin (5-HT) receptor antagonist methysergide. That 5-HT was involved in mediating the local spinal action of ketamine was further substantiated by reports demonstrating that intraperitoneal (i.p.) injections of ketamine produced methysergide-reversible elevations in TFL 1-2 days following spinal transection in rats (Pekoe and Smith, 1982). However, neither naloxone nor the a-adrenoceptor antagonist phentolamine effectively reversed the antinociceptive effects of i.p. ketamine in spinally transected rats. Thus, a prominent 5-HT component may mediate the spinal analgesic effects of ketamine (Pekoe and Smith, 1982; Crisp et al., 1987).
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The purpose of the present study was to more thoroughly identify the mediatory role of opioid, serotonergic and noradrenergic neuronal systems in spinal ketamine-induced analgesia. This was done by testing the ability of various subcutaneous (s.c.) doses of naloxone, methysergide and phentolamine to alter the antinociceptive effects of ketamine following intrathecal (i.t.) injections of the anesthetic.
2. Materials and methods
2.1. Animal model and surgical preparations Male Sprague-Dawley rats (350 _+ 50 g) were chronically implanted with i.t. polyethylene (PE-10) catheters (LoPachin et al., 1981) to allow for the direct spinal administration of ketamine. Under ketamine anesthesia, each rat was fitted with an indwelling spinal catheter which was inserted through a slit in the atlanto-occipital membrane and passed distally approximately 8.5 cm to the rostral edge of the lumbar enlargement. Rats were allowed at least 7 days to recuperate from surgery, and only those animals which displayed unimpaired motor function after i.t. catheterization were used in the experiments. An interval of at least 2 weeks was maintained between repeated injections, and rats were not injected more than twice. After the second injection, rats were killed and cannula placement was checked using methylene blue dye. Data obtained from animals with malpositioned catheters (e.g. tip location in the epidural space) were excluded from the study. Bilateral dorsolateral funiculus (DLF) lesions were made in rats previously cannulated with i.t. catheters using a lesioning technique described by Hayes et al. (1978). In short, a laminectomy was done to expose the cord at the level of T1-T4. The dura was removed and a pair of dissecting forceps were inserted into the cord and closed for 15-20 s bilaterally to lesion the DLF. After the lesions were made, the exposed cord was covered with Gelfoam, soaked with 0.9% saline, and the wound was then sutured. Analgesic tests were conducted in DLF-lesioned rats 24-48 h after the lesions were made, at a time when monoaminergic nerve terminals below the site of the lesion were still intact (Magnusson, 1973). The analgesic efficacy of ketamine administered i.t. was evaluated in rats with intact spinal cords, and in rats with bilateral D L F lesions.
2.2. Analgesiometric assay The spinal antinociceptive efficacy of different doses of ketamine was measured using a modified version of the tail-flick test (D'Amour and Smith, 1941). A Model 33 Analgesia Meter was used, and the rat's tail (blackened with India Ink) was placed in a depression over
the photocell in the tail-flick meter. The time (in s) required for the rat to remove its tail from the light source was automatically determined and was expressed as TFL. Routinely, four baseline values were obtained, and the mean of the last three was used as the predrug TFL. Preliminary experiments from this laboratory and others (Tung and Yaksh, 1981) have demonstrated that the analgesic action of spinal ketamine is only minimally detectable when using 'standard' T F L values (i.e. baseline T F L between 2-4 s, cut-off T F L values of 10 s). By decreasing the intensity of the light source on the tail-flick meter, we were able to increase the sensitivity of the tail-flick analgesiometric assay (Tung and Yaksh, 1981). Therefore, for these experiments, the intensity of the bulb on the tail-flick meter was adjusted to obtain predrug baseline values of 5-8 s, and a 15 s maximal exposure to the heat source was employed as the cut-off value to preclude tissue damage to the tail.
2.3. The effects of ketamine administered i.t. in rats Different doses of ketamine (0.3, 1 and 3 ~tmol/10 /~1) were administered i.t. in rats using a Harvard Model 975 Infusion Pump that was pre-set to deliver 10/zl of drug or saline per min. Each drug injection was followed by a 5/~1 saline flush to clear the spinal catheter of the drug. Drug-induced changes in T F L were obtained 5, 10, 15, 30 and 60 min post-ketamine injection, and dose-response curves were generated using individual groups of rats for each dose of ketamine. Ketamine-induced alterations in T F L were evaluated in rats with intact spinal cords and in rats with bilateral D L F lesions.
2.4. The relative participation of opiate, serotonergic and noradrenergic components mediating the spinal antinociceptive action of ketamine To determine if ketamine administered i.t. interacted with opioid a n d / o r monoaminergic neuronal systems to produce spinal analgesia, varying doses of naloxone (0.3-10 mg/kg), methysergide (0.001-0.1 mg/kg) or phentolamine (0.01-10 m g / k g ) were administered s.c. and tested for an ability to alter ketamine-induced spinal analgesia. In these experiments, the 3/~mol dose of ketamine was tested against the various antagonists because it produced consistent elevations in T F L without taking animals to cut-0ff. The range of inhibitory doses for the antagonists used in this study was obtained from previous experiments demonstrating the inhibitory efficacy of s.c. naloxone, methysergide and phentolamine on the spinal analgesic effects of morphine, 5-HT and norepinephrine (NE), respectively (Crisp and Smith, 1989). Naloxone was injected s.c. immediately prior to i.t. ketamine injections, whereas
169
methysergide and phentolamine were administered 15 min prior to spinal ketamine injections. In this manner, the maximal inhibitory effect of the antagonists would overlap with the time of ketamine's peak antinociceptive effect (established in the initial dose-response and duration experiments). Doses of naloxone, methysergide and phentolamine that were effective in reversing ketamine-induced spinal analgesia were injected alone in other experiments and tested for an ability to significantly alter T F L values by themselves. These values were compared to s.c. saline control T F L values.
2.5 Data analysis In order to analyze the ability of naloxone, methysergide and phentolamine to alter the spinal antinociceptive effects of ketamine, comparative assessments were made using apparent IDs0 values (defined as the dose of antagonist producing 50% inhibition of ketamine-induced analgesia; Tallarida and Murray, 1981). The probit analysis required that graded responses, obtained with a fixed dose of ketamine in the presence of various doses of each antagonist, be transformed to a quantal form. For this purpose, analgesia was defined as a post-drug T F L value that was greater than, or equal to, two S.D.s above the mean pre-drug latency of the study group (Crisp and Smith, 1989). All data were derived from experiments having an n of at least six animals per treatment group. Tail-flick latency values in figs. 1-5 are expressed as means + S.E.M. Statistical analyses of dose-response and duration data were conducted using a repeated measures analysis of variance (ANOVA) and a Dunnett's test as applicable to calculate significant differences between the means (P < 0.05). To statistically analyze the effects of different doses of naloxone, m e t h y s e r g i d e and p h e n t o l a m i n e on ketamine-induced analgesia, an ANOVA with no replications was used.
15-
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Fig. 1. Dose-responseand duration of i.t. ketamine((o) saline; (-) 0.3 #mol; (It) 1 #mol; (zx)3/xmol). Values represent the means+ S.E.M. of at least six animals/dose. * P < 0.05 versuspre-drug TFL values. with bilateral D L F lesions, the 3 /~mol i.t. dose of ketamine was no longer analgesic, and did not significantly elevate T F L values above pre-drug control values (fig. 2). Overt motor changes resulting from ketamine injections were assessed by placing rats on a 60 ° vertical wire mesh (Tung and Yaksh, 1981). In none of the rats tested did 0.3 or 1 /~mol dose of ketamine produce an inability to negotiate the mesh. The 3 /~mol dose produced a reversible hind limb paralysis within 3-4 min post-injection, but this effect lasted no longer than 8-10 min.
3.2. The effects of naloxone, methysergide and phentolamine administered s.c. on the spinal analgesic effects of ketamine In experiments designed to assess the involvement of opiate a n d / o r monoaminergic components mediating ketamine's spinal action, the 3 /~mol dose of ketamine was used. This dose produced a consistent analgesic 15'
3.
Results
,~
3.1. The effects of ketamine administered i.t. in rats In rats with intact spinal cords, ketamine injected i.t. produced a dose-dependent elevation in T F L that was significantly greater than pre-drug control values for 15 min post-injection (fig. 1). The onset of drug action occurred within 10 min, and the peak analgesic effect for the 1 and 3/~mol doses of ketamine was observed at 15 .m in post-injection. The T F L response to the 0.3 /~mol dose of the anesthetic was indistinguishable from pre-drug control values for the 60 min duration of the dose-response experiments. Ketamine-induced elevations in T F L were not significantly greater than controls after 15 min post-injection (fig. 1). Interestingly, in rats
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Fig. 2. Lack of response of DLF-lesioned rats to 3 /~mol ketamine. Values represent the means+S.E.M, of at least six animals. Fifteen minutes post-drug TFL values are compared to pre-drug controls.
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Fig. 3. High doses of naloxone administered s.c. reverse the analgesic action of i.t. ketamine. Values are the means + S.E.M. of at least six animals/treatment group. Time =15 min post-ketamine and 15 min post-naloxone. * P < 0.05 versus saline control, * * P < 0.05 versus 3 t~mol ketamine alone. response o n the tail-flick test w i t h o u t elevating T F L values a b o v e the 15 s cut-off point. T h e o p i a t e r e c e p t o r a n t a g o n i s t n a l o x o n e p r o d u c e d a d o s e - d e p e n d e n t reversal of k e t a m i n e ' s spinal a c t i o n (fig. 3), a n d h a d an a p p a r e n t IDs0 of 3.0 (2.11-4.28) m g / k g . V a r y i n g doses of m e t h y s e r g i d e were also tested against k e t a m i n e in a t t e m p t s to assess the relative i n v o l v e m e n t o f 5 - H T n e u r o n a l processes in k e t a m i n e - i n d u c e d spinal analgesia. M e t h y s e r g i d e dose d e p e n d e n t l y reversed k e t a m i n e - i n d u c e d elevations in T F L (fig. 4), having an a p p a r e n t IDs0 value of 0.008 (0.003-0.025) m g / k g . P h e n t o l a m i n e also decreased the spinal analgesic effects of k e t a m i n e in a d o s e - d e p e n d e n t m a n n e r (fig. 5). T h e a - a d r e n o c e p tor a n t a g o n i s t h a d an a p p a r e n t IDs0 of 0.88 (0.23-3.38) m g / k g s.c. D o s e s of naloxone, m e t h y s e r g i d e a n d p h e n 15'i --,
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Fig. 5. Phentolamine administered s.c. reversed the effects of ketamine in a dose-dependent manner. Values represent the means_+S.E.M. of at least six animals/treatment group. Time =15 min post-ketamine and 30 min post-phentolamine. * P < 0.05 versus i.t. saline controls, • * P < 0.05 versus 3 #mol ketamine alone.
t o l a m i n e t h a t b l o c k e d the spinal a n t i n o c i c e p t i v e effects of the 3 /~mol d o s e of k e t a m i n e also reversed the analgesic effects o f the 1 ~ m o l d o s e of k e t a m i n e a d m i n istered i.t. A d d i t i o n a l l y , w h e n s.c. doses of n a l o x o n e (10 m g / k g ) , m e t h y s e r g i d e (1 m g / k g ) a n d p h e n t o l a m i n e (10 m g / k g ) were a d m i n i s t e r e d alone in a n o t h e r g r o u p of animals, no significant changes in T F L were o b s e r v e d for 30 m i n p o s t - i n j e c t i o n when c o m p a r e d to i.t. saline values.
4. D i s c u s s i o n
T h e p r e s e n t d a t a suggest that the analgesic effects of k e t a m i n e a d m i n i s t e r e d i.t. are spinally m e d i a t e d . W h e n k e t a m i n e was injected i.c.v, in rats, even high doses failed to p r o d u c e a n a l g e s i a ( S m i t h et al., 1990), suggestm g that the r o s t r a l diffusion of i.t. k e t a m i n e to s u p r a s p i n a l sites was n o t requisite for an analgesic r e s p o n s e to b e evoked. F u r t h e r m o r e , the r a p i d onset of a n a l g e s i a (5-10 min) also suggests that local spinal processes are involved. T h e c u r r e n t s t u d y also c o n f i r m s the i n v o l v e m e n t of o p i o i d a n d m o n o a m i n e r g i c processes in the spinal action of k e t a m i n e . H o w e v e r , the m e c h a n i s m ( s ) u n d e r l y ing the i n t e r a c t i o n o f k e t a m i n e with these systems rem a i n unclear. T h e fact that b i l a t e r a l lesions of the D L F d i m i n i s h e d the a n t i n o c i c e p t i v e a c t i o n of i.t. k e t a m i n e does imply, however, that n e u r o n a l activity in descending p a i n i n h i b i t o r y nerves ( R i v o t et al., 1987; Crisp a n d Smith, 1989) is necessary for k e t a m i n e - i n d u c e d spinal a n a l g e s i a to b e expressed. I n this regard, it has been e s t a b l i s h e d that k e t a m i n e has the p o t e n t i a l to e n h a n c e m o n o a m i n e r g i c (5-HT, N E
171 and DA) neuronal transmission by inhibiting the high affinity monoaminergic uptake carriers (Azzaro and Smith, 1977; Smith et al., 1981). Perhaps the monoaminergic dependency of ketamine-induced analgesia in intact animals is related to a blockade of monoamine uptake and the enhancement of the spinal synaptic activity of 5-HT (Yaksh and Wilson, 1979) and NE (Reddy et al., 1980). An action such as this would require neuronal activity in the descending monoaminergic pathways to initiate the release of neurotransmitters, and would be interrupted by DLF lesions as observed in this study. This mechanism is also consistent with the suggestion by Hwang and Wilcox (1987) that the analgesic activity of various heterocyclic antidepressants (e.g. desipramine, protriptyline, fluoxetine and citalopram) is related to their ability to block monoaminergic uptake. Furthermore, preliminary experiments in this laboratory have demonstrated that the 5-HT uptake blocker fluoxetine produces antinociceptive effects similar to ketamine when administered i.t. using the extended baseline (Crisp, studies in progress). On the other hand, since ketamine also has the potential to interact at opiate receptors as an agonist (Smith et al., 1982), the inability to observe an antinociceptive action in the absence of tonic neuronal activity is more difficult to explain. Morphine-like opiate agonists produce analgesia in DLF-lesioned animals, presumably as a result of their interaction with intrinsic /x opiate receptors in the dorsal horn of the spinal cord (Crisp and Smith, 1989). In contrast however, it has been suggested that the # opiate receptor subtype preferred by morphine is not the same opiate receptor responsible for ketamine's opiate agonistic action (Smith et al., 1982; 1987; 1990). Data from the present study seem to confirm this suggestion since higher doses of naloxone were required to block the spinal effects of ketamine than were needed to reverse the antinociceprive effect of i.t. morphine (cf. Crisp and Smith, 1989). Thus, an opiate receptor subtype less sensitive to naloxone (e.g. K or putative phencyclidine (PCP)/o sites) may be involved (Zukin and Zukin, 1981; Contreras et al., 1986; Smith et al., 1987; Anis et al., 1983; Thompson et al., 1985). The inactivity of ketamine in DLF-lesioned rats may suggest that ketamine-sensitive opiate receptors lie on interneurons that modulate activity in tonically active descending antinociceptive neuronal pathways, as has been suggested by Monroe et al. (1986). On the other hand, the reliance of ketamine on opiate mechanisms may be secondary to actions on other systems. With regard to the latter case, Basbaum and Fields (1978) proposed the existence of enkephalinergic interneurons between descending serotonergic nerve terminals and primary afferent nerve terminals in the spinal cord. Such neuronal circuitry could provide a basis for a naloxone-reversible action of ketamine secondary to its
ability to block 5-HT uptake. Alternatively, the inactivity of ketamine in DLF-lesioned rats may suggest that it lacks meaningful interactions with opiate systems in the spinal cord as has been suggested by Tung and Yaksh (1981). Accordingly, the requirement for high doses of naloxone to reverse ketamine-induced analgesia in intact animals may be related to non-opiate neuronal effects of the antagonist. The spinal or epidural administration of ketamlne in man (Guinto-Enriquez et al., 1990; Schulman et al., 1990) and animals (Tung and Yaksh, 1981) is attended by sensory blockade as well as a shorter-rived reversible motor blockade at doses only slightly higher than those that are selectively antinociceptive. A similar pattern was also observed in the present study where a 1/~mol dose of ketamine produced only analgesia while most of the animals receiving a 3 ttmol dose experienced a short period of motor blockade. Although the mechanism underlying the motor effect is not clearly understood, most investigators suggest that it is similar to a reversible local anesthetic effect (Altura et al., 1990; Tung and Yaksh, 1981). There is no histological evidence of cellular neurotoxicity (Guinto-Enriquez et al., 1990). Because of the uncertainty regarding the neurochemical basis of this effect, in the present study analgesia was assessed in animals receiving the 3/~mol dose at a point and time when the motor effects had subsided and only sensory blockade was apparent. Thus, the monoaminergic dependence of ketamine analgesia is not likely to be artifactual as a consequence of altered neurochemical changes associated with the appearance of motor alterations. In support of this, the relative effectiveness of adrenoceptor, 5-HT and opioid receptor antagonists was similar at the lower ketamine dose (1 #mol) where the action was exclusively antinociceptive, but less consistent, and of such a short duration as to make extensive studies of that dose level difficult. Ketamine is being evaluated extensively as an epidural and i.t. analgesic in man (E1-Khateeb et al., 1990; Guinto-Enriquez et al., 1990; Schulman et al., 1990). The impetus for these clinical studies included the need to find an alternative to the classical narcotic analgesics that produce deleterious side effects such as delayed respiratory depression, urinary retention, nausea, vomiting and pruritis following administration at the spinal level. Furthermore, an alternative compound with a mechanism of action different from prototypical narcotics like morphine would be useful in patients who have developed tolerance. Therefore, it seems possible that ketamine may provide such an alternative since its spinal action seems to rely heavily on monoaminergically mediated components (present study; Tung and Yaksh, 1981; Pekoe and Smith, 1982) while morphineinduced spinal antinociception primarily involves an opiate (naloxone-reversible) component (Crisp and Smith, 1989).
172 Acknowledgements Supported by NIH Grants GM 30002 and 2 S07 RR 05433-26, West Virginia University Health Sciences Center (D.J.S.) and a Postdoctoral Fellowship from West Virginia University Health Sciences Center (T.C.).
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