Anesthetic effects on fictive locomotion in the rat isolated spinal cord.

General anesthetic mechanisms are poorly understood. Anesthetic immobilizing effects occur in the spinal ventral horn. However, a detailed analysis of anesthetic effects on ventral motor networks is lacking. We delivered isoflurane, desflurane, or propofol during NMDA/5-HT-induced, or noxious tail stimulus-evoked, fictive locomotion in neonatal rat isolated spinal cords. Anesthetics changed the frequency, amplitude, and regularity of fictive locomotion with little effect on phase-lag. Isoflurane abolished pharmacologically-induced versus noxious stimulus-induced motor output at similar concentrations. Propofol abolished pharmacologically-induced fictive locomotion through a γ-aminobutyric acid type A-receptor mechanism. Anesthetic effects on pharmacologically-elicted fictive locomotion appear clinically-relevant, and support a ventral horn immobilizing effect on locomotor rhythm generation.

Validation and insights of anesthetic action in an early vertebrate network: the isolated lamprey spinal cord.

BACKGROUND: The lamprey spinal cord is a well-characterized vertebrate network that could facilitate our understanding of anesthetic action. We tested several hypotheses concerning the lamprey's clinical application to anesthesia, and the sites/mechanisms of anesthetic action. METHODS: In isolated lamprey spinal cords, minimum immobilizing concentrations (MICs) were determined for halothane, isoflurane, sevoflurane, desflurane, propofol, or the nonimmobilizer F6 (1,2-dichlorohexafluorocyclobutane), applied during D-glutamate-induced fictive swimming or noxious tail stimulation. Isoflurane and propofol effects on fictive swimming were tested in the presence and absence of strychnine and/or picrotoxin. RESULTS: Volatile anesthetic MICs were clinically comparable. Isoflurane MIC for fictive swimming and noxious stimulus-evoked movement were the same. F6 did not produce immobility, but decreased the amplitude and phase lag of fictive swimming. Isoflurane decreased fictive swimming cycle frequency, amplitude, autocorrelation, rostrocaudal phase lag, and coherence. Strychnine and picrotoxin elicited only disorganized motor activity under isoflurane and caused small increases in MIC. The effects of propofol differed from isoflurane for all locomotor rhythm variables except amplitude. The propofol MIC was much larger in lampreys compared with mammals. However, picrotoxin reversed propofol-induced immobility by reinitiating coordinated locomotor activity and increasing MIC>8-fold. CONCLUSIONS: The lamprey spinal cord is a relevant and tractable vertebrate network model for anesthetic action. Isoflurane disrupts interneuronal locomotor networks. γ-Aminobutyric acid A and glycine receptors have marginal roles in isoflurane-induced immobility in lampreys. Propofol's selective γ-aminobutyric acid A receptor-mediated immobilizing mechanism is conserved in lampreys. The differential immobilizing mechanisms of isoflurane versus propofol reflect those in mammals, and further suggest different network modes of immobilizing action.

Analgesia mediated by soluble epoxide hydrolase inhibitors is dependent on cAMP.

Pain is a major health concern even though numerous analgesic agents are available. Side effects and lack of wide-spectrum efficacy of current drugs justify efforts to better understand pain mechanisms. Stabilization of natural epoxy-fatty acids (EFAs) through inhibition of the soluble epoxide hydrolase (sEH) reduces pain. However, in the absence of an underlying painful state, inhibition of sEH is ineffective. Surprisingly, a pain-mediating second messenger, cAMP, interacts with natural EFAs and regulates the analgesic activity of sEH inhibitors. Concurrent inhibition of sEH and phosphodiesterase (PDE) dramatically reduced acute pain in rodents. Our findings demonstrate a mechanism of action of cAMP and EFAs in the pathophysiology of pain. Furthermore, we demonstrate that inhibition of various PDE isozymes, including PDE4, lead to significant increases in EFA levels through a mechanism independent of sEH, suggesting that the efficacy of commercial PDE inhibitors could result in part from increasing EFAs. The cross-talk between the two major pathways-one mediated by cAMP and the other by EFAs-paves the way to new approaches to understand and control pain.

Naturally occurring monoepoxides of eicosapentaenoic acid and docosahexaenoic acid are bioactive antihyperalgesic lipids.

Beneficial physiological effects of long-chain n-3 polyunsaturated fatty acids are widely accepted but the mechanism(s) by which these fatty acids act remains unclear. Herein, we report the presence, distribution, and regulation of the levels of n-3 epoxy-fatty acids by soluble epoxide hydrolase (sEH) and a direct antinociceptive role of n-3 epoxy-fatty acids, specifically those originating from docosahexaenoic acid (DHA). The monoepoxides of the C18:1 to C22:6 fatty acids in both the n-6 and n-3 series were prepared and the individual regioisomers purified. The kinetic constants of the hydrolysis of the pure regioisomers by sEH were measured. Surprisingly, the best substrates are the mid-chain DHA epoxides. We also demonstrate that the DHA epoxides are present in considerable amounts in the rat central nervous system. Furthermore, using an animal model of pain associated with inflammation, we show that DHA epoxides, but neither the parent fatty acid nor the corresponding diols, selectively modulate nociceptive pathophysiology. Our findings support an important function of epoxy-fatty acids in the n-3 series in modulating nociceptive signaling. Consequently, the DHA and eicosapentaenoic acid epoxides may be responsible for some of the beneficial effects associated with dietary n-3 fatty acid intake.

Dorsal horn neurons expressing NK-1 receptors mediate scratching in rats.

Itch is thought to be signaled by pruritogen-responsive neurons in the superficial spinal dorsal horn. Many neurons here express the substance P NK-1 receptor. We investigated whether neurotoxic destruction of spinal NK-1-expressing neurons affected itch-related scratching behavior. Rats received intracisternal substance P conjugated to saporin (SP-SAP), or saporin (SAP) only (controls), and were subsequently tested for scratching behavior elicited by intradermal 5-hydroxytryptamine. SAP controls exhibited dose-related hindlimb scratching, which was significantly attenuated in SP-SAP-treated rats. There was a virtual absence of NK-1 immunoreactive neurons in superficial laminae of the upper cervical and medullary dorsal horn in SP-SAP-treated rats. These results indicate that superficial dorsal horn neurons expressing NK-1 receptors play a key role in spinal itch transmission.

Brainstem regions affecting minimum alveolar concentration and movement pattern during isoflurane anesthesia.

BACKGROUND: Spinal transection or selective delivery of volatile anesthetics to the spinal cord reduces minimum alveolar concentration (MAC), whereas precollicular decerebration does not. The authors sought to determine which brainstem regions influence anesthetic requirements and movement responses with isoflurane. METHODS: Movement (biceps femoris electromyogram) and MAC were measured in adult rats before and after decerebration at the precollicular, mid-collicular, pontine or medullary level, or decerebellation. Additional experiments assessed the effects of lidocaine inactivation of the mesencephalic locomotor region on MAC and the effects of isoflurane on nociceptive neuronal responses in this region. RESULTS: Transections placed at the level of the mid-colliculus, rostral pons, and pontomedullary junction significantly reduced MAC by approximately 10, 40, and 45%, respectively. MAC was decreased 9% after mid-medullary transections that were placed caudal to the nucleus raphe magnus but rostral to the dorsal reticular nucleus; however, only weak, single movements occurred. Caudal medullary transections at the obex decreased MAC by 60%. Bilateral inactivation of the mesencephalic locomotor region with lidocaine caused a reversible, 32% decrease in MAC and reduced the number and amplitude of movements at sub-MAC isoflurane concentrations. Neuronal responses of mesencephalic locomotor region neurons to supramaximal noxious tail clamp were reduced by 87% by 1.2 MAC isoflurane. CONCLUSIONS: The authors conclude that the mesencephalic locomotor region influences anesthetic requirements and promotes repetitive movement with sub-MAC isoflurane by facilitating ventral spinal locomotor circuits, where anesthetics seem to exert their key immobilizing effects. However, net brainstem influences on MAC seem to result from interaction among descending nociceptive and locomotor modulatory pathways.

Nitrous oxide-induced analgesia does not influence nitrous oxide's immobilizing requirements.

BACKGROUND: Nitrous oxide (N(2)O) acts on supraspinal noradrenergic neurons to produce analgesia, but it is unclear if analgesia contributes to N(2)O's immobilizing effects. We tested the hypothesis that N(2)O minimum alveolar anesthetic concentration (MAC) is unchanged after selective ablation of supraspinal noradrenergic neurons, or in naïve animals at N(2)O exposure timepoints when analgesia is absent. METHODS: We determined tailflick latency (TFL) and hindpaw withdrawal latency (HPL) under 70% N(2)O, N(2)O MAC, and isoflurane MAC before and after intracerebroventricular injections of anti-dopamine-beta hydroxylase conjugated to saporin (SAP-DBH; n = 7), or a control antibody conjugated to saporin (n = 5). In a separate group of naive rats (n = 8), N(2)O MAC was determined at 25-45 min after initiation of N(2)O exposure (during peak analgesia) and again at 120-140 min (after TFL and HPL returned to baseline). RESULTS: After 30 min of N(2)O exposure, TFL and HPL increased significantly but declined back to baseline within 120 min. N(2)O did not produce analgesia in rats that received SAP-DBH. However, N(2)O and isoflurane MAC were not significantly different between SAP-DBH and control-injected animals (Mean +/- sd for N(2)O: 1.7 +/- 0.1 atm vs 1.7 +/- 0.2 atm; isofurane: 1.6 +/- 0.2% vs 1.7 +/- 0.2%). In naïve animals, N(2)O MAC was not different at the 30 min period compared with the 120 min period (1.8 +/- 0.1 atm vs 1.8 +/- 0.2 atm). CONCLUSIONS: Destroying brainstem noradrenergic neurons or prolonged exposure to N(2)O removes its analgesic effects, but does not change MAC. The immobilizing mechanism of N(2)O is independent from its analgesic effects.

Rat dorsal horn nociceptive-specific neurons are more sensitive than wide dynamic range neurons to depression by immobilizing doses of volatile anesthetics: an effect partially reversed by the opioid receptor antagonist naloxone.

BACKGROUND: The mechanism and site of action within the spinal cord by which volatile anesthetics produce immobility are not well understood. Little work has been done directly comparing anesthetic effects on neurons with specific functional characteristics that mediate transfer of nociceptive information within the spinal cord. METHODS: Adult male rats were anesthetized and prepared for extracellular single-unit recordings from the lumbar dorsal horn. Nociceptive-specific (NS) and wide dynamic range (WDR) neurons were identified and noxious heat-evoked neuronal spike rates evaluated at 0.8 and 1.2 anesthetic minimum alveolar anesthetic concentration (MAC) halothane or isoflurane. In another group, noxious heat-evoked responses from NS neurons were evaluated at 0.8, 1.2 MAC halothane, and 1.2 MAC halothane plus IV naloxone (0.1 mg/kg). RESULTS: Increasing halothane from 0.8 to 1.2 MAC reduced the heat-evoked neuronal responses of NS neurons (n = 9) from 827 +/- 122 (mean +/- se) to 343 +/- 48 spikes/min (P < 0.05) but not WDR neurons (n = 9), 617 +/- 79 to 547 +/- 78 spikes/min. Increasing isoflurane from 0.8 to 1.2 MAC reduced the heat-evoked neuronal response of NS neurons (n = 9) from 890 +/- 339 to 188 +/- 97 spikes/min (P < 0.05) but did not alter the response of WDR neurons (n = 9) in which evoked spike rate went from 576 +/- 132 to 601 +/- 119 spikes/min. In a separate group, the response of NS neurons went from 282 +/- 60 to 74 +/- 32 spikes/min (P < 0.05) when halothane was increased from 0.8 to 1.2 MAC. IV administration of naloxone increased the heat-evoked response to 155 +/- 46 spikes/min (P < 0.05). CONCLUSIONS: NS but not WDR neurons in the lumbar dorsal horn are depressed by peri-MAC increases of halothane and isoflurane. This depression, at least with halothane, can be partially reversed by the opioid antagonist naloxone. Given that opioid receptors are not likely involved in the mechanisms by which volatile anesthetics produce immobility, this suggests that, although the neuronal depression is of substantial magnitude and occurs concurrent to the production of immobility, it may not play a major role in the production of this anesthetic end point.

Propofol produces immobility via action in the ventral horn of the spinal cord by a GABAergic mechanism.

BACKGROUND: We investigated the actions of propofol and isoflurane on nociceptive responses of neurons in the spinal cord. METHODS: We determined nociceptive responses of lumbar neurons in the dorsal horn (<1200 microm) and ventral horn (>1200 microm) of decerebrate rats before and during propofol (1 effective dose, ED(50)) or isoflurane (1 minimum alveolar concentration) anesthesia. During recording of ventral horn neurons, we administered picrotoxin by infusion to determine whether isoflurane and propofol differed in their effects at the gamma aminobutyric acid (GABA) Type A receptors. We also determined whether decerebration altered propofol requirements to produce immobility. RESULTS: Decerebration did not affect propofol requirements. The ED(50) for propofol was 497 +/- 58 microg x kg(-1) x min(-1) in intact rats and 420 +/- 65 microg x kg(-1) x min(-1) in decerebrated rats (P > 0.05), with corresponding propofol blood concentrations of 8.1 +/- 1.1 microg/mL and 7.3 +/- 1.1 microg/mL, respectively (P > 0.05). Propofol did not significantly depress dorsal horn neurons, but isoflurane depressed the responses to 56% of control (P < 0.05). Propofol depressed ventral horn neurons to 47% of control, whereas isoflurane depressed ventral horn neurons to 20% of control. Picrotoxin significantly reversed the depressant effect of propofol on ventral horn neuronal responses (79% of control, not significantly different from control). Pic- rotoxin, however, had no effect on isoflurane's depression of ventral horn neuronal responses (26% of control). CONCLUSIONS: Propofol acts in the spinal cord to produce immobility. This depressive effect occurs in the ventral horn and is mediated mainly by GABA(A) receptors. Isoflurane also depresses neurons in the ventral horn; however, isoflurane actions at the GABA(A) receptor are either weak or overridden by other effects in the ventral horn.

Soluble epoxide hydrolase and epoxyeicosatrienoic acids modulate two distinct analgesic pathways.

During inflammation, a large amount of arachidonic acid (AA) is released into the cellular milieu and cyclooxygenase enzymes convert this AA to prostaglandins that in turn sensitize pain pathways. However, AA is also converted to natural epoxyeicosatrienoic acids (EETs) by cytochrome P450 enzymes. EET levels are typically regulated by soluble epoxide hydrolase (sEH), the major enzyme degrading EETs. Here we demonstrate that EETs or inhibition of sEH lead to antihyperalgesia by at least 2 spinal mechanisms, first by repressing the induction of the COX2 gene and second by rapidly up-regulating an acute neurosteroid-producing gene, StARD1, which requires the synchronized presence of elevated cAMP and EET levels. The analgesic activities of neurosteroids are well known; however, here we describe a clear course toward augmenting the levels of these molecules. Redirecting the flow of pronociceptive intracellular cAMP toward up-regulation of StARD1 mRNA by concomitantly elevating EETs is a novel path to accomplish pain relief in both inflammatory and neuropathic pain states.

Increases in spinal cerebrospinal fluid potassium concentration do not increase isoflurane minimum alveolar concentration in rats.

BACKGROUND: Previous studies demonstrated that MAC for isoflurane directly correlates with the concentration of Na(+) in cerebrospinal fluid surrounding the spinal cord, the primary site for mediation of the immobility produced by inhaled anesthetics. If this correlation resulted from increased irritability of the cord, then infusion of increased concentrations of potassium (K(+)) might be predicted to act similarly. However, an absence of effect of K(+) might be interpreted to indicate that K(+) channels do not mediate the immobility produced by inhaled anesthetics whereas Na(+) channels remain as potential mediators. Accordingly, in the present study, we examined the effect of altering intrathecal concentrations of K(+) on MAC. METHODS: In rats prepared with chronic indwelling intrathecal catheters, we infused solutions deficient in K(+) and with an excess of K(+) into the lumbar space and measured MAC for isoflurane 24 h before, during, and 24 h after infusion. Rats similarly prepared were tested for the effect of altered osmolarity on MAC (accomplished by infusion of mannitol) and for the penetration of Na(+) into the cord. RESULTS: MAC of isoflurane never significantly increased with increasing concentrations of K(+) infused intrathecally. At infused concentrations exceeding 12 times the normal concentration of KCl, i.e., 29 mEq/L, rats moved spontaneously at isoflurane concentrations just below, and sometimes at MAC, but the average MAC in these rats did not exceed their control MAC. At the largest infused concentration (58.1 mEq/L), MAC significantly decreased and did not subsequently return to normal (i.e., such large concentrations produced injury). Infusions of lower concentrations of K(+) had no effect on MAC. Infusion of osmotically equivalent solutions of mannitol did not affect MAC. Na(+) infused intrathecally measurably penetrated the spinal cord. CONCLUSIONS: The results do not support a mediation or modulation of MAC by K(+) channels.

Volatile anesthetic effects on midbrain-elicited locomotion suggest that the locomotor network in the ventral spinal cord is the primary site for immobility.

BACKGROUND: Volatile anesthetics produce immobility primarily by action in the spinal cord; however, anesthetic effects among different neuronal classes located in different spinal regions, and how they relate to immobility, are not understood. METHODS: In decerebrated rats, effects of isoflurane and halothane on movement elicited by electrical microstimulation of the mesencephalic locomotor region (MLR) were assessed in relation to minimum alveolar concentration (MAC). Anesthetic effects on step frequency and isometric limb force were measured. The authors also examined effects of MLR stimulation on responses of nociceptive dorsal horn neurons and limb force responses to tail clamp. RESULTS: Mean isoflurane requirements to block MLR-elicited stepping were slightly but significantly greater than MAC by 10%. Mean halothane requirements to block MLR-elicited stepping were greater than those for isoflurane and exceeded MAC by 20%. From 0.4 to 1.3 MAC (but not 0.0 to 0.4 MAC), there was a dose-dependent reduction in the frequency and force of hind limb movements elicited by MLR stimulation during both anesthetics. MLR stimulation inhibited noxious stimulus evoked responses of dorsal horn neurons by approximately 80%. Aptly, MLR stimulation produced analgesia that outlasted the midbrain stimulus by at least 15 s, as indicated by an 81% reduction in hind limb force elicited noxious tail clamp. CONCLUSIONS: Because electrical stimulation of the MLR elicits movement independent of dorsal horn activation, the immobilizing properties of isoflurane and halothane are largely independent of action in the dorsal horn. The results suggest that volatile anesthetics produce immobility mainly by action on ventral spinal locomotor networks.

The effects of aromatic anesthetics on dorsal horn neuronal responses to noxious stimulation.

BACKGROUND: Gamma-aminobutyric acid type A receptor potentiation and/or N-methyl-d-aspartate (NMDA) receptor inhibition might explain the anesthetic properties of fluorinated aromatic compounds. We hypothesized that depression of dorsal horn neuronal responses to noxious stimulation would correlate with the magnitude of effect of benzene (BNZ), o-difluorobenzene, and hexafluorobenzene (HFB) on NMDA receptors. METHODS: Rats were anesthetized with desflurane. A T13-L1 laminectomy allowed extracellular recording of neuronal activity from the lumbar spinal cord. After discontinuing desflurane administration, MAC for each aromatic anesthetic was determined. A 5-s noxious mechanical stimulus was then applied to the hindpaw receptive field of nociceptive dorsal horn neurons, and single-neuron responses were recorded at 0.8 and 1.2 MAC. These responses were also recorded in decerebrate rats receiving BNZ and HFB at 0-1.2 MAC. RESULTS: In intact rats, depression of responses of dorsal horn neurons to noxious stimulation by peri-MAC increases in BZN, o-difluorobenzene, and HFB correlated directly with their in vitro capacity to block NMDA receptors. In decerebrate rats, 1.2 MAC BNZ depressed nociceptive responses by 60%, with a further percentage decrease continuing from 0.8 to 1.2 MAC approximately equal to that found in intact rats. In decerebrate rats, HFB caused a progressive dose-related decrease in MAC (maximum 25%), but in intact rats, an increase from 0.8 to 1.2 neuronal response caused an (insignificant) increase in neuronal response. CONCLUSIONS: The findings in intact rats suggest that NMDA blockade contributes to the depression of dorsal horn neurons to nociceptive stimulation by fluorinated aromatic anesthetics. These results, combined with the additional findings in decerebrate rats, suggest that supraspinal effects (perhaps on gamma-aminobutyric acid type A receptors) may have a supraspinal facilitatory effect on nociception for HFB.

Immobilizing doses of halothane, isoflurane or propofol, do not preferentially depress noxious heat-evoked responses of rat lumbar dorsal horn neurons with ascending projections.

BACKGROUND: The spinal cord is an important site where volatile anesthetics decrease sensation and produce immobility. Beyond this knowledge, our understanding of a site of anesthetic action is limited. Previous evidence suggests that dorsal horn neurons with ascending projections may be more susceptible to depression by general anesthetics than local spinal interneurons. In this study we evaluated the effects of volatile and injectable general anesthetics on lumbar dorsal horn neurons with and without ascending projections. METHODS: Thirty-seven adult male rats underwent laminectomies at C1, for placement of a stimulating electrode, and T13/L1, for extracellular recording from the spinal cord dorsal horn. Neuronal responses to heat were evaluated under two doses of halothane, isoflurane, or propofol anesthesia. RESULTS: Under both halothane and isoflurane anesthesia, increasing the dose from 0.8 to 1.2 minimum alveolar concentration (MAC) had no significant effect on heat-evoked responses in neurons that had ascending projections identified via antidromic stimulation (AD) or those without ascending projections (nAD). Heat responses in AD neurons 1 min after i.v. administration of 3 and 5 mg/kg of propofol were reduced to 60% +/- 18% (mean +/- SE) and 39% +/- 14% of control respectively. Similarly, in nAD neurons responses were reduced to 56% +/- 14% and 50% +/- 10% of control by 3 and 5 mg/kg propofol respectively. CONCLUSIONS: Our findings suggest, at peri-MAC concentrations, these general anesthetics do not preferentially depress lumbar dorsal horn neurons with ascending projections compared to those with no identifiable ascending projections.

Glutamate receptor blockade in the rostral ventromedial medulla reduces the force of multisegmental motor responses to supramaximal noxious stimuli.

The rostral ventromedial medulla (RVM) has been established as part of a descending pain-modulatory pathway. While the RVM has been shown to modulate homosegmental nociceptive reflexes such as tail flick or hindpaw withdrawal, it is not known what role the RVM plays in modulating the magnitude of multisegmental, organized motor responses elicited by noxious stimuli. Using local blockade of glutamate receptors with the non-specific glutamate receptor antagonist kynurenate (known to selectively block nociceptive facilitatory ON-cells), we tested the hypothesis that the RVM facilitates the magnitude of multi-limb movements elicited by intense noxious stimuli. In male Sprague-Dawley rats, we determined the minimum alveolar concentration (MAC) of isoflurane necessary to block multi-limb motor responses to noxious tail clamp. MAC was determined so that all animals were anesthetized at an equipotent isoflurane concentration (0.7 MAC). Supramaximal mechanical stimulation of the hindpaw or electrical stimulation of the tail elicited synchronous, repetitive movements in all four limbs that ceased upon, or shortly after (<5 s) termination of the stimulus. Kynurenate microinjection (2 nmol) into the RVM significantly attenuated, by 40-60%, the peak and integrated limb forces elicited by noxious mechanical stimulation of the hindpaw (p<0.001; two-way ANOVA; n=8) or electrical stimulation of the tail (peak force: p<0.011, two-way ANOVA; n=8), with significant recovery 40-60 min following injection. The results suggest that glutamatergic excitation of RVM neurons, presumably ON-cells, facilitates organized, multi-limb escape responses to intense noxious stimuli.

Neurons in the ventral spinal cord are more depressed by isoflurane, halothane, and propofol than are neurons in the dorsal spinal cord.

BACKGROUND: Volatile anesthetics act primarily in the spinal cord to produce immobility but their exact site of action is unclear. Between 0.8 and 1.2 minimum alveolar anesthetic concentration (MAC), isoflurane does not depress neurons in the dorsal horn, suggesting that it acts at a more ventral site within the spinal cord such as in premotor interneurons and motoneurons. We hypothesized that isoflurane, halothane, and propofol would exert a greater depressant effect on nociceptive responses of ventral horn neurons when compared with dorsal horn neurons. METHODS: Rats were anesthetized with isoflurane or halothane and responses of dorsal (<1200 microm deep) and ventral (>1200 microm) lumbar neurons to noxious mechanical stimulation of the hindpaw were determined at 0.8 and 1.2 MAC. In a third group anesthetized with isoflurane at 0.8 MAC, we administered 5 mg/kg propofol while recording responses from dorsal horn or ventral horn neurons. RESULTS: Dorsal horn neuronal responses were not significantly affected when either isoflurane or halothane was increased from 0.8 to 1.2 MAC; propofol also had no significant effect. On the other hand, with increased isoflurane or halothane concentration, responses of ventral horn neurons were depressed by 60% and 45%, respectively. Propofol profoundly depressed (>90%) ventral horn neurons. CONCLUSIONS: These data suggest that, in the peri-MAC range, isoflurane, halothane, and propofol have little or no effect on neuronal responses to noxious mechanical stimulation in the spinal dorsal horn but depress such responses in the ventral horn. Immobility produced in the 0.8-1.2 MAC range by these anesthetics appears to result from a depressant action in the ventral horn.

Inhibition of soluble epoxide hydrolase reduces LPS-induced thermal hyperalgesia and mechanical allodynia in a rat model of inflammatory pain.

Soluble epoxide hydrolases catalyze the hydrolysis of epoxides in acyclic systems. In man this enzyme is the product of a single copy gene (EPXH-2) present on chromosome 8. The human sEH is of interest due to emerging roles of its endogenous substrates, epoxygenated fatty acids, in inflammation and hypertension. One of the consequences of inhibiting sEH in rodent inflammation models is a profound decrease in the production of pro-inflammatory and proalgesic lipid metabolites including prostaglandins. This prompted us to hypothesize that sEH inhibitors may have antinociceptive properties. Here we tested if sEH inhibitors can reduce inflammatory pain. Hyperalgesia was induced by intraplantar LPS injection and sEH inhibitors were delivered topically. We found that two structurally dissimilar but equally potent sEH inhibitors can be delivered through the transdermal route and that sEH inhibitors effectively attenuate thermal hyperalgesia and mechanical allodynia in rats treated with LPS. In addition we show that epoxydized arachidonic acid metabolites, EETs, are also effective in attenuating thermal hyperalgesia in this model. In parallel with the observed biological activity metabolic analysis of oxylipids showed that inhibition of sEH resulted with a decrease in PGD2 levels and sEH generated degradation products of linoleic and arachidonic acid metabolites with a concomitant increase in epoxides of linoleic acid. These data show that inhibition of sEH may become a viable therapeutic strategy to attain analgesia.

Enhancement of antinociception by coadministration of nonsteroidal anti-inflammatory drugs and soluble epoxide hydrolase inhibitors.

Combination therapies have long been used to treat inflammation while reducing side effects. The present study was designed to evaluate the therapeutic potential of combination treatment with nonsteroidal anti-inflammatory drugs (NSAIDs) and previously undescribed soluble epoxide hydrolase inhibitors (sEHIs) in lipopolysaccharide (LPS)-challenged mice. NSAIDs inhibit cyclooxygenase (COX) enzymes and thereby decrease production of metabolites that lead to pain and inflammation. The sEHIs, such as 12-(3-adamantan-1-yl-ureido)-dodecanoic acid butyl ester (AUDA-BE), stabilize anti-inflammatory epoxy-eicosatrienoic acids, which indirectly reduce the expression of COX-2 protein. Here we demonstrate that the combination therapy of NSAIDs and sEHIs produces significantly beneficial effects that are additive for alleviating pain and enhanced effects in reducing COX-2 protein expression and shifting oxylipin metabolomic profiles. When administered alone, AUDA-BE decreased protein expression of COX-2 to 73 +/- 6% of control mice treated with LPS only without altering COX-1 expression and decreased PGE(2) levels to 52 +/- 8% compared with LPS-treated mice not receiving any therapeutic intervention. When AUDA-BE was used in combination with low doses of indomethacin, celecoxib, or rofecoxib, PGE(2) concentrations dropped to 51 +/- 7, 84 +/- 9, and 91 +/- 8%, respectively, versus LPS control, without disrupting prostacyclin and thromboxane levels. These data suggest that these drug combinations (NSAIDs and sEHIs) produce a valuable beneficial analgesic and anti-inflammatory effect while prospectively decreasing side effects such as cardiovascular toxicity.

Isoflurane disrupts central pattern generator activity and coordination in the lamprey isolated spinal cord.

BACKGROUND: Although volatile anesthetics such as isoflurane can depress sensory and motor neurons in the spinal cord, movement occurring during anesthesia can be coordinated, involving multiple limbs as well as the head and trunk. However, it is unclear whether volatile anesthetics depress locomotor interneurons comprising central pattern generators or disrupt intersegmental central pattern generator coordination. METHODS: Lamprey spinal cords were excised during anesthesia and placed in a bath containing artificial cerebrospinal fluid and D-glutamate to induce fictive swimming. The rostral, middle, and caudal regions were bath-separated using acrylic partitions and petroleum jelly, and in each compartment, the authors recorded ventral root activity. The authors selectively delivered isoflurane (0.5, 1, and 1.5%) only to the middle segments of the spinal cord. Spectral analyses were then used to assess isoflurane effects on central pattern generator activity and coordination. RESULTS: Isoflurane dose-dependently reduced fictive locomotor activity in all three compartments, with 1.5% isoflurane nearly eliminating activity in the middle compartment and reducing spectral amplitudes in the anesthetic-free rostral and caudal compartments to 23% and 31% of baseline, respectively. Isoflurane decreased burst frequency in the caudal compartment only, to 53% of baseline. Coordination of central pattern generator activity between the rostral and caudal compartments was also dose-dependently decreased, to 42% of control at 1.5% isoflurane. CONCLUSION: Isoflurane disrupts motor output by reducing interneuronal central pattern generator activity in the spinal cord. The effects of isoflurane on motor output outside the site of isoflurane application were presumably independent of effects on sensory or motor neurons.

Drastic decrease in isoflurane minimum alveolar concentration and limb movement forces after thoracic spinal cooling and chronic spinal transection in rats.

BACKGROUND: Individuals with spinal cord injury may undergo multiple surgical procedures; however, it is not clear how spinal cord injury affects anesthetic requirements and movement force under anesthesia during both acute and chronic stages of the injury. METHODS: The authors determined the isoflurane minimum alveolar concentration (MAC) necessary to block movement in response to supramaximal noxious stimulation, as well as tail-flick and hind paw withdrawal latencies, before and up to 28 days after thoracic spinal transection. Tail-flick and hind paw withdrawal latencies were measured in the awake state to test for the presence of spinal shock or hyperreflexia. The authors measured limb forces elicited by noxious mechanical stimulation of a paw or the tail at 28 days after transection. Limb force experiments were also conducted in other animals that received a reversible spinal conduction block by cooling the spinal cord at the level of the eighth thoracic vertebra. RESULTS: A large decrease in MAC (to /= 90%) in both chronic and acute cold-block spinal animals. CONCLUSIONS: The immobilizing potency of isoflurane increases substantially after spinal transection, despite the absence of a baseline motor depression, or "spinal shock." Therefore, isoflurane MAC is determined by a spinal depressant action, possibly counteracted by a supraspinal facilitatory action. The partial recovery in MAC at later time points suggests that neuronal plasticity after spinal cord injury influences anesthetic requirements.