Differential Block of Fast and Slow Inactivating Tetrodotoxin-sensitive Sodium Channels by Droperidol in Spinal Dorsal Horn Neurons

Background: Dorsal horn neurons of the spinal cord participate in neuronal pain transmission. During spinal and epidural anesthesia, dorsal horn neurons are exposed to local anesthetics and opioids. Droperidol is usually given with opioids to avoid nausea and vomiting. A recently developed method of "entire soma isolation" has made it possible to study directly the action of droperidol on different components of Na+ current in dorsal horn neurons.

Methods: Using a combination of the whole-cell patch-clamp recording from spinal cord slices and the entire soma isolation method, we studied the direct action of droperidol on two types of Na+ currents in dorsal horn neurons of young rats.

Results: The tetrodotoxin-sensitive Na+ current in isolated somata consisted of a fast inactivating ([tau]F, 0.5-2 ms; 80-90% of the total amplitude) and a slow inactivating ([tau]S, 6-20 ms; 10-20% of the total amplitude) component. Droperidol, at concentrations relevant for spinal and epidural anesthesia, selectively and reversibly suppressed the fast component with a half-maximum inhibiting concentration (IC50) of 8.3 [mu]m. The slow inactivating component was much less sensitive to droperidol; the estimated IC50 value was 809 [mu]m.     

Suppression of potassium conductance by droperidol has influence on excitability of spinal sensory neurons

BACKGROUND: During spinal and epidural anesthesia with opioids, droperidol is added to prevent nausea and vomiting. The mechanisms of its action on spinal sensory neurons are not well understood. It was previously shown that droperidol selectively blocks a fast component of the Na+ current. The authors studied the action of droperidol on voltage-gated K+ channels and its effect on membrane excitability in spinal dorsal horn neurons of the rat. METHODS: Using a combination of the patch-clamp technique and the "entire soma isolation" method, the action of droperidol on fast-inactivating A-type and delayed-rectifier K+ channels was investigated. Current-clamp recordings from intact sensory neurons in spinal cord slices were performed to study the functional meaning of K+ channel block for neuronal excitability. RESULTS: Droperidol blocked delayed-rectifier K+ currents in isolated somata of dorsal horn neurons with a half-maximum inhibiting concentration of 20.6 microm. The A-type K+ current was insensitive to up to 100 microm droperidol. At droperidol concentrations insufficient for suppression of an action potential, the block of delayed-rectifier K+ channels led to an increase in action potential duration and, as a consequence, to lowering of the discharge frequency in the neuron. CONCLUSIONS: Droperidol blocks delayed-rectifier K+ channels in a concentration range close to that for suppression of Na+ channels. The block of delayed-rectifier K+ channels by droperidol enhances the suppression of activity in spinal sensory neurons at drug concentrations insufficient for complete conduction block.     

Suppression of Potassium Conductance by Droperidol Has Influence on Excitability of Spinal Sensory Neurons

Background: During spinal and epidural anesthesia with opioids, droperidol is added to prevent nausea and vomiting. The mechanisms of its action on spinal sensory neurons are not well understood. It was previously shown that droperidol selectively blocks a fast component of the Na+ current. The authors studied the action of droperidol on voltage-gated K+ channels and its effect on membrane excitability in spinal dorsal horn neurons of the rat.

Methods: Using a combination of the patch-clamp technique and the "entire soma isolation" method, the action of droperidol on fast-inactivating A-type and delayed-rectifier K+ channels was investigated. Current-clamp recordings from intact sensory neurons in spinal cord slices were performed to study the functional meaning of K+ channel block for neuronal excitability.

Results: Droperidol blocked delayed-rectifier K+ currents in isolated somata of dorsal horn neurons with a half-maximum inhibiting concentration of 20.6 [mu]m. The A-type K+ current was insensitive to up to 100 [mu]m droperidol. At droperidol concentrations insufficient for suppression of an action potential, the block of delayed-rectifier K+ channels led to an increase in action potential duration and, as a consequence, to lowering of the discharge frequency in the neuron.     

Blockade of Na sup + and K sup + Currents by Local Anesthetics in the Dorsal Horn Neurons of the Spinal Cord

Background: The dorsal horn of the spinal cord is a pivotal point for transmission of neuronal pain. During spinal and epidural anesthesia, the neurons of the dorsal horn are exposed to local anesthetics. Unfortunately, little is known about the action of local anesthetics on the major ionic conductances in dorsal horn neurons. In this article, the authors describe the effects of bupivacaine, lidocaine, and mepivacaine on voltage-gated Na sup + and K sup + currents in the membranes of these neurons.

Methods: The patch-clamp technique was applied to intact dorsal horn neurons from laminae I-III identified in 200-micro meter slices of spinal cord from newborn rats. Under voltage-clamp conditions, the whole-cell Na sup + and K sup + currents activated by depolarization were recorded in the presence of different concentrations of local anesthetics.

Results: Externally applied bupivacaine, lidocaine, and mepivacaine produced tonic block of Na sup + currents with different potencies. Half-maximum inhibiting concentrations (IC50) were 26, 112, and 324 micro Meter, respectively. All local anesthetics investigated also showed a phasic, that is, a use-dependent, block of Na sup + channels. Rapidly inactivating K sup + currents (KA currents) also were sensitive to the blockers with IC50 values for tonic blocks of 109, 163, and 236 micro Meter, respectively. The block of KA currents was not use dependent. In contrast to Na sup + and KA currents, delayed-rectifier K sup + currents were almost insensitive to the local anesthetics applied.     

Meperidine suppresses the excitability of spinal dorsal horn neurons

BACKGROUND: In addition to local anesthetics, meperidine has been successfully used for local anesthesia. When applied intrathecally, the dorsal horn neurons of the superficial laminae are exposed to high concentrations of meperidine. These cells represent an important point for the transmission of pain information. This study investigated the blocking effects of meperidine on different ionic currents of spinal dorsal horn neurons and, in particular, its impact on the generation of action potentials. METHODS: Using a combination of the patch clamp technique and the entire soma isolation method, the action of meperidine on voltage-gated Na+ and K+ currents in spinal dorsal horn neurons of rats was described. Current clamp recordings from intact neurons showed the functional relevance of the ion current blockade for the generation of action potentials. RESULTS: Externally applied meperidine reversibly blocked voltage-gated Na+ currents with a half-maximum inhibiting concentration (IC50) of 112 microM. During repetitive stimulation, a slight phasic block occurred. In addition, A-type K+ currents and delayed-rectifier K+ currents were affected in a dose-dependent manner, with IC50 values of 102 and 52 microM, respectively. In the current clamp mode, single action potentials were suppressed by meperidine. The firing frequency was lowered to 54% at concentrations (100 microM) insufficient for the suppression of a single action potential. CONCLUSIONS: Meperidine inhibits the complex mechanism of generating action potentials in spinal dorsal horn neurons by the blockade of voltage-gated Na+ and K+ channels. This can contribute to the local anesthetic effect of meperidine during spinal anesthesia.     

Meperidine Suppresses the Excitability of Spinal Dorsal Horn Neurons

Background: In addition to local anesthetics, meperidine has been successfully used for local anesthesia. When applied intrathecally, the dorsal horn neurons of the superficial laminae are exposed to high concentrations of meperidine. These cells represent an important point for the transmission of pain information. This study investigated the blocking effects of meperidine on different ionic currents of spinal dorsal horn neurons and, in particular, its impact on the generation of action potentials.

Methods: Using a combination of the patch clamp technique and the entire soma isolation method, the action of meperidine on voltage-gated Na+ and K+ currents in spinal dorsal horn neurons of rats was described. Current clamp recordings from intact neurons showed the functional relevance of the ion current blockade for the generation of action potentials.

Results: Externally applied meperidine reversibly blocked voltage-gated Na+ currents with a half-maximum inhibiting concentration (IC50) of 112 [mu]m. During repetitive stimulation, a slight phasic block occurred. In addition, A-type K+ currents and delayed-rectifier K+ currents were affected in a dose-dependent manner, with IC50 values of 102 and 52 [mu]m, respectively. In the current clamp mode, single action potentials were suppressed by meperidine. The firing frequency was lowered to 54% at concentrations (100 [mu]m) insufficient for the suppression of a single action potential.     

Characteristics of ropivacaine block of Na+ channels in rat dorsal root ganglion neurons

When used for epidural anesthesia, ropivacaine can produce a satisfactory sensory block with a minor motor block. We investigated its effect on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) Na(+) currents in rat dorsal root ganglion (DRG) neurons to elucidate the mechanisms underlying the above effects. Whole-cell patch-clamp recordings were made from enzymatically dissociated neurons from rat DRG. A TTX-S Na(+) current was recorded preferentially from large DRG neurons and a TTX-R Na(+) current preferentially from small ones. Ropivacaine shifted the activation curve for the TTX-R Na(+) channel in the depolarizing direction and the inactivation curve for both types of Na(+) channel in the hyperpolarizing direction. Ropivacaine blocked TTX-S and TTX-R Na(+) currents, but its half-maximum inhibitory concentration (IC(50)) was significantly lower for the latter current (116 +/- 35 vs 54 +/- 14 microM; P: < 0.01); similar IC(50) values were obtained with the (R)-isomer of ropivacaine. Ropivacaine produced a use-dependent block of both types of Na(+) channels. Ropivacaine preferentially blocks TTX-R Na(+) channels over TTX-S Na(+) channels. We conclude that because TTX-R Na(+) channels exist mainly in small DRG neurons (which are responsible for nociceptive sensation), such selective action of ropivacaine could underlie the differential block observed during epidural anesthesia with this drug. Implications: Whole-cell patch-clamp recordings of tetrodotoxin-sensitive and tetrodotoxin-resistant Na(+) currents in rat dorsal root ganglion neurons showed ropivacaine preferentially blocked tetrodotoxin-resistant Na(+) channels over tetrodotoxin-sensitive Na(+) channels. This could provide a desirable differential sensory blockade during epidural anesthesia using ropivacaine.     

全身麻醉药在脊髓内的作用机制

脊髓是全身麻醉药抑制伤害性刺激体动反应和抗伤害效应的重要作用部位,含有不同配体门控离子受体等多个可能介导麻醉效应的靶点.不同药物在脊髓内经各自特异靶点通过多种分子机制发挥作用.现就全身麻醉药制动和镇痛效应在脊髓内的作用位点和分子机制作一综述.     

Synaptic blockade plays a major role in the neural disturbance of experimental spinal cord compression

We analyzed dynamic processes of neural excitation propagation in the experimentally compressed spinal cord using a high-speed optical recording system. Transverse slices of the juvenile rat cervical spinal cord were stained with a voltage-sensitive dye (di-4-ANEPPS). Two components were identified in the depolarizing optical responses to dorsal root electrical stimulation: a fast component of short duration corresponding to pre-synaptic excitation and a slow component of long duration corresponding to post-synaptic excitation. In the directly compressed dorsal horn, the slow component was attenuated more (attenuated to 37.4 +/- 9.1% of the control) than the fast component (to 70.5 +/- 14.9%) (p < 0.01) at 400 msec after stimulation. Depolarizing optical responses to compression and to chemical synaptic blockade were similar. There was a regional difference between white matter (attenuated to 86.2 +/- 10.5%) and gray matter (to 72.6 +/- 10.4%) (p < 0.03) in compression-induced changes of the fast components; neural activity in the white matter was resistant to compression, especially in the dorsal root entry zone. Depolarizing optical signals in the region adjacent to the directly compressed site were also attenuated; the fast component was attenuated to 77.6 +/- 10.4% and the slow component to 31.8 +/- 11.3% of the control signals (p < 0.01). Spinal cord dysfunction induced by purely mechanical compression without tissue destruction was virtually restored with early decompression. We suggest that a disturbance of synaptic transmission plays an important role in the pathophysiological mechanisms of spinal cord compression, at least under in vitro experimental conditions of juvenile rats.     

Spinally administered epinephrine suppresses noxiously evoked activity of WDR neurons in the dorsal horn of the spinal cord

This study was designed to determine if spinally administered epinephrine is capable of suppressing noxiously evoked activity of wide dynamic range (WDR) neurons in the dorsal horn of the spinal cord. Extracellular activity was recorded from single WDR neurons in the dorsal horn of decerebrate, spinal cord-transected (T-12) cats. Activity was evoked by the presentation of a noxious radiant heat stimulus (51 degrees C) to the cells' receptive fields on the hind paws. Evoked activity was monitored both before and after the spinal administration of either 50 micrograms (n = 6) or 100 micrograms (n = 6) epinephrine. Both doses of epinephrine produced a significant suppression of noxiously evoked activity, which was dose-dependent. In addition, the 100-microgram dose produced a suppression that was of longer duration than that seen following the 50-microgram dose. Recovery from suppression was recorded following both the 50- and 100-microgram dose. These results indicate that spinally administered epinephrine is capable of suppressing noxiously evoked activity of WDR neurons in the dorsal horn of the spinal cord. Since WDR neurons have been identified as cells of origin for the spinothalamic tract, such an action may block the central transmission of afferent pain information. This may be a mechanism by which spinally administered epinephrine enhances the duration or intensity of spinal anesthesia produced by local anesthetics and may also explain spinal analgesia resulting from the spinal administration of adrenergic agonists. Interactions between spinally administered epinephrine and spinally administered opioids also were studied.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anatomy,physiology and pharmacology of pain

Nociception is conveyed from the periphery to the brain at three levels: the peripheral nociceptor, the spinal cord, and the supra-spinal (brain) levels. Physiological (first or ‘fast’) pain is produced by stimulation of high threshold thermo/mechanical nociceptors, which transmit via fast conducting myelinated A delta fibres. These enter the dorsal horn of the spinal cord and synapse at laminae I and V. Pathophysiological (second or ‘slow’) pain originates from stimulation of the high threshold polymodal nociceptors (free endings) present in all tissues. The nociceptors respond to mechanical, chemical and thermal stimuli and are transmitted via slow conducting unmyelinated C fibres. These synapse at laminae II and III (substantia gelatinosa) of the dorsal horn. The second order neurons are either nociceptive specific (substantia gelatinosa) or wide dynamic range (WDR) neurons (in laminae V and VI) that respond to a wide range of noxious and non-noxious input. Both pathways ascend up the spinal cord via the spinothalamic tracts to the thalamus, which synapse and project on to the somatosensory cortex. Inhibitory inter-neurons in the substantia gelatinosa prevent activation of the dorsal root ganglia. Interneurons can be activated by A beta and inhibited by A beta and C fibre activity. Pain can be ‘gated-out’ by stimulating the large A beta fibres in the painful area. This is the working mechanism behind transcutaneous electrical nerve stimulation. The descending inhibition pathways originate at the level of the cortex and thalamus, and descend via the brainstem (periaqueductal grey) and the dorsal columns to terminate at the dorsal horn of the spinal cord. Neurotransmitters noradrenaline, serotonin (5-HT) and the endogenous opioids are released to provide antinociception.     

The changes in human spinal sympathetic preganglionic neurons after spinal cord injury

We have applied conventional histochemical, immunocytochemical and morphometric techniques to study the changes within the human spinal sympathetic preganglionic neurons (SPNs) after spinal cord injury. SPNs are localized within the intermediolateral nucleus (IML) of the lateral horn at the thoraco-lumbar level of the spinal cord and are the major contributors to central cardiovascular control. SPNs in different thoracic segments in the normal spinal cord were similar in soma size. SPNs in the IML were also identified using immunoreactivity to choline acetyltransferase. Soma area of SPNs was 400.7+15 microm2 and 409.9+/-22 microm2 at the upper thoracic (T3) and middle thoracic (T7) segments, respectively. In the spinal cord obtained from a person who survived for 2 weeks following a spinal cord injury at T5, we found a significant decrease in soma area of the SPNs in the segments below the site of injury: soma area of SPNs at T8 was 272.9+/-11 microm2. At T1 the soma area was 418+/-19 microm2. In the spinal cord obtained from a person who survived 23 years after cord injury at T3, the soma area of SPNs above (T1) and below (T7) the site of injury was similar (416.2+/-19 and 425.0+/-20 microm2 respectively). The findings demonstrate that the SPNs in spinal segments caudal to the level of the lesion undergo a significant decrease of their size 2 weeks after spinal cord injury resulting in complete transection of the spinal cord. The impaired cardiovascular control after spinal cord injury may be accounted for, in part, by the described changes of the SPNs. The SPNs in spinal segments caudal to the injury were of normal size in the case studied 23 years after the injury, suggesting that the atrophy observed at 2 weeks is transient. More studies are necessary to establish the precise time course of these morphological changes in the spinal preganglionic neurons.     

Effects of dorsal root entry zone lesion on spinal cord potentials evoked by segmental,ascending and descending volleys

Summary The spinal cord potentials (SCPs) were recorded from the dorsal root entry zone (DREZ) and posterior epidural space in patients before and after dorsal root entry zone lesion (DREZL) during general anaesthesia. The SCPs from the DREZ activated by segmental, ascending and descending volleys were basically the same in fundamental waveform as those recorded from the posterior epidural space. Segmentally activated slow negative (N₁) wave, reflecting synchronized activities of dorsal horn neurones, and positive (P₂) wave, thought to indicate primary afferent depolarization, were affected by DREZL in all 4 subjects tested, even by contralateral stimulation, suggesting that these components of the segmental SCPs in man partly reflect the activities of the contralateral dorsal horn. The spike-like potentials activated by ascending volleys were not affected by DREZL, while the subsequent slow components were decreased in the lesioned level. This may indicate that ascending spinal cord tracts are not affected by the operation, and suggests that the origin of the slow components by ascending volleys lies at least in part in the segmental dorsal horn. The slow negative and positive components, recorded at a remote segment from DREZL, in response to the descending volleys, were augmented after DREZL, suggesting that activation of ascending or descending inhibition through a feedback loop via the supraspinal structures might occur at least transiently following DREZL. All components of the SCPs activated by descending volleys were decreased or disappeared in recording from the lesioned level, as expected. Thus, intra-operative recording of the SCPs during DREZL might be beneficial for monitoring and studying human spinal cord function.     

The effects of isoflurane on conditioned inhibition by dorsal column stimulation

Spinal dorsal column stimulation (DCS) modulates sensory transmission, including pain, at the dorsal horn of the cord. However, the mechanisms of DCS modulatory actions and the effects of anesthetics on these mechanisms remain to be investigated. We studied the effects of isoflurane (1.0% and 2.0%) on conditioned inhibition, the amplitude decrease of the spinal cord potentials (SCPs) after a conditioning volley (DCS), in the ketamine-anesthetized rat by recording the sharp negative (N) and slow positive (P) waves of the SCPs evoked by conditioning dorsal column (DC) and testing segmental stimulations. The N wave is believed to be the synchronized activity of the dorsal horn neurons, and the P wave, primary afferent depolarization (PAD), reflecting presynaptic inhibition. The P potentials evoked by either DC or segmental stimulation were depressed by isoflurane, whereas the N waves remained unchanged, indicating that the pharmacological characteristics of these N and P waves are similar between DC-evoked and segmentally evoked SCPs. The conditioned inhibition of segmental N and P waves by DC stimulation was almost completely suppressed by 2.0% isoflurane. The conditioned inhibition of the segmental N wave was not changed by spinal cord transection, whereas the conditioned inhibition of the segmental P wave was decreased. The results indicate that isoflurane depresses presynaptic inhibition without affecting the synchronized activity of dorsal horn neurons and, most profoundly, depresses the conditioned inhibition by DC stimulation of the dorsal horn neurons and PAD. Further, the results indicate that conditioned inhibition by DC stimulation of PAD receives a facilitatory influence from the supraspinal structures, whereas that of the synchronized activity of the dorsal horn neurons does not. IMPLICATIONS: To investigate how anesthetics affect supraspinal modulation of sensory transmission in the spinal cord, the spinal cord potential (SCP) evoked by dorsal cord stimulation (DCS) and segmentally evoked SCP conditioned by DCS were recorded in intact and spinal cord-transected rats during isoflurane anesthesia.     

Droperidol inhibits GABA(A) and neuronal nicotinic receptor activation

BACKGROUND: Droperidol is used in neuroleptanesthesia and as an antiemetic. Although its antiemetic effect is thought to be caused by dopaminergic inhibition, the mechanism of droperidol's anesthetic action is unknown. Because gamma-aminobutyric acid type A (GABAA) and neuronal nicotinic acetylcholine receptors (nAChRs) have been implicated as putative targets of other general anesthetic drugs, the authors tested the ability of droperidol to modulate these receptors. METHODS: gamma-Aminobutyric acid type A alpha1beta1gamma2 receptor, alpha7 and alpha4beta2 nAChRs were expressed in Xenopus oocytes and studied with two-electrode voltage clamp recording. The authors tested the ability of droperidol at concentrations from 1 nm to 100 microm to modulate activation of these receptors by their native agonists. RESULTS: Droperidol inhibited the GABA response by a maximum of 24.7 +/- 3.0%. The IC50 for inhibition was 12.6 +/- 0.47 nm droperidol. At high concentrations, droperidol (100 microm) activates the GABAA receptor in the absence of GABA. Inhibition of the GABA response is significantly greater at hyperpolarized membrane potentials. The activation of the alpha7 nAChR is also inhibited by droperidol, with an IC50 of 5.8 +/- 0.53 microm. The Hill coefficient is 0.95 +/- 0.1. Inhibition is noncompetitive, and membrane voltage dependence is insignificant. CONCLUSIONS: Droperidol inhibits activation of both the GABAA alpha1beta1gamma2 and alpha7 nAChR. The submaximal GABA inhibition occurs within a concentration range such that it might be responsible for the anxiety, dysphoria, and restlessness that limit the clinical utility of high-dose droperidol anesthesia. Inhibition of the alpha7 nAChR might be responsible for the anesthetic action of droperidol.     

Isoflurane reduces glutamatergic transmission in neurons in the spinal cord superficial dorsal horn: evidence for a presynaptic site of an analgesic action

The minimum alveolar concentration (MAC) of a volatile anesthetic defines anesthetic potency in terms of a suppressed motor response to a noxious stimulus. Therefore, the MAC of an anesthetic might in part reflect depression of motor neuron excitability. In the present study we evaluated the effect of isoflurane (ISO) on neurons in the substantia gelatinosa driven synaptically by putative nociceptive inputs in an in vitro spinal cord preparation of the rat. Whole-cell patch-clamp recordings were performed in neurons with their soma in the substantia gelatinosa of transverse rat spinal cord slices. We investigated the effect of ISO on excitatory postsynaptic currents (EPSC) evoked by dorsal root stimulation (eEPSC), spontaneous (sEPSC), and miniature (mEPSC) EPSC. ISO reversibly reduced the amplitude of eEPSC to 39% +/- 22% versus control. ISO decreased the frequency of sEPSC and mEPSC to 39% +/- 26% and 63% +/- 7%. Neither the amplitudes nor the kinetics of mEPSC and sEPSC were altered by ISO. We conclude that ISO depresses glutamatergic synaptic transmission of putative nociceptive primary-afferent inputs, presumably by reducing the release of the excitatory transmitter. This effect may contribute to an antinociceptive action of volatile anesthetics at the spinal cord level. IMPLICATIONS: The present electrophysiological in vitro experiments provide evidence that the volatile anesthetic isoflurane reduces excitatory transmitter release at the first site of synaptic integration of nociceptive inputs, the spinal cord superficial dorsal horn. This effect may contribute to the anesthetic action of volatile anesthetics at the spinal cord level.     

Inhibitory action of sensory transmission by inhalational anesthetics in the spinal cord

This review article focuses on the suppression of sensory transmission by inhalational anesthetics at the spinal cord level. Volatile anesthetics (e.g. halothane and isoflurane) suppress neuronal responses evoked by both noxious and non-noxious stimuli. This suppression is mediated largely by activation of GABAA and glycine receptors systems in the spinal dorsal horn. Depression of spinal glutamate receptor systems is also probably involved. The analgesic action of nitrous oxide is produced by activation of supra-spinal descending inhibitory systems, not by direct action on the spinal cord. Activation of the descending inhibitory systems by nitrous oxide causes release of noradrenaline in the spinal dorsal horn, and activates alpha 2 adrenergic receptor systems, resulting in depression of neuronal responses evoked by noxious stimuli. GABAA and glycine receptor systems in the spinal dorsal horn are also important components of nitrous oxide anesthesia in depressing neuronal responses evoked by non-noxious stimuli. Although excitation or inhibition of GABAA, glycine, alpha 2 adrenergic and glutamate receptors systems is an important action of inhalational anesthetics, influence of inhalational anesthetics on interactions among these receptor systems has yet to be studied.     

Halothane and isoflurane have additive minimum alveolar concentration (MAC) effects in rats

Studies suggest that at concentrations surrounding MAC (the minimum alveolar concentration suppressing movement in 50% of subjects in response to noxious stimulation), halothane depresses dorsal horn neurons more than does isoflurane. Similarly, these anesthetics may differ in their effects on various receptors and ion channels that might be anesthetic targets. Both findings suggest that these anesthetics may have effects on movement in response to noxious stimulation that would differ from additivity, possibly producing synergism or even antagonism. We tested this possibility in 20 rats. MAC values for halothane and (separately) for isoflurane were determined in duplicate before and after testing the combination (also in duplicate; six determinations of MAC for each rat). The sum of the isoflurane and halothane MAC fractions for individual rats that produced immobility equaled 1.037 +/- 0.082 and did not differ significantly from a value of 1.00. That is, the combination of halothane and isoflurane produced immobility in response to tail clamp at concentrations consistent with simple additivity of the effects of the anesthetics. These results suggest that the immobility produced by inhaled anesthetics need not result from their capacity to suppress transmission through dorsal horn neurons. IMPLICATIONS: Despite differences in their capacities to inhibit spinal dorsal horn cells, isoflurane and halothane are additive in their ability to suppress movement in response to a noxious stimulus.     

Bupivacaine inhibits activation of neuronal spinal extracellular receptor-activated kinase through selective effects on ionotropic receptors

BACKGROUND: Central terminals of primary nociceptors release neurotransmitters glutamate and substance P, which bind to ionotropic or metabotropic receptors on spinal neurons to induce cellular responses. Extracellular signal-regulated kinases are activated by these receptors and are important modulators of pain at the dorsal horn. The authors investigated these pathways as potential targets for antinociceptive actions of local anesthetics. METHODS: The effects of bupivacaine on the activation of extracellular receptor-activated kinase (phosphorylation to pERK) in rat spinal cord slices, induced by presynaptic release (capsaicin), by presynaptic or postsynaptic ionotropic or metabotropic receptor activation, or by activation of intracellular protein kinase C or protein kinase A and also by a receptor-independent Ca2+ ionophore, were quantitated by immunohistochemistry, counting pERK-positive neurons in the superficial dorsal horn. RESULTS: Capsaicin (3 microm, 10 min)-stimulated pERK was reduced by bupivacaine (IC50 approximately 2 mm, approximately 0.05%), which similarly suppressed pERK induced by the ionotropic glutamate receptors for N-methyl-D-aspartate and (S)-alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid but not that induced by the metabotropic receptors for glutamate, bradykinin, or substance P. Extracellular receptor-activated kinase activation by the Ca2+ ionophore ionomycin was also sensitive to bupivacaine, but direct activation by protein kinase A or protein kinase C was not. CONCLUSIONS: Bupivacaine inhibits pERK activation resulting from different modes of Ca2+ influx through the plasma membrane. This represents a postsynaptic mechanism of analgesia that occurs in parallel with impulse inhibition during neuraxial blockade.     

Functional mu opioid receptors are reduced in the spinal cord dorsal horn of diabetic rats

BACKGROUND: The mechanisms of decreased spinal analgesic potency of morphine in neuropathic pain are not fully known. Agonist-stimulated [35S]GTPgammaS receptor autoradiography has been used to measure receptor activation of G proteins in vitro. Using this technique, we determined changes in the functional mu opioid receptors in the spinal dorsal horn in diabetic rats. METHODS: Rats were rendered diabetic with an intraperitoneal injection of streptozotocin. The lumbar spinal cord was obtained from age-matched normal and diabetic rats 4 weeks after streptozotocin treatment. [D-Ala2,N-MePhe4,Gly5-ol]-enkephalin (DAMGO, 10 microm)-stimulated [35S]GTPgammaS binding was performed in both tissue sections and isolated membranes. RESULTS: The DAMGO-stimulated [35S]GTPgammaS binding in the spinal dorsal horn was significantly reduced (approximately 37%) in diabetic rats compared with normal rats. However, [35S]GTPgammaS bindings in the spinal dorsal horn stimulated by other G protein-coupled receptor agonists, including [D-Pen2,D-Pen5]-enkephalin, R(-)N6-(2-phenylisopropyl)-adenosine, and WIN-55212, were not significantly altered in diabetic rats. The basal [35S]GTPgammaS binding in the spinal dorsal horn was slightly (approximately 13%) but significantly increased in diabetic rats. Western blot analysis revealed no significant difference in the expression of the alpha subunits of G(i) and G(o) proteins in the dorsal spinal cord between normal and diabetic rats. CONCLUSIONS: These data suggest that the functional mu opioid receptors in the spinal cord dorsal horn of diabetic rats are reduced. The impaired functional mu opioid receptors in the spinal cord may constitute one of the mechanisms underlying the reduced spinal analgesic effect of mu opioids in diabetic neuropathic pain.