Tomohiro Yamakura

Effects of gaseous anesthetics nitrous oxide and xenon on ligand-gated ion channels. Comparison with isoflurane and ethanol.

BackgroundLigand-gated ion channels are considered to be potential general anesthetic targets. Although most general anesthetics potentiate the function of &ggr;-aminobutyric acid receptor type A (GABAA), the gaseous anesthetics nitrous oxide and xenon are reported to have little effect on GABAA receptors but inhibit N-methyl-d-aspartate (NMDA) receptors. To define the spectrum of effects of nitrous oxide and xenon on receptors thought to be important in anesthesia, the authors tested these anesthetics on a variety of recombinant brain receptors. MethodsThe glycine, GABAA, GABA receptor type C (GABAC), NMDA, &agr;-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), kainate, 5-hydroxytryptamine3 (5-HT3), and nicotinic acetylcholine (nACh) receptors were expressed in Xenopus oocytes and effects of nitrous oxide and xenon, and as equipotent concentrations of isoflurane and ethanol, were studied using the two-electrode voltage clamp. ResultsNitrous oxide (0.58 atmosphere [atm]) and xenon (0.46 atm) exhibited similar effects on various receptors. Glycine and GABAA receptors were potentiated by gaseous anesthetics much less than by isoflurane, whereas nitrous oxide inhibited GABAC receptors. Glutamate receptors were inhibited by gaseous anesthetics more markedly than by isoflurane, but less than by ethanol. NMDA receptors were the most sensitive among glutamate receptors and were inhibited by nitrous oxide by 31%. 5-HT3 receptors were slightly inhibited by nitrous oxide. The nACh receptors were inhibited by gaseous and volatile anesthetics, but ethanol potentiated them. The sensitivity was different between &agr;4&bgr;2 and &agr;4&bgr;4 nACh receptors; &agr;4&bgr;2 receptors were inhibited by nitrous oxide by 39%, whereas &agr;4&bgr;4 receptors were inhibited by 7%. The inhibition of NMDA and nACh receptors by nitrous oxide was noncompetitive and was slightly different depending on membrane potentials for NMDA receptors, but not for nACh receptors. ConclusionsNitrous oxide and xenon displayed a similar spectrum of receptor actions, but this spectrum is distinct from that of isoflurane or ethanol. These results suggest that NMDA receptors and nACh receptors composed of &bgr;2 subunits are likely targets for nitrous oxide and xenon.

SUBUNIT- AND SITE-SPECIFIC PHARMACOLOGY OF THE NMDA RECEPTOR CHANNEL

N-Methyl-D-aspartate (NMDA) receptor channels play important roles in various physiological functions such as synaptic plasticity and synapse formation underlying memory, learning and formation of neural networks during development. They are also important for a variety of pathological states including acute and chronic neurological disorders, psychiatric disorders, and neuropathic pain syndromes. cDNA cloning has revealed the molecular diversity of NMDA receptor channels. The identification of multiple subunits with distinct distributions, properties and regulation, implies that NMDA receptor channels are heterogeneous in their pharmacological properties, depending on the brain region and the developmental stage. Furthermore, mutation studies have revealed a critical role for specific amino acid residues in certain subunits in determining the pharmacological properties of NMDA receptor channels. The molecular heterogeneity of NMDA receptor channels as well as their dual role in physiological and pathological functions makes it necessary to develop subunit- and site-specific drugs for precise and selective therapeutic intervention. This review summarizes from a molecular perspective the recent advances in our understanding of the pharmacological properties of NMDA receptor channels with specific references to agonists binding sites, channel pore regions, allosteric modulation sites for protons, polyamines, redox agents, Zn2+ and protein kinases, phosphatases.

Different sensitivities of NMDA receptor channel subtypes to non-competitive antagonists

Four kinds of heteromeric N-methyl-D-aspartate (NMDA) receptor channels, the epsilon 1/zeta 1, epsilon 2/zeta 1, epsilon 3/zeta 1 and epsilon 4/zeta 1 channels, were expressed in Xenopus oocytes and their sensitivities to various non-competitive antagonists were examined. The epsilon 1/zeta 1 and epsilon 2/zeta 1 channels were more sensitive to (+)MK-801 (dizocilpine) than the epsilon 3/zeta 1 and epsilon 4/zeta 1 channels, whereas the sensitivities to phencyclidine (PCP), ketamine and N-allylnormetazocine (SKF-10,047) were only slightly variable among the four epsilon/zeta channels. Furthermore, the replacement by glutamine or arginine of the conserved asparagine residue in segment M2 of the epsilon 2 and zeta 1 NMDA receptor channel subunits reduced the sensitivities to PCP, ketamine and SKF-10,047, though to different extents. These results, together with previous findings, suggest that these non-competitive antagonists as well as (+)MK-801 and Mg2+ act on a common site.

THE EFFECTIVENESS OF PREEMPTIVE ANALGESIA VARIES ACCORDING TO THE TYPE OF SURGERY: A RANDOMIZED, DOUBLE-BLIND STUDY

UNLABELLED The reliability of preemptive analgesia is controversial. Its effectiveness may vary among anatomical areas or surgical types. We evaluated preemptive analgesia by epidural morphine in six surgery types in a randomized, double-blind manner. Pain intensity was rated using a visual analog scale, a verbal report, and a measurement of postsurgical morphine consumption. Preemptive analgesia was effective in limb surgery and mastectomy, but ineffective for gastrectomy, hysterectomy, herniorrhaphy, and appendectomy. Relief of postsurgical pain in hemiorrhaphy was more rapid than that in the other surgery types. Preemptive analgesia was effective in limb surgery and mastectomy, but not in surgeries involving laparotomy, regardless of whether the surgery was major (gastrectomy and hysterectomy) or minor (herniorrhaphy and appendectomy). These results suggest that viscero-peritoneal nociception is involved in postsurgical pain. The abdominal viscera and peritoneum are innervated both heterosegmentally (in duplicate or triplicate by the vagus and/or phrenic nerves) and segmentally (by the spinal nerves). Therefore, supraspinal and/or cervical spinal neurons might be sensitized, despite the blockade of the segmental nerves with epidural morphine. The rapid retreat of the pain after hemiorrhaphy suggests that central sensitization remits soon after minor surgery, but that in appendicitis, it may be protracted by additional noxious stimuli, such as infection. IMPLICATIONS Epidural preemptive analgesia was reliably effective in limb and breast surgeries but ineffective in abdominal surgery, suggesting involvement of the brainstem and cervical spinal cord via the vagus and phlenic nerves.

Preemptive analgesia by intravenous low-dose ketamine and epidural morphine in gastrectomy: a randomized double-blind study.

Background Morphine and ketamine may prevent central sensitization during surgery and result in preemptive analgesia. The reliability of preemptive analgesia, however, is controversial. Methods Gastrectomy patients were given preemptive analgesia consisting of epidural morphine, intravenous low-dose ketamine, and combinations of these in a randomized, double-blind manner. Postsurgical pain intensity was rated by a visual analog scale, a categoric pain evaluation, and cumulative morphine consumption. Results Preemptive analgesia by epidural morphine and by intravenous low-dose ketamine were significantly effective but not definitive. With epidural morphine, a significant reduction in visual analog scale scores at rest was observed at 24 and 48 h, and morphine consumption was significantly lower at 6 and 12 h, compared with control values. With intravenous ketamine, visual analog scale scores at rest and morphine consumption were significantly lower at 6, 12, 24, and 48 h than those in control subjects. The combination of epidural morphine and intravenous ketamine provided definitive preemptive analgesia: Visual analog scale scores at rest and morphine consumption were significantly the lowest at 6, 12, 24, and 48 h, and the visual analog scale score during movement and the categoric pain score also were significantly the lowest among the groups. Conclusion The results suggest that for definitive preemptive analgesia, blockade of opioid and N-methyl-d-aspartate receptors is necessary for upper abdominal surgery such as gastrectomy; singly, either treatment provided significant, but not definitive, postsurgical pain relief. Epidural morphine may affect the spinal cord segmentally, whereas intravenous ketamine may block brain stem sensitization via the vagus nerve during upper abdominal surgery.

Subunit-dependent inhibition of human neuronal nicotinic acetylcholine receptors and other ligand-gated ion channels by dissociative anesthetics ketamine and dizocilpine.

Background: The neuronal mechanisms responsible for dissociative anesthesia remain controversial. N-methyl-D-aspartate (NMDA) receptors are inhibited by ketamine and related drugs at concentrations lower than those required for anesthetic effects. Thus, the authors studied whether ligand-gated ion channels other than NMDA receptors might display a sensitivity to ketamine and dizocilpine that is consistent with concentrations required for anesthesia. Methods: Heteromeric human neuronal nicotinic acetylcholine receptors (hnAChR channels &agr;2&bgr;2, &agr;2&bgr;4, &agr;3&bgr;2, &agr;3&bgr;4, &agr;4&bgr;2 and &agr;4&bgr;4), 5-hydroxytryptamine3 (5-HT3), &agr;1&bgr;2&ggr;2S&ggr;-aminobutyric acid type A (GABAA) and &agr;1 glycine receptors were expressed in Xenopus oocytes, and effects of ketamine and dizocilpine were studied using the two-electrode voltage-clamp technique. Results: Both ketamine and dizocilpine inhibited hnAChRs in a noncompetitive and voltage-dependent manner. Receptors containing &bgr;4 subunits were more sensitive to ketamine and dizocilpine than those containing &bgr;2 subunits. The inhibitor concentration for half-maximal response (IC50) values for ketamine of hnAChRs composed of &bgr;4 subunits were 9.5–29 &mgr;M, whereas those of &bgr;2subunits were 50–92 &mgr;M. Conversely, 5-HT3 receptors were inhibited only by concentrations of ketamine and dizocilpine higher than the anesthetic concentrations. This inhibition was mixed (competitive/noncompetitive). GABAA and glycine receptors were very resistant to dissociative anesthetics. Conclusions: Human nAChRs are inhibited by ketamine and dizocilpine at concentrations possibly achieved in vivo during anesthesia in a subunit-dependent manner, with &bgr; subunits being more critical than &agr; subunits. Conversely, 5-HT3, GABAA, and glycine receptors were relatively insensitive to dissociative anesthetics.

Involvement of the carboxyl-terminal region in modulation by TPA of the NMDA receptor channel.

The epsilon 1/zeta 1 and epsilon 2/zeta 1 heteromeric N-methyl-D-aspartate (NMDA) receptor channels expressed in Xenopus oocytes, but not the epsilon 3/zeta 1 and epsilon 4/zeta 1 channels, are positively modulated by the treatment with 12-O-tetradecanoyl phorbol 13-acetate (TPA). Failure of potentiation in the presence of staurosporine suggests the involvement of protein kinases in the TPA effect. To identify the structural domain involved in the modulation of the NMDA receptor channel by the TPA treatment, we constructed chimeric subunits between the epsilon 2 and epsilon 3 subunits. Functional analysis of heteromeric channels containing chimeric epsilon subunits has shown that the carboxyl-terminal region of the epsilon 2 subunit is responsible for the activation of the epsilon 2/zeta 1 channel by the TPA treatment.

Headache after attempted epidural block: the role of intrathecal air.

Background Postmeningeal puncture headache (PMPH) is typically attributed to the loss of cerebrospinal fluid (CSF). However, when it occurs after an attempted epidural puncture, it may be due to either CSF loss or, potentially, to the subarachnoid injection of air used as a part of “loss‐of‐resistance” testing. This study was performed to examine the relation between intrathecal air and PMPH. Methods Using a loss‐of‐resistance test with an air‐filled (n = 1,812; air group) or saline‐filled (n = 1,918; saline group) syringe, epidural block was performed in patients with acute or chronic pain. The dura was judged to be perforated not only when backflow of CSF was recognized in the needle but also when signs and symptoms solely attributable to meningeal perforation were seen, such as high spinal blockade or severe motor blockade. The incidence, onset time, and duration of PMPH in the air and saline groups were compared. In all patients with signs of meningeal perforation, brain computed tomography was examined. Results The incidence of PMPH in the air group (32 cases) was significantly higher than that in the saline group (5 cases), although the occurrences of meningeal perforation between the air (48 cases) and saline (51 cases) groups did not differ significantly. Intrathecal air bubbles were detected on brain computed tomography in both the deep supraspinal structures such as the ventricles, Silvian fissures and cisterns, and the superficial subarachnoid space in 30 of 32 patients with PMPH in the air group, whereas no intrathecal air bubbles were seen in the saline group. PMPH was significantly more rapid in onset and shorter in duration in the air group than that in the saline group. Conclusions The use of air for loss‐of‐resistance testing during epidural block was associated with a higher incidence of PMPH, which might be attributable to subarachnoid air injection and CSF leakage.

Effects of propofol on various AMPA-, kainate- and NMDA-selective glutamate receptor channels expressed in Xenopus oocytes☆

Effects of a general intravenous anesthetic 2,6-diisopropylphenol (propofol) on various glutamate receptor (GluR) channels were examined on the alpha 1 and alpha 1/alpha 2 GluR channels selective for alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), the beta 2/gamma 2 GluR channels selective for kainate, and the epsilon 2/zeta 1 and epsilon 3/zeta 1 N-methyl-D-aspartate (NMDA) receptor channels expressed in Xenopus oocytes. Propofol suppressed the current responses of the alpha 1/alpha 2, beta 2/gamma 2, epsilon 2/zeta 1 and epsilon 3/zeta 1 channels in a dose-dependent manner, whereas it enhanced the current responses of the alpha 1 channel. The extents of inhibition were in the order epsilon 2/zeta 1 > epsilon 3/zeta 1 > beta 2/gamma 2 > alpha 1/alpha 2 channels. During perfusion of 500 microM propofol, the alpha 1/alpha 2, beta 2/gamma 2, epsilon 2/zeta 1 and epsilon 3/zeta 1 channels were progressively suppressed. Furthermore, 10 min perfusion of 20 microM propofol inhibited the epsilon 2/zeta 1 channel by 24%. These results suggest that clinical concentrations (approximately 35 microM) of propofol suppress the NMDA receptor channels slightly.

Action of dexmedetomidine on the substantia gelatinosa neurons of the rat spinal cord

Dexmedetomidine is a highly specific, potent and selective α2‐adrenoceptor agonist. Although intrathecal and epidural administration of dexmedetomidine has been found to produce analgesia, whether this analgesia results from an effect on spinal cord substantia gelatinosa (SG) neurons remains unclear. Here, we investigated the effects of dexmedetomidine on postsynaptic transmission in SG neurons of rat spinal cord slices using the whole‐cell patch‐clamp technique. In 92% of the SG neurons examined (n = 84), bath‐applied dexmedetomidine induced outward currents at −70 mV in a concentration‐dependent manner, with the value of effective concentration producing a half‐maximal response (0.62 μm). The outward currents induced by dexmedetomidine were suppressed by the α2‐adrenoceptor antagonist yohimbine, but not by prazosin, an α1‐, α2B‐ and α2C‐adrenoceptor antagonist. Moreover, the dexmedetomidine‐induced currents were partially suppressed by the α2C‐adrenoceptor antagonist JP‐1302, while simultaneous application of JP‐1302 and the α2A‐adrenoceptor antagonist BRL44408 abolished the current completely. The action of dexmedetomidine was mimicked by the α2A‐adrenoceptor agonist oxymetazoline. Plots of the current–voltage relationship revealed a reversal potential at around −86 mV. Dexmedetomidine‐induced currents were blocked by the addition of GDP‐β‐S [guanosine‐5′‐O‐(2‐thiodiphosphate)] or Cs+ to the pipette solution. These findings suggest that dexmedetomidine hyperpolarizes the membrane potentials of SG neurons by G‐protein‐mediated activation of K+ channels through α2A‐ and α2C‐adrenoceptors. This action of dexmedetomidine might contribute, at least in part, to its antinociceptive action in the spinal cord.