Munehiro Shiraishi

Effects of Anesthetics on Mutant N-Methyl-d-Aspartate Receptors Expressed in Xenopus Oocytes

Alcohols, inhaled anesthetics, and some injectable anesthetics inhibit the function of N-methyl-d-aspartate (NMDA) receptors, but the mechanisms responsible for this inhibition are not fully understood. Recently, it was shown that ethanol inhibition of NMDA receptors was reduced by mutation of residues in the transmembrane (TM) segment 3 of the NR1 subunit (F639A) or in TM4 of the NR2A subunit (A825W), suggesting putative ethanol binding sites. We hypothesized that the actions of other anesthetics might also require these amino acids and evaluated the effects of anesthetics on the NMDA receptors expressed in Xenopus oocytes with two-electrode voltage-clamp recording. Effects of hexanol, octanol, isoflurane, halothane, chloroform, cyclopropane, 1-chloro-1,2,2-trifluorocyclobutane, and xenon were reduced or eliminated in the mutant NMDA receptors, whereas the inhibitory effects of nitrous oxide, ketamine, and benzene were not affected by these mutations. Rapid applications of glutamate and glycine by a T-tube device provided activation time constants, which suggested different properties of ketamine and isoflurane inhibition. Thus, amino acids in TM3 and TM4 are important for the actions of many anesthetics, but nitrous oxide, benzene, and ketamine seem to have distinct mechanisms for inhibition of the NMDA receptors.

The inhibitory effects of tramadol on muscarinic receptor-induced responses in Xenopus oocytes expressing cloned M(3) receptors.

Tramadol is a widely used analgesic, but its mechanism of action is not completely understood. Muscarinic receptors are involved in neuronal function in the brain and autonomic nervous system, and much attention has been paid to these receptors as targets of analgesic drugs in the central nervous system. In this study, we investigated the effects of tramadol on type-3 muscarinic (M3) receptors using the Xenopus oocyte expression system. Tramadol (10 nM–100 &mgr;M) inhibited acetylcholine-induced currents in oocytes expressing M3 receptor. Although GF109203X, a protein kinase C inhibitor, increased the basal current, it had little effect on the inhibition of acetylcholine-induced currents by tramadol. Moreover, tramadol inhibited the specific binding sites of [3H]quinuclidinyl benzilate. These findings suggest that tramadol at clinically relevant concentrations inhibits M3 function via quinuclidinyl benzilate-binding sites. This may explain the modulation of neuronal function and the anticholinergic effects of tramadol.

Gargling with sodium azulene sulfonate reduces the postoperative sore throat after intubation of the trachea

Postoperative sore throat (POST) is a complication that remains to be resolved in patients undergoing endotracheal intubation. In this study, we investigated whether preoperative gargling with sodium 1,4-dimethyl-7-isopropylazulene-3-sulfonate monohydrate (sodium azulene sulfonate, Azunol) reduces POST after endotracheal intubation. Forty patients scheduled for elective surgery under general anesthesia were randomized into Azunol and control groups. In the Azunol group, patients gargled with 4 mg Azunol diluted with 100 mL tap water (40 &mgr;g/mL). In the control group, patients gargled with 100 mL of tap water. After emergence from general anesthesia, the patients with POST were counted and POST was evaluated using a verbal analog pain scale. There were no significant differences between the two groups by age, height, body weight, gender distribution, or duration of anesthesia and surgery. In the control group, 13 patients (65%) complained of POST, which remained 24 h later in nine patients (45%). In the Azunol group, five patients (25%) also complained of POST, which completely disappeared by 24 h later. The incidence of POST and verbal analog pain scale scores in the Azunol group decreased significantly compared with the control group. We demonstrated that gargling with Azunol effectively attenuated POST with no adverse reactions.

The inhibitory effects of tramadol on 5-hydroxytryptamine type 2C receptors expressed in Xenopus oocytes

Although tramadol is widely available as an analgesic, its mechanism of antinociception remains unresolved. Serotonin (5-hydroxytryptamine, 5-HT) is a monoaminergic neurotransmitter that modulates numerous sensory, motor, and behavioral processes. The 5-HT type 2C receptor (5-HT2CR) is one of the major 5-HT receptor subtypes and is implicated in many important effects of 5-HT, including pain, feeding, and locomotion. In this study, we used a whole-cell voltage clamp to examine the effects of tramadol on 5-HT-induced Ca2+-activated Cl− currents mediated by 5-HT2CR expressed in Xenopus oocytes. Tramadol inhibited 5-HT-induced Cl− currents at pharmacologically relevant concentrations. The protein kinase C (PKC) inhibitor, bisindolylmaleimide I (GF109203×), did not abolish the inhibitory effects of tramadol on the 5-HT2CR-mediated events. We also studied the effects of tramadol on [3H]5-HT binding to 5-HT2CR expressed in Xenopus oocytes, and found that it inhibited the specific binding of [3H]5-HT to 5-HT2CR. Scatchard analysis of [3H]5-HT binding revealed that tramadol altered the apparent dissociation constant for binding without changing maximal binding, indicating competitive inhibition. The results suggest that tramadol inhibits 5-HT2CR function, and the mechanism of this inhibitory effect seems to involve competitive displacement of the 5-HT binding to the 5-HT2CR, rather than via activation of the PKC pathway.

Inhibitory effects of tramadol on nicotinic acetylcholine receptors in adrenal chromaffin cells and in Xenopus oocytes expressing α7 receptors

Tramadol has been used clinically as an analgesic; however, the mechanism of its analgesic effects is still unknown. We used bovine adrenal chromaffin cells to investigate effects of tramadol on catecholamine secretion, nicotine‐induced cytosolic Ca2+ concentration ([Ca2+]i) increases and membrane current changes. We also investigated effects of tramadol on α7 nicotinic acetylcholine receptors (AChRs) expressed in Xenopus oocytes. Tramadol concentration‐dependently suppressed carbachol‐induced catecholamine secretion to 60% and 27% of the control at the concentration of 10 and 100 μM, respectively, whereas it had little effect on veratridine‐ or high K+‐induced catecholamine secretion. Tramadol also suppressed nicotine‐induced ([Ca2+]i) increases in a concentration‐dependent manner. Tramadol inhibited nicotine‐induced inward currents, and the inhibition was unaffected by the opioid receptor antagonist naloxone. Tramadol inhibited nicotinic currents carried by α7 receptors expressed in Xenopus oocytes. Tramadol inhibited both α‐bungarotoxin‐sensitive and ‐insensitive nicotinic currents in bovine adrenal chromaffin cells. In conclusion, tramadol inhibits catecholamine secretion partly by inhibiting nicotinic AChR functions in a naloxone‐insensitive manner and α7 receptors are one of those inhibited by tramadol.

Inhibitory effects of nicorandil on rat mesangial cell proliferation via the protein kinase G pathway.

We investigated the effects of nicorandil, which is a hybrid between a nitrate and an ATP-sensitive potassium channel (KATP) opener, on cultured rat mesangial cell proliferation. Nicorandil (1 µM to 1 mM inhibited [3H]thymidine incorporation into rat mesangial cells in a concentration-dependent manner. Nicorandil (1 µM to 1 mM) also inhibited the number of cells. Nicorandil increased cyclic guanosine 3′,5′-cyclic monophosphate accumulation in mesangial cells. A protein kinase G inhibitor, KT5823, partially eliminated the inhibition of mesangial cell proliferation by nicorandil. Methylene blue, a guanylate cyclase inhibitor, blocked the inhibitory effect of nicorandil on mesangial cell proliferation. We also examined the effects of KATP mediators. Cromakalim, a KATP activator, and glibenclamide, a KATP inhibitor, had little effect on the proliferation of mesangial cells. These results suggest that the inhibitory effects of nicorandil on mesangial cell proliferation are mediated via the protein kinase G pathway.

The effects of the tramadol metabolite O-desmethyl tramadol on muscarinic receptor-induced responses in Xenopus oocytes expressing cloned M1 or M3 receptors.

O-desmethyl tramadol is one of the main metabolites of tramadol. It has been widely used clinically and has analgesic activity. Muscarinic receptors are involved in neuronal functions in the brain and autonomic nervous system, and much attention has been paid to these receptors as targets for analgesic drugs in the central nervous system. We have reported that tramadol inhibits the function of type-1 muscarinic (M1) receptors and type-3 muscarinic (M3) receptors, suggesting that muscarinic receptors are sites of action of tramadol. However, the effects of O-desmethyl tramadol on muscarinic receptor functions have not been studied in detail. In this study, we investigated the effects of O-desmethyl tramadol on M1 and M3 receptors, using the Xenopus oocyte expression system. O-desmethyl tramadol (0.1–100 &mgr;M) inhibited acetylcholine (ACh)-induced currents in oocytes expressing the M1 receptors (half-maximal inhibitory concentration [IC50] = 2 ± 0.6 &mgr;M), whereas it did not suppress ACh-induced currents in oocytes expressing the M3 receptor. Although GF109203X, a protein kinase C inhibitor, increased the ACh-induced current, it had little effect on the inhibition of ACh-induced currents by O-desmethyl tramadol in oocytes expressing M1 receptors. The inhibitory effect of O-desmethyl tramadol on M1 receptor was overcome when the concentration of ACh was increased (KD with O-desmethyl tramadol = 0.3 &mgr;M). O-desmethyl tramadol inhibited the specific binding of [3H]quinuclidinyl benzilate ([3H]QNB) to the oocytes expressed M1 receptors (IC50 = 10.1 ± 0.1 &mgr;M), whereas it did not suppress the specific binding of [3H]QNB to the oocytes expressed M3 receptors. Based on these results, O-desmethyl tramadol inhibits functions of M1 receptors but has little effect on those of M3 receptors. This study demonstrates the molecular action of O-desmethyl tramadol on the receptors and may help to explain its neural function.

Effects of the Intravenously Administered Anaesthetics Ketamine, Propofol, and Thiamylal on the Cortical Renal Blood Flow in Rats

Intravenous anaesthetics such as ketamine, propofol, and thiamylal are widely used, although the direct effects of these anaesthetics on the renal blood flow (RBF) have not been well elucidated. In this study, we examined the effects of bolus and continuous administrations of ketamine, propofol, and thiamylal on cortical RBF and the effects of noradrenaline (NA) on RBF under continuous administration of these anaesthetics. We used laser Doppler flowmetry to measure the effects of bolus injection and continuous infusion of ketamine, propofol, and thiamylal on cortical RBF in male Wistar rats. We also examined the effects of the anaesthetics on mean arterial blood pressure (MAP) and heart rate (HR). Bolus injections of ketamine, propofol, or thiamylal (1–8 mg/kg each, n = 10) at clinically relevant concentrations did not affect MAP, HR, or RBF. Continuous administration of ketamine, propofol, or thiamylal (1–8 mg/kg/h each, n = 10) did not affect MAP, HR or RBF. Exogenous NA (2 µg/kg) caused an increase in MAP and a decrease in RBF and HR. In experiments with continuous infusions of propofol or thiamylal (1–8 mg/kg/h each, n = 10), similar results were observed without infusion of any anaesthetics. However, bolus injection of NA did not result in a decrease in RBF during continuous ketamine infusion (98.8 ± 6.7% of control, n = 6, p < 0.05), while ketamine did not affect the NA-induced increase in MAP. In conclusion, bolus and continuous administrations of ketamine, propofol, and thiamylal did not affect the RBF. From our present findings, ketamine would be useful for maintaining the RBF.

The Prolonged Effect of a Muscle Relaxant in a Patient with Chronic Inflammatory Demyelinating Polyradiculoneuropathy

I n 1975, Dyck et al. (1) first described chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). This disease is characterized as a predominantly symmetric sensorimotor neuropathy with a relapsing, chronic monophasic, or steadily progressive course, without the presence of an associated causal disease (2). Patients with this disease often have symptoms such as paresthesia and weakness of the proximal and distal muscles, including the respiratory muscles. No reports mention the anesthetic management of patients with CIPD. We found that vecuronium had a prolonged effect in a patient with CIDP during general anesthesia. In this report, we discuss the problems of anesthetic management for patients with CIDP.

The Tramadol Metabolite, O-Desmethyl Tramadol, Inhibits 5-Hydroxytryptamine Type 2C Receptors Expressed in Xenopus Oocytes

Purpose: Tramadol is widely used clinically as an analgesic, yet the mechanism by which it produces antinociception remains unclear. O-Desmethyl tramadol, the main metabolite of tramadol, is a more potent analgesic than tramadol. We reported previously that tramadol inhibits the 5-hydroxytryptamine (5-HT) type 2C receptor (5-HT2CR), a G-protein-coupled receptor that is expressed widely within brain and that mediates several effects of 5-HT, including nociception, feeding, and locomotion. The effects of O-desmethyl tramadol on 5-HT2CR have not been studied. In this study, we investigated the effect of O-desmethyl tramadol on 5-HT2CR expressed in Xenopus oocytes. Methods: We examined the effect of O-desmethyl tramadol on 5-HT2CR using the Xenopus oocyte expression system. Furthermore, we investigated the effects of O-desmethyl tramadol on the binding of [3H]5-HT by 5-HT2CR. Results: O-Desmethyl tramadol, at pharmacologically relevant concentrations, inhibited 5-HT-evoked Ca2+-activated Cl– currents in oocytes that expressed 5-HT2CR. The inhibitory effect of O-desmethyl tramadol on 5-HT2CR was overcome at higher concentrations of 5-HT. Bisindolylmaleimide I (GF109203X), a protein kinase C inhibitor, increased 5-HT-evoked currents but had little effect on the inhibition of 5-HT-evoked currents by O-desmethyl tramadol. O-Desmethyl tramadol inhibited the specific binding of [3H]5-HT by 5-HT2CR expressed in oocytes. O-Desmethyl tramadol altered the apparent dissociation constant for binding of [3H]5-HT by 5-HT2CR without changing maximum binding, which indicated competitive inhibition. Conclusion: These results suggest that O-desmethyl tramadol inhibits 5-HT2CR, which provides further insight into the pharmacological properties of tramadol and O-desmethyl tramadol.