Pharmacokinetic Study on Excretion of Inorganic Fluoride Ion, a Metabolite of Sevoflurane

Blood and urinary inorganic fluoride ion concentration was determined in six healthy volunteers after inhalation of 2% sevoflurane for one hour. The serum inorganic fluoride ion concentration increased 30 min after discontinuation of inhalation to 14.8 ± 3.0 μmol/liter, which was about 10 times higher than the level before inhalation. The serum elimination constant of inorganic fluoride was calculated to be 0.000467 and the half-life was 1,487 min. The urinary excretion rate of inorganic fluoride ion was the highest ( 452 nmol/min) after 12-24 hr. The urinary excretion rate constant of inorganic fluoride was calculated to be 0.000268 and the half-life was 2,583 min. The distribution volume of inorganic fluoride excreted in the urine was calculated to be 127 liters. This value showed that fluoride ion produced in the cell cannot readily pass through the cell membrane due to its polarity, resulting in a delay of the maximum excretion rate of inorganic fluoride until the first or second day after inhalation of the anesthetic.This study was supported in part by a Grant-in-aid for Science Research from the Ministry of Education, Science and Culture of Japan and a Grant-in-aid from the Association for the Advancement of Medicine of the Tsuchiya Foundation

Urinary Excretion of Inorganic and Organic Fluoride after Inhalation of Sevoflurane

This study was designed to investigate the defluorination of sevoflurane in patients. Five patients, scheduled for orthopedic surgery, were administered sevoflurane for 60 min during NLA-nitrous oxide-oxygen. The end-tidal concentration of sevoflurane was adjusted at 0.6% throughout the entire inhalation period. The serum concentration of inorganic F- increased significantly 15 min after the onset of inhalation and reached a plateau at 45 min with a mean value about 15 μM. The serum organic fluoride level increased significantly 45 min after the onset of inhalation and did not change significantly 4 hr later with a mean value of about 140 μM. The elimination half-lives and rate constants were calculated from urinary data to be 2040 min and 0.00034 for inorganic fluoride and 1800 min and 0.00038 for organic fluoride respectively. The ratio organic/inorganic fluoride was calculated to be 2.3.This study was supported in part by a Grant-in-aid for Science Research from the Ministry of Education, Science and Culture of Japan

Clinical Evaluation and Metabolism of Sevoflurane in Patients

Sevoflurane was submitted to Phase II studies in patients following Phase I studies. Sevoflurane, 2% inspired during maintenance, was administered with 50% N2O in oxygen to produce surgical anesthesia in 9 orthopedic patients of ASA Physical Status I. Under controlled ventilation, endotracheal concentration of sevoflurane was recorded. The blood concentration of sevoflurane was measured during and after the inhalation. Serum, urinary inorganic fluoride, and glucuronide of hexafluoroisopropanol were analysed with ion chromatographic analyzer. The patient inhaled sevoflurane for 3.5 ± 1.6 hr. All the patients were anesthetized and operated uneventfully. Postoperative laboratory findings showed no unexplainable abnormality. The end expiratory concentration of sevoflurane reached a plateau in 4.0 ± 0.8 min and fell rapidly after discontinuation of sevoflurane. Blood concentration of sevoflurane was about 500 μM during inhalation. It decreased promptly after termination of sevoflurane and was not correlated with anesthetic time. The time for verbal response after discontinuation was 11.8 ± 4.2 min. The serum concentration of inorganic fluoride increased after inhalation and reached a plateau (13.7 ± 8.2 μM) in 120 min. The level lasted for 120 min after anesthesia and fell by half at 12 hr after anesthesia. Urinary fluoride concentration varied from 20 to 3,000 μM during the first 12 hr urine, and showed its maximum in the first postoperative 12 or 24 hr urine. The findings that sevoflurane with nitrous oxide and oxygen produced surgical anesthesia without any sequelae and that the serum fluoride level did not exceed the nephrotoxic level warrent the further clinical evaluation in a wider range of subjects.A part of this work was supported by a Research Grant from the Japanese Ministry of Education, Science and Culture and presented at the 8th European Congress of Anaesthesiology, Vienna, Austria, in September, 1986

Genetic screening for malignant hyperthermia and comparison of clinical symptoms in Japan

Malignant hyperthermia (MH) is an anaesthetic complication that causes an abnormal hypermetabolic state. RYR1 encoding ryanodine receptors of the sarcoplasmic reticulum and CACNA1S encoding α subunits of dihydropyridine receptors are known to be associated with MH pathogenicity. We performed genetic screening using next-generation sequencing to evaluate the prevalence of genes associated with MH pathogenicity and clinical symptoms. This was a retrospective cohort study wherein next-generation sequencing data of 77 families diagnosed with MH predisposition by calcium-induced calcium release (CICR) tests from 1995 to 2019 was used to search for RYR1 and CACNA1S variants. Furthermore, the clinical symptoms and predisposition tests in participants with RYR1 and CACNA1S variants were compared. In the 77 families, 44.2%, 7.8%, and 48.1% individuals had RYR1, CACNA1S, and neither RYR1 nor CACNA1S variants, respectively. Clinically significant differences were found in the maximum body temperature, maximum elevated body temperature for 15 min, creatinine kinase level, and CICR rate between the RYR1 and CACNA1S groups. The prevalence of pathogenic CACNA1S variants appears to be prominent in Japan. The severity of clinical symptoms and the CICR rate were greater in individuals with RYR1 variants than in those with CACNA1S variants, likely due to more direct regulation of calcium levels by ryanodine receptors than by dihydropyridine receptors. Genetic analysis of MH in future studies may help identify other genes associated with MH, which will further clarify the relationship between genotypes and MH symptoms and contribute to safer anaesthesia practice.This study was supported by a Grant-in-Aid for Young Scientists (grant number: 17K16733 to Y.N. and 20K17783 to R.K.) from the Japan Society for the Promotion of Science and by the Takeda Science Foundation (H.K.)

Genetic Analysis with Calcium-induced Calcium Release Test in Japanese Malignant Hyperthermia Susceptible (MHS) Families

Some genetic studies have shown a linkage between malignant hyperthermia susceptibility (MHS) and chromosome 19q or the skeletal muscle ryanodine receptor (RYR1) gene. Some types of MHS seem to be caused by an abnormality of calcium-induced calcium release (CICR). We analyzed the linkage of RYR 1 gene polymorphisms in Japanese MHS families and investigated the correlation between genetic evidence of RYR1 gene mutations and an accelerated rate of CICR.　　 We studied 63 subjects who were referred to our institute for investigation of MHS. CICR rates were measured by the skinned fiber method in 23 subjects. DNA samples were collected from 63 individuals belonging to 22 unrelated families. Restriction fragment length polymorphism (RFLP) analyses on the RYR1 locus and hypervariable microsatellite analysis were performed.　　 We found one family with a linkage between acceleration of the CICR mechanism and a group of RFLPs. In CICR tests, ten of the 11 patients who had presented with fulminant MH showed accelerated rates of CICR. Analysis for the mutation C1840T, which was performed in 63 samples, did not demonstrate an alteration in any of the patients. Although we found heterozygotes in RFLP studies, we did not recognize a specific relationship between the acceleration of CICR and the RFLPs.　　 We suggest a linkage between the acceleration of CICR and an abnormal human RYR1 gene in MHS. These results also suggest that heterogeneity exists for MH. We conclude that genetic tests cannot replace CICR tests or caffeine-halothane contracture tests with muscle biopsy as a diagnosing test for MH in the near future.This work was partly supported by grants-in-aid from the Ministry of Education, Science and Culture of Japan (No.08407052: Osafumi Yuge, No.09771157: Yasuhiro Maehara) and by a research grant from the Labor Welfare Corporation of Japan

Remimazolam-based total intravenous anesthesia in a patient with a confirmed diagnosis of malignant hyperthermia: a case report

Abstract Background Malignant hyperthermia (MH) is a rare, life-threatening disorder of calcium homeostasis in skeletal muscle cells that is triggered by volatile anesthetics and succinylcholine, leading to a hypermetabolic reaction. The pathogenic ryanodine receptor 1 (RYR1) gene variant is critical. Patients susceptible to MH should avoid triggering agents, and total intravenous anesthesia (TIVA) is preferred. Remimazolam is safe in patients with suspected MH. Case presentation We present the first case of remimazolam treatment in a genetically confirmed patient with MH without MH development. A 72-year-old man with a family history of MH underwent remimazolam-based TIVA. After informed consent was obtained, a muscle biopsy and genetic testing were performed. Intraoperatively and postoperatively, the patient exhibited no signs of MH. An enhanced function of the RYR1 channel into releasing calcium was indicated, and the genetic testing revealed a pathogenic variant of RYR1. Conclusions Remimazolam-based TIVA is safe in patients confirming the diagnosis of MH

Effects of Remimazolam and Propofol on Ca2+ Regulation by Ryanodine Receptor 1 with Malignant Hyperthermia Mutation

Background. We investigated the potential safety of remimazolam and propofol in malignant hyperthermia- (HM-) susceptible patients using ryanodine receptor 1- (RYR1-) expressing human embryonic kidney- (HEK-) 293 cells. Methods. We compared the enhanced responsiveness of HEK-293 cells expressing wild-type RYR1 with that of mutant RYR1 to caffeine following perfusion with remimazolam or propofol. Furthermore, we investigated whether RYR1 enhanced the responsiveness of cells to remimazolam or propofol and compared the median effective concentration (EC50; i.e., the concentration required to reach half-maximal activation) using an unpaired two-tailed t-test while a P<0.05 was considered significant. Results. Remimazolam and propofol did not promote the caffeine-induced increase in intracellular Ca2+ levels in HEK-293 cells expressing mutant RYR1 even with exposure to approximately 100-fold the clinically used concentration. In wild-type RYR1, EC50 values of remimazolam following refusion vs. nonperfusion were 2.86 mM vs. 2.75 mM (P=0.76) while for propofol perfusion vs. nonperfusion, they were 2.76 mM vs. 2.75 mM, respectively (P=0.83). In mutant RYR1, EC50 values of remimazolam refusion vs. nonperfusion were 1.58 mM vs. 1.71 mM, respectively (P=0.63) while for propofol perfusion vs. nonperfusion, they were 1.65 mM vs. 1.71 mM, respectively (P=0.73). Remimazolam and propofol increased intracellular Ca2+ levels in a concentration-dependent manner, but the effect was not enhanced by RYR1. EC50 values of remimazolam with non-RYR1 vs. wild-type RYR1 were 1.00 mM vs. 0.92 mM, respectively (P=0.91) while those of propofol were 1.09 mM vs. 1.05 mM, respectively (P=0.84). Conclusions. The increase in intracellular Ca2+ concentration caused by remimazolam or propofol was not considered an RYR1-mediated reaction. We conclude that remimazolam and propofol can be safely used as an anesthetic in MH-susceptible patients with RYR1-mutation without causing MH and may be safely substituted for an MH-triggering anesthetic when RYR1-mediated MH occurs