Studies on the mechanism of general anesthesia.

Inhaled anesthetics are a chemically diverse collection of hydrophobic molecules that robustly activate TWIK-related K+ channels (TREK-1) and reversibly induce loss of consciousness. For 100 y, anesthetics were speculated to target cellular membranes, yet no plausible mechanism emerged to explain a membrane effect on ion channels. Here we show that inhaled anesthetics (chloroform and isoflurane) activate TREK-1 through disruption of phospholipase D2 (PLD2) localization to lipid rafts and subsequent production of signaling lipid phosphatidic acid (PA). Catalytically dead PLD2 robustly blocks anesthetic TREK-1 currents in whole-cell patch-clamp recordings. Localization of PLD2 renders the TRAAK channel sensitive, a channel that is otherwise anesthetic insensitive. General anesthetics, such as chloroform, isoflurane, diethyl ether, xenon, and propofol, disrupt lipid rafts and activate PLD2. In the whole brain of flies, anesthesia disrupts rafts and PLDnull flies resist anesthesia. Our results establish a membrane-mediated target of inhaled anesthesia and suggest PA helps set thresholds of anesthetic sensitivity in vivo.

Global optimization of the Michaelis-Menten parameters using physiologically-based pharmacokinetic (PBPK) modeling and chloroform vapor uptake data in F344 rats.

Objective: To quantify metabolism, a physiologically based pharmacokinetic (PBPK) model for a volatile compound can be calibrated with the closed chamber (i.e. vapor uptake) inhalation data. Here, we introduce global optimization as a novel component of the predictive process and use it to illustrate a procedure for metabolic parameter estimation.Materials and methods: Male F344 rats were exposed in vapor uptake chambers to initial concentrations of 100, 500, 1000, and 3000 ppm chloroform. Chamber time-course data from these experiments, in combination with optimization using a chemical-specific PBPK model, were used to estimate Michaelis-Menten metabolic constants. Matlab® simulation software was used to integrate the mass balance equations and to perform the global optimizations using MEIGO (MEtaheuristics for systems biology and bIoinformatics Global Optimization - Version 64 bit, R2016A), a toolbox written for Matlab®. The cost function used the chamber time-course data and least squares to minimize the difference between data and simulation values.Results and discussion: The final values estimated for Vmax (maximum metabolic rate) and Km (affinity constant) were 1.2 mg/h and a range between 0.0005 and 0.6 mg/L, respectively. Also, cost function plots were used to analyze the dose-dependent capacity to estimate Vmax and Km within the experimental range used. Sensitivity analysis was used to assess identifiability for both parameters and show these kinetic data may not be sufficient to identify Km.Conclusion: In summary, this work should help toxicologists interested in optimization techniques understand the overall process employed when calibrating metabolic parameters in a PBPK model with inhalation data.

Pharmacological Characterization and Raman Spectroscopy Evaluation of Oral and Maxillofacial Surgery-Related Carnoy´S Solution Modified by Different Viscosity Agents.

BACKGROUND: There are several lesions of odontogenic and non-odontogenic origin in the oral cavity, such as odontogenic keratocyst, as well as many treatment options for such lesions. In order to reduce recurrence due to conservative treatments and less aesthetic and functional impairment of the patient (radical therapies), Carnoy's solution has been used as an adjuvant to surgery, showing satisfactory results. Its application is not standardized, presenting risks to adjacent tissues. Thus, we characterized the Carnoy's solution with different viscosity agents to enhance its applicability. MATERIAL AND METHODS: All solutions prepared (Carnoy with and without chloroform) were added with viscosity agent: ethyl cellulose, propylene glycol, and glycerol totaling eight solutions. The pharmacological characterization of the solutions was performed by determining the mass density and relative density (using a clean and dry pycnometer), pH (using pH meter), and concentration of Fe3+ (using ultraviolet/visible spectroscopy). The analyses of the inorganic components were determined by Raman micro spectrometry. Data were analyzed with statistical program BIOESTAT 5.3. RESULTS: Solutions with ethyl cellulose were discarded due to precipitate formation and suspension of the viscosity agent. In the other solutions, viscosity increase (propylene glycol solutions) and acidic pH were observed mainly in the glycerol group. The ferric chloride characterized as a hemostatic agent had its concentration increased with the use of thickening agents, theoretically favoring its action. CONCLUSION: The similarity of the propylene glycol and glycerol molecules justifies the Raman spectra of these substances to be similar and the difficulty in obtaining a "fingerprint".
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Chloroform and desflurane immobilization with recovery of viable Drosophila larvae for confocal imaging.

Imaging of living, intact Drosophila larvae is challenged if normal bodily function must be observed or when healthy larvae must be recovered for subsequent studies. Here, we describe a simple and short protocol that employs transient airborne chloroform or desflurane (1,2,2,2-tetrafluoroethyl difluoromethyl ether) to efficiently immobilize larvae without the use of manipulation devices, vaporizers or imaging chambers. This non-lethal method allows the use of anesthetics while allowing tracking of individual Drosophila into adulthood for follow-up experiments. At dosages sufficient to immobilize larvae, Desflurane, but not chloroform reduced the central nervous system response to auditory stimulus. Desflurane doses were sufficient to arrest the heart, however significant rapid recovery was observed. With our method, chloroform provided more rapid anesthesia but slower recovery than Desflurane. Without specialized hardware, this technique allows for repeated imaging of living Drosophila larvae.

Assessment of indirect inhalation exposure to formaldehyde evaporated from water.

Volatilization volumes and health risks associated with indirect inhalation exposure to formaldehyde evaporated from water have not been investigated quantitatively. We experimentally investigated formaldehyde volatility, compared with chloroform volatility, predicted formaldehyde inhalation exposure concentrations in Japanese bathrooms, and then re-evaluated drinking water quality standards. Although the Henry's law constant of formaldehyde is 1/104 that of chloroform, with a 30-min exposure period, the formaldehyde non-equilibrium partition coefficient (K'd) was 1/500th the chloroform value because of formaldehyde's faster volatilization rate. We used this ratio to estimate the cumulative probability distribution of formaldehyde concentrations in bathroom air. For a formaldehyde concentration in water of ≤2.6 mg/L-water (WHO tolerable concentration), the probability that the incremental formaldehyde concentration due to volatilization would exceed 100 µg/m3-air (WHO indoor air quality guideline) was low. However, major sources of formaldehyde in indoor air are building materials and furniture. We therefore calculated the allowable concentration in water by allocating a small percentage of the indoor air guideline value to indirect inhalation exposure via volatilization from tap water. With an allocation factor of 20% (10%), the allowable concentration was 0.52 (0.26) mg/L-water. These concentrations are similar to the Health Canada guideline concentration but they are 3-6 times the Japanese water quality standard.

A Newly Discovered Manuscript of Charles T. Jackson, MD, on the Preparation and Administration of Anesthetics for Humans and Animals.

A newly discovered handwritten manuscript of Charles T. Jackson, MD, contains instructions for the preparation and administration of sulfuric ether, information on Jackson's preferred mixture of ether and chloroform, an account of his experiments with other potential anesthetic agents, and his comments on etherizing cattle and other animals. Jackson's nine-page manuscript is believed to have been written in the autumn of 1851, around the time that he submitted his memorial on the discovery of etherization to Baron von Humboldt, and made a separate submission to the US Congress.

The Perfect and Famous Anesthetic Known as Methyl in Boston in 1895.

Extravagant claims were made for proprietary dental anesthetics in Boston, MA, in the late 1800s. For instance, in 1883, Urial K. Mayo introduced an inhaled Vegetable Anaesthetic comprised of nitrous oxide that had been uselessly pretreated with botanical material. This misguided concept may have been inspired by homeopathy, but it was also in line with the earlier false belief of Elton R. Smilie, Charles T. Jackson, and William T.G. Morton that sulfuric ether could volatilize opium at room temperature. In 1895, the Dental Methyl Company advertised an agent they called Methyl, a supposedly perfect topical anesthetic for painless dental extraction. The active ingredient was probably chloroform. Anesthetic humbug did not cease in Boston on Ether Day of October 16, 1846.

Hazards of Improper Dispensary: Literature Review and Report of an Accidental Chloroform Injection.

Several clear, transparent solutions are used in endodontics. Inappropriate dispensing methods can lead to accidental injection or accidental irrigation. These accidents can cause permanent tissue damage including damage to the bone, periodontium, nerves, and vasculature. This article reports on the consequences of an accidental chloroform injection. Nonsurgical retreatment of tooth #8 was planned as part of a restorative treatment plan in a 69-year-old woman. The dentist accidentally injected chloroform instead of local anesthesia because chloroform was loaded into the anesthetic syringe. The patient experienced severe pain and swelling and soft tissue necrosis and suffered permanent sensory and motor nerve damage. A review of the literature was performed on accidents caused by improper dispensary, namely accidental injections and accidental irrigations. The data were extracted and summarized. Sodium hypochlorite, chlorhexidine, formalin, formocresol, 1:1000 adrenaline, benzalkonium chloride, and lighter fuel were accidentally injected as an intraoral nerve block or as infiltration injections. Bone and soft tissue necrosis, tooth loss, and sensory nerve damage (anesthesia and paresthesia) were the most common consequences reported. Such disastrous events can be prevented by appropriate labeling and separate dispensing methods for each solution. There is a need for disseminating information on toxicity and biocompatibility of materials/solutions used in endodontics. The authors recommend training dental students and endodontic residents on immediate and long-term therapeutic management of patients when an accidental injection or accidental irrigation occurs.

Anesthetized by chloroform before hanging.

We present a unique case of suicidal hanging. The deceased was a 31-year-old male who was found hanging from a tree in a dense thicket, with his lower limbs in contact with the ground (partial suspension). There was an apparatus similar to a facial mask placed around his nose and mouth. A strong chemical smell was emanating from the apparatus, which was identified as chloroform (Formyl trichloride/CHCl3). A ligature with a soft cloth beneath it was around his neck. A ligature mark was present around the neck. The decedent's blood alcohol levels were 112 mg/dl. The blood and stomach contents were negative for chloroform. A complete death investigation, including scene investigation and complete autopsy examination, confirmed the cause of death as hanging. The manner of death was suicide. This case highlights how the deceased had used several methods whilst committing suicide to minimize pain, including the inhalation of chloroform, which would have also resulted in the inability to engage in protective actions during the act.

Noteworthy Chemistry of Chloroform.

Inhaled chloroform anesthesia was introduced in 1847. Soon thereafter, the chemical reactivity of aerobically heated chloroform permitted John Snow and Claude Bernard to do seminal experiments in the assay of drug levels and drug metabolism. However, it was not widely appreciated until a clinical mishap in 1899 that thermal decomposition generated significant levels of toxic phosgene from air-polluting quantities of chloroform in poorly ventilated operating rooms that were illuminated by flames. Phosgene is also generated metabolically from chloroform. A clue appeared in the 1950s when subanesthetic traces of inhaled chloroform proved accidentally lethal to strains of male mice spontaneously expressing high levels of chloroform-metabolizing enzymes. Furthermore, in microbial experiments of 1967, the reactive chloroform molecule was inadvertently discovered to selectively inactivate vitamin B12-dependent enzymes. Chloroform can also activate enzymes. As a solvent, it was serendipitously found in 1903 to activate what is now known as plasminogen to plasmin.

Metabolite profiles of rats in repeated dose toxicological studies after oral and inhalative exposure.

The MetaMap(®)-Tox database contains plasma-metabolome and toxicity data of rats obtained from oral administration of 550 reference compounds following a standardized adapted OECD 407 protocol. Here, metabolic profiles for aniline (A), chloroform (CL), ethylbenzene (EB), 2-methoxyethanol (ME), N,N-dimethylformamide (DMF) and tetrahydrofurane (THF), dosed inhalatively for six hours/day, five days a week for 4 weeks were compared to oral dosing performed daily for 4 weeks. To investigate if the oral and inhalative metabolome would be comparable statistical analyses were performed. Best correlations for metabolome changes via both routes of exposure were observed for toxicants that induced profound metabolome changes. e.g. CL and ME. Liver and testes were correctly identified as target organs. In contrast, route of exposure dependent differences in metabolic profiles were noted for low profile strength e.g. female rats dosed inhalatively with A or THF. Taken together, the current investigations demonstrate that plasma metabolome changes are generally comparable for systemic effects after oral and inhalation exposure. Differences may result from kinetics and first pass effects. For compounds inducing only weak changes, the differences between both routes of exposure are visible in the metabolome.

Evaluation and modeling of the impact of coexposures to VOC mixtures on urinary biomarkers.

CONTEXT: Urinary biomarkers are widely used among biomonitoring studies because of their ease of collection and nonintrusiveness. Chloroform and TEX (i.e., toluene, ethylbenzene, and m-xylene) are chemicals that are often found together because of common use. Although interactions occurring among TEX are well-known, no information exists on possible kinetic interactions between these chemicals and chloroform at the level of parent compound or urinary biomarkers. OBJECTIVE: The objective of this study was therefore to study the possible interactions between these compounds in human volunteers with special emphasis on the potential impact on urinary biomarkers. MATERIALS AND METHODS: Five male volunteers were exposed by inhalation for 6 h to single, binary, and quaternary mixtures that included chloroform. Exhaled air and blood samples were collected and analyzed for parent compound concentrations. Urinary biomarkers (o-cresol, mandelic, and m-methylhippuric acids) were quantified in urine samples. Published PBPK model for chloroform was used, and a Vmax of 3.4 mg/h/kg was optimized to provide a better fit with blood data. Adapted PBPK models from our previous study were used for parent compounds and urinary biomarkers for TEX. RESULTS: Binary exposures with chloroform resulted in no significant interactions. Experimental data for quaternary mixture exposures were well predicted by PBPK models using published description of competitive inhibition among TEX components. However, no significant interactions were observed at levels used in this study. CONCLUSION: PBPK models for urinary biomarkers proved to be a good tool in quantifying exposure to VOC.

An Examination of Horace Wells' Life as a Manifestation of Major Depressive and Seasonal Affective Disorders.

Horace Wells was a Hartford, Connecticut, dentist whose practice flourished because of his clinical skills. He had an imaginative mind that propelled him to the forefront in several aspects of dentistry. Unfortunately, he suffered a recurrent "illness" that began in the winter and resolved in the spring. These symptoms were compatible with both major depressive disorder and seasonal affective disorder as a qualifier. Wells' introduction of nitrous oxide as an anesthetic was also associated with self-inhalation. This led to periods of hypomania, followed by depression. With the progression to ether, then chloroform, there was an episode of mania in January 1848, followed by depression and suicide.

Horace Wells and His House on 120 Chambers St in New York City.

On January 24, 1848, delirious from chloroform, Horace Wells rushed from his house and office on 120 Chambers St into the street and threw acid on 2 alleged prostitutes. He was arrested and committed to New York's infamous Tombs Prison (currently Manhattan Detention Complex), where he committed suicide. Remodeled and reconstructed, this house, 120 Chambers St, is still standing in Tribeca District.

Systemic inflammatory response due to chloroform intoxication--an uncommon complication.

Well-known adverse effects of chloroform are drowsiness, nausea, and liver damage. Two cases with an uncommon complication due to chloroform intoxication are presented. In the first case, a general physician, because of nausea and dyspnea, admitted a 34-year-old woman to hospital. She developed a toxic pulmonary edema requiring mechanical ventilation for a few days, and a systemic inflammatory response syndrome (SIRS) with elevated white blood cell counts, a moderate increase of C-reactive protein, and slightly elevated procalcitonin levels. There were inflammatory altered skin areas progressing to necrosis later on. However, bacteria could be detected neither in blood culture nor in urine. Traces of chloroform were determined from a blood sample, which was taken 8 h after admission. Later, the husband confessed to the police having injected her chloroform and put a kerchief soaked with chloroform over her nose and mouth. In the second case, a 50-year-old man ingested chloroform in a suicidal attempt. He was found unconscious in his house and referred to a hospital. In the following days, he developed SIRS without growth of bacteria in multiple blood cultures. He died several days after admission due to multi-organ failure. SIRS in response to chloroform is a rare but severe complication clinically mimicking bacterial-induced sepsis. The mechanisms leading to systemic inflammation after chloroform intoxication are currently unclear. Possibly, chloroform and/or its derivates may interact with pattern recognition receptors and activate the same pro-inflammatory mediators (cytokines, interleukins, prostaglandins, leukotrienes) that cause SIRS in bacterial sepsis.

Estimation of chloroform inhalation dose by other routes based on the relationship of area under the blood concentration-time curve (AUC)-inhalation dose to chloroform distribution in the blood of rats.

The present study investigated the time-course changes of concentration of chloroform (CHCl3) in the blood during and after exposure of male rats to CHCl3 by inhalation. Increasing the dose of CHCl3 in the inhalation exposed groups caused a commensurate increase in the concentration of CHCl3 in the blood and the area under the blood concentration-time curve (AUC). There was good correlation (r = 0.988) between the inhalation dose and the AUC/kg body weight. Based on the AUC/kg body weight-inhalation dose curve and the AUC/kg body weight after oral administration, inhalation equivalent doses of orally administered CHCl3 were calculated. Calculation of inhalation equivalent doses allows the body burden due to CHCl3 by inhalation exposure and oral exposure to be directly compared. This type of comparison facilitates risk assessment in humans exposed to CHCl3 by different routes. Our results indicate that when calculating inhalation equivalent doses of CHCl3, it is critical to include the AUC from the exposure period in addition to the AUC after the end of the exposure period. Thus, studies which measure the concentration of volatile organic compounds in the blood during the inhalation exposure period are crucial. The data reported here makes an important contribution to the physiologically based pharmacokinetic (PBPK) database of CHCl3 in rodents.

Asphyxial suicide by inhalation of chloroform inside a plastic bag.

Asphyxia suicide by placing a plastic bag over the head in addition with inhalation of gases or use of sedative substances is an unusual method of committing suicide, but frequently referenced by right to die groups in the Internet. This article reports 2 suicides in which chloroform was used to induce unconsciousness and subsequent asphyxia by placing the head in a plastic bag. Case histories of 2 males, ages 23 and 28, are described with special emphasis on characteristics death related to suffocation using plastic bags and chloroform. The final remarkable point in both cases is that the victims previously searched the WEB for instructions of suicide methods. The importance of the phenomenon of misuse of Internet by young people who commit suicide is stressed.

Relative source allocation of TDI to drinking water for derivation of a criterion for chloroform: a Monte-Carlo and multi-exposure assessment.

Drinking water quality standard (DWQS) criteria for chemicals for which there is a threshold for toxicity are derived by allocating a fraction of tolerable daily intake (TDI) to exposure from drinking water. We conducted physiologically based pharmacokinetic model simulations for chloroform and have proposed an equation for total oral-equivalent potential intake via three routes (oral ingestion, inhalation, and dermal exposures), the biologically effective doses of which were converted to oral-equivalent potential intakes. The probability distributions of total oral-equivalent potential intake in Japanese people were estimated by Monte Carlo simulations. Even when the chloroform concentration in drinking water equaled the current DWQS criterion, there was sufficient margin between the intake and the TDI: the probability that the intake exceeded TDI was below 0.1%. If a criterion that the 95th percentile estimate equals the TDI is regarded as both providing protection to highly exposed persons and leaving a reasonable margin of exposure relative to the TDI, then the chloroform drinking water criterion could be a concentration of 0.11mg/L. This implies a daily intake equal to 34% of the TDI allocated to the oral intake (2L/d) of drinking water for typical adults. For the highly exposed persons, inhalation exposure via evaporation from water contributed 53% of the total intake, whereas dermal absorption contributed only 3%.