Kinetic and Thermodynamic Aspects of the Mechanism of General Anesthesia in a Model System of Firefly Luminescence in Vitro

Ueda, Issaku; Kamaya, Hiroshi

doi:10.1097/00000542-197305000-00002

Cited by 100 publications

(35 citation statements)

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“…The preparation of cell-free extract from firefly tails in arsenate buffer and the technique for measurement of initial flash intensity have been described (5). The final concentration of the extract was 2.5 mg of dried firefly tail (Calbiochem) per 1 ml of 0.1 M sodium arsenate buffer (pH 7.5) with 0.04 MgSO4.…”

Section: Methodsmentioning

confidence: 99%

“…We have previously shown that volatile general anesthetics (5) and dissociable local anesthetics (6) inhibit flash intensities of firefly luminescence in vitro. Firefly luciferin and luciferase can be solubilized from the tail lantern of fireflies and emit light in vitro when ATP is added.…”

mentioning

confidence: 99%

“…The flash intensity and total light output is proportional to the amount of added ATP. The adequacy of bioluminescence as a model system for the anesthesia mechanism has been previously discussed (5). With the cell-free firefly luminescent system, the problem of a diffusion barrier is eliminated.…”

mentioning

confidence: 99%

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Combined effects of dissociable and undissociable local anesthetics upon ATP-induced firefly bioluminescence.

Kamaya¹,

Ueda²,

Eyring³

1976

Proc. Natl. Acad. Sci. U.S.A.

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Combined effects of two drugs present simultaneously are usually expressed as summation, synergism or potentiation, and antagonism. When the sum of the effects of each drug present separately equals the combined effect of the two drugs present simultaneously, the action is called additive or summation. However, the expected value of the sum of each effect of drugs present alone has not been well defined. In this report, the theoretical value of the expected sum of each effect of two inhibitors is given and a graphical metho is presented to visualize summation, synergism, and antagonism. The inhibitory effects of a dissociable local anesthetic, tetracaine, and an undissociable local anesthetic, benzyl alcohol, upon a soluble firefly luminescent system were analyzed according to the above theory. The results clearly indicate that the action of these two classes of local anesthetics is pure additive or summation.Local anesthetics inhibit propagation of nerve impulses at synapses and at axons. A number of dissociable and undissociable compounds of diverse structure show local anesthetic activities.Clinically used local anesthetics possess a lipophilic benzene at one end and a tertiary amine on the other. The amine terminal is protonated to form a quaternary nitrogen and becomes highly hydrophilic with its positive charge. The extent of the positive charge is a function of the pka of the compound and the pH of the medium. At physiological pH these compounds exist in both uncharged and charged forms. It has been a matter of discussion as to which is the pharmacologically active form.Alcohols and volatile general anesthetics do not form charged molecules and show local anesthetic activities in excised nerve preparations. Barbiturates dissociate into negatively charged and uncharged molecules and show local anesthetic activities. The uncharged molecule should interact with nerve cells hydrophobically, whereas the charged molecule should bind to nerve cells electrostatically or by hydrogen bonding.Ritchie and Greengard (1) and later Narahashi et al. (2) proposed that local anesthetics penetrate the nerve cell membrane in its uncharged form and inhibit the nerve action by binding to the effector sites from the inside of the cell membrane in its positively charged form. This theory suggests that there are at least two separate modes of action among agents that show local anesthetic action: the predominant force of interaction of dissociable anesthetics is ionic, whereas the mode of action of undissociable local anesthetics is not. However, Agin et al. (3) and Hersh (4) suggested that dissociable and undissociable local anesthetics may interact with living and model systems in an identical way.We have previously shown that volatile general anesthetics (5) and dissociable local anesthetics (6) inhibit flash intensities of firefly luminescence in vitro. Firefly luciferin and luciferase can be solubilized from the tail lantern of fireflies and emit light in vitro when ATP is added. The flash intensity and total light o...

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

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Combined effects of dissociable and undissociable local anesthetics upon ATP-induced firefly bioluminescence.

Kamaya¹,

Ueda²,

Eyring³

1976

Proc. Natl. Acad. Sci. U.S.A.

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“…[1][2][3][4][5][6] The effect of hydrophobic molecules, such as terminally-substituted normal alkyl compounds, on the bacterial luciferase (BL) bioluminescence has been reported in our previous studies. 5,6 The BL reaction requires a reduced flavin mononucleotide (FMNH2) as one of the substrates.…”

Section: Introductionmentioning

confidence: 96%

Electrochemical Control of Bioluminescence by Blocking the Adsorption of the Bacterial Luciferase Using a Mercaptobipyridine Self-assembled Monolayer

et al. 2017

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An N-butyl-N′-(4-mercaptobutyl)-4,4′-bipyridinium (4BMBP) was modified on a gold electrode to improve the electrochemical control of the bacterial luciferase (BL) luminescence system. The 4BMBP-modified gold electrode (4BMBP/Au) was able to prevent the adsorption of BL on the electrode surface, and enhanced the electrochemical regeneration rate of the reduced flavin mononucleotide (FMNH2), which is one of the substrates of the BL luminescence reaction. By using the 4BMBP/Au, the luminescence intensity increased by about 27% compared to that of a bare gold electrode (bare Au). Moreover, the modified electrode improved the time required for analysis because the modified layer prevented BL adsorption. Even without a refreshing procedure for each measurement, a constant luminescence intensity could be observed, and the analysis time was reduced to half (about 10 min) for one sample. The 4BMBP/Au is not only useful to control of the BL luminescence system, but also for electrochemical measurements in the presence of proteins.

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“…Among them, FFL has been widely used because the bioluminescence of FFL acts as a sensitive probe for the inhibition of the enzyme reaction by GAs. [10][11][12][13] In our recent studies, we have examined the applicability of the luminescence enzyme bacterial luciferase (BL) to analyze the action of hydrophobic molecules on proteins because of its high sensitivity and the easy traceability of the enzyme reaction.…”

Section: Introductionmentioning

confidence: 99%

Bioluminescence Inhibition of Bacterial Luciferase by Aliphatic Alcohol, Amine and Carboxylic Acid: Inhibition Potency and Mechanism

2013

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The inhibitory effects of hydrophobic molecules on the bacterial luciferase, BL, luminescence reaction were analyzed using an electrochemically-controlled BL luminescence system. The inhibition potency of alkyl amines, CnNH2, and fatty acids, CmCOOH (m = n -1), on the BL reaction increased with an increase in the alkyl chain-length of these aliphatic compounds. CmCOOH showed lower inhibition potency than CnNH2 and alkyl alcohols, CnOH, data for which have been previously reported. To make clear the inhibition mechanisms of the aliphatic compounds on the BL reaction, the initial rate of the BL reaction was measured and analyzed using the Dixon plot and Cornish-Bowden plot. The C12OH inhibited the BL reaction in competition with the substrate C11CHO, while C12NH2 and C11COOH inhibited in an uncompetitive manner with the C11CHO. These results suggest that the alkyl chain-length and the terminal unit of the aliphatic compound determine the inhibition potency and the inhibition mechanism, respectively.

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Kinetic and Thermodynamic Aspects of the Mechanism of General Anesthesia in a Model System of Firefly Luminescence in Vitro

Cited by 100 publications

References 0 publications

Combined effects of dissociable and undissociable local anesthetics upon ATP-induced firefly bioluminescence.

Combined effects of dissociable and undissociable local anesthetics upon ATP-induced firefly bioluminescence.

Electrochemical Control of Bioluminescence by Blocking the Adsorption of the Bacterial Luciferase Using a Mercaptobipyridine Self-assembled Monolayer

Bioluminescence Inhibition of Bacterial Luciferase by Aliphatic Alcohol, Amine and Carboxylic Acid: Inhibition Potency and Mechanism

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