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Anesth Analg 2006;102:772-778
© 2006 International Anesthesia Research Society
doi: 10.1213/01.ane.0000196536.60320.f9

ANESTHETIC PHARMACOLOGY

Intravenous Anesthetics Are More Effective than Volatile Anesthetics on Inhibitory Pathways in Rat Hippocampal CA1

Takehisa Asahi, MD*, Koki Hirota, MD, PhD*, Rika Sasaki, MD, PhD*, Yamazaki Mitsuaki, MD, PhD*, and Sheldon H. Roth, PhD{dagger}

*Department of Anesthesiology, University of Toyama, Japan; and {dagger}Departments of Pharmacology & Therapeutics and Anaesthesia, Faculty of Medicine, University of Calgary, Alberta, Canada

Address correspondence and reprint requests to Takehisa Asahi, MD, Department of Anesthesiology, University of Toyama, 2630 Sugitani, Toyama-city, Toyama, 930-0194 Japan. Address e-mail to take3260{at}ms.toyama-mpu.ac.jp.


   Abstract
Top
 Abstract
Introduction
 Methods
 Results
 Discussion
 References
 
In this study, we have examined the effects of both volatile and IV general anesthetics on excitatory synaptic transmission, with and without recurrent inhibition, to clarify whether excitatory or inhibitory synapses are the major targets of action. Field population spike amplitudes (fPSs) of CA1 pyramidal neurons were recorded in rat hippocampal slices. Schaffer-collateral-commissural fibers (Sch) were stimulated orthodromically, and the evoked fPSs (PS[Sch]) in CA1 area were measured. In addition, the fPSs (PS[Alv+Sch]) elicited by stimulation of the Sch after antidromic stimulation of the alveus hippocampi (Alv) to produce recurrent inhibition were determined. It was observed that sevoflurane (0.5%–5%) and isoflurane (0.5%–5%) primarily inhibited PS[Sch] and also produced additive inhibition on the PS[Alv+Sch] in a concentration-dependent manner. The calculated 50% effective concentration (EC50) values for PS[Sch] and PS[Alv+Sch] were 5.3 vol% and 3.9 vol% (sevoflurane) and 1.7 vol% and 1.1 vol% (isoflurane), respectively. In comparison, thiopental (2.0 x 10–5–5.0 x 10–4 mol/L) reduced both the PS[Sch] and PS[Alv+Sch] in a concentration-dependent manner. The calculated EC50 values for thiopental on PS[Sch] and PS[Alv+Sch] were 3.4 x 10–4 and 5.7 x 10–5 mol/L, respectively. Propofol (2.0 x 10–5–3.5 x 10–4 mol/L) had little effect on the PS[Sch] but reduced PS[Alv+Sch] with a calculated EC50 value of 5.1 x 10–4 mol/L. The effects of the IV anesthetics with recurrent inhibition were antagonized in the presence of the {gamma}-aminobutyric acid-A-receptor antagonist bicuculline methiodide. In addition, all anesthetics prolonged recurrent inhibition from 100 ms (sevoflurane and isoflurane) to 400 ms (propofol). The results suggest that sevoflurane and isoflurane inhibit mainly on glutamate-mediated orthodromic pathways, whereas thiopental and propofol enhance {gamma}-aminobutyric acid-A-mediated recurrent inhibitory pathways in CA1 neurons, thus providing further evidence that the mechanisms of general anesthetics are drug- and pathway-specific.


   Introduction
Top
 Abstract
 Introduction
Methods
 Results
 Discussion
 References
 
The mechanisms of anesthetic actions occur via specific and multiple sites and that the effects seem to be both pathway and drug specific (1). Many electrophysiological studies have examined the effects of general anesthetics on both excitatory and inhibitory synaptic transmission. Nishikawa and MacIver (2,3) demonstrated that volatile anesthetics decrease excitatory glutaminergic synaptic transmission (2) and enhance inhibitory {gamma}-aminobutyric acid (GABA)-mediated synaptic transmission (3) in rat CA1 hippocampal neurons. They concluded that volatile anesthetics have multisite and drug-specific mechanisms. Previous studies have demonstrated that IV anesthetics can inhibit glutamate-mediated excitatory synaptic transmission (4), whereas others have shown that these anesthetics can modulate GABA-mediated inhibition (5,6). Few reports have compared the effects of anesthetics on both excitatory and inhibitory synaptic transmission on the same preparation (7). In the present study, we compared the effects of both volatile and IV anesthetics on excitatory synaptic transmission in rat hippocampal slices with and without recurrent inhibition to elucidate whether excitatory and inhibitory synapses are the major targets of action.


   Methods
Top
 Abstract
 Introduction
 Methods
Results
 Discussion
 References
 
Ethical approval was obtained from the Animal Research Committee of Toyama Medical and Pharmaceutical University. Rat hippocampal brain slices were prepared using identical techniques previously described by Hirota and Roth (8). Electrophysiological experiments were performed on hippocampal slices from young male Wistar rats (50–200g). The animals were deeply anesthetized with sevoflurane (SEV) and then decapitated. The brain was rapidly removed, and 400-µm transverse slices were prepared from the dissected hippocampus in cold, oxygenated artificial cerebrospinal fluid (ACSF) using a Rotorslicer DTY-7700 (DSK, Osaka, Japan). The slices were placed on a nylon mesh screen at the interface of ACSF liquid (90 mL/h) and humidified 95% O2/5% CO2 gas (1 L/min) phases in a recording chamber. Slices were warmed to 37°C slowly and then allowed to equilibrate for 90–120 min without electrical stimulation.

A glass extracellular recording microelectrode (3–5 M{Omega} filled with 2 M of NaCl) was placed in the cell body region of CA1 pyramidal neurons. One bipolar Nichrome-stimulating electrode was placed in the region of Schaffer-collateral-commissural fibers (Sch) to stimulate the input to CA1 neurons, and a second electrode was placed in the region of alveus hippocampi (Alv) to activate inhibitory interneurons of the CA1 (Fig. 1).


Figure 116
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  		Figure 1. Rat hippocampal slice preparation. A glass extracellular microelectrode was placed in the cell body regions of CA1 pyramidal neurons for recording field potentials. Two bipolar Nichrome-stimulating electrodes were placed on Schaffer collateral fiber (Sch) input to CA1 pyramidal neurons and on alveus hippocampi (Alv) input to CA1, respectively, in the same slice. (a and b) Glutamate-mediated excitatory synapses. (c) {gamma}-aminobutyric acid (GABA)-mediated inhibitory synapse. Int = inhibitory interneuron; mf = mossy fiber; dent = dentate gyrus; pp = perforant path.

 

Square-wave stimuli (5–10V, 0.05ms, 0.1Hz), generated with a SEN-3301 stimulator (Nihon Kohden, Tokyo, Japan), were delivered to both pathways alternately. The minimal stimulus intensity (5–10 volt) that elicited the maximal population spike (PS) amplitude (maximal stimulus) was used. Initially, stimulations to Sch were applied as test-pulses, and field PS amplitudes (fPSs) as PS[Sch]-evoked responses in the CA1 area were recorded. Next, the region of Alv was stimulated as a pre-pulse before the application of the test-pulse. Examples of recorded PS[Alv] and PS[Alv+Sch] in the CA1 area are shown in Figure 2. Because the antidromic pre-pulse activates not only CA1 pyramidal cells but also inhibitory interneurons (Fig. 1), PS[Alv+Sch], the fPS in the CA1 area, is recorded in the presence of recurrent inhibition.


Figure 216
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  		Figure 2. Example of recorded field potential amplitude (fPS) elicited with orthodromic stimulation to Sch (PS[Sch]) (upper tracing). Example of recorded fPSs elicited with a pre-pulse antidromic stimulation to Alv (PS[Alv]) to activate inhibitory interneurons of the CA1 followed by a test-pulse to Sch (PS [Alv +Sch]) with an interstimulus interval (ISI) of 40 ms (middle tracing). Recording similar to middle tracing with ISI of 10 ms (lower tracing). Sch = Schaffer collateral fiber; Alv = Alveus hippocampi; PS = population spike; ACSF = artificial cerebrospinal fluid; p = pre-pulse, t = test-pulse.

 

The amplitudes of PS[Sch] and PS[Alv+Sch] were compared in the absence of anesthetic (control) to estimate physiological recurrent inhibitory characteristics in this area. The ratios of PS[Alv+Sch]/PS[Sch] were <1.0 when pre- and test-pulse intervals were shorter than 40 ms (Figs. 2 and 3); therefore, the concentration-dependent effects of the anesthetics were examined under fixed 40-ms interstimulus interval (ISI) conditions that would not be influenced by physiological recurrent inhibition. For the experiments examining the antagonistic effects of bicuculline methiodide (BMI) and prolongation of recurrent inhibition by anesthetics, approximate effective concentration of 50% (EC50) values (except for propofol) were used.


Figure 316
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  		Figure 3. Relationship of population spike amplitudes (PS[Alv+Sch]/PS[Sch]) elicited with orthodromic or antidromic (with pre-pulse) stimulation paradigms and length of interstimulus intervals (ISI) in control solution (ACSF) and the presence of 1.0 x 10–5 mol/L of bicuculline methiodide (BMI). Each point represents the mean ± sd (n = 7). ACSF = artificial cerebrospinal fluid.

 
All preparations used in the present study showed control variability <5% during the initial data acquisition period and after washout of anesthetic drugs. Recovery responses were recorded at least 25 min after washout of anesthetic-equilibrated ACSF from the chamber. Volatile anesthetics, SEV and isoflurane (ISO), were applied as vapors in the prewarmed carrier gas (95% O2/5% CO2) above the slices using calibrated commercial vaporizers such as Tec 3 (Omeda, Steeton, West Yorkshire, UK) and Forawick (Muraco, Tokyo, Japan), respectively. Concentrations, expressed as volume percent (vol%) refer to the dial settings on the vaporizers. Concentrations of volatile anesthetics in the perfusate of the recording chamber were determined using gas chromatography (Shimazu, Kyoto, Japan). It was found that there was a linear relationship (0.64 and 0.55 mM per 1 vol% of SEV and ISO, respectively) up to 5.0%.

Thiopental and BMI were dissolved in ACSF at required concentrations before use. Propofol was dissolved in dimethyl sulfoxide first and then diluted in ACSF before it was perfused into the chamber. The concentrations of dimethyl sulfoxide used in the experiments did not affect the field potentials. All anesthetics were applied for a minimum of 20 min to reach equilibrium, as previously demonstrated (7). The composition of the ACSF was NaCl 124 mM, KCl 5 mM, CaCl2 2 mM, NaH2PO4 1.25 mM, MgSO4 2 mM, NaHCO3 26 mM, and glucose 10 mM prepared in purified water. The ACSF was precooled (8°C–10°C) and kept saturated with 95% O2/5% CO2 gas mixture before use (pH 7.1–7.4).

SEV and ISO were donated by Dinabot (Osaka, Japan) and Maruishi Pharmaceutical Co (Osaka, Japan), respectively. All of the other chemicals used were purchased from Sigma (St. Louis, MO).

Evoked fPSs were amplified with a MEZ-8301 amplifier (Nihon Kohden, Japan) and filtered 1 Hz–10 kHz. Analog-digital conversions of data were made at a rate of 100 kHz using InstruNet (GW, Somerville, MA). The results were stored on the hard drive of a Macintosh computer (Apple, Cupertino, CA) and analyzed using SuperScopeTM software (GW).

The PS amplitudes were determined from peak positive to peak negative of the waveform (Fig. 2). Results were collected in groups of five, averaged, and stored as a single record. Concentration-response relationships for each anesthetic were expressed as percent of control (absence of anesthetic drug), and prolongation effects of recurrent inhibition were expressed as PS[Alv+Sch]/PS [Sch] < 1.0. The mean ± sd were calculated based on results from at least 5 slices.

Statistical differences among groups of control and different anesthetics were tested by repeated-measures analysis of variance (ANOVA) and differences between paired sets of data were compared by the Bonferroni/Dunn test. A P value < 0.05 was considered significantly different. Curve fitting analysis was performed using SigmaPlotTM (Jandel Scientific, San Rafael, CA) software.


   Results
Top
 Abstract
 Introduction
 Methods
 Results
Discussion
 References
 
To gain a better understanding of the characteristics of the physiological recurrent inhibition in the CA1 hippocampal region, we recorded both PS[Sch] and PS[Alv+Sch] in the absence of general anesthetics (Fig. 2). The ratio of PS[Alv+Sch]/PS[Sch] was <1.0 in conditions where ISI was shorter than 40 ms. Therefore, we conducted subsequent experiments of concentration-response effects on recurrent inhibition under fixed 40-ms intervals that would not be influenced by this physiological characteristic. Recurrent inhibition was antagonized by application of BMI (1. 0 x 10–5 mol/L) (Fig. 3).

Representative recordings of the effects of the anesthetics on both PS[Sch] and PS[Alv+Sch] are shown in Figure 4A. The major effects of SEV (4.0 vol%) and ISO (1.0 vol%) were depression of PS[Sch] and PS[Alv+Sch]. The depressant effects were similar for both responses.


Figure 416
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  		Figure 4. (A) The effects of 4.0% sevoflurane (SEV) and 1.0% isoflurane (ISO) on evoked-population field potentials compared with control solution (artificial cerebrospinal fluid [ACSF]). Reduction of the PS[Alv+Sch] was slightly more than the decrease in PS[Sch]. However, this is not significant. See text for explanation of stimulation properties (upper tracing). In comparison, 5.0 x 10–5 mol/L of thiopental (THIO) and 3.5 x 10–4 mol/L of propofol (PRO) produced a greater depression on PS[Alv+Sch] compared to PS[Sch] (lower tracing). p = pre-pulse; t = test-pulse. (B) The depressant effects of the general anesthetics on the evoked population spikes tested are concentration dependent. All data were obtained with interstimulus intervals (ISI) of 40 ms. The depressant effects of the volatile anesthetics (SEV and ISO) seem to be equal on PS[Sch] and PS[Alv+Sch]; however, the IV anesthetics exhibit greater effects on the PS[Alv+Sch]. Each data point represents the mean ± sd (n = 6).

 

In comparison, thiopental (5.0 x 10–5 mol/L) depressed both PS[Sch] and PS[Alv+Sch]; however, the effects were greater on the PS[Alv+Sch]. Propofol (3.5 x 10–4 mol/L) also had a smaller effect on PS[Sch] than PS[Alv+Sch].

Concentration-response relationships of both PS[Sch] and PS[Alv+Sch] for the anesthetics are shown in Figure 4B. SEV (0.5%–5%) and ISO (0.5%–5%) significantly depressed PS[Sch] and PS[Alv+Sch] in a concentration-dependent and reversible manner. There was no significant difference between the degree of inhibition observed on either PS[Sch] and PS[Alv+Sch] in the presence of each concentration of both volatile anesthetics. The continuous line showed in Figure 4B is the best fit using the following equation:



Formula 1

where C is the concentration (vol% or mol/L) of anesthetic, k is the concentration that induced 50% depression of amplitude, and n is the Hill coefficient. The EC50 values and Hill coefficient for depression of both PS[Sch] and PS[Alv+Sch] were calculated from data in Figure 4B and shown in Table 1. There were no significant differences between the calculated Hill coefficients (SEV, 1.5 and 1.6; ISO, 1.2 and 1.3) and EC50s (SEV, 5.3 vol% and 3.9 vol%; ISO, 1.7 vol% and 1.1 vol%) for either PS[Sch] or PS[Alv+Sch] (Table 1).


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  		Table 1. Hill Coefficient and EC50 of Anesthetics in the Preparation

 

As shown in Figure 4B, thiopental (2.0 x 10–5–2.0 x 10–4 mol/L) depressed PS[Sch] and PS[Alv+Sch] in a concentration-dependent and reversible manner. There were significant differences between the degree of inhibition observed on PS[Sch] compared with PS[Alv+Sch] at the larger concentrations of thiopental (mean ± sd, 81.5% ± 16.8% and 52.6% ± 23.0% in 5.0 x 10–5 mol/L, 75.9% ± 13.4% and 30.4% ± 22.1% in 1.0 x 10–4 mol/L, and 62.3% ± 26.1% and 12.0% ± 12.6% in 2.0 x 10–4 mol/L). Calculated EC50 values for thiopental for PS[Sch] and for PS[Alv+Sch] were 3.4 x 10–4 mol/L and 5.7 x 10–5 mol/L, respectively. In comparison, propofol (2.0 x 10–5–3.5 x 10–4 mol/L) had little effect on PS[Sch] but depressed PS[Alv+Sch] (Figure 4B) in a concentration-dependent and reversible manner. The calculated EC50 value for propofol on the PS[Alv+Sch] was 5.1 x 10–4mol/L (Table 1). Because the effects of propofol at the EC50 value were not reversible, we did not apply this concentration. Instead, we used 3.5 x 10–4 mol/L of propofol to investigate the antagonistic effects of BMI and prolongation of recurrent inhibition by propofol.

As shown in Figure 5, BMI antagonized the IV anesthetic effects on recurrent inhibition completely. The antagonistic effects of BMI were not significant for volatile anesthetics because the effects of 4.0% SEV and 1.0% ISO on recurrent inhibition were relatively weak. These results indicate that anesthetic-enhancing effects of recurrent inhibition in the CA1 area may be mediated via the GABAA receptor.


Figure 516
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  		Figure 5. The effects of 5.0 x 10–5 mol/L of thiopental (THIO) and 3.5 x 10–4 mol/L of propofol (PRO) on recurrent inhibition as expressed by the ratios of (PS[Alv+Sch]/PS[Sch]) are antagonized by 1.0 x 10–5 mol/L of bicuculline methiodide (BMI). The antagonistic effects of BMI are not significant for volatile anesthetics. Each column represents the mean ± sd (n = 5). GABA = {gamma}-aminobutyric acid.

 

We also examined the effects of anesthetics on both PS[Sch] and PS[Alv+Sch] under various durations of ISI (10 ms–1000ms). PS[Alv+Sch]/PS[Sch] ratios were significantly smaller than those in ACSF when ISI was less than 100 ms for SEV and ISO, 200 ms for thiopental, and 400 ms for propofol (Fig. 5). The anesthetics also prolonged the effects of recurrent inhibition.


   Discussion
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
References
 
The hippocampus is a well-defined structure, and its comparatively simple synaptic organization consisting of glutamate-mediated trisynaptic excitatory pathways and GABA-mediated inhibitory interneurons (3,9) may be a target for general anesthetics (10). Thus, the hippocampal slice preparation is a reasonable model for the study of anesthetic action on synaptic transmission in the central nervous system. We previously reported that halothane and ISO did not alter antidromically evoked PS[Alv] but depressed orthodromically evoked PS[Sch] in rat hippocampal slice preparations and concluded that volatile anesthetics primarily act on synaptic events (11). In the present study, we observed a lack of effect on PS[Alv], thus providing further support for synaptic targets.

The orthodromic pathway (Sch to CA1) is excitatory, monosynaptic, and mediated by glutamate and its receptor (Fig. 1, a) (2,9). Alteration of PS[Sch] amplitudes by the anesthetics are therefore a result of action on excitatory synaptic transmission. However, the pre-pulse on Alv evokes antidromically (nonsynaptic) PS[Alv] in the CA1 area (12). Pre-pulse also activates recurrent inhibitory interneurons by glutamate and its receptor (Fig. 1, b), thus hyperpolarizing CA1 pyramidal cells by GABA and its receptor (9) (Fig. 1, c). Inhibition of the PS[Alv+Sch] thus reflects anesthetic actions on both GABAergic and glutaminergic synaptic transmission.

Many studies have examined the effects of general anesthetics using paired-pulse stimulation (PPS) paradigms to evoke recurrent inhibition in this region (2,7,8,12,13). Inhibitory interneurons are activated by the discharge produced in response to the first pulse (PS1) and compete with excitatory postsynaptic potential facilitation to modulate responses of the second pulse (PS2). ISO and halothane inhibit PS1 rather than PS2, whereas pentobarbital and propofol decrease PS2 with minimal change to PS1 (7). There are, however, some problems associated with using this method to determine the action of anesthetics on both excitatory and inhibitory synapses. First, the magnitude of input to the inhibitory interneuron depends on PS1, i.e., when PS1 itself is decreased by anesthetics, input to the inhibitory interneuron could be reduced. Second, excitatory synaptic transmission has the potential to produce paired-pulse facilitation (PPF) or paired-pulse depression via paired-pulse stimulation (14). PS2 might also be affected by similar properties as well as recurrent inhibition. Because antidromical PS[Alv] was not altered in the presence of anesthetic, similar outputs could be produced as CA1. We have confirmed that PPF and paired-pulse depression did not occur in the present study (Fig. 3).

In control conditions (no anesthetic), it is evident that PS[Alv+Sch]/PS[Sch] ratios are <1.0 if ISI is less than 40 ms (Figs. 2 and 3). It still remains unclear how recurrent inhibition functions in physiological synaptic networks; therefore, we studied the effects of anesthetics using a 40-ms ISI to not involve physiological recurrent inhibition.

Vincent and Steven (15) recently pointed out that BMI is not a specific GABAA receptor antagonist. Thus, the effects of BMI (Figs. 3 and 5) may not be specific to the inhibition on GABAA. As seen in Figure 3, however, the reduction of PS[Alv+Sch]/PS[Sch] in ACSF disappeared in the presence of BMI, indicating that BMI blocks the inhibition of interneurons. In the present study, we used BMI as a pharmacological tool for blocking the recurrent inhibitory pathways.

A few previous reports have shown that volatile anesthetics inhibit glutaminergic synaptic transmission via both pre- and postsynaptic mechanisms (2). Other reports have demonstrated that volatile anesthetics at clinically relevant concentrations also enhance GABAergic synaptic inhibition via both pre- and postsynaptic mechanisms (16,17). The present results provide further evidence that volatile anesthetics act at several discrete sites within central nervous system structures.

Because we did not observe a significant difference between EC50s and Hill coefficients of SEV and ISO when accompanied by recurrent inhibition, it is possible that SEV and ISO may have also depressed the glutaminergic excitatory synaptic input from CA1 to inhibitory interneurons (Fig. 1, b) (9). This would tend to diminish the role of enhanced inhibitory interneurons by volatile anesthetics (17). Because the increase in inhibitory synaptic transmission is lessened, we may conclude that the primary effect of SEV and ISO is depression of excitatory synaptic transmission at clinically relevant concentrations, and that the augmentation of inhibitory synaptic transmission is not a major effect but may be additive. In addition, we have demonstrated that thiopental and propofol reduced the excitation of CA1, primarily by increasing recurrent inhibition. The EC50 of thiopental for inhibition of PS[Sch] was approximately 10-fold more than for inhibition of PS[Alv.+Sch], and propofol exhibited little effect on excitatory synaptic transmission. The IV anesthetics also extended the duration of recurrent inhibition in comparison to the volatile anesthetics (Fig. 6).


Figure 616
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  		Figure 6. Prolongation effects of {gamma}-aminobutyric acid (GABA)ergic recurrent inhibition by general anesthetics. General anesthetics not only increase the effect of recurrent inhibition, but also prolong it. Each data point represents the mean ± sd (n = 7). SEV = sevoflurane: 4.0 vol%; ISO = isoflurane: 1.0 vol%; THIO = thiopental: 5.0 x 10–5 mol/L; PRO = propofol: 3.5 x 10–4 mol/L; ACSF = artificial cerebrospinal fluid; ISI = interstimulus intervals.

 

Zhu et al. (18) have reported that thiopental (6.0 x 10–4 mol/L) can inhibit N-methyl-d-aspartate (NMDA) and {alpha}-amino-3-hydroxy-5-methyl-4-isoxazoleproprionate (AMPA)-mediated glutamate excitotoxicity. In the present study, thiopental (2.0 x 10–4 mol/L) altered GABAergic and glutamine-mediated synaptic transmission, thus supporting the hypothesis that barbiturates block NMDA receptors at larger concentrations (18,19). Similar to the volatile anesthetics, thiopental can also depress the glutaminergic excitatory synaptic input to inhibitory interneurons, and thus tends to diminish the effects of these pathways. However, the phenomenon is not pronounced, because the effect of thiopental on excitatory synaptic transmission may require considerably larger concentrations. These results suggest that the major target of thiopental is GABAA-mediated synaptic transmission.

Propofol suppresses the responses of the AMPA-, kainate-, and NMDA-receptor channels at large concentrations and slightly depresses the NMDA receptor channels at clinical concentrations (4,20). In addition, propofol did not inhibit PS1 in studies using PPS techniques (6,7). The results of these electrophysiologic studies indicate that the effects of propofol on glutamate receptors require considerably larger concentrations than those on GABAA receptors (21). Propofol may not inhibit excitatory inputs to interneurons (Fig. 1, b), thus allowing persistent activation of inhibitory interneurons. This is supported by the observations of the present study showing that propofol enhanced and prolonged recurrent inhibition (Fig. 6).

Although all the anesthetics tested in the present study, except propofol, were effective at clinical concentrations, similar to our previous reports (7,22), it was required to use concentrations of propofol approximately 10 times larger than clinical serum levels of 2.0 x 10–5 mol/L (6,23). In addition, considering the high protein binding of propofol, its effect site concentration may be smaller than clinical serum concentrations. Bieda and MacIver (24) reported that the propofol concentration in brain slice is significantly smaller than the applied perfusate concentration. They pointed out that slices often exhibit a high diffusional/binding barrier for entry of propofol, and the effects often continue to increase after 40–60 minutes or more. Pittson et al. (16) have reported that lipophilic IV anesthetics might take several hours to diffuse the 200–300 µm into brain slices, unlike volatile anesthetics that have a high aqueous solubility. In addition, there are obvious differences between diffusion characteristics in brain slices compared with intact brain tissue (6). In the present study, the application time for propofol was relatively short, similar to numerous other studies (6,7,18,24); therefore, the actual concentration in the slice was probably much smaller than the applied concentration.

In conclusion, we have shown that SEV and ISO inhibit mainly on glutamate-mediated orthodromic pathways, whereas thiopental and propofol enhance primarily on GABAA-mediated recurrent inhibitory pathways in the rat hippocampal CA1 region. These results also provide further support for the idea that the mechanisms of action of general anesthetics are drug- and pathway-specific.


   Footnotes
 
Supported, in part, by Grants-In-Aid for Scientific Research from Ministry of Education, Culture, Sports, Science, & Technology of Japan.

Accepted for publication October 19, 2005.


   References
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 

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Lippincott, Williams & Wilkins Anesthesia & Analgesia® is published for the International Anesthesia Research Society® by Lippincott Williams & Wilkins with the assistance of Stanford University Libraries' HighWire Press®. Copyright 2006 by the International Anesthesia Research Society. Online ISSN: 1526-7598   Print ISSN: 0003-2999 HighWire Press