Free
Meeting Abstracts  |   September 1996
Riluzole Anesthesia: Use-Dependent Block of Presynaptic Glutamate Fibers
Author Notes
  • (MacIver) Assistant Professor of Neurophysiology, Stanford Anesthesia.
  • (Amagasu) M.S. Graduate Student in Biological Sciences, Stanford University.
  • (Mikulec) M.D. Student, Stanford University School of Medicine.
  • (Monroe) Research Assistant, Stanford Anesthesia.
  • Received from the Stanford Neuroscience Program and Neuropharmacology Laboratory, Department of Anesthesia, Stanford University School of Medicine, Stanford, California. Submitted for publication December 5, 1995. Accepted for publication May 20, 1995. Supported in part by National Institutes of Health grant GM49811. Presented at the annual meeting of the American Society of Anesthesiologists, Atlanta, Georgia, October 21-25, 1995.
  • Address reprint requests to Dr. MacIver: Department of Anesthesia, Stanford, California 94305-5117. Address electronic mail to: bruce. maciver@forsythe.stanford.edu.
Article Information
Meeting Abstracts   |   September 1996
Riluzole Anesthesia: Use-Dependent Block of Presynaptic Glutamate Fibers
Anesthesiology 9 1996, Vol.85, 626-634.. doi:
Anesthesiology 9 1996, Vol.85, 626-634.. doi:
Key words: Animals: rats. Anesthetics: thiopental. Brain: hippocampus, CA 1; cortex; cortical. Neurophysiology: action potential; conduction; postsynaptic; nerve; sodium channel. Pharmacology: gamma-aminobutyric acid; depressant; glutamate; hypnotic; narcotic; riluzole. Synapse: excitation; inhibition; transmission.
DEPRESSION of glutamate-mediated excitatory synaptic transmission is thought to play an important role in the central nervous system depression produced by several anesthetics. [1] Depressed glutamate responses at N-methyl-D-aspartate receptors appear to be a major mechanism of action for dissociative anesthetics like ketamine. [2,3] Similarly, volatile anesthetics and barbiturates have been shown to depress glutamate-mediated synapse, [4-6] and this could combine with enhanced gamma-amino butyric acid (GABA)-mediated inhibition to produce central nervous system depression. [7,8] Recent interest has focused on anesthetics that reduce excitatory synaptic transmission by depressing glutamate release from nerve terminals. [9] Riluzole (2-amino-6-trifluoro-methoxy benzothiazole, RP 54274, PK 26124) has been shown to depress glutamate release at mammalian central nervous system excitatory synapses [10,11] and produces potent hypnotic, anesthetic, and minimum alveolar concentration-sparing effects in rats. [12] Riluzole's mechanism of action is not known; however, effects on presynaptic sodium channels, [13] N-methyl-D-aspartate-induced CA sup ++ currents, [14-16] GABA uptake, [17] and glutamate release [11] have been suggested. In the current study, we investigated riluzole's effects on: (1) synaptically evoked population spike responses, (2) GABA-mediated paired pulse inhibition, (3) glutamate-mediated synaptic excitation, and (4) presynaptic action potential conduction (Figure 1), to determine which actions contribute to depressed synaptically evoked discharge of CA 1 pyramidal neurons in rat hippocampal slices.
Figure 1. Drawing of a hippocampal slice (A) showing the relative positions of a bipolar tungsten stimulating electrode (STIMULATE), and extracellular glass recording electrodes (RECORD). The recording electrode position was different for population spikes (PS) than for fiber volleys (FV) and excitatory postsynaptic potentials (EPSP). The three major excitatory pathways in hippocampal slices are shown: the perforant path (pp) originates in entorhinal cortex and projects to granule cells of the dentate gurus (DG), the mossy fiber pathway (mf) comprises granule cell axons and projects to pyramidal cells of CA 3, and Schaffer collateral (sc) fibers from CA 3 neurons form excitatory synapses with dendrites of CA 1 neurons. Synaptically evoked discharge (PS) was used to measure riluzole effects on the output of the CA 1 circuit (B). Circled numbers correspond to the four measures used to determine the major sites of action for riluzole in this circuit (see beginning of article).
Figure 1. Drawing of a hippocampal slice (A) showing the relative positions of a bipolar tungsten stimulating electrode (STIMULATE), and extracellular glass recording electrodes (RECORD). The recording electrode position was different for population spikes (PS) than for fiber volleys (FV) and excitatory postsynaptic potentials (EPSP). The three major excitatory pathways in hippocampal slices are shown: the perforant path (pp) originates in entorhinal cortex and projects to granule cells of the dentate gurus (DG), the mossy fiber pathway (mf) comprises granule cell axons and projects to pyramidal cells of CA 3, and Schaffer collateral (sc) fibers from CA 3 neurons form excitatory synapses with dendrites of CA 1 neurons. Synaptically evoked discharge (PS) was used to measure riluzole effects on the output of the CA 1 circuit (B). Circled numbers correspond to the four measures used to determine the major sites of action for riluzole in this circuit (see beginning of article).
Figure 1. Drawing of a hippocampal slice (A) showing the relative positions of a bipolar tungsten stimulating electrode (STIMULATE), and extracellular glass recording electrodes (RECORD). The recording electrode position was different for population spikes (PS) than for fiber volleys (FV) and excitatory postsynaptic potentials (EPSP). The three major excitatory pathways in hippocampal slices are shown: the perforant path (pp) originates in entorhinal cortex and projects to granule cells of the dentate gurus (DG), the mossy fiber pathway (mf) comprises granule cell axons and projects to pyramidal cells of CA 3, and Schaffer collateral (sc) fibers from CA 3 neurons form excitatory synapses with dendrites of CA 1 neurons. Synaptically evoked discharge (PS) was used to measure riluzole effects on the output of the CA 1 circuit (B). Circled numbers correspond to the four measures used to determine the major sites of action for riluzole in this circuit (see beginning of article).
×
Materials and Methods
Slice Preparation
Experiments were performed on brain slices isolated from male Sprague-Dawley rats (80-120 g). Experimental protocols were approved by the Institutional Animal Care Committee at Stanford University and adhered to published guidelines of the National Institutes of Health, Society for Neuroscience, and the American Physiological Society. Rats were anesthetized with diethyl ether, and their hearts were stopped with a blow to the back of the thorax. Brains were removed and placed in cold (1-2 degrees Celsius), oxygenated artificial cerebrospinal fluid (ACSF, see Materials section). Brains were sectioned in the coronal plane into 450-micro meter thick slices using a vibratome (Vibraslice, Boston, MA). Slices were then hemisected and placed on filter papers at the interface of a humidified carbogen (95% Oxygen2-5% CO2) gas phase and ACSF liquid phase. Slices were allowed at least 1 h for recovery before submersion in ACSF in a recording chamber. The ACSF was saturated with carbogen gas and perfused at a rate of 2.5 ml/min at room temperature (21-24 degrees Celsius). Rapid and accurate solution changes were made using a ValveBank8 computerized perfusion system (AutoMate Scientific, Oakland, CA). Thiopental concentrations were measured using high-pressure liquid chromatography. [18] 
Electrophysiology
Fiber volleys, population spikes (PS), and field excitatory postsynaptic potentials (EPSP) were evoked (0.5-15 V, 250 micro second, 0.1 Hz) with a bipolar tungsten stimulating electrode placed in stratum radiatum of area CA 1 (Figure 1). Stimulus pulses were delivered from constant-current isolation units (PSIU6) driven by a digitally timed controller (S 8800, Grass Medical Instruments, Quincy, MA). Fiber volleys were recorded with high resistance (1-2 M Omega) glass electrodes filled with ACSF and placed directly in line with stimulating electrodes in stratum radiatum, but separated from the stimulating electrode by 1.0-1.5 mm. Fiber volleys were isolated by blocking synaptic transmission using low calcium (< 0.2 mM) ACSF. Population spikes and EPSP responses were recorded with glass electrodes (0.2-1.0 M Omega) filled with ACSF and placed in stratum oriens or radiatum, respectively. Signals were amplified (5,000-20,000 times), filtered (DC to 10 KHz; Brownlee Precision, Santa Clara, CA) and digitized at 50 KHz using DataWave Technologies software (Longmont, CO) to control an analog to digital converter.
Data Analysis
All responses were measured using DataWave software, analyzed and graphed using IGOR Pro (WaveMetrics, Lake Oswego, OR) software. Fiber volley responses were measured as the peak negative to peak positive amplitude. Excitatory postsynaptic potentials amplitudes were measured as the peak negativity from baseline or as the slope of the early response between 20% and 80% of maximum. Population spike responses were measured as the peak negative to peak positive amplitude. All responses were normalized as a percent of control based on the average amplitude during a 10-min recording immediately before riluzole application. Statistical significance was determined by comparing riluzole effects with time-matched control responses using analysis of variance (AXUM, TriMetrix, Seattle, WA).
Materials
Rats were obtained from Simonsen Laboratories (Gilroy, CA). Riluzole was provided by Rhone-Poulenc Rorer (Collegeville, PA). Thiopental was obtained from Sigma (St. Louis, MO). All solutions were made up in high-pressure liquid chromatography and spectrophotometric grade water (OmniSolv) supplied by EM Science (Gibbstown, NJ). The ACSF had the following ionic composition (in mM): Sodium sup +, 151.25; Potassium sup +, 3.5; Calcium sup ++, 2.0; Magnesium sup ++, 2.0; Chlorine sup -, 130.5; HCO3sup -, 26; SO4sup -, 2.0; H2PO4sup -, 1.25; and glucose, 10. Chemicals for the ACSF were reagent grade or better and obtained from J.T. Baker (Philadelphia, PA).
Results
Riluzole Produced a Potent Depression of Population Spike Amplitudes
Evoked population spike amplitudes were depressed by low concentrations of riluzole (0.5-10 micro Meter), and a half maximal depression (54.2 plus/minus 12.6%; mean plus/minus SD, n = 9) occurred at 5.0 micro Meter (P < 0.001, analysis of variance compared with control). Initial effects were observed 5-8 min after application, and the half-time for onset was 13 plus/minus 1.7 min, determined from a first-order exponential fit. Steady-state effects were observed by approximately 25 min at a concentration of 5.0 micro Meter. Faster onset times were observed at 10 micro Meter (9 plus/minus 2.9 min), and complete PS depression occurred within 10 min at 20 micro Meter. Recovery from the riluzole-induced depression required more than 2 h. Riluzole-induced PS depression was accompanied by an increase in spike latency (approximately 2.0 ms) and decreases in initial and late positive components (Figure 2), indicative of effects on synaptic input and postsynaptic discharge.
Figure 2. Field recordings of evoked population spikes (PS). Records shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Note the increase in PS latency, evident in the overlay, that accompanied the depression produced by riluzole. The bottom is a time versus amplitude graph of population spikes showing the stability of responses during the control (predrug) 20-min period. Riluzole produced a marked reduction (approximately 50% of control) in PS amplitude, expressed as a percent of control.
Figure 2. Field recordings of evoked population spikes (PS). Records shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Note the increase in PS latency, evident in the overlay, that accompanied the depression produced by riluzole. The bottom is a time versus amplitude graph of population spikes showing the stability of responses during the control (predrug) 20-min period. Riluzole produced a marked reduction (approximately 50% of control) in PS amplitude, expressed as a percent of control.
Figure 2. Field recordings of evoked population spikes (PS). Records shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Note the increase in PS latency, evident in the overlay, that accompanied the depression produced by riluzole. The bottom is a time versus amplitude graph of population spikes showing the stability of responses during the control (predrug) 20-min period. Riluzole produced a marked reduction (approximately 50% of control) in PS amplitude, expressed as a percent of control.
×
Riluzole Did Not Alter gamma-Amino Butyric Acid-mediated Paired Pulse inhibition
Riluzole produced comparable depression of both the first and second PS responses to an identical pair of stimuli (Figure 3). The ratio of second/first PS response was 1.5 in control conditions and 1.9 in the presence of 5.0 micro Meter riluzole. Thiopental produced a greater depression of the second of a pair of PS responses (ratio = 0.65; Figure 3) at all effective concentrations (10-100 micro Meter). The PS depression produced by thiopental was not accompanied by an alteration in the early positivity. Riluzole, however, slowed the rate of rise and depressed peak amplitudes (Figure 3), suggesting that PS depression resulted from depressed excitatory synaptic input.
Figure 3. Comparison of effects produced by riluzole or thiopental on paired pulse recordings revealed differences in effects on recurrent inhibition. Riluzole produced an equivalent reduction of both population spikes after paired stimulus pulses. Thiopental produced a greater depression of the second population spike, indicating that enhanced gamma-amino butyric acid-mediated inhibition occurred. The bottom traces are expanded views of recordings showing that riluzole decreased the rate of rise for the early positive synaptic response, whereas thiopental did not.
Figure 3. Comparison of effects produced by riluzole or thiopental on paired pulse recordings revealed differences in effects on recurrent inhibition. Riluzole produced an equivalent reduction of both population spikes after paired stimulus pulses. Thiopental produced a greater depression of the second population spike, indicating that enhanced gamma-amino butyric acid-mediated inhibition occurred. The bottom traces are expanded views of recordings showing that riluzole decreased the rate of rise for the early positive synaptic response, whereas thiopental did not.
Figure 3. Comparison of effects produced by riluzole or thiopental on paired pulse recordings revealed differences in effects on recurrent inhibition. Riluzole produced an equivalent reduction of both population spikes after paired stimulus pulses. Thiopental produced a greater depression of the second population spike, indicating that enhanced gamma-amino butyric acid-mediated inhibition occurred. The bottom traces are expanded views of recordings showing that riluzole decreased the rate of rise for the early positive synaptic response, whereas thiopental did not.
×
Riluzole Depressed Excitatory Postsynaptic Potentials Responses
To determine whether riluzole depressed excitatory synaptic inputs to CA 1 neurons, EPSP responses from dendritic fields in stratum radiatum were recorded. Riluzole depressed EPSP amplitudes (Figure 4), and the second EPSP of a paired pulse response was depressed more (52 plus/minus 9.4% of control; n = 10) compared with the first (46.6 plus/minus 19.8%) in the presence of 5 micro Meter riluzole (P < 0.001, for both effects compared with control, P < 0.01 for the first EPSP effect compared with the second. To determine whether this degree of EPSP depression could account for the PS reduction produced by riluzole, the EPSP versus PS relation was studied (Figure 5). In control conditions, the slope for the EPSP versus PS curve was 1.35, so the depression of EPSP amplitudes observed can fully account for reduced PS responses (47 * 1.35 [nearly equal] 63%, compared with the observed 55% depression).
Figure 4. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) showing paired pulse facilitation. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Two time-versus-amplitude graphs of EPSP amplitudes show a marked reduction in both the first and second pulse responses after a 20-min application of riluzole (heavy line). Points represent the mean plus/minus SEM for five determinations from separate slices. Note the decrease in paired pulse facilitation that accompanied EPSP depression.
Figure 4. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) showing paired pulse facilitation. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Two time-versus-amplitude graphs of EPSP amplitudes show a marked reduction in both the first and second pulse responses after a 20-min application of riluzole (heavy line). Points represent the mean plus/minus SEM for five determinations from separate slices. Note the decrease in paired pulse facilitation that accompanied EPSP depression.
Figure 4. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) showing paired pulse facilitation. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Two time-versus-amplitude graphs of EPSP amplitudes show a marked reduction in both the first and second pulse responses after a 20-min application of riluzole (heavy line). Points represent the mean plus/minus SEM for five determinations from separate slices. Note the decrease in paired pulse facilitation that accompanied EPSP depression.
×
Figure 5. An overlay of 70-ms records shows change of population spike (PS) amplitude and excitatory postsynaptic potential (EPSP) slope at increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 2.0 to 7.0 V. Time versus PS amplitude graph (B) is shown by the solid line and the left axis; time versus EPSP slope is shown by the dotted line and the right axis. Both responses show a progressive decrease in amplitude as stimulus intensity was reduced, followed by increases as stimulus intensity was subsequently increased in steps back to initial stimulus strength. Arrows correspond to responses shown above in A. Population spike amplitude versus corresponding EPSP slope is seen in C; both values expressed as percentage of maximum response. A best fit of the data was found to have a slope of 1.35 over the linear region. Therefore, a 20% reduction in EPSP slope would result in a 27% decrease in population spike amplitude.
Figure 5. An overlay of 70-ms records shows change of population spike (PS) amplitude and excitatory postsynaptic potential (EPSP) slope at increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 2.0 to 7.0 V. Time versus PS amplitude graph (B) is shown by the solid line and the left axis; time versus EPSP slope is shown by the dotted line and the right axis. Both responses show a progressive decrease in amplitude as stimulus intensity was reduced, followed by increases as stimulus intensity was subsequently increased in steps back to initial stimulus strength. Arrows correspond to responses shown above in A. Population spike amplitude versus corresponding EPSP slope is seen in C; both values expressed as percentage of maximum response. A best fit of the data was found to have a slope of 1.35 over the linear region. Therefore, a 20% reduction in EPSP slope would result in a 27% decrease in population spike amplitude.
Figure 5. An overlay of 70-ms records shows change of population spike (PS) amplitude and excitatory postsynaptic potential (EPSP) slope at increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 2.0 to 7.0 V. Time versus PS amplitude graph (B) is shown by the solid line and the left axis; time versus EPSP slope is shown by the dotted line and the right axis. Both responses show a progressive decrease in amplitude as stimulus intensity was reduced, followed by increases as stimulus intensity was subsequently increased in steps back to initial stimulus strength. Arrows correspond to responses shown above in A. Population spike amplitude versus corresponding EPSP slope is seen in C; both values expressed as percentage of maximum response. A best fit of the data was found to have a slope of 1.35 over the linear region. Therefore, a 20% reduction in EPSP slope would result in a 27% decrease in population spike amplitude.
×
Excitatory postsynaptic potential depression was accompanied by a depression of Schaffer-collateral fiber volleys (Figure 6). Fiber volleys provide a measure of action potential conduction in presynaptic axons. Fiber volley amplitude was depressed by 36 plus/minus 17% (n = 13, P < 0.001) in the presence of 5 micro Meter riluzole, and the second response to a paired pulse was reduced by 47 plus/minus 9%. The fiber volley versus EPSP relation had a slope of 1.2 (Figure 7), so depressed action potential conduction could fully account for riluzole's reduction of EPSP amplitudes (36 * 1.2 [nearly equal] 43%, close to the observed 47% depression). Similarly, the increased depression observed for the second fiber volley correlated well with the increased riluzole effect on the second EPSP (56% calculated vs. 53% depression observed).
Figure 6. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) and fiber volleys showing that EPSP depression occurred with a decrease in fiber volley amplitude. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Bottom trace is an expanded view of the overlay's early portion, to highlight the decrease in fiber volley amplitude with 5.0 micro Meter riluzole (solid line) compared with control (dotted line).
Figure 6. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) and fiber volleys showing that EPSP depression occurred with a decrease in fiber volley amplitude. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Bottom trace is an expanded view of the overlay's early portion, to highlight the decrease in fiber volley amplitude with 5.0 micro Meter riluzole (solid line) compared with control (dotted line).
Figure 6. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) and fiber volleys showing that EPSP depression occurred with a decrease in fiber volley amplitude. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Bottom trace is an expanded view of the overlay's early portion, to highlight the decrease in fiber volley amplitude with 5.0 micro Meter riluzole (solid line) compared with control (dotted line).
×
Figure 7. An overlay of 60-ms records shows the relation of FV amplitude to excitatory postsynaptic potential (EPSP) amplitude with increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 1.5 to 5.0 V. Time versus EPSP amplitude is described by the solid line and the left axis (EPSP); time versus FV amplitude of the same preparation is shown by the dotted line and the right axis (FV). At 32 min, the stimulus intensity was reduced in steps and then gradually increased back to initial strengths (42 to 60 min). Excitatory postsynaptic potential amplitude versus corresponding fiber volley amplitude with increasing stimulus intensity is shown in C, both values expressed as a percentage of maximum response. A best fit to the data was found to have a slope of 1.2 over the linear region. A 20% decrease in FV amplitude would result in a 24% decrease in EPSP amplitude.
Figure 7. An overlay of 60-ms records shows the relation of FV amplitude to excitatory postsynaptic potential (EPSP) amplitude with increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 1.5 to 5.0 V. Time versus EPSP amplitude is described by the solid line and the left axis (EPSP); time versus FV amplitude of the same preparation is shown by the dotted line and the right axis (FV). At 32 min, the stimulus intensity was reduced in steps and then gradually increased back to initial strengths (42 to 60 min). Excitatory postsynaptic potential amplitude versus corresponding fiber volley amplitude with increasing stimulus intensity is shown in C, both values expressed as a percentage of maximum response. A best fit to the data was found to have a slope of 1.2 over the linear region. A 20% decrease in FV amplitude would result in a 24% decrease in EPSP amplitude.
Figure 7. An overlay of 60-ms records shows the relation of FV amplitude to excitatory postsynaptic potential (EPSP) amplitude with increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 1.5 to 5.0 V. Time versus EPSP amplitude is described by the solid line and the left axis (EPSP); time versus FV amplitude of the same preparation is shown by the dotted line and the right axis (FV). At 32 min, the stimulus intensity was reduced in steps and then gradually increased back to initial strengths (42 to 60 min). Excitatory postsynaptic potential amplitude versus corresponding fiber volley amplitude with increasing stimulus intensity is shown in C, both values expressed as a percentage of maximum response. A best fit to the data was found to have a slope of 1.2 over the linear region. A 20% decrease in FV amplitude would result in a 24% decrease in EPSP amplitude.
×
Riluzole Depressed Fiber Volleys in a Use-dependent Manner
The increased sensitivity observed for the second fiber volley suggested that a use-dependent depression of axonal conduction could contribute to riluzole's effect. To determine whether riluzole depressed axonal conduction, effects on isolated fiber volleys were studied (Figure 8) after block of synaptic responses (see Methods section). Riluzole depressed isolated fiber volleys in a concentration-dependent manner (0.5-20 micro Meter), with an EC50of 7.5 micro Meter. In the presence of 2 micro Meter riluzole, fiber volleys were depressed by 21 plus/minus 8% for first pulse responses (P < 0.001). The second and subsequent fiber volleys in a 30-Hz train showed a marked increased in sensitivity to riluzole (Figure 8). The second response in a train was depressed an additional 10% (to approximately 30%), and the 12th response was depressed by a further 20% compared with the first, for a total depression of approximately 40%. Onset of the riluzole effect was faster for responses near the end of the train, and recovery was slower.
Figure 8. Recordings of 12 consecutive fiber volleys separated by interpulse intervals of 30 ms (A) demonstrating a use-dependent decrease in amplitude produced by riluzole (2.0 micro Meter). The lower four traces (B) are expanded views of the first and last fiber volley in a train, highlighting the greater decrease in FV amplitude produced by riluzole on the last response compared with control. A graph of fiber volley amplitudes versus experimental time (C) shows the slow onset and recovery times after a 20-min application of riluzole. Later responses in the train were depressed sooner, but required more time for recovery, compared with early fiber volleys. Only responses to the first, fourth, eighth, and twelfth stimulus in each train were plotted. Values are expressed as a percent of control fiber volley amplitudes.
Figure 8. Recordings of 12 consecutive fiber volleys separated by interpulse intervals of 30 ms (A) demonstrating a use-dependent decrease in amplitude produced by riluzole (2.0 micro Meter). The lower four traces (B) are expanded views of the first and last fiber volley in a train, highlighting the greater decrease in FV amplitude produced by riluzole on the last response compared with control. A graph of fiber volley amplitudes versus experimental time (C) shows the slow onset and recovery times after a 20-min application of riluzole. Later responses in the train were depressed sooner, but required more time for recovery, compared with early fiber volleys. Only responses to the first, fourth, eighth, and twelfth stimulus in each train were plotted. Values are expressed as a percent of control fiber volley amplitudes.
Figure 8. Recordings of 12 consecutive fiber volleys separated by interpulse intervals of 30 ms (A) demonstrating a use-dependent decrease in amplitude produced by riluzole (2.0 micro Meter). The lower four traces (B) are expanded views of the first and last fiber volley in a train, highlighting the greater decrease in FV amplitude produced by riluzole on the last response compared with control. A graph of fiber volley amplitudes versus experimental time (C) shows the slow onset and recovery times after a 20-min application of riluzole. Later responses in the train were depressed sooner, but required more time for recovery, compared with early fiber volleys. Only responses to the first, fourth, eighth, and twelfth stimulus in each train were plotted. Values are expressed as a percent of control fiber volley amplitudes.
×
Discussion
Riluzole was more potent than other anesthetics at depressing CA 1 neuron population spike amplitudes, consistent with the higher potency for riluzole observed in vivo. [12,19] Riluzole depressed responses at lower concentrations (5 micro Meter) than halothane (EC50= 230 micro Meter), [20] thiopental (EC50= 50 micro Meter, Figure 2)* or propofol (EC50= 20 micro Meter). [21] Riluzole exhibited slower onset kinetics compared with these other anesthetics. The long duration of riluzole effects (160 min, Figure 8(C), approximately 4 times longer than thiopental) were also comparable with those seen in vivo, although the markedly different pharmacokinetic properties of the two systems make it difficult to relate drug effects seen in brain slices to those observed in vivo. Sleep times in vivo were more than 3.5 times longer after intraperitoneal riluzole (25 mg/kg; 178 plus/minus 23 min) compared with thiopental (25 mg/kg; 46 plus/minus 4 min) or ketamine (80 mg/kg; 47 plus/minus 2 min). [12] Halothane, in contrast, produced steady-state effects in brain slices within 2-3 min, and recovery occurred within 15 min, comparable with in vivo response times. [20-22] A slow onset and long duration of action are consistent with a use-dependent mechanism of action, where drug binding is dependent on a transient conformation of a receptor/channel. Results from the current study indicate that riluzole depressed CA 1 neuron discharge by blocking presynaptic conduction in a use-dependent manner.
Fiber Volley Depression Was Necessary and Sufficient for Riluzole-induced Population Spike Block
Riluzole-induced depression of synaptically evoked CA 1 neuron discharge was paralleled by an equivalent reduction in EPSP and fiber volley amplitudes (Figure 9). Input/output analyses of fiber volley versus EPSP versus PS relations demonstrated that depressed fiber volley amplitudes fully accounted for the depression of EPSP responses (Figure 7). Excitatory postsynaptic potential depression, in turn, could fully account for the depressed postsynaptic discharge observed (Figure 2, Figure 5, and Figure 6). Therefore, riluzole-induced depression of CA 1 neurons appeared to result from an action at a presynaptic site to block excitatory synaptic input. Enhanced recurrent inhibition was not produced by riluzole, because the ratio of second/first PS responses was not decreased (Figure 5). The second PS response is known to be preferentially depressed by some anesthetics because recurrent GABA-mediated inhibition, resulting from the first population discharge, is enhanced. [23-25] Although volatile anesthetics and barbiturates increase paired pulse inhibition, additional actions at several sites are required to account for depression of CA 1 neuron discharge produced by these anesthetics. These sites include depressed fiber conduction, reduced excitatory input, increased spike threshold, and enhanced GABA-mediated inhibition (Figure 1). [24-28] Riluzole appears to be unique among anesthetics because depressed CA 1 discharge can be attributed to a single site of action: blocked action potential conduction in presynaptic glutamate fibers.
Figure 9. Concentration-effect relations for riluzole-induced depression of fiber volley, excitatory postsynaptic potential (EPSP), and population spike amplitudes. All three responses were significantly depressed (P < 0.01, analysis of variance) by 2.0 micro Meter riluzole, and maximal effects occurred at approximately 20 micro Meter. Population spikes were depressed to a greater extent compared with EPSP and fiber volley responses. Each point represents the mean plus/minus SD (n greater or equal to 4), and curves are least squares fits to the means at each concentration. Note the similar slopes for each curve, consistent with a single effect site (depressed presynaptic conduction) leading to depression of EPSP and population spike responses.
Figure 9. Concentration-effect relations for riluzole-induced depression of fiber volley, excitatory postsynaptic potential (EPSP), and population spike amplitudes. All three responses were significantly depressed (P < 0.01, analysis of variance) by 2.0 micro Meter riluzole, and maximal effects occurred at approximately 20 micro Meter. Population spikes were depressed to a greater extent compared with EPSP and fiber volley responses. Each point represents the mean plus/minus SD (n greater or equal to 4), and curves are least squares fits to the means at each concentration. Note the similar slopes for each curve, consistent with a single effect site (depressed presynaptic conduction) leading to depression of EPSP and population spike responses.
Figure 9. Concentration-effect relations for riluzole-induced depression of fiber volley, excitatory postsynaptic potential (EPSP), and population spike amplitudes. All three responses were significantly depressed (P < 0.01, analysis of variance) by 2.0 micro Meter riluzole, and maximal effects occurred at approximately 20 micro Meter. Population spikes were depressed to a greater extent compared with EPSP and fiber volley responses. Each point represents the mean plus/minus SD (n greater or equal to 4), and curves are least squares fits to the means at each concentration. Note the similar slopes for each curve, consistent with a single effect site (depressed presynaptic conduction) leading to depression of EPSP and population spike responses.
×
Use-dependent Block of Sodium Channels and Riluzole-induced Depression
Riluzole, like some anesthetics, prevented the memory loss and hippocampal neuron degeneration after ischemia produced by transient carotid artery occlusion in rodents. [29] This neuroprotective effect was also seen in hippocampal slices and was suggested to result from depressed glutamate release, secondary to a block of voltage-activated sodium channels or altered function at N-methyl-D-aspartate receptors. [16,30,31] In Xenopus oocytes, 30 micro Meter riluzole depressed rat-brain-derived sodium channels by stabilizing inactivated channels. [15] Similarly, riluzole blocked sodium channel conductance in frog nerve with an EC50of approximately 50 micro Meter, but with an apparent dissociation constant of 0.29 micro Meter for block of inactivated channels. [32] This selective interaction of riluzole with inactivated sodium channels could explain the use-dependent depression of sodium channel-mediated conduction observed in the current study, although our results cannot address this issue directly. By analogy to the modulated receptor theory, these results suggest that riluzole binds selectively to open or inactivated sodium channels, or both, in a manner analogous to the open channel block produced by some local anesthetics. [15,33,34] The high degree of selectivity for riluzole block of sodium channels (more than 300 times greater than for Potassium sup + channel depression) [13,35] could explain why riluzole may be more useful for general anesthesia compared with traditional local anesthetics. [33] Local anesthetic-induced Potassium sup + channel block leads to hyperexcitable neuronal responses, which appear to counteract the depressant effects produced by Sodium sup + channel block in the central nervous system. [36] The consequences of such channel selectivity for general anesthesia remains to be determined for riluzole and other channel-blocking agents.
The authors thank Heath Lukatch, Catherine Hagan, and Sky Pittson for comments on the manuscript.
*Lukatch HS, Monroe FA, MacIver MB: Thiopental blocks paired-pulse facilitation in hippocampal CA1 neurons (abstract). Society for Neuroscience 1993; 19:275.
REFERENCES
Pocock G, Richards CD: Cellular mechanisms in general anaesthesia. Br J Anaesth 1991; 66:116-28.
Yamamura T, Harada K, Okamura A, Kemmotsu O: Is the site of action of ketamine anesthesia the N-methyl-D-aspartate receptor? ANESTHESIOLOGY 1990; 72:704-10.
Daniell LC: The noncompetitive N-methyl-D-aspartate antagonists, MK-801, phencyclidine and ketamine, increase the potency of general anesthetics. Pharmacol Biochem Behav 1990; 36:111-5.
Landau EM, Richter J, Cohen S: Differential solubilities in subregions of the membrane: A nonsteric mechanism of drug specificity. J Med Chem 1979; 22:325-7.
Kendig JJ, MacIver MB, Roth SH: Anesthetic actions in the hippocampal formation. Ann N Y Acad Sci 1991; 625:37-53.
Perouansky M, Baranov D, Salman M, Yaari Y: Effects of halothane on glutamate receptor-mediated excitatory postsynaptic currents. ANESTHESIOLOGY 1995; 83:109-19.
Tanelian DL, Kosek P, Mody I, MacIver MB: The role of the GABA sub A receptor/chloride channel complex in anesthesia. ANESTHESIOLOGY 1993; 78:757-76.
Macdonald RL, Olsen RW: GABA sub A receptor channels. Annu Rev Neurosci 1994; 17:569-602.
Schlame M, Hemmings HC: Inhibition by volatile anesthetics of endogenous glutamate release from synaptosomes by a presynaptic mechanism. ANESTHESIOLOGY 1995; 82:1406-16.
Cheramy A, Barbeito L, Godeheu G, Glowinski J: Riluzole inhibits the release of glutamate in the caudate nucleus of the cat in vivo. Neurosci Lett 1992; 147:209-12.
Martin D, Thompson MA, Nadler JV: The neuroprotective agent riluzole inhibits release of glutamate and aspartate from slices of hippocampal area CA1. Eur J Pharmacol 1993; 250:473-6.
Mantz J, Cheramy A, Thierry AM, Glowinski J, Desmonts JM: Anesthetic properties of riluzole (54274 RP), a new inhibitor of glutamate neurotransmission. ANESTHESIOLOGY 1992; 76:844-8.
Benoit E, Escande D: Riluzole specifically blocks inactivated Sodium channels in myelinated nerve fibre. Pflugers Arch 1991; 419:603-9.
Debono MW, Le Guern J, Canton T, Doble A, Pradier L: Inhibition by riluzole of electrophysiological responses mediated by rat kainate and NMDA receptors expressed in Xenopus oocytes. Eur J Pharmacol 1993; 235:283-9.
Hebert T, Drapeau P, Pradier L, Dunn RJ: Block of the rat brain IIA sodium channel alpha subunit by the neuroprotective drug riluzole. Mol Pharmacol 1994; 45:1055-60.
Malgouris C, Daniel M, Doble A: Neuroprotective effects of riluzole on N-methyl-D-aspartate- or veratridine-induced neurotoxicity in rat hippocampal slices. Neurosci Lett 1994; 177:95-9.
Mantz J, Laudenbach V, Lecharny JB, Henzel D, Desmonts JM: Riluzole, a novel antiglutamate, blocks GABA uptake by striatal synaptosomes. Eur J Pharmacol 1994; 257:R7-8.
Ebling WF, Mills-Williams L, Harapat SR, Stanski DR: High-performance liquid chromatographic method for determining thiopental concentrations in twelve rat tissues: Application to physiologic modeling of disposition of barbiturate. J Chromatogr 1989; 490:339-53.
Stutzmann JM, Lucas M, Blanchard JC, Laduron PM: Riluzole, a glutamate antagonist, enhances slow wave and REM sleep in rats. Neurosci Lett 1988; 88:195-200.
MacIver MB, Roth SH: Inhalation anaesthetics exhibit pathway-specific and differential actions on hippocampal synaptic responses in vitro. Br J Anaesth 1988; 60:680-91.
Travis VL, MacIver MB: Propofol enhances GABA sub A,slow feed forward inhibition in CA1 neuron dentrites (abstract). ANESTHESIOLOGY 1995; 83:A751.
MacIver MB, Tauck DL, Kendig JJ: General anaesthetic modification of synaptic facilitation and long-term potentiation in hippocampus. Br J Anaesth 1989; 62:301-10.
Mody I, Tanelian DL, MacIver MB: Halothane enhances tonic neuronal inhibition by elevating intracellular calcium. Brain Res 1991; 538:319-23.
MacIver MB, Tanelian DL, Mody I: Two mechanisms for anesthetic-induced enhancement of GABA sub A -mediated neuronal inhibition. Ann N Y Acad Sci 1991; 625:91-6.
MacIver MB, Roth SH: Barbiturate effects on hippocampal excitatory synaptic responses are selective and pathway specific. Can J Physiol Pharmacol 1987; 65:385-94.
MacIver MB, Kendig JJ: Anesthetic effects on resting membrane potential are voltage-dependent and agent-specific. ANESTHESIOLOGY 1991; 74:83-8.
Lukatch HS, MacIver MB: Prolonged GABA sub A chloride currents underlie the thiopental induced slowing of EEG (abstract). ANESTHESIOLOGY 1994; 81:A798.
Mikulec AA, Amagasu SM, Monroe FA, MacIver MB: Three sites of action are necessary and sufficient for halothane-induced depression of hippocampal CA 1 neurons (abstract). ANESTHESIOLOGY 1995; 83:A1266.
Malgouris C, Bardot F, Daniel M, Pellis F, Rataud J, Uzan A, Blanchard JC, Laduron PM: Riluzole, a novel antiglutamate, prevents memory loss and hippocampal neuronal damage in ischemic gerbils. J Neurosci 1989; 9:3720-7.
Pratt J, Rataud J, Bardot F, Roux M, Blanchard JC, Laduron PM, Stutzmann JM: Neuroprotective actions of riluzole in rodent models of global and focal cerebral ischaemia. Neurosci Lett 1992; 140:225-30.
Hubert JP, Delumeau JC, Glowinski J, Premont J, Doble A: Antagonism by riluzole of entry of calcium evoked by NMDA and veratridine in rat cultured granule cells: Evidence for a dual mechanism of action. Br J Pharmacol 1994; 113:261-7.
Boireau A, Miquet JM, Dubedat P, Meunier M, Doble A: Riluzole and experimental parkinsonism: Partial antagonism of MPP(+)-induced increase in striatal extracellular dopamine in rats in vivo. Neuroreport 1994; 5:2157-60.
Butterworth JT, Strichartz GR: Molecular mechanisms of local anesthesia: A review. ANESTHESIOLOGY 1990; 72:711-34.
Guo XT, Castle NA, Chernoff DM, Strichartz GR: Comparative inhibition of voltage-gated cation channels by local anesthetics. Ann N Y Acad Sci 1991; 625:181-99.
Benoit E, Escande D: Fast Potassium channels are more sensitive to riluzole than slow Potassium channels in myelinated nerve fibre. Pflugers Arch 1993; 422:536-8.
Brown DL, Ransom DM, Hall JA, Leicht CH, Schroeder DR, Offord KP: Regional anesthesia and local anesthetic-induced systemic toxicity: Seizure frequency and accompanying cardiovascular changes. Anesth Analg 1995; 81:321-8.
Figure 1. Drawing of a hippocampal slice (A) showing the relative positions of a bipolar tungsten stimulating electrode (STIMULATE), and extracellular glass recording electrodes (RECORD). The recording electrode position was different for population spikes (PS) than for fiber volleys (FV) and excitatory postsynaptic potentials (EPSP). The three major excitatory pathways in hippocampal slices are shown: the perforant path (pp) originates in entorhinal cortex and projects to granule cells of the dentate gurus (DG), the mossy fiber pathway (mf) comprises granule cell axons and projects to pyramidal cells of CA 3, and Schaffer collateral (sc) fibers from CA 3 neurons form excitatory synapses with dendrites of CA 1 neurons. Synaptically evoked discharge (PS) was used to measure riluzole effects on the output of the CA 1 circuit (B). Circled numbers correspond to the four measures used to determine the major sites of action for riluzole in this circuit (see beginning of article).
Figure 1. Drawing of a hippocampal slice (A) showing the relative positions of a bipolar tungsten stimulating electrode (STIMULATE), and extracellular glass recording electrodes (RECORD). The recording electrode position was different for population spikes (PS) than for fiber volleys (FV) and excitatory postsynaptic potentials (EPSP). The three major excitatory pathways in hippocampal slices are shown: the perforant path (pp) originates in entorhinal cortex and projects to granule cells of the dentate gurus (DG), the mossy fiber pathway (mf) comprises granule cell axons and projects to pyramidal cells of CA 3, and Schaffer collateral (sc) fibers from CA 3 neurons form excitatory synapses with dendrites of CA 1 neurons. Synaptically evoked discharge (PS) was used to measure riluzole effects on the output of the CA 1 circuit (B). Circled numbers correspond to the four measures used to determine the major sites of action for riluzole in this circuit (see beginning of article).
Figure 1. Drawing of a hippocampal slice (A) showing the relative positions of a bipolar tungsten stimulating electrode (STIMULATE), and extracellular glass recording electrodes (RECORD). The recording electrode position was different for population spikes (PS) than for fiber volleys (FV) and excitatory postsynaptic potentials (EPSP). The three major excitatory pathways in hippocampal slices are shown: the perforant path (pp) originates in entorhinal cortex and projects to granule cells of the dentate gurus (DG), the mossy fiber pathway (mf) comprises granule cell axons and projects to pyramidal cells of CA 3, and Schaffer collateral (sc) fibers from CA 3 neurons form excitatory synapses with dendrites of CA 1 neurons. Synaptically evoked discharge (PS) was used to measure riluzole effects on the output of the CA 1 circuit (B). Circled numbers correspond to the four measures used to determine the major sites of action for riluzole in this circuit (see beginning of article).
×
Figure 2. Field recordings of evoked population spikes (PS). Records shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Note the increase in PS latency, evident in the overlay, that accompanied the depression produced by riluzole. The bottom is a time versus amplitude graph of population spikes showing the stability of responses during the control (predrug) 20-min period. Riluzole produced a marked reduction (approximately 50% of control) in PS amplitude, expressed as a percent of control.
Figure 2. Field recordings of evoked population spikes (PS). Records shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Note the increase in PS latency, evident in the overlay, that accompanied the depression produced by riluzole. The bottom is a time versus amplitude graph of population spikes showing the stability of responses during the control (predrug) 20-min period. Riluzole produced a marked reduction (approximately 50% of control) in PS amplitude, expressed as a percent of control.
Figure 2. Field recordings of evoked population spikes (PS). Records shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Note the increase in PS latency, evident in the overlay, that accompanied the depression produced by riluzole. The bottom is a time versus amplitude graph of population spikes showing the stability of responses during the control (predrug) 20-min period. Riluzole produced a marked reduction (approximately 50% of control) in PS amplitude, expressed as a percent of control.
×
Figure 3. Comparison of effects produced by riluzole or thiopental on paired pulse recordings revealed differences in effects on recurrent inhibition. Riluzole produced an equivalent reduction of both population spikes after paired stimulus pulses. Thiopental produced a greater depression of the second population spike, indicating that enhanced gamma-amino butyric acid-mediated inhibition occurred. The bottom traces are expanded views of recordings showing that riluzole decreased the rate of rise for the early positive synaptic response, whereas thiopental did not.
Figure 3. Comparison of effects produced by riluzole or thiopental on paired pulse recordings revealed differences in effects on recurrent inhibition. Riluzole produced an equivalent reduction of both population spikes after paired stimulus pulses. Thiopental produced a greater depression of the second population spike, indicating that enhanced gamma-amino butyric acid-mediated inhibition occurred. The bottom traces are expanded views of recordings showing that riluzole decreased the rate of rise for the early positive synaptic response, whereas thiopental did not.
Figure 3. Comparison of effects produced by riluzole or thiopental on paired pulse recordings revealed differences in effects on recurrent inhibition. Riluzole produced an equivalent reduction of both population spikes after paired stimulus pulses. Thiopental produced a greater depression of the second population spike, indicating that enhanced gamma-amino butyric acid-mediated inhibition occurred. The bottom traces are expanded views of recordings showing that riluzole decreased the rate of rise for the early positive synaptic response, whereas thiopental did not.
×
Figure 4. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) showing paired pulse facilitation. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Two time-versus-amplitude graphs of EPSP amplitudes show a marked reduction in both the first and second pulse responses after a 20-min application of riluzole (heavy line). Points represent the mean plus/minus SEM for five determinations from separate slices. Note the decrease in paired pulse facilitation that accompanied EPSP depression.
Figure 4. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) showing paired pulse facilitation. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Two time-versus-amplitude graphs of EPSP amplitudes show a marked reduction in both the first and second pulse responses after a 20-min application of riluzole (heavy line). Points represent the mean plus/minus SEM for five determinations from separate slices. Note the decrease in paired pulse facilitation that accompanied EPSP depression.
Figure 4. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) showing paired pulse facilitation. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Two time-versus-amplitude graphs of EPSP amplitudes show a marked reduction in both the first and second pulse responses after a 20-min application of riluzole (heavy line). Points represent the mean plus/minus SEM for five determinations from separate slices. Note the decrease in paired pulse facilitation that accompanied EPSP depression.
×
Figure 5. An overlay of 70-ms records shows change of population spike (PS) amplitude and excitatory postsynaptic potential (EPSP) slope at increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 2.0 to 7.0 V. Time versus PS amplitude graph (B) is shown by the solid line and the left axis; time versus EPSP slope is shown by the dotted line and the right axis. Both responses show a progressive decrease in amplitude as stimulus intensity was reduced, followed by increases as stimulus intensity was subsequently increased in steps back to initial stimulus strength. Arrows correspond to responses shown above in A. Population spike amplitude versus corresponding EPSP slope is seen in C; both values expressed as percentage of maximum response. A best fit of the data was found to have a slope of 1.35 over the linear region. Therefore, a 20% reduction in EPSP slope would result in a 27% decrease in population spike amplitude.
Figure 5. An overlay of 70-ms records shows change of population spike (PS) amplitude and excitatory postsynaptic potential (EPSP) slope at increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 2.0 to 7.0 V. Time versus PS amplitude graph (B) is shown by the solid line and the left axis; time versus EPSP slope is shown by the dotted line and the right axis. Both responses show a progressive decrease in amplitude as stimulus intensity was reduced, followed by increases as stimulus intensity was subsequently increased in steps back to initial stimulus strength. Arrows correspond to responses shown above in A. Population spike amplitude versus corresponding EPSP slope is seen in C; both values expressed as percentage of maximum response. A best fit of the data was found to have a slope of 1.35 over the linear region. Therefore, a 20% reduction in EPSP slope would result in a 27% decrease in population spike amplitude.
Figure 5. An overlay of 70-ms records shows change of population spike (PS) amplitude and excitatory postsynaptic potential (EPSP) slope at increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 2.0 to 7.0 V. Time versus PS amplitude graph (B) is shown by the solid line and the left axis; time versus EPSP slope is shown by the dotted line and the right axis. Both responses show a progressive decrease in amplitude as stimulus intensity was reduced, followed by increases as stimulus intensity was subsequently increased in steps back to initial stimulus strength. Arrows correspond to responses shown above in A. Population spike amplitude versus corresponding EPSP slope is seen in C; both values expressed as percentage of maximum response. A best fit of the data was found to have a slope of 1.35 over the linear region. Therefore, a 20% reduction in EPSP slope would result in a 27% decrease in population spike amplitude.
×
Figure 6. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) and fiber volleys showing that EPSP depression occurred with a decrease in fiber volley amplitude. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Bottom trace is an expanded view of the overlay's early portion, to highlight the decrease in fiber volley amplitude with 5.0 micro Meter riluzole (solid line) compared with control (dotted line).
Figure 6. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) and fiber volleys showing that EPSP depression occurred with a decrease in fiber volley amplitude. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Bottom trace is an expanded view of the overlay's early portion, to highlight the decrease in fiber volley amplitude with 5.0 micro Meter riluzole (solid line) compared with control (dotted line).
Figure 6. Field recordings of evoked excitatory postsynaptic potentials (EPSPs) and fiber volleys showing that EPSP depression occurred with a decrease in fiber volley amplitude. Traces shown are for control, 5.0 micro Meter riluzole, and an overlay for comparison. Bottom trace is an expanded view of the overlay's early portion, to highlight the decrease in fiber volley amplitude with 5.0 micro Meter riluzole (solid line) compared with control (dotted line).
×
Figure 7. An overlay of 60-ms records shows the relation of FV amplitude to excitatory postsynaptic potential (EPSP) amplitude with increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 1.5 to 5.0 V. Time versus EPSP amplitude is described by the solid line and the left axis (EPSP); time versus FV amplitude of the same preparation is shown by the dotted line and the right axis (FV). At 32 min, the stimulus intensity was reduced in steps and then gradually increased back to initial strengths (42 to 60 min). Excitatory postsynaptic potential amplitude versus corresponding fiber volley amplitude with increasing stimulus intensity is shown in C, both values expressed as a percentage of maximum response. A best fit to the data was found to have a slope of 1.2 over the linear region. A 20% decrease in FV amplitude would result in a 24% decrease in EPSP amplitude.
Figure 7. An overlay of 60-ms records shows the relation of FV amplitude to excitatory postsynaptic potential (EPSP) amplitude with increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 1.5 to 5.0 V. Time versus EPSP amplitude is described by the solid line and the left axis (EPSP); time versus FV amplitude of the same preparation is shown by the dotted line and the right axis (FV). At 32 min, the stimulus intensity was reduced in steps and then gradually increased back to initial strengths (42 to 60 min). Excitatory postsynaptic potential amplitude versus corresponding fiber volley amplitude with increasing stimulus intensity is shown in C, both values expressed as a percentage of maximum response. A best fit to the data was found to have a slope of 1.2 over the linear region. A 20% decrease in FV amplitude would result in a 24% decrease in EPSP amplitude.
Figure 7. An overlay of 60-ms records shows the relation of FV amplitude to excitatory postsynaptic potential (EPSP) amplitude with increasing stimulus intensities (A). The lower case letters shown are for different stimulus intensities and correspond to the time points indicated in B. Stimulus intensity ranged from 1.5 to 5.0 V. Time versus EPSP amplitude is described by the solid line and the left axis (EPSP); time versus FV amplitude of the same preparation is shown by the dotted line and the right axis (FV). At 32 min, the stimulus intensity was reduced in steps and then gradually increased back to initial strengths (42 to 60 min). Excitatory postsynaptic potential amplitude versus corresponding fiber volley amplitude with increasing stimulus intensity is shown in C, both values expressed as a percentage of maximum response. A best fit to the data was found to have a slope of 1.2 over the linear region. A 20% decrease in FV amplitude would result in a 24% decrease in EPSP amplitude.
×
Figure 8. Recordings of 12 consecutive fiber volleys separated by interpulse intervals of 30 ms (A) demonstrating a use-dependent decrease in amplitude produced by riluzole (2.0 micro Meter). The lower four traces (B) are expanded views of the first and last fiber volley in a train, highlighting the greater decrease in FV amplitude produced by riluzole on the last response compared with control. A graph of fiber volley amplitudes versus experimental time (C) shows the slow onset and recovery times after a 20-min application of riluzole. Later responses in the train were depressed sooner, but required more time for recovery, compared with early fiber volleys. Only responses to the first, fourth, eighth, and twelfth stimulus in each train were plotted. Values are expressed as a percent of control fiber volley amplitudes.
Figure 8. Recordings of 12 consecutive fiber volleys separated by interpulse intervals of 30 ms (A) demonstrating a use-dependent decrease in amplitude produced by riluzole (2.0 micro Meter). The lower four traces (B) are expanded views of the first and last fiber volley in a train, highlighting the greater decrease in FV amplitude produced by riluzole on the last response compared with control. A graph of fiber volley amplitudes versus experimental time (C) shows the slow onset and recovery times after a 20-min application of riluzole. Later responses in the train were depressed sooner, but required more time for recovery, compared with early fiber volleys. Only responses to the first, fourth, eighth, and twelfth stimulus in each train were plotted. Values are expressed as a percent of control fiber volley amplitudes.
Figure 8. Recordings of 12 consecutive fiber volleys separated by interpulse intervals of 30 ms (A) demonstrating a use-dependent decrease in amplitude produced by riluzole (2.0 micro Meter). The lower four traces (B) are expanded views of the first and last fiber volley in a train, highlighting the greater decrease in FV amplitude produced by riluzole on the last response compared with control. A graph of fiber volley amplitudes versus experimental time (C) shows the slow onset and recovery times after a 20-min application of riluzole. Later responses in the train were depressed sooner, but required more time for recovery, compared with early fiber volleys. Only responses to the first, fourth, eighth, and twelfth stimulus in each train were plotted. Values are expressed as a percent of control fiber volley amplitudes.
×
Figure 9. Concentration-effect relations for riluzole-induced depression of fiber volley, excitatory postsynaptic potential (EPSP), and population spike amplitudes. All three responses were significantly depressed (P < 0.01, analysis of variance) by 2.0 micro Meter riluzole, and maximal effects occurred at approximately 20 micro Meter. Population spikes were depressed to a greater extent compared with EPSP and fiber volley responses. Each point represents the mean plus/minus SD (n greater or equal to 4), and curves are least squares fits to the means at each concentration. Note the similar slopes for each curve, consistent with a single effect site (depressed presynaptic conduction) leading to depression of EPSP and population spike responses.
Figure 9. Concentration-effect relations for riluzole-induced depression of fiber volley, excitatory postsynaptic potential (EPSP), and population spike amplitudes. All three responses were significantly depressed (P < 0.01, analysis of variance) by 2.0 micro Meter riluzole, and maximal effects occurred at approximately 20 micro Meter. Population spikes were depressed to a greater extent compared with EPSP and fiber volley responses. Each point represents the mean plus/minus SD (n greater or equal to 4), and curves are least squares fits to the means at each concentration. Note the similar slopes for each curve, consistent with a single effect site (depressed presynaptic conduction) leading to depression of EPSP and population spike responses.
Figure 9. Concentration-effect relations for riluzole-induced depression of fiber volley, excitatory postsynaptic potential (EPSP), and population spike amplitudes. All three responses were significantly depressed (P < 0.01, analysis of variance) by 2.0 micro Meter riluzole, and maximal effects occurred at approximately 20 micro Meter. Population spikes were depressed to a greater extent compared with EPSP and fiber volley responses. Each point represents the mean plus/minus SD (n greater or equal to 4), and curves are least squares fits to the means at each concentration. Note the similar slopes for each curve, consistent with a single effect site (depressed presynaptic conduction) leading to depression of EPSP and population spike responses.
×