At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes

<p>Abstract</p> <p>Background</p> <p>Many sedative agents, including anesthetics, produce explicit memory impairment by largely unknown mechanisms. Sharp-wave ripple (SPW-R) complexes are network activity thought to represent the neuronal substrate for information transfer from the hippocampal to neocortical circuits, contributing to the explicit memory consolidation. In this study we examined and compared the actions of two barbiturates with distinct amnesic actions, the general anesthetic thiopental and the anticonvulsant phenobarbital, on in vitro SPW-R activity.</p> <p>Results</p> <p>Using an in vitro model of SPW-R activity we found that thiopental (50–200 μM) significantly and concentration-dependently reduced the incidence of SPW-R events (it increased the inter-event period by 70–430 %). At the concentration of 25 μM, which clinically produces mild sedation and explicit memory impairment, thiopental significantly reduced the quantity of ripple oscillation (it reduced the number of ripples and the duration of ripple episodes by 20 ± 5%, n = 12, <it>P </it>< 0.01), and suppressed the rhythmicity of SPWs by 43 ± 15% (n = 6, <it>P </it>< 0.05). The drug disrupted the synchrony of SPWs within the CA1 region at 50 μM (by 19 ± 12%; n = 5, <it>P </it>< 0.05). Similar effects of thiopental were observed at higher concentrations. Thiopental did not affect the frequency of ripple oscillation at any of the concentrations tested (10–200 μM). Furthermore, the drug significantly prolonged single SPWs at concentrations ≥50 μM (it increased the half-width and the duration of SPWs by 35–90 %). Thiopental did not affect evoked excitatory synaptic potentials and its results on SPW-R complexes were also observed under blockade of NMDA receptors. Phenobarbital significantly accelerated SPWs at 50 and 100 μM whereas it reduced their rate at 200 and 400 μM. Furthermore, it significantly prolonged SPWs, reduced their synchrony and reduced the quantity of ripples only at the clinically very high concentration of 400 μM, reported to affect memory.</p> <p>Conclusion</p> <p>We hypothesize that thiopental, by interfering with SPW-R activity, through enhancement of the GABA_Areceptor-mediated transmission, affects memory processes which involve hippocampal circuit activation. The quantity but not the frequency of ripple oscillation was affected by the drug.</p

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-5

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p>eceptor antagonist CPP (10 μM, blue bar) produced reversible reduction in the rate of SPWs, a similar effect to that observed when the anesthetic was applied alone (compare with Fig. 3A). Inserts show examples of SPWs (traces on the top, low-pass filtered) and ripples (traces on the bottom, band-pass filtered) selected from times before and during thiopental application (arrows indicating the panel on the left and right respectively). Note that under thiopental SPWs are prolonged and ripple episode was shortened, containing decreased number of ripple waves. Calibration bars in the inserts: 50 μV, 25 ms for SPW; 25 μV, 50 ms for ripples. . Histogram of instantaneous period of SPW-R activity recorded from the CA1 st. pyramidale of a slice which received 100 μM of thiopental (red bar) in the presence of CPP (blue bar). Inserts show the power spectra, at the ripple range, generated from epochs of 2 min taken from the different conditions (i.e. in CPP, in thiopental and after washing-out thiopental) as indicated by arrows. . Collective data of the effects of thiopental on SPWs' (C) and Ripples' (D) measures in the presence of CPP. Measures of SPWs and Ripples were obtained from 9 and 7 slices respectively, at the concentration of 50 μM and from 8 and 5 slices respectively at the concentration of 100 μM. Asterisks indicate statistically significant drug effects (paired t-test, at < 0.05). Error bars not shown were smaller than the size of the symbol

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-2

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p>-event period. Horizontal bars in "b" denote the time of application of consecutively larger drug concentrations. Relatively large data values of period during 100 and 200 μM of the drug are curtailed for reasons of clarity of lower values. Note the great value dispersion of instant period during drug application at ≥50 μM. Data in "a" and "b" were obtained from two different slices. The collective data plot (percent changes) of period is shown in (c). The asterisk here, as in the subsequent collective plots in the figure, denotes the lowest concentration with statistically significant drug action (paired t-test, < 0.001 at 50 μM and < 0.05 at 100–200 μM). The number of slices studied at the five concentrations, of 10, 25, 50, 100 and 200 μM, were respectively 12, 18, 22, 12 and 8. Error bars not shown were smaller than the size of the symbol. . Example of auto-correlation before and during 50 μM thiopental (a) and collective auto-correlation plot (b) showing the disruption of rhythmicity at concentrations ≥25 μM (paired t-test, < 0.05 at 25 μM and < 0.01 at ≥50 μM). Numbers of slices studied at the five concentrations were 5, 6, 12, 8 and 4. . Example of thiopental-induced prolongation of SPWs (a) at two different concentrations. Averages of low-pass filtered sweeps taken from a two minute epoch are shown. Dotted traces are control sweeps. The corresponding collective plot (b) shows that significant prolongation of single SPWs occurred at drug concentrations ≥50 μM (paired t-test, < 0.01). . Effect of thiopental on SPWs' synchronization along the CA1 region. Single SPWs simultaneously recorded from two locations along the CA1 st. pyramidale, measuring 1 mm, are shown in "a", before (traces on the left) and during application of 50 μM thiopental (traces on the right). Sweeps were low-pass filtered in order to make feasible the time-discrimination of their peaks. Note the increased phase-lag (marked by dotted lines) during drug application (9.25 ms) compared with control condition (3.8 ms). The cross-correlation plot from another slice (b) illustrates the large decrease of function value during drug application at 100 μM. The collective plot (c) shows that significant suppression of synchrony started at 25 μM and reached a plateau at 100 μM (< 0.05 at 50 μM and < 0.01 at ≥50 μM). Numbers of slices studied at the five concentrations were 3, 5, 5, 4 and 4

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-4

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p>dium (left panels) and in medium containing 50 or 100 μM of thiopental (right panels). fEPSPs and spontaneous activity were simultaneously recorded from two neighboring slices in "A" and from the same slice in "B". Note that the drug did not affect fEPSPs despite its suppressive effect on spontaneous activity. . Grouped data of fEPSP slope for the two concentrations of thiopental. The drug did not significantly change the slope of fEPSP at any concentration

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-0

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p>ottom). Data were obtained from a slice four hours after its placement on the recording chamber. Note the persistency and stability over time of both characteristics of SPWs. . Record of spontaneous activity obtained from a different slice, presented at a faster sweep speed. . A wide band record of a single SPW-R event (C1) and the filtered sweeps are shown, illustrating the distinct components of the event. Low-pass (0.5–30 Hz) and band-pass (100–300 Hz) filtering reveals the slow wave (C2) and the associated ripple oscillation (C3) respectively. Band-pass filtering at very high frequencies (0.5–1 kHz, C4) discloses the burst-like multiunit activity occurring mostly during the rising and peaking phases of the slow positive potential. . FFT from 1 min raw record of spontaneous activity showing the difference in power between SPWs (upper diagram) and ripples (lower diagram, same power spectrum shown at a greater magnification). SPW dominant peak is at 5–10 Hz. Ripple activity is at ~150 Hz. . Histogram of period (inter-event interval) of SPWs calculated from a population of slices, showing that most of the slices displayed a period between 0.2–0.6 sec, which correspond to a frequency range of 1.5–5.0 Hz. . Auto-correlogram from a 5 min low-pass record from a distinct experiment, showing the typical degree of rhythmicity of SPWs

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-6

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p> activity was increased at 50 and 100 μM and reduced at 200 and 400 μM (asterisk denotes the lowest drug concentration with statistical significance effect, paired t-test, < 0.001 paired t t-test, < 0.05). In the plot of instantaneous period (b) note that even at the highest concentration (400 μM) phenobarbital did not abolish spontaneous activity. Measures for constructing the plot of collective data (c) were obtained from 3, 6, 10, 12 and 18 slices respectively for the five drug concentrations. . Example of auto-correlation analysis for SPWs recorded in one slice before and during application of 100 μM of phenobarbital (diagram on the left). The collective data plot (diagram on the right) illustrates that only at 400 μM did the drug significantly reduce rhythmicity of SPWs (asterisk, < 0.05). Data were obtained from 6, 10, 10 and 12 slices respectively for the four drug concentrations. . Averages of filtered SPWs recorded from an epoch lasting two minutes (a) obtained from a slice bathed in normal medium (trace with the smallest amplitude) and increasing concentrations of phenobarbital (50, 200 and 400 μM, traces with consecutively increasing amplitude). Note the concomitant increase in the falling phase and the amplitude of the potentials. The drug significantly increased half-width of SPWs at the highest concentration of 400 μM (b). However, the drug did not significantly change the amplitude of SPWs at any concentration (c). Data in collective plots were obtained from 6, 10, 12 and 18 slices respectively for the four drug concentrations in half-width and from 3, 6, 10, 12 and 18 slices for the five concentrations in amplitude. . Phenobarbital significantly decreased synchronization only at 400 μM, as illustrated in the example (on the left) and collective diagram (on the right, paired t-test, < 0.05). The concentration of 25 μM is missing from the plots of auto-correlation, half-width and cross-correlation because measures of these variables were obtained from only one experiment

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-1

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p>r (b). The dotted line in "a" marks the temperature of 32°C. In "b", the three representative temperatures (32°C, 27°C and 37°C) are indicated for purpose of comparison. During the increase of the temperature from 34–35°C to higher values, the histogram of the period bifurcated. This deviation of the values was due to the fact that SPWs were frequently generated in a burst-like fashion (shown in B), where consecutive waves appeared at short intervals, whereas at the same time the period between bursts of events increased. . Continuous recording of SPW-R complexes (left column) and power spectra at the high frequency range (right column) obtained at different temperatures. Single ripple episodes (band-pass filtered traces) are shown as inserts in spectral plots; calibration bars: 10 μV, 10 ms. At 37°C SPWs appeared frequently as bursts of several events. Ripple frequency changed monotonically with temperature. Note that at 37°C ripple frequency exceeded 200 Hz. Data in "A" an "B" were collected from two different slices. . Collective date of the period of SPWs and of the frequency of ripple oscillation at three different temperatures. Asterisks indicate significant differences with reference to the group of 32°C (paired t-test, at < 0.05, n = 4)

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-7

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p>ests, < 0.05). . Power spectra obtained from different times of a single experiment illustrating the reduction in ripple power only at high drug concentrations (200 and 400 μM). . Diagrams of collective data for the different measures of the ripple oscillation. Note that the reducing effect of phenobarbital on number, duration and ripple power was statistically significant (asterisks, < 0.05) only at the highest concentration of 400 μM. Measures of the above variables (except power) at the five drug concentrations were obtained from 2, 3, 7, 5 and 9 slices. Values of power were obtained from 4, 7, 12, 9 and 12 slices

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-3

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p> of ripple episode by thiopental. Note that even at 100 μM of the drug, which strongly suppressed the amount of ripples, the oscillation frequency remained unchanged. . Power spectra from three experiments (a-c) showing the reduction of ripple power at 50 and 100 μM but not at lower concentrations. . Plots of collective data of drug effects (percent changes) on the various measures of ripple oscillation. Asterisks mark the lowest concentration with statistically significant drug action (see text for details of statistical analysis). Note that ripple number and ripple episode duration were more sensitive than ripple power and ripple amplitude. Measures of number, duration and amplitude for each of the five drug concentrations (10, 25, 50, 100 and 200 μM) were respectively collected from 10, 12, 17, 10 and 8 slices. The corresponding numbers of slices for ripple power measures were respectively 12, 14, 22, 15, and 9. . Recordings of ripples (upper traces) and multiunits (middle traces), revealed after filtering original records, showing the thiopental-induced reduction in the rate of multiunits during ripple episodes. Dotted line indicates the threshold level set in this experiment for spike detection. Histograms of multiunit rate (bottom panel) triggered by peaks of SPWs (not shown) used to measure the mean rate under the different experimental conditions

At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes-8

<p><b>Copyright information:</b></p><p>Taken from "At clinically relevant concentrations the anaesthetic/amnesic thiopental but not the anticonvulsant phenobarbital interferes with hippocampal sharp wave-ripple complexes"</p><p>http://www.biomedcentral.com/1471-2202/8/60</p><p>BMC Neuroscience 2007;8():60-60.</p><p>Published online 31 Jul 2007</p><p>PMCID:PMC1950312.</p><p></p>ottom). Data were obtained from a slice four hours after its placement on the recording chamber. Note the persistency and stability over time of both characteristics of SPWs. . Record of spontaneous activity obtained from a different slice, presented at a faster sweep speed. . A wide band record of a single SPW-R event (C1) and the filtered sweeps are shown, illustrating the distinct components of the event. Low-pass (0.5–30 Hz) and band-pass (100–300 Hz) filtering reveals the slow wave (C2) and the associated ripple oscillation (C3) respectively. Band-pass filtering at very high frequencies (0.5–1 kHz, C4) discloses the burst-like multiunit activity occurring mostly during the rising and peaking phases of the slow positive potential. . FFT from 1 min raw record of spontaneous activity showing the difference in power between SPWs (upper diagram) and ripples (lower diagram, same power spectrum shown at a greater magnification). SPW dominant peak is at 5–10 Hz. Ripple activity is at ~150 Hz. . Histogram of period (inter-event interval) of SPWs calculated from a population of slices, showing that most of the slices displayed a period between 0.2–0.6 sec, which correspond to a frequency range of 1.5–5.0 Hz. . Auto-correlogram from a 5 min low-pass record from a distinct experiment, showing the typical degree of rhythmicity of SPWs