The Journal of Neuroscience, June 28, 2006, ():
Temperature-Dependent Shift of Balance among the Components of Short-Term Plasticity in Hippocampal Synapses
J. Neurosci. Klyachko and Stevens
26: 6945
Supplemental data
Files in this Data Supplement:
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Supplementary Fig. 1. Minimal stimulation confirms the coexistence of different forms of STP in putative single synapses. A. Synaptic responses evoked by 100-stimulus train at 60 Hz were recorded at 33°C with a minimal stimulation defined as in (Dobrunz and Stevens, 1997). Average EPSC amplitudes during 45 presentations of the stimulus train in one cell are plotted as a function of the stimulus number. Dotted line represents the average control value. B. The normalized changes in the average EPSC amplitude after the stimulus train are plotted vs. time and reflect the decay of augmentation. Parameters of augmentation determined from this plot are very similar to those observed in fEPSP recordings (Fig. 5). Similar to the example shown in this figure, we observed a mixture of depression and one or both components of potentiation in all 5 cells recorded.
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Supplementary Fig. 2. Traces of fEPSP and EPSC responses on an expanded timescale. A. Traces of fEPSP responses to 40Hz 150-stimulus trains from Fig.1 are shown on an expanded time scale for 23°C (black trace) and 33°C (red trace). B. Same as in (A), but for EPSC traces at 23°C, 33C and 38°C.
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Supplementary Fig. 3. Whole-cell recordings confirm that the changes in apparent depression with temperature reflect interactions of various STP components, rather than the fast component of depression alone. A. Apparent depression during high-frequency trains of 2-40 Hz was calculated as described in the legend of Fig. 3 for 23°C (in black) and 33°C (in red) (n = 7-11 cells). B. Amplitude and timecourse of the fast component of depression was determined with whole-cell recordings from n = 5 cells as described in Fig. 3. Whole-cell recordings revealed no significant change in the amplitude and a ∼2.5-fold faster timecourse of depression at higher temperatures, similar to the results reported by fEPSP recordings.
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Supplementary Fig. 4. Receptor desensitization and receptor saturation do not play a significant role in the observed temperature-dependent effects. A. Receptor desensitization and receptor saturation were reduced with cyclothiazide (CTZ, 100 µM) and a low-affinity AMPA antagonist γ-DGG (1 mM) in n = 7 cells (2 mM γ-DGG were also tested in 2 cells with no significant additional effects on train responses evident). EPSC responses to 150-stimulus, 40 Hz trains were measured both at 23 and 33°C in each cell (4-5 train presentations at each temperature per cell). The average normalized EPSC amplitudes for all experiments are plotted vs. stimulus number in the train (A). 5 Controls at 0.2 Hz and 10 test pulses at 0.2 Hz were recorded before and after the train respectively. Inset: The effect of CTZ and γ-DGG application on the shape of EPSCs. Two EPSCs from the same cell (normalized to their peaks) are shown before and after the application of the blockers. Downward part of the stimulus artifact was removed for clarity. Scale bars are 10 ms and 0.2 (normalized EPSC) B. A large shift in apparent depression with temperature (values for 40 Hz, 150-stimulus trains are shown) is similar in control experiments (shown in Fig. 1) and in the presence of CTZ and γ-DGG. *** - P < 10-5.
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Supplementary Fig. 5. No hysteresis effects are observed in STP temperature-dependence. A. B. Synaptic responses to a natural stimulus pattern evoked in the same slice first at 23°C and then at 33°C are compared with the responses evoked in the opposite order of temperatures in a different slice. Independently of the direction in which the temperature was varied, responses were highly reproducible at 33°C, much less reproducible at 23°C (A), and responses at 23°C and 33°C were weakly (and negatively) correlated with each other (B). C. No significant changes in the parameters of facilitation (left panels) augmentation (middle panels) and depression (right panels) were recorded at each temperature in the transition range of 26-29°C when temperature was varied in opposite directions. This indicates the absence of hysteresis effects in STP temperature-dependence (n = 6).
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Supplementary Fig. 6. EGTA-AM does not completely block augmentation during high-frequency trains.
The effects of slice incubation with 250μM EGTA-AM for various durations (30 min - 4 hours) were evaluated during 150-stimulus trains at 40Hz. Control (black trace), 30 min of EGTA-AM (blue trace) and 2-4 hours of EGTA-AM (dark blue trace) are plotted vs. stimulus number in the train. Although paired-pulse facilitation was completely blocked after 30-min incubation (inset), even 4-hour incubation was not sufficient to block augmentation (n = 6). The effects of EGTA-AM, which included the slower increase in synaptic response during trains and prolonged decay of augmentation, saturated after ∼ 2 hours of incubation and responses corresponding to the longer incubation times were averaged together.
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Supplementary Fig. 7. Lowering extracellular calcium reveals augmentation by minimizing synaptic depression. A. Responses to 150-stimulus trains at 10Hz were recorded in 0.75 mM Ca2+o/2.5mM Mg2+o at 23°C (black trace) and 33°C (red trace). Depression was greatly reduced at both temperatures and test pulses after the end of the train revealed pure augmentation (B). The temperature-dependence of augmentation remained qualitatively similar to that previously found at 2mM Ca2+o as it underwent a large increase in amplitude and 5-fold acceleration of decay (C). * - P<0.05, ** - P<0.01.
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Supplementary Fig. 8. Ba2+ application did not increase augmentation but caused prolonged depression in hippocampal slices. A. Responses to 2Hz (left panel) and 20Hz (middle panel) 150-stimulus train before (red traces) and after (blue traces) application of 0.6mM Ba2+ are plotted vs. stimulus number. Ba2+ application resulted in a strong and long-lasting depression during trains of all frequencies tested, 2-40Hz (right panel). B. Ba2+ application also produced a strong depression during natural stimulation. These effects were observed in a wide range of Ba2+ concentrations, 0.6-2 mM in n = 6 slices.
