Article Figures & Data
Figures
- Figure 1.
Bath application of APV or MK-801 reduces the number of action potentials generated in response to 5 and 100 Hz synaptic stimulation. A, Recordings from a representative cell in response to stimulation at 5 Hz before (Pre-), during (APV), and after (Wash) washout of APV (50 μm). Of 30 traces recorded in each condition, only eight are shown superimposed for clarity. B, Recordings from a representative cell in response to 100 Hz stimulation in the presence of APV and after washout (Wash). Compiled data from the same experiments are shown to the right of traces from each frequency. In the 100 Hz case, a 5 Hz test stimulation was used to assess washin of the drug and the number of action potentials in the presence of drug compared with that in the absence of drug (washout). Means ± SEMs from 5-10 CA1 pyramidal neurons are shown. C, Left, Recordings from a representative cell in response to stimulation at 5 Hz before and during application of MK-801 (20 μm). Of 30 traces recorded, only eight are shown superimposed for clarity. Middle, Recordings from the same cell in response to 100 Hz stimulation in the presence of MK-801. Right, Compiled data from the same experiments. Five hertz stimulation was used to assess action potentials in the predrug condition, and the action potentials were induced with either 5 or 100 Hz in the presence of drug. Means ± SEMs from 10 CA1 pyramidal neurons are shown. Washout of MK-801 was not assessed. Error bars represent SEMs.
- Figure 2.
Bicuculline restores action potentials blocked by CPP. Bath application of bicuculline (10 μm) prevents the loss of action potentials with CPP (20 μm). A, Recordings from a representative cell in response to 5 and 100 Hz stimulation before (Pre-) and during (CPP+Bic.) application of bicuculline and CPP. B, Compiled data from the same experiments showing that action potentials are restored in both 5 and 100 Hz stimulation cases. Means ± SEMs from 14 CA1 pyramidal neurons are shown. Error bars represent SEMs.
- Figure 3.
Bicuculline restores immunostaining for p-ERK blocked by NMDAR antagonists. A, p-ERK staining in response to “control” 5 Hz stimulation is blocked with 50 μm APV (+APV) or 20 μm CPP (+CPP) but is restored when 10 μm bicuculline (+Bic.) is used in addition to the NMDAR antagonists. B, Experiments similar to those in A, except that 100 Hz synaptic stimulation was used instead of 5 Hz. Stratum pyramidale and stratum radiatum in area CA1 of the hippocampus are shown. The DAB reaction product appears dark, with pyramidal neurons and their apical dendrites showing stain. Note the apical dendrites of pyramidal cells coming off of the cell bodies. Scale bar, 100 μm. Regions immediately adjacent to the stimulating electrode were chosen for the +APV and +CPP cases to show the few cells that stained positive for p-ERK. p-ERK staining induced with TBS is not sensitive to APV alone (Dudek and Fields, 2001). C, Left, Immunoblot showing that bicuculline has no effect on basal ERK phosphorylation. Blots were probed with the same antibody as was used in A and B. Right, Immunoblot showing an experiment similar to A. In both nuclear and cytoplasmic fractions, ERK phosphorylation induced with 5 Hz stimulation, blocked with 50 μm APV, is restored with 10 μm bicuculline.
- Figure 4.
NMDAR antagonists have no effect on action potentials when TBS is used. A, Two 1 s recordings during TBS in the presence of APV (50 μm) and after washout (Wash) of the drug. B, Two superimposed 1 s recordings during TBS in the presence of CPP (20 μm) and after washout (Wash). C, The number of action potentials from experiments like those illustrated in A and B were counted, and the compiled results are shown. Means ± SEMs from 13 CA1 pyramidal neurons are shown from each frequency. Error bars represent SEMs.
- Figure 5.
The effects of NMDAR antagonists on action potentials are dose dependent. A, Dose-response relationship of APV comparing the effect when 5 Hz stimulation (closed diamonds) or TBS (open squares) was used (n = 9, 12, 10, 11, 11, and 10 at 5, 10, 20, 30, 40, and 50 μm, respectively, for 5 Hz; n = 13, 10, and 10 at 50, 100, and 200 μm, respectively, for TBS). The number of action potentials is presented as a percentage of the predrug control (5 Hz) or washout control (TBS) conditions. B, Dose-response relationship of CPP (n = 13, 13, 13, 13, and 9 at 10, 12, 15, 18, and 20 μm, respectively, for 5 Hz; n = 9, 14, 10, and 9 at 20, 50, 100, and 200 μm, respectively, for TBS) as in A.
- Figure 6.
Resistance of TBS-evoked action potentials to APV is caused by temporal summation within bursts. A, As intervals between pulses within bursts are increased, the number of action potentials evoked (in 50 μm APV) decreases (n = 11). Two traces are superimposed showing action potentials induced with TBS consisting of 10, 20, or 40 ms intervals between pulses. B, As intervals between bursts are increased, the number of action potentials evoked remains unchanged (in 50 μm APV) (n = 10). Two traces are superimposed showing action potentials induced with TBS consisting of 200, 333, or 500 ms intervals between bursts. Note the different time scales for each set of traces.
- Figure 7.
Summary of whole-cell ERK activation by synaptic activity. Different presynaptic stimulation (5 Hz, 100 Hz, or TBS) results in action potentials that are either dependent (5 and 100 Hz) or independent (TBS) on NMDARs. ERK is activated under either condition, provided action potentials are evoked. Stim., Stimulation.
Additional Files
Supplemental data
Files in this Data Supplement:
- supplemental material - Supplementary Figure 1. Bath application of CPP reduces the number of action potentials generated in response to 5 Hz and 100 Hz synaptic stimulation. (A) Recordings from a representative cell in response to stimulation at 5 Hz prior to, during, and after washout of CPP (20 |*mu*|M). Of thirty traces recorded in each condition, only eight are shown superimposed for clarity of the figure. (B) Recordings from a representative cell in response to 100 Hz stimulation in the presence of CPP and after washout. Compiled data from the same experiments are shown to the right of traces from each frequency. In the 100 Hz case, a 5 Hz test stimulation was used to assess wash-in of the drug, and the number of action potentials in the presence of drug compared with that in the absence of drug (washout). Shown are means +/- SEMs from 11 cells (5 Hz) and 11 cells (100 Hz).
- supplemental material - Supplementary Figure 2. CPP, at a concentration that blocks action potential generation, has no effect on AMPA currents. Bath application of 20 |*mu*|M CPP has no effect on AMPA- mediated EPSCs.(A) Data are means +/- SEMs from 14 cells. Representative recordings of EPSCs with and without 20 |*mu*|M CPP are shown superimposed . (B) Shown are the averages of 10 traces.
- supplemental material - Supplementary Figure 3. Intracellular MK801 prevents action potential generation induced with 5 Hz stimulation. Shown are traces from recordings immediately after break-in (A) and 5 minutes after break-in (B) when 20 |*mu*|M MK801 was included in the internal solution of the patch pipette.
- supplemental material - Supplementary Figure 4. Increasing stimulus intensity fails to fully compensate for the effects of NMDAR antagonists. Bath application of 50 |*mu*|M APV reduces the number of action potentials generated with 5 Hz stimulation. The number of action potentials induced with 30 stimuli are plotted against stimulation intensity. Shown are means +/- SEMs from 6-20 cells.
