D-Serine differently modulates NMDA receptor function in rat CA1 hippocampal pyramidal cells and interneurons

Abstract

The organization of the neuronal hippocampal network depends on the tightly regulated interaction between pyramidal cells (PCs) and interneurons (Ints). NMDA receptor (NMDAR) activation requires the binding of glutamate and co-activation of the 'glycine site'. It has been reported that D-serine is a more potent endogenous agonist than glycine for that site. While many studies have focused on NMDAR function in PCs, little is known regarding the modulation of NMDARs in Ints. We studied the modulatory effect of D-serine on NMDAR EPSCs in PCs and in stratum radiatum Ints using whole-cell patch-clamp recording in rat acute hippocampal slices. We found that D-serine enhances NMDAR function and differently modulates NMDAR currents in both cell types. The augmentation of NMDAR currents by D-serine was significantly larger in PCs compared with Ints. Moreover, we found differences in the kinetics of NMDAR currents in PCs and Ints. Our findings indicate that regulation of NMDAR through the 'glycine site' depends on the cell types. We speculate that the observed differences arise from assemblies of diverse NMDAR subunits. Overall, our data suggest that D-serine may be involved in regulation of the excitation-inhibition balance in the CA1 hippocampal region.

Figure 1. Contrasting morphological and physiological properties of pyramidal cells (PCs) and interneurons (Ints) in CA1 region of the hippocampus

A1 and B1, confocal images of one PC and one Int of CA1 pyramidal layer and stratum radiatum, respectively. The cells were recorded at the positions indicated in A2 and B2 (asterisks; confocal and DIC image superimposed). A3 and B3, confocal images of the boxed dendritic regions shown in A1 and B1, respectively. Note the spiny dendritic segment of the PC (A3) in contrast with the aspiny (B3) one of the Int. The arrowheads indicate dendritic spines. A4 and B4 show the voltage responses of the same PC and Int in A1 and B1, to a series of intracellular current pulses. The current was applied at rest (−78 and −65 mV, respectively). Abbreviations: alv, alveus; so, stratum oriens; sp, stratum pyramidale; sr, stratum radiatum; slm, stratum lacunosum moleculare. Calibration bars in A2 and A4 are applied to B2 and B4, respectively.

Figure 2. Effect of d-serine on the NMDAR currents in PCs and Ints

Neurons were recorded in a low-Mg²⁺ ACSF (0.1 mm) in the presence of NBQX (20 μm), picrotoxin (50 μm), CGP 52432 (10 μm) and strychnine (0.5 μm). Each trace is an average of 20 traces. A and B, responses evoked by bipolar electrical stimuli at V_m=−70 mV in a pyramidal cell (PC) and in an interneuron (Int). A1 and B1, the NMDAR currents recorded in low-Mg²⁺ ACSF (thin line), during application of d-serine (10 μm; thick line), and after recovery from the drug (dotted line), are superimposed. A2 and B2, decay time course of NMDAR currents in low-Mg²⁺ ACSF (thin line) and during application of d-serine (10 μm; thick line). The traces of the NMDAR currents in low-Mg²⁺ ACSF (thin line) are normalized to the amplitude of the traces after the application of d-serine (thick line). The decay of the NMDAR currents in Ints was best fitted with two exponentials (τ_f and τ_s, fast and slow decay components). Note that the decay of the NMDAR currents in the PC during the application of d-serine (10 μm) is slower than in low-Mg²⁺ ACSF. The same was observed for the τ_f in the Int. A3 and B3, histograms of the effect of d-serine (10 μm) on NMDAR currents (51.66 ± 7.89 % in PCs, n = 17 and 25.07 ± 5.65 % in Ints, n = 12). The asterisk indicates that the difference in percentage due to the application of d-serine on PCs compared with Ints is significant (P < 0.005). The values of the Ints’τ_f and τ_s are in the range of 30 to 52.4 ms and 43 to 387 ms for control condition, and 39 to 86.8 ms and 55 to 400 ms following the application of d-serine.

Figure 3. Effect of d-serine on the voltage-dependent properties of the NMDAR currents in PCs and Ints

Neurons were recorded in normal ACSF in the presence of picrotoxin (50 μm), CGP 52432 (10 μm), strychnine (0.5 μm) and NBQX (20 μm). Each trace is an average of five traces. A1 and B1, the NMDAR currents of one PC and one Int shown before (thin line) and during the application of d-serine (10 μm; thick line), at the indicated membrane potentials, are superimposed. A2 and B2, peak current-voltage relations are shown for PCs and Ints before (▪) and during (•) the application of d-serine. The peak current-voltage relations are averages of three PCs and three Ints.

Figure 4. Effect of d-serine on the NMDAR currents in PCs and Ints as a function of time

Neurons were recorded in a low-Mg²⁺ ACSF (0.1 mm) in the presence of NBQX (20 μm), picrotoxin (50 μm), CGP 52432 (10 μm) and strychnine (0.5 μm). A1 and B1, graphs plotting the normalized NMDAR-mediated current (NMDAR-mc) amplitude (average of the control) as a function of time for PCs (n = 8) and Ints (n = 6). A2 and B2, one PC and one Int showing the effect of d-serine over a period of 40 min. d-Serine was applied at time zero (arrowhead).

Figure 5. Effect of d-serine on the responses of one PC and one Int to local pressure application of NMDA

The NMDA (100 μm) was applied through a patch pipette positioned directly above the proximal dendrites. TTX (0.5 μm) was present throughout the experiments. The cells were voltage-clamped at V_m=−70 mV. A and B, NMDA-evoked responses observed in low-Mg²⁺ ACSF (thick line), during application of d-serine (10 μm; thin line), and after recovery from the drug (dotted line) are superimposed for one PC and one Int, respectively. Each trace is an average of four traces. Time base is 1 s for A and 500 ms for B. The inset represents the same traces in B with a slower time base.

Figure 6. Effect of d-serine on paired pulse stimulation

A, responses were evoked by bipolar electrical stimuli at V_m = −70 mV in PCs. Two pulses of identical intensity were delivered with an interval of 60 ms. Glutamatergic-evoked responses observed in normal ACSF (thick line), during application of d-serine (10 μm; thin line), and after recovery from the drug (dotted line) are superimposed. Each trace is an average of 20 traces. B, histogram of the ratio between the amplitude of the second and the first evoked response (2.03 ± 0.25 and 2.25 ± 0.32, n = 4, in control and in d-serine, respectively) is shown. This difference was not statistically significant (P > 0.5).

Figure 7. d-Serine (10 μm) has no effect when the ‘glycine site’ of the NMDARs on PCs (A) and Ints (B) is blocked by 7-chlorokynurenic acid (50 μm)

Neurons were recorded in a low-Mg²⁺ ACSF (0.1 mm) in the presence of NBQX (20 μm), picrotoxin (50 μm), CGP 52432 (10 μm) and strychnine (0.5 μm). A and B, graphs plotting the normalized NMDAR-mediated current (NMDAR-mc) amplitude as a function of time for PCs (n = 5) and Ints (n = 3). 7-Chlorokynurenic acid (50 μm) was applied at time zero and maintained throughout the experiment. d-Serine (10 μm) was applied 3 min after the application of 7-chlorokynurenic acid, for a period of 6 min. Note that in both PCs and Ints, in the presence of 7-chlorokynurenic acid, d-serine did not enhance the NMDAR currents. The insets show traces of NMDAR currents at V_m = −70 mV in low-Mg²⁺ ACSF and in the presence of 7-chlorokynurenic acid plus d-serine. Every trace is the average of 20 traces. The current bar applies to each trace.