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Differences in GluN2B-Containing NMDA Receptors Result in Distinct Long-Term Plasticity at Ipsilateral Versus Contralateral Cortico-Striatal Synapses

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Differences in GluN2B-Containing NMDA Receptors Result in Distinct Long-Term Plasticity at Ipsilateral Versus Contralateral Cortico-Striatal Synapses

Wei Li et al. eNeuro.

Abstract

Excitatory neurons in the primary motor cortex project bilaterally to the striatum. However, whether synaptic structure and function in ipsilateral and contralateral cortico-striatal pathways is identical or different remains largely unknown. Here, we describe that excitatory synapses in the mouse contralateral pathway have higher levels of NMDA-type of glutamate receptors (NMDARs) than those in the ipsilateral pathway, although both synapses utilize the same presynaptic vesicular glutamate transporter (VGLUT). We also show that NMDARs containing the GluN2B subunit, but not GluN2A, contribute to this difference. The altered NMDAR subunit composition in these two pathways results in opposite synaptic plasticity induced by θ-burst stimulus: long-term depression in the ipsilateral pathway and long-term potentiation (LTP) in the contralateral pathway. The standard long-term depression (LTD)-inducing protocol using paired postsynaptic and presynaptic activity triggers synaptic depression at ipsilateral pathway synapses, but not at those of the contralateral pathway. Altogether, our results provide novel and unexpected evidence for the lack of bilaterality of NMDAR-mediated synaptic transmission at cortico-striatal pathways due to differences in the expression of GluN2B subunits, which results in differences in bidirectional synaptic plasticity.

Keywords: GluN2A; GluN2B; NMDARs; cortico-striatal synapses; synaptic plasticity.

Figures

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Figure 1.
Figure 1.
Morphologic and functional features of ipsilateral and contralateral cortico-striatal pathways. A, EYFP immunostaining shows the bilateral projection of M1 cortical neurons in the striatum. Asterisk shows the injection site of AAV-CaMKIIα-ChR2-EYFP in M1; white boxes in the ipsilateral and contralateral dorsolateral striatum are enlarged below. Scale bars: 500 µm (top) and 50 µm (bottom). B, Double VGLUT1 and EYFP immunostaining (left) shows co-localized puncta (arrowheads) in the striatum of both hemispheres, ipsilateral and contralateral to the AAV-CaMKIIα-ChR2-EYFP injection site in M1. Double VGLUT2 and EYFP immunostaining (right) shows a lack of co-localization. Scale bar: 5 µm. C, Representative EPSCs (from D) evoked by blue light (470 nm) in ipsilateral (top) and contralateral (bottom) MSNs in striatal slices from mice expressing CaMKIIα-ChR2-EYFP in M1. Traces are from baseline, 10 min after NBQX, and 10 min after D,L-APV. D, Average of the NMDAR component of the blue light-evoked EPSC in ipsilateral and contralateral MSNs (n = 9, Ipsi MSNs; n = 7, Contra MSNs; p < 0.0001, unpaired Student’s t test). E, Representative EPSCs (from F–H) evoked by blue light with different durations (1, 2, 3, and 4 ms) in ipsilateral (top) and contralateral (bottom) MSNs. MSNs were held at +60 mV and perfused with aCSF containing normal Mg2+. Traces are from baseline, 10 min after NBQX, and 10 min after D,L-APV. F–H, I/O relationship between light pulse duration, and total EPSC amplitude (F; n = 6, Ipsi MSNs; n = 6, Contra MSNs; p = 0.14, two-way ANOVA with Bonferroni post hoc tests) or NMDAR EPSC amplitude (G; p = 0.036) or the ratio of NMDAR EPSC amplitude to total EPSC amplitude (H; p < 0.0001).
Figure 2.
Figure 2.
Differences in subunit composition of synaptic NMDARs between the ipsilateral and contralateral cortico-striatal pathways. A, Double tdTomato and EGFP immunostaining shows segregation of D1 and D2 MSNs in the striatum of Drd1a-tdTomato::Drd2-EGFP double transgenic mice. B, Representative NMDAR EPSCs (from C) in D1 (top) and D2 MSNs (bottom) evoked by electrical stimulation of the cortico-striatal pathway. Traces are from baseline (1), 10 min after ifenprodil (2), 10 min after TCN-201 (3), and 10 min after D,L-APV. C, Average effects of ifenprodil, TCN-201, and D,L-APV on NMDAR EPSC amplitudes (n = 7, D1 MSNs; n = 6, D2 MSNs; p = 0.56, two-way ANOVA). D, Representative EPSCs (from E) evoked by blue light in ipsilateral (top) and contralateral (bottom) MSNs. Traces are from baseline (1), 10 min after ifenprodil (2), and 10 min after TCN-201 (3). E, Average effects of ifenprodil and TCN-201 on blue light-evoked NMDAR EPSCs (n = 19, Ipsi MSNs; n = 16, Contra MSNs; p = 0.0013, two-way ANOVA). F, Average percentage of the ifenprodil-sensitive and TCN-201-sensitive components of NMDAR-mediated EPSCs. The ifenprodil-sensitive component of the NMDAR EPSCs was calculated by subtracting EPSCs after 10 min in ifenprodil from baseline EPSCs; the TCN-201-sensitive component of the NMDAR EPSCs was calculated by subtracting EPSCs after 10 min in TCN-201 from EPSCs in the presence of ifenprodil (***p < 0.0001, unpaired Student’s t test). G, tdTomato and GFP immunostaining shows ipsilateral Chrimson expression and contralateral Chronos expression. Scale bars: 100 µm (left) and 10 µm (right). H, Representative EPSCs (from I) evoked by either Chrimson-activating red light (625 nm, top) or Chronos-activating blue light (470 nm, bottom). Traces are from baseline (1), 10 min after ifenprodil (2), and 10 min after TCN-201 (3). l, Average effects of ifenprodil and TCN-201 on red light-evoked and blue light-evoked NMDAR EPSCs (n = 19, Ipsi MSNs; n = 16, Contra MSNs; p = 0.028, two-way ANOVA).
Figure 3.
Figure 3.
The role of GluN2A-containing and GluN2B-containing NMDARs in TBS-induced synaptic plasticity at the ipsilateral and contralateral cortico-striatal pathways. A, EYFP immunostaining shows ChETA expression in M1. Arrowheads indicate soma of pyramidal neurons expressing ChETA. Scale bars: 10 µm. B, Schematic diagram of TBS. C, TBS pattern of blue light (470 nm) pulses evokes spiking in a ChETA-expressing M1 pyramidal neuron (top) and temporally summating subthreshold EPSPs in a MSN (bottom). D, Representative EPSPs (from E) evoked by a single blue light pulse before (1) and 25 min after TBS of blue light pulses (2) in control or slices treated with either ifenprodil or TCN-201. E, TBS of blue light (bar) induced LTD of EPSPs in ipsilateral MSNs, which was blocked by either ifenprodil or TCN-201 (n = 7, CTL; n = 8, ifenprodil; n = 9, TCN-201; p < 0.0001 CTL vs ifenprodil and TCN-201, two-way ANOVA). LTD in ipsilateral MSNs was sensitive to AM-251 treatment (n = 5; p < 0.0001 CTL vs AM-251, two-way ANOVA). F, Analysis of EPSP amplitude at 25–30 min after TBS in ipsilateral MSNs (p < 0.0001 CTL vs ifenprodil and TCN-201, one-way ANOVA). G, Representative EPSPs (from H) evoked by a single blue light pulse before (1) and 25 min after TBS of blue light pulses (2) in control or slices treated with either ifenprodil or TCN-201. H, TBS of blue light (bar) induced LTP of EPSPs in contralateral MSNs, which was blocked by either ifenprodil or TCN-201 (n = 7, CTL; n = 7, ifenprodil; n = 10, TCN-201; p = 0.0023 ifenprodil vs CTL; p = 0.0049 TCN-201 vs CTL; two-way ANOVA). I, Analysis of EPSP amplitude at 25–30 min after TBS in contralateral MSNs (p < 0.01 CTL vs ifenprodil; p < 0.05 CTL vs TCN-201, one-way ANOVA). *p < 0.05, **p < 0.01, ***p < 0.0001.
Figure 4.
Figure 4.
The role of GluN2A-containing and GluN2B-containing NMDARs in postsynaptic-presynaptic STDP pairing-induced synaptic plasticity at the ipsilateral and contralateral cortico-striatal pathways. A, Schematic diagram of STDP protocol, where postsynaptic depolarization is followed by presynaptic light stimulus. B, Representative trace shows spikes induced by postsynaptic current injections and subsequent EPSPs (arrowheads) induced by delayed presynaptic stimulus. C, Representative EPSPs (from D) evoked by a single blue light pulse before (1) and 25 min after paring (2) in control or slices treated with either ifenprodil or TCN-201. D, STDP pairing (bar) induced LTD of EPSPs in ipsilateral MSNs, which was blocked by either ifenprodil or TCN-201 (n = 7, CTL; n = 6, ifenprodil; n = 8, TCN-201; p = 0.034 CTL vs ifenprodil; p = 0.029 CTL vs TCN-201; two-way ANOVA). E, Analysis of EPSP amplitude at 25–30 min after pairing in ipsilateral MSNs (p < 0.05 CTL vs ifenprodil; p < 0.05 CTL vs TCN-201; one-way ANOVA). F, Representative EPSPs (from G) evoked by a single blue light pulse before (1) and 25 min after pairing (2) in control or slices treated with either ifenprodil or TCN-201. G, Pairing (bar) did not induce synaptic plasticity in contralateral MSNs, and ifenprodil or TCN-201 had no effect on EPSPs (n = 7, CTL; n = 6, ifenprodil; n = 5, TCN-201; p = 0.29 CTL vs ifenprodil; p = 0.66 CTL vs TCN-201; two-way ANOVA). H, Analysis of EPSP amplitude at 25–30 min after pairing in contralateral MSNs (p > 0.05 CTL vs ifenprodil; p > 0.05 CTL vs TCN-201; one-way ANOVA). *p < 0.05.

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