NMDA receptor activation induces long-term potentiation of glycine synapses

Abstract

Of the fast ionotropic synapses, glycinergic synapses are the least well understood, but are vital for the maintenance of inhibitory signaling in the brain and spinal cord. Glycinergic signaling comprises half of the inhibitory signaling in the spinal cord, and glycinergic synapses are likely to regulate local nociceptive processing as well as the transmission to the brain of peripheral nociceptive information. Here we have investigated the rapid and prolonged potentiation of glycinergic synapses in the superficial dorsal horn of young male and female mice after brief activation of NMDA receptors (NMDARs). Glycinergic inhibitory postsynaptic currents (IPSCs) evoked with lamina II-III stimulation in identified GABAergic neurons in lamina II were potentiated by bath-applied Zn2+ and were depressed by the prostaglandin PGE2, consistent with the presence of both GlyRα1- and GlyRα3-containing receptors. NMDA application rapidly potentiated synaptic glycinergic currents. Whole-cell currents evoked by exogenous glycine were also readily potentiated by NMDA, indicating that the potentiation results from altered numbers or conductance of postsynaptic glycine receptors. Repetitive depolarization alone of the postsynaptic GABAergic neuron also potentiated glycinergic synapses, and intracellular EGTA prevented both NMDA-induced and depolarization-induced potentiation of glycinergic IPSCs. Optogenetic activation of trpv1 lineage afferents also triggered NMDAR-dependent potentiation of glycinergic synapses. Our results suggest that during peripheral injury or inflammation, nociceptor firing during injury is likely to potentiate glycinergic synapses on GABAergic neurons. This disinhibition mechanism may be engaged rapidly, altering dorsal horn circuitry to promote the transmission of nociceptive information to the brain.

Glutamate, d-(−)-2-Amino-5-Phosphonopentanoic Acid, and N-Methyl-d-Aspartate Do Not Directly Modulate Glycine Receptors

“…To test the theory that an enhancement of GlyR mIPSCs could be observed under our recording conditions when the recorded neuron had a low intracellular buffering capacity, we recorded mIPSCs using an intracellular solution that contained 0.6 mM EGTA + 0 mM Ca 2+ ( Figure 4B). NMDAR dependent increases in glycine eIPSCs has been observed in mouse spinal cord slices when recorded with 0.6mM EGTA-containing intracellular solution (Kloc et al, 2019). While we did not detect a consistent increase in mIPSC amplitude (normalized amp (NMDA/control) = 1.13, 95% CI: 0.99 to 1.27, P = 0.10), we did detect a small enhancement in charge transfer (normalized charge (NMDA/control) = 1.23, 95% CI: 1.09 to 1.38, P = 0.01) accompanied by an increase in mIPSC 20-80% decay (control = 7.74 ms, 95% CI: 5.91 to 9.56 vs NMDA = 8.54 ms, CI: 7.10 to 9.98, P = 0.04).…”

“…solution contained: (in mM): 140 CsCl, 1 MgCl 2 , 0.6 EGTA, 4 Mg-ATP, 5 QX314 [N-(2,6dimethylphenylcarbamoylmethyl)triethylammonium-chloride], and 10 HEPES, (pH 7.4 with CsOH, 300 ± 5 mOsm). In Figure 4B, neurons were exposed to NMDA for > 5 minutes before mIPSC were analyzed (Kloc et al, 2019).…”

“…This form of potentiation is indirect (in contrast to the effect reported by (Liu et al, 2010)) as it is dependent on calcium/calmodulin-dependent protein kinase II (CAMKII) (Xu et al, 2000) ( Yamanaka et al, 2013) and is prevented by including calcium chelators such as EGTA and BAPTA in the intracellular solutions (Kloc et al, 2019;Xu et al, 2000)}. The Ca 2+ -dependent GlyR current enhancement occurs over time-frames compatible with the results reported by Lui et al (2000) and can be stimulated by NMDA and NMDAR activation (Kloc et al, 2019) (Fucile et al, 2000;Lévi et al, 2008;Xu et al, 2000). Our intracellular solution contained 1 mM BAPTA + 10 mM EGTA + 1 mM Ca 2+ , which strongly limits intracellular calcium transients.…”

Glycine receptors are not directly modulated by glutamate, AP5 or NMDA

“…Intracellular calcium increases are a common feature of neuronal signalling and many reports have shown that various neuronal activators cause intracellular calcium dependent increases in GlyR (Fucile et al, 2000;Kloc et al, 2019;Xu et al, 2000) (Lévi et al, 2008) and GABAR (Stelzer and Wong, 1989) signalling . In essence this means that any process that leads to elevations in intracellular Ca 2+ will enhance GlyRs.…”

“…A recently published paper investigating the NMDA receptorintracellular Ca 2+ link to GlyR signalling (Kloc et al, 2019) recorded glycine eIPSCs in the presence and absence of NMDA or D-AP5. Although these experiment were not specifically designed to test the findings reported in Liu et al 2010, this data demonstrates that glycine eIPSCs are not enhanced by NMDA (50 µM) when NMDAR are blocked by the non-competitive antagonist 7-chlorokynurenic acid or when 15 mM EGTA is included in the intracellular pipette solution (Kloc et al, 2019). This data is consistent with our finding that NMDA does not have a direct effect on glycine receptors.…”