NMDA-INDUCED BURST DISCHARGE IN GUINEA-PIG TRIGEMINAL MOTONEURONS IN-VITRO

1. The responses of guinea pig trigeminal motoneurons (TMNs) to N-meth yl-D,L-aspartate (NMA) were studied using brain stem slice preparation s and whole cell patch-clamp (n = 89) or conven tional microelectrode (n = 22) recording techniques. The primary goals of this study were to determine whether N-methyl-D-aspartate (NMDA) receptor activation wou ld produce spontaneous bursting activity in TMNs and, if so, the under lying mechanisms responsible for the generation of these bursts. 2. Ba th-applied NMA (100-300 mu M, n = 80) in standard per fusion medium el icited depolarization, increase in apparent input resistance (R(inp)), and rhythmic burst discharges (1-90 a in duration) from TMNs. These e ffects were blocked by the NMDA receptor antagonist DL-2-amino-5-phosp honopentanoic acid (AP5, 30 mu M, n = 6), but not by the non-NMDA rece ptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 5-10 mu M, n = 10). Furthermore, the burst-inducing effect of NMA was not mimick ed by the non-NMDA receptor agonists kainate (KA, 5-10 mu M, n = 6) an d lpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA, 5-10 mu M, n = 5). 3. In tetrodotoxin (TTX) treatment conditions (n = 13), NMA elicited depolarization, an increase in apparent R(inp), and rhyth mic membrane potential oscillations without action potential bursts (i .e., plateau potentials), suggesting that the effects of NMA observed in the TTX-free condition resulted from activation of postsynaptic NMD A receptors. 4. Graded depolarization of neurons (n = 20) by intracell ular direct current injection generally led to a graded increase in fr equency and duration of the NMA-induced bursts and plateau potentials until these rhythmic events eventually became transformed into continu ous spike discharge and maintained depolarization, respectively. Remov al of Mg2+ from the perfusion medium (n = 11) also turned the bursts a nd plateau potentials into continuous spike discharge and maintained d epolarization, respectively. 5. The effects of NMA on the current-volt age (I-V) curve after a depolarizing ramp voltage-clamp command (15-20 mV/s) were examined (n = 40). Under NMA (100-300 mu M) conditions, th e I-V relationship exhibited a region of negative slope conductance (N SC) between -60 and -35 mV, thus making the I-V relationship N-shaped. The NSC was abolished by AP5 (30 mu M, n = 8), but not by CNQX (5-10 mu M, n = 6). The I-V relationship in AMPA (3-10 mu M n = 5) or KA (3- 10 mu M, n = 5) was almost linear between -80 and -30 mV. In perfusion medium lacking Mg2+, the NMA-induced NSC was abolished, leaving the I -V relationship linear in the region between -80 and -30 mV (n = 6). 6 . The role of Ca2+ in the generation and termination of individual NMA -induced bursts was examined (n = 13). Application of zero- or low-Ca2 + medium either 1) blocked completely the generation of NMA-induced pl ateau potentials (n = 6), 2) increased the duration of plateau potenti als (n = 5), or 3) prevented the termination (i.e., repolarization) of plateau potentials (n = 2). The I-V curves examined in low-Ca2+ mediu m were almost linear in the vast majority of the cases (7 of 8) becaus e of the reduction of NMA-induced inward current in the region of the NSC. 7. The role of Ca2+-dependent K+ currents in terminating individu al NMA-induced bursts was assessed using the bee venom apamin and the scorpion venom iberiotoxin, which block the SK and BK types of K+ chan nels, respectively. Although apamin (200 nM, n = 5) completely blocked the afterhyperpolarization after each action potential, it (100 nM, n = 2; 200 nM, n = 3) failed to prevent the termination of NMA-induced bursts; only a slight to moderate increase of the duration and the und erlying depolarization of the bursts was apparent during application o f apamin. Iberiotoxin (30 nM) had no obvious effects on either the bur sts or the action potential waveform (n = 5). 8. The data from this st udy demonstrate that selective activation of postsynaptic NMDA recepto rs induces rhythmic burst discharges in TMNs. Additionally, the data s uggest that the NMA-induced bursts arise from activation of an Mg2+- a nd voltage-dependent NMDA conductance that is manifest as a region of NSC in the NMA I-V relationship. Furthermore, the bursts are dependent on the presence of Ca2+ in the ACSF in a subpopulation of TMNs. 9. Th erefore it is concluded that any models of rhythmic oral-motor activit y must include the unique properties of the NMDA channel.