CA2-DEPENDENT INACTIVATION OF NMDA RECEPTORS - FAST KINETICS AND HIGHCA2+ SENSITIVITY IN RAT DORSAL HORN NEURONS()

1. Ca2+-dependent inactivation (CDI) of NMDA receptors was studied usi ng bath cultured embryonic rat dorsal horn neurons and acutely dissoci ated postnatal rat dorsal horn neurons. The perforated patch recording method was employed in order to preserve intracellular Ca2+ buffers a nd other cellular constituents. In this way, the kinetics of intracell ular Ca2+ concentration ([Ca2+](i)) transients and other second messen ger signalling systems were maintained in a relatively normal conditio n. 2. Continuous application of 30 mu M NMDA to cultured dorsal horn n eurons voltage clamped at -70 mV evoked currents that inactivated to a bout 40% of the peak value with time constants between 200 and 600 ms. CDI with similar kinetics was also observed in acutely dissociated po stnatal rat dorsal horn neurons 3. When NMDA was applied in a low (20 mu M) Ca2+ bath or when dorsal horn neurons mere held at +70 mV, inact ivation was either very weak or absent. The peaks of NMDA currents wer e significantly suppressed when preceded by voltage steps to 0 mV or b y evoked action potentials. The suppression was dependent on the prese nce of Ca2+ in the extracellular solution. Voltage steps to +100 mV we re ineffective in suppressing NMDA responses. Therefore, the observed inactivation was caused by an increase in [Ca2+](i) following Ca2+ ent ry through NMDA channels or through voltage-gated Ca2+ channels. 4. Ca ffeine application reduced currents evoked by subsequent NMDA applicat ions. This reduction was not dependent on the presence of extracellula r Ca2+ release from intracellular stores also induced CDI. 5. Simultan eous measurements of somal [Ca2+](i) and of currents evoked by somal N MDA applications showed that the magnitude of CDI was correlated with [Ca2+](i) levels and that [Ca2+](i) elevations of 100-300 nM were usua lly sufficient to inactivate NMDA currents by more than 30%. 6. Dose-r esponse curves of non-inactivated and inactivated NMDA responses showe d that the apparent-receptor affinity for NMDA is not different under the two conditions. CDI is caused instead by non-competitive inhibitio n of NMDA receptors. CDI was not overcome by increasing glycine concen tration, suggesting that it is not mediated by glycine dissociation fr om the receptor. 7. These results show that, with an intact intracellu lar environment, CDI in dorsal horn neurons constitutes a potent, inhi bitory control of NMDA currents with a faster onset than previously de monstrated. CDI is induced by a variety of [Ca2+](i)-elevating stimuli of physiological relevance including Ca2+ entry through ligand- and v oltage-gated channels and Ca2+ release from intracellular stores. Our demonstration that CDI is strongly expressed in neurons maturing in vi vo supports the hypothesis that CDI may regulate, in part, the Postsyn aptic integration of excitatory input in the mature or maturing nervou s system.