N-methyl-D-aspartate-activated channels of mouse central neurones in magnesium-free solutions

Abstract

1. The whole-cell and outside-out configurations of the patch-clamp method were used to investigate the properties of the channels activated by N-methyl-D-aspartate (NMDA channels) in mouse central neurones in culture. Recording was made in Mg2+-free solutions. 2. In the whole-cell recording mode the currents induced by both NMDA and L-glutamate were accompanied by a large increase in noise. In both cases the noise power spectra were well fitted by single Lorentzian functions and the corresponding mean time constant, tau, was about 6 ms at room temperature. The single-channel conductance, gamma n, estimated from the ratio of the noise variance to the total current, varied between 22 and 40 pS. 3. Endogenous amino acids known to activate NMDA receptors (L-glutamate, L-aspartate, L-cysteine sulphinate and quinolinate) as well as exogenous NMDA agonists such as ibotenate and trans-2,3-piperidine dicarboxylate (trans-PDA) all produced similar responses. In particular, analysis of the current noise yielded tau values between 4 and 8 ms in all cases. 4. NMDA responses were antagonized by 2-amino-5-phosphonovalerate (APV) without any effect on gamma n or tau values measured by noise analysis; NMDA responses were also diminished by D-alpha-aminoadipate and cis-2,3-piperidine dicarboxylate. 5. In outside-out patches, it was observed that the single-channel current amplitude varies linearly as a function of membrane potential between -80 and +60 mV. The reversal potential is near 0 mV. NMDA channels are permeable to Na+, K+ and Cs+, but blocked by choline. The single-channel conductance, gamma e, varies between 40 and 50 pS at room temperature. 6. The NMDA channels open in bursts of short openings interrupted by brief closures. At -60 mV, the closures had a mean duration, tc, of 0.4 +/- 0.2 ms. The mean channel open time, to, was 5.9 +/- 1.0 ms for NMDA and 5.3 +/- 1.7 ms for L-glutamate. The mean burst duration, tb, was 10.5 +/- 0.7 ms for NMDA and 8.5 +/- 2.0 ms for L-glutamate. 7. When the temperature was increased between 14 and 24 degrees C, the NMDA channel conductance increased with a Q10 of 1.6 while the mean open time decreased with a Q10 close to 2. 8. The NMDA channel showed, in addition to the 'main' conductance state (40-50 pS), smaller conductance states of 15 and 35 pS.(ABSTRACT TRUNCATED AT 400 WORDS)