Requirement of NMDA receptor/channels for intracellular high-energy phosphates and the extent of intraneuronal calcium buffering in cultured mouse hippocampal neurons.
Whole-cell patch-clamp recordings were undertaken in cultured mouse hippocampal neurons in order to investigate time-dependent changes in: (i) currents evoked by L-aspartic acid (Asp) and kainic acid (KAI), two excitatory amino acids active at N-methyl-D-aspartic acid (NMDA) and KAI receptor sites respectively, and (ii) tetrodotoxin (TTX) resistant voltage-dependent inward currents carried by Ca2+. Consistent with previous observations, Ca2+ currents gradually run down unless a support system containing Mg-ATP, phosphocreatine and creatinine phosphokinase is added to the intracellular medium. Here we report that, in addition to suppressing the rundown of currents through voltage-gated Ca2+ channels, such a support system is also necessary to prevent rundown of ionic currents through excitatory amino acid-gated channels of the NMDA type. When this support system was omitted from the recording pipette, currents induced by Asp, but not KAI, progressively declined over a period of 20 min and stabilized at values of about 50% of the initial. This progressive decline occurred regardless of the extent of intraneuronal Ca2+ buffering, indicating that it was not due to accumulation of cytosolic Ca2+. After the rundown, reversal potentials of ASP-induced currents were the same whether recorded with or without the intracellular support system and the Asp induced currents could be blocked by the specific NMDA channel blocker ketamine. We conclude that ionic currents through NMDA gated channels have two components: one requires high-energy phosphates and will run down if these are not supplied; the other requires no such supply and remains steady.
Mody
I
I
Playfair Neuroscience Unit, Toronto Hospital, Ont., Canada.
Salter
M W
MW
MacDonald
J F
JF
D000255:Adenosine Triphosphate / Q000378:metabolism / Q000502:physiology*; D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000502:physiology*; D002478:Cells, Cultured; D006624:Hippocampus / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D007608:Kainic Acid / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D016194:Receptors, N-Methyl-D-Aspartate; D017981:Receptors, Neurotransmitter / Q000378:metabolism / Q000502:physiology*
D000596:Amino Acids; D016194:Receptors, N-Methyl-D-Aspartate; D017981:Receptors, Neurotransmitter; D000255:Adenosine Triphosphate; D007608:Kainic Acid; D002118:Calcium