NMDAR in cultured astrocytes: Flux-independent pH sensor and flux-dependent regulator of mitochondria and plasma membrane-mitochondria bridging

FASEB J. 2020 Dec;34(12):16622-16644. doi: 10.1096/fj.202001300R. Epub 2020 Nov 1.

Abstract

Glutamate N-methyl-D-aspartate (NMDA) receptor (NMDAR) is critical for neurotransmission as a Ca2+ channel. Nonetheless, flux-independent signaling has also been demonstrated. Astrocytes express NMDAR distinct from its neuronal counterpart, but cultured astrocytes have no electrophysiological response to NMDA. We recently demonstrated that in cultured astrocytes, NMDA at pH6 (NMDA/pH6) acting through the NMDAR elicits flux-independent Ca2+ release from the Endoplasmic Reticulum (ER) and depletes mitochondrial membrane potential (mΔΨ). Here we show that Ca2+ release is due to pH6 sensing by NMDAR, whereas mΔΨ depletion requires both: pH6 and flux-dependent NMDAR signaling. Plasma membrane (PM) NMDAR guard a non-random distribution relative to the ER and mitochondria. Also, NMDA/pH6 induces ER stress, endocytosis, PM electrical capacitance reduction, mitochondria-ER, and -nuclear contacts. Strikingly, it also produces the formation of PM invaginations near mitochondria along with structures referred to here as PM-mitochondrial bridges (PM-m-br). These and earlier data strongly suggest PM-mitochondria communication. As proof of the concept of mass transfer, we found that NMDA/pH6 provoked mitochondria labeling by the PM dye FM-4-64FX. NMDA/pH6 caused PM depolarization, cell acidification, and Ca2+ release from most mitochondria. Finally, the MCU and microtubules were not involved in mΔΨ depletion, while actin cytoskeleton was partially involved. These findings demonstrate that NMDAR has concomitant flux-independent and flux-dependent actions in cultured astrocytes.

Keywords: NMDAR; astrocyte; calcium; flux-independent; mitochondria; organelle communication.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Astrocytes / metabolism*
  • Calcium / metabolism
  • Cell Membrane / metabolism*
  • Cells, Cultured
  • Endoplasmic Reticulum / metabolism
  • Endoplasmic Reticulum Stress / physiology
  • Hippocampus / metabolism
  • Hydrogen-Ion Concentration
  • Membrane Potential, Mitochondrial / physiology
  • Mitochondria / metabolism*
  • N-Methylaspartate / metabolism*
  • Neurons / metabolism
  • Rats
  • Receptors, N-Methyl-D-Aspartate / metabolism*
  • Signal Transduction / physiology

Substances

  • Receptors, N-Methyl-D-Aspartate
  • N-Methylaspartate
  • Calcium