(R293) NMDAR in cultured astrocytes: Flux-independent pH sensor and flux-dependent regulator of mitochondria and plasma  membrane-mitochondria bridging

Pavel Montes de Oca B (Instituto de Fisiologia Celular-Lab BL-305)| Mauricio  Matus Núñez (UNAM)| Arturo Picones (Instituto de Fisiologia Celular-Lab BL-305)| Arturo Hernandez-Cruz (Instituto de Fisiologia Celular-Lab BL-305)

Glutamate N-methyl-D-aspartate (NMDA) receptor (NMDAR) is critical for neu-
rotransmission as a Ca2+ channel. Nonetheless, flux-independent signaling has also 
been demonstrated. Astrocytes express NMDAR distinct from its neuronal counter-
part, but cultured astrocytes have no electrophysiological response to NMDA. We 
recently demonstrated that in cultured astrocytes, NMDA at pH6 (NMDA/pH6) act-
ing 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 deple-
tion requires both: pH6 and flux-dependent NMDAR signaling. Plasma membrane 
(PM) NMDAR guard a non-random distribution relative to the ER and mitochon-
dria. 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 sug-
gest 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 demon-
strate that NMDAR has concomitant flux-independent and flux-dependent actions in 
cultured astrocytes. This work has been recently accepted for publication in The FASEB J.