[Abstract Title]. - Society for Neuroscience

Poster
242. Glial Neuronal Interactions: Glutamate and Calcium
Time: Sunday, November 16, 2008, 1:00 pm - 5:00 pm
Program#/Poster#: 242.21/J4
Topic: B.11.b. Cell biology and signalling
Support: Swiss National Science Foundation grant # 3100A0-119827/1
Title: Sodium-dependent glutamate uptake induces mitochondrial matrix acidification in
astrocytes
Authors: G. AZARIAS 1 , D. POBURKO 2 , N. DEMAUREX 2 , *J.-Y. CHATTON 1 ;
1 Dept Physiol & DBCM, Univ. Lausanne, Lausanne, Switzerland; 2 Dept Cell Physiol. and
Metabolism, Univ. Geneva, Geneva, Switzerland
Abstract: An increasing body of evidence collected in vitro and in vivo suggests that
synaptically-released glutamate taken-up by astrocytes enhances the formation of lactate from
aerobic glycolysis. Astrocytic sodium homeostasis has been identified as a major factor linking
neuronal activity with astrocyte metabolism, but the mechanisms regulating the lactate shuttle
remain to be elucidated. Since astrocytic processes ensheathing synapses contain mitochondria,
we hypothesized that mitochondrial physiology is altered during glutamate capture. To tackle
this issue, we made use of Mito SypHer, a new genetically encoded pH sensor targeted to the
mitochondrial matrix. Cortical astrocytes in primary culture transfected with Mito SypHer
exhibited a typical mitochondrial labeling colocalized with CoroNa Red, a mitochondrial specific
dye. In situ calibration revealed that the ratiometric Mito SypHer signal increased monotonically
with mitochondrial pH. Short-term (1.5 min) application of glutamate evoked substantial
mitochondrial acidification in a concentration-dependent manner (e.g. delta pH = -0.30±0.02 for
200uM glutamate). Na + -coupled glutamate transporters seemed to be mainly responsible for the
mitochondrial acidification, as similar effects were obtained using D-aspartate and the glutamate
effect was inhibited by the glutamate transporter inhibitor DL-threo-beta-benzyloxyaspartate. To
investigate the mechanisms responsible for mitochondrial acidification, we first challenged the
role of the mitochondrial Na + /H + exchanger. Inhibition of this exchanger by ethyl-isopropyl
amiloride decreased the rate but not the amplitude of glutamate-evoked mitochondrial
acidification. We found that the cytosolic acidification associated with plasma-membrane
glutamate transport activity was responsible for the mitochondrial acidification. Compensating
cytosolic acidification using tri-ethylammonium during a pulse of glutamate almost abolished
glutamate-evoked mitochondrial acidification, without markedly altering cytosolic and
mitochondrial Na + responses. This study suggests that plasma membrane glutamate transport, in
addition to producing a substantial energy load, causes mitochondrial pH to drop, likely
weakening the driving force for mitochondrial ATP production.
Disclosures: G. Azarias, None; J. Chatton , None; D. Poburko, None; N. Demaurex, None.