Calcium Signaling in Brain Mitochondria

Abstract

Ca2؉ signaling in mitochondria has been mainly attributed to Ca 2؉ entry to the matrix through the Ca 2؉ uniporter and activation of mitochondrial matrix dehydrogenases. However, mitochondria can also sense increases in cytosolic Ca 2؉ and activation of mitochondrial dehydrogenases (mitDH) (1). However, mitochondria can also sense increases in cytosolic Ca 2ϩ through a mechanism that involves the aspartate-glutamate carriers (AGCs) and not the CaU (2-5). Aralar (Slc25a12), also named aralar1, the AGC isoform with predominant expression in brain (6 -8), is a component of the NADH malateaspartate shuttle (MAS), which in brain is activated by extramitochondrial Ca 2ϩ (S 0.5 324 nM) (4). Immunocytochemistry and in situ hybridization data and mRNA levels in acutely isolated brain cells indicate that aralar is localized preferentially in neurons (8 -12). This is consistent with a higher MAS activity in neuronal than astrocyte cultures (8) and with aralar being one of the more enriched proteins during differentiation of P19 cells to a neuronal phenotype (13). It is also consistent with the higher levels of aralar in total than in synaptosome-free mitochondrial fractions (9).Studies in cultured neurons, which have aralar as only AGC isoform, showed that small Ca 2ϩ signals that have limited access to mitochondria are able to activate the aralar-MAS pathway (4). However, large Ca 2ϩ signals that induce robust mitochondrial Ca 2ϩ transients fail to activate the pathway (4). This suggested that in neuronal mitochondria the aralar-MAS pathway is inhibited under conditions in which the CaUmitDH pathway is activated. This surprising result indicates that Ca 2ϩ activation of the malate aspartate shuttle and tricarboxylic acid cycle activity are somehow mutually exclusive in neurons. We have now studied the interplay between the AGG-MAS and CaU-mitDH pathways in brain mitochondria under Ca 2ϩ -stimulation conditions. Our results show that the shared metabolite ␣KG controls the relationship between the second transporter of MAS, the oxoglutarate carrier (OGC, Slc25a11), and ␣KGDH in the Krebs cycle, by virtue of the effects of Ca 2ϩ on the kinetics of ␣KGDH. Interestingly, the inhibition of OGC and MAS is fully reversible, in parallel with Ca 2ϩ egress from mitochondria. Behaviorally evoked brain activation results in an increased cerebral blood flow and increased glucose utilization, but paradoxically, this is not accompanied with an equivalent increase in oxygen utilization (14, 15). As a consequence, the oxygen glucose index, which is close to 6 when glucose is fully oxidized in resting conditions, falls to about 5 (16). This is accompanied by an increase in brain lactate production (14, 16 -19). Our results suggest that MAS inhibition during Ca 2ϩ -induced Krebs cycle activation would drive pyruvate to lactate formation and may play a role in lactate formation during brain activation. EXPERIMENTAL PROCEDURESAnimals and Materials-3-Month-old C57BL/6xSv129 mice were housed with a 12-h light cycle and fed ad libitum on standa...

CypD−/− hearts have altered levels of proteins involved in Krebs cycle, branch chain amino acid degradation and pyruvate metabolism

“…The MAS assay was adapted from Contreras and Satrustegui [24]. Briefly, 100 μg of mitochondria were mixed with 10 mM Tris (pH 7.4), 300 mM mannitol, 10 mM K 2 HPO 4 , 5 mM MgCl 2 and 10 μM KCl prior to adding 4 units/mL aspartate aminotransferase, 6 units/mL malate dehydrogenase, 140 μM NADH, 5 mM aspartate, and 0.5 mM ADP.…”

Glutamate–oxaloacetate transaminase activity promotes palmitate lipotoxicity in rat hepatocytes by enhancing anaplerosis and citric acid cycle flux

“…Glutamate can produce ␣KG through direct deamination by Glud1 or through transamination to produce NEAAs such as alanine or aspartate. Of particular interest is the GOT family of enzymes because they play a key role in the malate/aspartate shuttle, a critical redox shuttle whose activity can be influenced by alterations in intracellular calcium (23,24). GOT catalyzes the conversion of glutamate to ␣KG via transamination of oxaloacetate to aspartate.…”

Astrocytic energetics during excitatory neurotransmission: What are contributions of glutamate oxidation and glycolysis?

“…Determination of the capacity and regulation of the malate-aspartate shuttle is very important because it can help explain the rise in lactate production during brain activation. The malate-aspartate shuttle is stimulated by low cytoplasmic calcium levels and inhibited by greater influx of calcium into mitochondria, indicating that neuronal calcium dynamics during brain activation may be involved in regulation of neuronal lactate production (Contreras et al, 2007; Satrustegui et al, 2007; Bak et al, 2009; Contreras and Satrustegui, 2009; Bak et al, 2012). …”

“…Experimental evidence that considerable lactate production during activation may be neuronal (Ueda and Ikemoto, 2007; Caesar et al, 2008; Contreras and Satrustegui, 2009; Ivannikov et al, 2010; Bak et al, 2012) and that neurons and astrocytes can oxidize both glucose and lactate (Zielke et al, 2007, 2009) has led to serious challenges of the validity of the ANL transport-oxidation model, a synopsis of which is presented below.…”