Introduction Initially postulated to represent the main
Initially postulated to represent the main source of ATP production in eukaryotic cells, mitochondria are recently recognized to perform manifold essential functions beyond energy production, impacting most areas of cell biology and medicine . Mitochondrial metabolism is both the basis for and target of nutrient signals that ultimately orchestrate an integrated physiological response. These organelles appear to be involved in the conserved energy sensing intracellular signaling pathways AMP-activated protein kinase (AMPK) signaling , adrenergic signaling  and apoptosis . Mitochondrial functions have been found as essential parts of energy metabolism of different organs such as heart, liver, kidneys and brain [5,6]. Moreover, it was shown that oxidative phosphorylation is the main mechanism initially providing energy to power neuronal activity, that the main subcellular mechanisms underlying information processing, i.e., presynaptic action potentials, neurotransmitter release, postsynaptic currents, and postsynaptic action potentials, all consume oxygen, and that an astrocyte–neuron lactate shuttle is not needed for oxidative phosphorylation to occur . Indeed, the dependence of neurotransmission on energy supply is not surprising given the especial role of ATP in maintaining synaptic membrane potential, and fueling numerous steps of the vesicle cycle. It can be shared in several essential processes. First of all, the presynapse may experience large intracellular calcium rises acting locally to elicit vesicle fusion, or globally to influence various signaling pathways upon stimulation . Mitochondria are thought to modulate these processes by regulating calcium levels although plasma membrane Ca2+-ATPases, and the endoplasmic reticulum (ER) yield more contribution in them . Thereby, the synchronization between mitochondrial conductivity and calcium dynamics in the presynaptic terminal suggests an active role of mitochondria in synaptic plasticity . Secondly, the nucleotide release can cause some special signaling pathways for instance the synapse relies on ATP for priming and to drive NSF-mediated disassembly of soluble NSF attachment protein receptor complexes . Relying on these observations it would be very reasonable to suppose direct involvement of mitochondria into a cell signaling. In order to be a part of some pathway the different receptors have to be located on inner or outer mitochondrial membranes. Indeed, typical plasma membrane receptors have been found in mitochondrial membrane to date: nicotine NVP-BSK805 receptor , canabioid receptor CB1 , a group of thyroid and steroid hormones receptors . These discoveries declare clearly an existence of entire “cellular” receptor system in mitochondria, with one of the purposes to co-ordinate cell and mitochondria operation. If the hypothesis of mitochondrial participation in cell signaling pathways is true then other famous receptor systems for mediators has to observed in the organelle membranes as well. Thus, in this work, the presence of three more receptors in mitochondria is demonstrated, that are generally recognized as plasma membrane receptors of nervous cells: N-methyl-d-aspartate receptor (NMDAR), γ-aminobutyric acid type A (GABAAR) and type B (GABABR) receptors. NMDAR has recently been found in synaptic brain mitochondria . Following indirect signs, we assume NMDAR presence in non-synaptic brain  and also in cardiac mitochondria . In this paper, NMDAR and GABAR existence in rat heart mitochondria is proved by direct methods of immunogold labeling and immunoblot.
NMDAR detection in mitochondria
Discussion The obtained results argue for NMDAR and GABAR presence in rat heart mitochondria. These receptors are known to play a key role in higher nervous activity [21,22]. Their role in mitochondria is not clear at present time, but one can make an assumption about NMDAR operation in hear mitochondria based on literature data. NMDAR is a cation selective channel activated by glutamate. For this type of receptor glycine acts as a co-agonist and the pore is penetrable to calcium ions. Ca++ is known to reinforce mitochondrial energetic, including FAD-glycerol phosphate dehydrogenase, pyruvate dehydrogenase, NAD-isocitrate dehydrogenase and oxoglutarate dehydrogenase activation , as well as FO F1 -ATPase . MitoNMDAR being in relation to myocardial contractile activity could be an attractive supposition. Thereupon, one should mention the presence of NMDAR2B subunit presence in myocardial cells of neonatal rats . It is found in Z-disk and co-localized with ryanodine receptor (RyR2) which provides Ca++ release from endoplasmic reticulum. One can believe that a newly discovered mitoNMDAR also keeps company with RyR2 or with inositol-3-phosphate receptor (IP3), which is placed in contact membranes (MAMs) between mitochondria and endoplasmic reticulum . It should be emphasized that, in addition to the data from Ref. , we precisely determine NMDAR subcellular location (in mitochondria) and demonstrate the presence of both receptor subunits in mature rats. NMDAR role in mitochondria requires an independent extensive study. At this point, a model can be used similar to excitation–signaling coupling model [27,28], that states ATP synthesis rate to be synchronized with cardiac rhythm through Ca++ flux. In our model, one can assume the mitochondrial Ca++ uptake to be provided under the auspices of MAMs-localized mitoNMDAR and VDAC. Ca++ is known to promote mitochondrial reactive oxygen species (ROS) production under certain conditions. Therefore, glutamate-dependent H2O2 synthesis detected previously in our laboratory [16,17] can be considered as one of the effects resulting from mitoNMDAR operation. Taking into account that the development of oxidative stress in patients with ischemic stroke is caused by primary disruption of bioenergetic processes during the reduction of antioxidant system activity  glutamate-induced excess of ROS outcome yields the reversible and irreversible mitochondrial events under micro level forming an ischemic area in different organs . In the present work, glutamate NMDAR in mitochondria is revealed to be accompanied by GABAR. GABAergic system is found in the heart in sinoatrial node . Furthermore, an expression of two genes is observed in cardiomyocytes, that produce enzymes implementing GABA synthesis from succinate [32,33]. Since GABAR agonist (γ-aminobutyric acid) presence is possible in heart cells, the corresponding receptor is also operable. One can also surmise mitoGABAAR to conduct a superoxide, similarly to other chloride channels .