Na⁺, K⁺-ATPase in the Brain: Structure and Function

Abstract:

Maintenance of the Na⁺ and K⁺ gradients between the intracellular and extracellular compartments of animal cells is a prerequisite for basic cellular homeostasis and for diverse functions of specialized cells. Na⁺, K⁺-ATPase (sodium- and potassium-activated adenosine 5′-triphosphatase), also called Na⁺ pump or Na⁺, K⁺-pump, is an ubiquitous membrane transport protein in mammalian cells, responsible to establish and maintain high K⁺ and low Na⁺ concentration in the cytoplasm; it is essential for normal resting membrane potentials and diverse cellular activities. As all ATPases, it hydrolyzes ATP and occludes ions within the membrane inserted segment of the protein during the translocation process. This system couples the hydrolysis of one molecule of ATP to exchange three sodium for two potassium ions, thus maintaining the normal gradient of these cations in animal cells. It acts as an electrogenic ion transporter, which is autophosphorylated on an aspartic acid residue by the gamma phosphate group of the ATP molecule that it hydrolyzes. Oxidative metabolism is very active in brain, where large amounts of chemical energy as ATP molecules are consumed, mostly required to maintain cellular Na⁺/K⁺ gradients that underlie resting and action potentials, which are involved in nerve impulse propagation, neurotransmitter release and cation homeostasis.

Na⁺, K⁺-ATPase is an olygomeric enzyme consisting of α and β subunits, both required for enzyme function. The α subunit is the catalytic one which crosses ten times the membrane; the binding sites for ATP and the inhibitor ouabain as well as ion occlusion occur in this subunit. Three isoforms with cell-type and development-specific expression patterns are present in brain. Subunits α1, α2, and α3 bind ouabain with low, intermediate and high affinity, respectively; the last two isoforms are associated to neurons whereas the first is related to glial cells. The β subunit regulates the activity and conformational stability of the α subunit, is highly glycosylated and presents a single-transmembrane span. It seems to participate in the modulation of enzyme affinity for K⁺ and Na⁺ and is important for ATP hydrolysis, ion transport, and binding of inhibitors such as ouabain. This subunit must interact with α subunit in order to accomplish ion transport. In association with the αβ dimmer there is a third subunit which belongs to the FXYD family proteins. It modulates transport function of the enzyme, seems not essential for functional Na⁺, K⁺-ATPase but most likely plays a regulatory role in a tissue-specific manner. These small proteins are considered as channels or regulators of ion channels; they are hydrophobic type I proteins with a single-transmembrane span. The mammalian FXYD proteins from FXYD1 to FXYD7 exhibit tissue-specific distribution. FXYD7 is expressed exclusively in the brain, it is associated with α1-β isozymes and is most likely involved in neuronal excitability. Phospholemman (FXYD1) is highly expressed in selected structures in the CNS.

Due to changing physiological needs, diverse regulatory mechanisms are operative to ensure not only appropriate expression of Na⁺, K⁺-ATPase but also required enzyme activity. Multiple mechanisms can regulate Na⁺, K⁺-ATPase activity, coherent with the diverse functional roles in different conditions, which leads this protein to be vulnerable to pathogenic insults and a target for therapeutics. Besides its dependence for ATP, this enzyme activity is regulated by phosphorylation state, endogenous ouabain-like substances, neurotransmitters, oxidant stress such as reactive oxygen species, and diverse peptides.

In addition to pumping ions, Na⁺, K⁺-ATPase seems to act as a signal transducer. Binding of ouabain to Na⁺, K⁺-ATPase changes the interaction of the enzyme with neighboring membrane proteins inducing the formation of multiple signaling modules, leading to Src kinase activation, transactivation of the epidermal growth factor receptor and enhanced formation of reactive oxygen species. Interaction of such signals results in the activity of several other cascades, including phospholipase C activation. The inhibition of Src blocks many of the ouabain-activated signaling pathways. Src binds to Na⁺, K⁺-ATPase directly and ouabain modulates the interaction between Na⁺, K⁺-ATPase and Src, leading to Src activation. The possibility that signaling Na⁺, K⁺-ATPase is concentrated in an separate pool on the plasma membrane has been advanced and potential interaction between Na⁺, K⁺-ATPase and caveolins studied, due to enzyme concentration in caveloae/rafts.

Apoptosis has been recognized in a wide range of disease states, and Na⁺, K⁺-ATPase deficiency seems to be a contributor to apoptosis and pathogenesis. Physiological concentration of intracellular K⁺ acts as a repressor of apoptotic effectors. The pro-apoptotic disruption of K⁺ homeostasis can be mediated by over-activated K⁺ channels or ionotropic glutamate receptor, accompanied by decreased K⁺ uptake provoked by Na⁺, K⁺-ATPase disfunction.

Na⁺, K⁺-ATPase activity is reduced or is insufficient to maintain adequate ionic balances during and after episodes of hypoglycemia, ischemia or epilepsy, as well as after administration of excitotoxins like glutamate agonists. Besides, a relationship between endogenous Na⁺, K⁺-ATPase inhibitors with central dysfunction as occurs in Parkinson's disease, CNS glioma, schizophrenia and epilepsy has been established.