Calcium Export from Neurons and Multi-Kinase Signaling Cascades Contribute to Ouabain Neuroprotection in Hyperhomocysteinemia

Abstract

Pathological homocysteine (HCY) accumulation in the human plasma, known as hyperhomocysteinemia, exacerbates neurodegenerative diseases because, in the brain, this amino acid acts as a persistent N-methyl-d-aspartate receptor agonist. We studied the effects of 0.1–1 nM ouabain on intracellular Ca2+ signaling, mitochondrial inner membrane voltage (φmit), and cell viability in primary cultures of rat cortical neurons in glutamate and HCY neurotoxic insults. In addition, apoptosis-related protein expression and the involvement of some kinases in ouabain-mediated effects were evaluated. In short insults, HCY was less potent than glutamate as a neurotoxic agent and induced a 20% loss of φmit, whereas glutamate caused a 70% decrease of this value. Subnanomolar ouabain exhibited immediate and postponed neuroprotective effects on neurons. (1) Ouabain rapidly reduced the Ca2+ overload of neurons and loss of φmit evoked by glutamate and HCY that rescued neurons in short insults. (2) In prolonged 24 h excitotoxic insults, ouabain prevented neuronal apoptosis, triggering proteinkinase A and proteinkinase C dependent intracellular neuroprotective cascades for HCY, but not for glutamate. We, therefore, demonstrated here the role of PKC and PKA involving pathways in neuronal survival caused by ouabain in hyperhomocysteinemia, which suggests existence of different appropriate pharmacological treatment for hyperhomocysteinemia and glutamate excitotoxicity.

Research Progress in Pharmacological Activities and Applications of Cardiotonic Steroids

“…At present, more and more studies have found that some cardiotonic steroid drugs also show significant pharmacological activities in neurological diseases. For example, CTS are involved in psychophysiological processes such as regulation of mental symptoms and protection of nerve cells through the regulation of neurotransmitters and homocysteine (HCY) (Sibarov et al, 2012;Lopachev et al, 2019;Ivanova et al, 2020).…”

“…Therefore, the control of nerve damage caused by HCY is an important means to prevent the occurrence of cerebrovascular diseases and alleviate the symptoms of Parkinson's disease and Alzheimer's disease. HCY-induced neurodegeneration is caused by neuronal permanent plasma membrane depolarization and cytosolic Ca 2+ overload of neurons, known as excitotoxic stress (Ivanova et al, 2020). Studies have found that ouabain can effectively antagonize neuronal damage caused by high HCY (Ivanova et al, 2020).…”

Roles of N-Methyl-D-Aspartate Receptors (NMDARs) in Epilepsy

“…Redox modulation of cysteine residues is one of the post-translational modifications of NMDAR. HCY accumulation in the human plasma, known as hyperhomocysteinemia, can exacerbate neurodegenerative diseases and act as a persistent NMDAR agonist (Ivanova M. et al, 2020). Meanwhile, HCY activates GluN2 subunit-dependent redox regulation of NMDAR by the reduction of NMDAR disulfide (Sibarov et al, 2020).…”

GluN2 Subunit-Dependent Redox Modulation of NMDA Receptor Activation by Homocysteine

“…It is well established that HCY activates N-methyl-d-aspartate receptors (NMDARs) [10,11] and group I metabotropic receptors [12,13]. The NMDAR activation by HCY and related intracellular calcium accumulation is thought to provide a major contribution to the HCY neurotoxicity [11,14,15]. NMDARs represent tetramer protein complexes composed of two GluN2 subunits, which form an agonist binding site for endogenous amino acids glutamate and HCY, and two GluN1 subunits bearing co-agonist binding sites for glycine [16] or d-serine [17,18].…”

“…HCY activates the GluN1/2A subtype of NMDARs with the highest affinity among NMDARs but provokes desensitization of GluN1/2B receptors [19]. This is a pharmacological feature determining the GluN2A-subunit dependence of the HCY neurotoxicity [15,[20][21][22][23]. NMDAR competitive antagonist (AP-5) and channel blockers (MK-801 and memantine) prevent HCY-induced neuronal damage both in vitro [7,11] and in vivo [24] during mild and intermediate hyperhomocysteinemia.…”