Involvement of the Na+, K+-ATPase α1 Isoform and Endogenous Cardiac Steroids in Depression- and Manic-like Behaviors.

Bipolar disorder (BD) is a severe and common chronic mental illness characterized by recurrent mood swings between depression and mania. The biological basis of the disease is poorly understood, and its treatment is unsatisfactory. Na+, K+-ATPase is a major plasma membrane transporter and signal transducer. The catalytic α subunit of this enzyme is the binding site for cardiac steroids. Three α isoforms of the Na+, K+-ATPase are present in the brain. Previous studies have supported the involvement of the Na+, K+-ATPase and endogenous cardiac steroids (ECS) in the etiology of BD. Decreased brain ECS has been found to elicit anti-manic and anti-depressive-like behaviors in mice and rats. However, the identity of the specific α isoform involved in these behavioral effects is unknown. Here, we demonstrated that decreasing ECS through intracerebroventricular (i.c.v.) administration of anti-ouabain antibodies (anti-Ou-Ab) decreased the activity of α1+/- mice in forced swimming tests but did not change the activity in wild type (wt) mice. This treatment also affected exploratory and anxiety behaviors in α1+/- but not wt mice, as measured in open field tests. The i.c.v. administration of anti-Ou-Ab decreased brain ECS and increased brain Na+, K+-ATPase activity in wt and α1+/- mice. The serum ECS was lower in α1+/- than wt mice. In addition, a study in human participants demonstrated that serum ECS significantly decreased after treatment. These results suggest that the Na+, K+-ATPase α1 isoform is involved in depressive- and manic-like behaviors and support that the Na+, K+-ATPase/ECS system participates in the etiology of BD.

Endogenous Cardiac Steroids in Bipolar Disorder: State of the Art.

Bipolar disorder (BD) is a severe psychiatric illness with a poor prognosis and problematic, suboptimal, treatments. Treatments, borne of an understanding of the pathoetiologic mechanisms, need to be developed in order to improve outcomes. Dysregulation of cationic homeostasis is the most reproducible aspect of BD pathophysiology. Correction of ionic balance is the universal mechanism of action of all mood stabilizing medications. Endogenous sodium pump modulators (collectively known as endogenous cardiac steroids, ECS) are steroids which are synthesized in and released from the adrenal gland and brain. These compounds, by activating or inhibiting Na+, K+-ATPase activity and activating intracellular signaling cascades, have numerous effects on cell survival, vascular tone homeostasis, inflammation, and neuronal activity. For the past twenty years we have addressed the hypothesis that the Na+, K+-ATPase-ECS system may be involved in the etiology of BD. This is a focused review that presents a comprehensive model pertaining to the role of ECS in the etiology of BD. We propose that alterations in ECS metabolism in the brain cause numerous biochemical changes that underlie brain dysfunction and mood symptoms. This is based on both animal models and translational human results. There are data that demonstrate that excess ECS induce abnormal mood and activity in animals, while a specific removal of ECS with antibodies normalizes mood. There are also data indicating that circulating levels of ECS are lower in manic individuals, and that patients with BD are unable to upregulate synthesis of ECS under conditions that increase their elaboration in non-psychiatric controls. There is strong evidence for the involvement of ion dysregulation and ECS function in bipolar illness. Additional research is required to fully characterize these abnormalities and define future clinical directions.

Synthesis and Biological Evaluation of Novel Bufalin Derivatives.

Bufalin and other cardiac steroids (CS) have been used for centuries for the treatment of congestive heart failure, arrhythmias, and other maladies. However, toxicity and the small therapeutic window of this family of steroids limit their use. Therefore, attempts to synthesize a potent, but less toxic, CS are of major importance. In the present study, two novel bufalin derivatives were synthesized and some of their pharmacological properties were characterized. The reaction of bufalin with Ishikawa's reagent resulted in the production of two novel bufalin derivatives: bufalin 2,3-ene and bufalin 3,4-ene. The compounds were purified with TLC and HPLC and their structure was verified with UV, NMR, and MS analyses. The biological activities of these compounds were evaluated by testing their ability to inhibit the Na+, K+-ATPase activity of the brain microsomal fraction to induce cytotoxic activity against the NCI-60 human tumor cell line panel and non-cancer human cells, and to increase the force of contraction of quail embryonic heart muscle cells in culture. The two steroids exhibited biological activities similar to those of other CS in the tested experimental systems, but with reduced cytotoxicity, advocating their development as drugs for the treatment of heart failure and arrhythmias.

Chemical Element Profiling in the Sera and Brain of Bipolar Disorders Patients and Healthy Controls.

Bipolar Disorder (BD) is a severe recurrent affective mood disorder characterized by a wide range of lifelong mood swings, varying between depressive and manic states. BD affects more than 1% of the world's population irrespective of nationality, ethnic origin, or socioeconomic status and is one of the main causes of disability among young people, leading to cognitive and functional impairment and raised mortality, particularly death by suicide. Trace elements play a vital role in many biochemical and physiological processes. Compelling evidence shows that element toxicity might play a crucial role in the onset and progression of neurodegenerative disorders, but their involvement in mood disorders has been scarcely studied. In the present investigation, we determined the concentration of 26 elements in the serum of BD patients before and after treatment and in postmortem brain samples from BD patients and compared them with matched controls. The only element that was reduced significantly in the serum following treatment was vanadium (V). Furthermore, the concentration of Al, B, Cu, K, Mg and V were significantly lower in the pre-frontal cortex of BD patients compared with those of the controls. A comparison of Spearman's rank correlation coefficients between the elements in the serum and brain of BD patients and control groups pointed to boron and aluminum as being involved in the disease. These results suggest that there is a disturbance in the elements' homeostasis and the inter-elements' relationship in the brain of BD patients and advocate a thorough examination of the possible involvement of chemical elements in different stages of the disease.

Vanadium in Bipolar Disorders-Reviving an Old Hypothesis.

Bipolar disorder (BD) is a severe and common chronic mental illness. The biological basis of the disease is poorly understood and its treatment is unsatisfactory. Our previous studies supported the notion that alterations in Na+, K+-ATPase activity were involved in the etiology of BD. As various chemical elements inhibit Na+, K+-ATPase, we determined the concentration of 26 elements in the serum of BD patients before and after treatment and in postmortem brain samples from BD patients, and compared them with matched controls. The only element that was reduced significantly in the serum following treatment was vanadium (V). Furthermore, the concentration of V was significantly lower in the pre-frontal cortex of BD patients compared with that of the controls. Intracerebroventricular administration of V in mice elicited anxiolytic and depressive activities, concomitantly inhibited brain Na+, K+-ATPase activity, and increased extracellular signal-regulated kinase phosphorylation. A hypothesis associating V with BD was set forth decades ago but eventually faded out. Our results are in accord with the hypothesis and advocate for a thorough examination of the possible involvement of chemical elements, V in particular, in BD.

Na+, K+-ATPase α Isoforms and Endogenous Cardiac Steroids in Prefrontal Cortex of Bipolar Patients and Controls.

Bipolar disorder is a chronic multifactorial psychiatric illness that affects the mood, cognition, and functioning of about 1-2% of the world's population. Its biological basis is unknown, and its treatment is unsatisfactory. The α1, α2, and α3 isoforms of the Na+, K+-ATPase, an essential membrane transporter, are vital for neuronal and glial function. The enzyme and its regulators, endogenous cardiac steroids like ouabain and marinobufagenin, are implicated in neuropsychiatric disorders, bipolar disorder in particular. Here, we address the hypothesis that the α isoforms of the Na+, K+-ATPase and its regulators are altered in the prefrontal cortex of bipolar disease patients. The α isoforms were determined by Western blot and ouabain and marinobufagenin by specific and sensitive immunoassays. We found that the α2 and α3 isoforms were significantly higher and marinobufagenin levels were significantly lower in the prefrontal cortex of the bipolar disease patients compared with those in the control. A positive correlation was found between the levels of the three α isoforms in all samples and between the α1 isoform and ouabain levels in the controls. These results are in accordance with the notion that the Na+, K+-ATPase-endogenous cardiac steroids system is involved in bipolar disease and suggest that it may be used as a target for drug development.

Na⁺, K⁺-ATPase Signaling and Bipolar Disorder.

Bipolar disorder (BD) is a severe and common chronic mental illness characterized by recurrent mood swings between depression and mania. The biological basis of the disease is poorly understood and its treatment is unsatisfactory. Although in past decades the "monoamine hypothesis" has dominated our understanding of both the pathophysiology of depressive disorders and the action of pharmacological treatments, recent studies focus on the involvement of additional neurotransmitters/neuromodulators systems and cellular processes in BD. Here, evidence for the participation of Na⁺, K⁺-ATPase and its endogenous regulators, the endogenous cardiac steroids (ECS), in the etiology of BD is reviewed. Proof for the involvement of brain Na⁺, K⁺-ATPase and ECS in behavior is summarized and it is hypothesized that ECS-Na⁺, K⁺-ATPase-induced activation of intracellular signaling participates in the mechanisms underlying BD. We propose that the activation of ERK, AKT, and NFκB, resulting from ECS-Na⁺, K⁺-ATPase interaction, modifies neuronal activity and neurotransmission which, in turn, participate in the regulation of behavior and BD. These observations suggest Na⁺, K⁺-ATPase-mediated signaling is a potential target for drug development for the treatment of BD.

Essential Opposite Roles of ERK and Akt Signaling in Cardiac Steroid-Induced Increase in Heart Contractility.

Interaction of cardiac steroids (CS) with the Na(+), K(+)-ATPase elicits, in addition to inhibition of the enzyme's activity, the activation of intracellular signaling such as extracellular signal-regulated (ERK) and protein kinase B (Akt). We hypothesized that the activities of these pathways are involved in CS-induced increase in heart contractility. This hypothesis was tested using in vivo and ex vivo wild type (WT) and sarcoplasmic reticulum Ca(2+) atpase1a-deficient zebrafish (accordion, acc mutant) experimental model. Heart contractility was measured in vivo and in primary cardiomyocytes in WT zebrafish larvae and acc mutant. Ca(2+) transients were determined ex vivo in adult zebrafish hearts. CS dose dependently augmented the force of contraction of larvae heart muscle and cardiomyocytes and increased Ca(2+) transients in WT but not in acc mutant. CS in vivo increased the phosphorylation rate of ERK and Akt in the adult zebrafish heart of the two strains. Pretreatment of WT zebrafish larvae or cardiomyocytes with specific MAPK inhibitors completely abolished the CS-induced increase in contractility. On the contrary, pretreatment with Akt inhibitor significantly enhanced the CS-induced increase in heart contractility both in vivo and ex vivo without affecting CS-induced Ca(2+) transients. Furthermore, pretreatment of the acc mutant larvae or cardiomyocytes with Akt inhibitor restored the CS-induced increase in heart contractility also without affecting Ca(2+) transients. These results support the notion that the activity of MAPK pathway is obligatory for CS-induced increases in heart muscle contractility. Akt activity, on the other hand, plays a negative role, via Ca(2+) independent mechanisms, in CS action. These findings point to novel potential pharmacological intervention to increase CS efficacy.

Endogenous cardiac steroids in animal models of mania.

OBJECTIVES: Bipolar disorder (BD) is a complex psychiatric disorder characterized by mania and depression. Alterations in brain Na(+) , K(+) -ATPase and cardiac steroids (CSs) have been detected in BD, raising the hypothesis of their involvement in this pathology. The present study investigated the behavioral and biochemical consequences of a reduction in endogenous brain CS activity in animal models of mania. METHODS: Amphetamine (AMPH)-induced hyperactivity in BALB/c and black Swiss mice served as a model of mania. Behavior was evaluated in the open-field test in naïve mice or in mice treated with anti-ouabain antibodies. CS levels were determined by enzyme-linked immunosorbent assay (ELISA), using sensitive and specific anti-ouabain antibodies. Extracellular signal-regulated kinase (ERK) and protein kinase B (Akt) phosphorylation levels in the frontal cortex were determined by western blot analysis. RESULTS: Administration of AMPH to BALB/c and black Swiss mice resulted in a marked increase in locomotor activity, accompanied by a threefold increase in brain CSs. The lowering of brain CSs by the administration of anti-ouabain antibodies prevented the hyperactivity and the increase in brain CS levels. AMPH caused an increase in phosphorylated ERK (p-ERK) and phosphorylated Akt (p-Akt) levels in the frontal cortex, which was significantly reduced by administration of the antibodies. A synthetic 'functional antagonist' of CSs, 4-(3'α-15'ß-dihydroxy-5'ß-estran-17'ß-yl) furan-2-methyl alcohol, also resulted in attenuation of AMPH-induced hyperactivity. CONCLUSIONS: These results are in accordance with the notion that malfunctioning of the Na(+) , K(+) -ATPase/CS system may be involved in the manifestation of mania and identify this system as a potential new target for drug development.

Reduction in maternal circulating ouabain impairs offspring growth and kidney development.

Ouabain, a steroid present in the circulation and in various tissues, was shown to affect the growth and viability of various cells in culture. To test for the possible influence of this steroid on growth and viability in vivo, we investigated the involvement of maternal circulating ouabain in the regulation of fetal growth and organ development. We show that intraperitoneal administration of anti-ouabain antibodies to pregnant mice resulted in a >80% decline in the circulating ouabain level. This reduction caused a significant decrease in offspring body weight, accompanied by enlargement of the offspring heart and inhibition of kidney and liver growth. Kidney growth inhibition was manifested by a decrease in the size and number of nephrons. After the reduction in maternal circulating ouabain, kidney expression of cyclin D1 was reduced and the expression of the α1 isoform of the Na(+), K(+)-ATPase was increased. In addition, the elevation of proliferation signals including ERK1/2, p-90RSK, Akt, PCNA, and Ki-67, and a reduction in apoptotic factors such as Bax, caspase-3, and TUNEL were detected. During human pregnancy, the circulating maternal ouabain level increased and the highest concentration of the steroid was found in the placenta. Furthermore, circulating ouabain levels in women with small-for-gestational age neonates were significantly lower than the levels in women with normal-for-gestational age newborns. These results support the notion that ouabain is a growth factor and suggest that a reduction in the concentration of this hormone during pregnancy may increase the risk of impaired growth and kidney development.