Hyperforin--a key constituent of St. John's wort specifically activates TRPC6 channels.

Hyperforin, a bicyclic polyprenylated acylphloroglucinol derivative, is the main active principle of St. John's wort extract responsible for its antidepressive profile. Hyperforin inhibits the neuronal serotonin and norepinephrine uptake comparable to synthetic antidepressants. In contrast to synthetic antidepressants directly blocking neuronal amine uptake, hyperforin increases synaptic serotonin and norepinephrine concentrations by an indirect and yet unknown mechanism. Our attempts to identify the molecular target of hyperforin resulted in the identification of TRPC6. Hyperforin induced sodium and calcium entry as well as currents in TRPC6-expressing cells. Sodium currents and the subsequent breakdown of the membrane sodium gradients may be the rationale for the inhibition of neuronal amine uptake. The hyperforin-induced cation entry was highly specific and related to TRPC6 and was suppressed in cells expressing a dominant negative mutant of TRPC6, whereas phylogenetically related channels, i.e., TRPC3 remained unaffected. Furthermore, hyperforin induces neuronal axonal sprouting like nerve growth factor in a TRPC6-dependent manner. These findings support the role of TRPC channels in neurite extension and identify hyperforin as the first selective pharmacological tool to study TRPC6 function. Hyperforin integrates inhibition of neurotransmitter uptake and neurotrophic property by specific activation of TRPC6 and represents an interesting lead-structure for a new class of antidepressants.

Involvement of oxidative stress induction in Na+ toxicity and its relation to the inhibition of a Ca2+ -dependent but calcineurin-independent mechanism in Saccharomyces cerevisiae.

Uridine 5'-hexadecylphosphate (UMPC16) inhibited the growth of Saccharomyces cerevisiae under a hypersaline stress condition with Na+ more strongly than the calcineurin inhibitor cyclosporine A (CsA). Additional Ca2+ supplementation similarly suppressed the inhibitory activities of UMPC16 and CsA on yeast cell growth in a medium with Na+. UMPC16, but not CsA, accelerated mitochondrial reactive oxygen species (ROS) generation in combination with Na+, suggesting its inhibition of a Ca2+ -dependent but calcineurin-independent mechanism for protection against Na+ toxicity.

Momordica charantia extracts inhibit uptake of monosaccharide and amino acid across rat everted gut sacs in-vitro.

The inhibitory effects of Momordica charantia extracts were studied on the uptake of glucose and tyrosine across rat everted gut sacs in vitro. The aqueous extract of the plant was found to inhibit primarily the uptake of glucose in a dose-dependent manner. Uptake of tyrosine was affected at high substrate concentrations only. The extract was also found to decrease the absorptive capacity of fluid across the small intestine and sodium ions. It is hypothesized that the effects of Momordica could involve a washout of glucose from the blood stream.

Mechanism of regulation of Na+ transport by angiotensin II in primary renal cells.

BACKGROUND: Angiotensin II (Ang II) has a dose-dependent, biphasic effect on the activity of the Na+/H+ antiport system in the renal proximal tubule (RPT). The aim of the present study was to further delineate the signaling pathways involved in Ang II action. METHODS: To examine Ang II signaling, 22Na+ uptake studies were conducted with a primary rabbit RPT cell culture system. The activation of phospholipase A2 (PLA2) was assessed by measuring the release of [3H]-arachidonic acid (AA), and changes in intracellular calcium levels were determined by means of confocal microscopy. RESULTS: Low dosages of Ang II (<10-10 mol/L) stimulated Na+ uptake, whereas high dosages of Ang II (>10-10 mol/L) inhibited Na+ uptake. Ang II (>10-10 mol/L) also caused an increase in AA release associated with an increase in intracellular calcium. Not only did exogenous AA inhibit Na+ uptake, but two PLA2 inhibitors (mepacrine and AACOCF3) blocked the Ang II-mediated inhibition of Na+ uptake. However, the cytochrome P450-dependent epoxygenase inhibitor econazole also blocked the Ang II-induced inhibition of Na+ uptake. Inhibition of Na+ uptake was obtained by the metabolic product of the epoxygenase 5,6-EET. In turn, the inhibitory effect of 5,6-EET was blocked by indomethacin. CONCLUSIONS: The results indicate the involvement of a calcium-dependent PLA2 in mediating the inhibitory effect of Ang II on Na+ uptake. The AA, which is released following PLA2 activation, acts indirectly, through its own metabolism, via a cytochrome P450 epoxygenase pathway and ultimately cyclooxygenase itself.

Mechanisms of sodium transport at the blood-brain barrier studied with in situ perfusion of rat brain.

The mechanism of unidirectional transport of sodium from blood to brain in pentobarbital-anesthetized rats was examined using in situ perfusion. Sodium transport followed Michaelis-Menten saturation kinetics with a Vmax of 50.1 nmol/g/min and a Km of 17.7 mM in the left frontal cortex. The kinetic analysis indicated that, at a physiologic sodium concentration, approximately 26% of sodium transport at the blood-brain barrier (BBB) was carrier mediated. Dimethylamiloride (25 microM), an inhibitor of Na+/H+ exchange, reduced sodium transport by 28%, whereas phenamil (25 microM), a sodium channel inhibitor, reduced the transfer constant for sodium by 22%. Bumetanide (250 microM) and hydrochlorothiazide (1.5 mM), inhibitors of Na(+)-K(+)-2Cl-/NaCl symport, were ineffective in reducing blood to brain sodium transport. Acetazolamide (0.25 mM), an inhibitor of carbonic anhydrase, did not change sodium transport at the BBB. Finally, a perfusate pH of 7.0 or 7.8 or a perfusate PCO2 of 86 mm Hg failed to change sodium transport. These results indicate that 50% of transcellular transport of sodium from blood to brain occurs through Na+/H+ exchange and a sodium channel in the luminal membrane of the BBB. We propose that the sodium transport systems at the luminal membrane of the BBB, in conjunction with Cl-/HCO3- exchange, lead to net NaCl secretion and obligate water transport into the brain.

Calcium retention and increased vascular reactivity caused by a hypothalamic sodium transport inhibitor.

1. Using a previously established method of isolating an active-sodium-transport inhibitor (ASTI) from hypothalamic cell culture medium, the inhibitor was isolated and partially purified from sequential passages through Sephadex G-25 and h.p.l.c., and its effects on de-endothelialized rabbit aortic strips were investigated. 2. ASTI caused a cumulative concentration-dependent increase in tension which reversed slowly after wash, and the wash showed an identical effect on fresh strips. 3. Ouabain, used as a control, also caused a concentration-dependent increase in tension which reached a plateau at a concentration of 10 mmol/l. Both ouabain and ASTI caused a significant potentiation of the vasoconstrictor effect of noradrenaline at concentrations of 1 nmol/l-0.1 mmol/l. 4. Both ASTI and ouabain caused a significantly greater (P less than 0.01) calcium retention than control medium in aortic strips. 5. Incubation of ASTI with prolidase, chymotrypsin and carboxypeptidase A destroyed the vasoconstrictor effects as well as its inhibitory effects on sodium, potassium-dependent adenosine triphosphatase and sodium efflux from erythrocytes, but leucine aminopeptidase was ineffective. 6. These studies suggest that hypothalamic cells in culture release a peptidic inhibitor of active sodium transport which increases vascular reactivity, potentiates vasoconstrictor effects of noradrenaline and causes calcium retention.

An active sodium transport inhibitor released from spontaneously hypertensive and normotensive rat fetal hypothalamic cells in culture.

An inhibitor of active sodium transport (Na+ + K+-ATPase inhibitor), partially purified from the culture medium of fetal rat hypothalamic cells, has been shown to possess vasoactive properties. In order to explore whether fetal hypothalamic neurons from spontaneously hypertensive rats produce higher concentrations of the inhibitor than produced by those from normotensive rats, we cultured hypothalamic cells from both sources. An average of 10(6) cells per hypothalamus was obtained, and heat-treated (80 degrees C for 10 minutes) culture medium (120 ml) after lyophilization yielded 0.8 g of material. After Sephadex G-15 chromatography, 0.5 g of lyophilized medium from fetal hypothalamic neurons of spontaneously hypertensive rats yielded 254 +/- 47 arbitrarily defined units of Na+ + K+-ATPase inhibitory activity compared with 238 +/- 59 units from identical material of normotensive source. These studies show that the production of the hypothalamic Na+ + K+-ATPase inhibitor is not increased at the fetal stage in the spontaneously hypertensive rats.

Sodium pump inhibitor in the serum of patients with essential hypertension and its partial purification from hemofiltrate.

A 3-fold higher concentration of "endogenous digitalis" is found in the serum of patients with essential hypertension than in the serum of normotensives, whose concentration was determined in 22 normotensive probands by an receptor assay using isolated (Na+ + K+)-ATPase as 76.3 +/- 9.3 nM ouabain equivalents. Since the concentration of "endogenous digitalis" was 7-19 fold higher in 2 patients, who had become uremic due to a malignant hypertension and since their serum levels fell 3-fold by hemodialysis, the hemofiltrate was used for partial purification of the substance. This was possible by hydrophobic chromatography on Amberlite XAD-2, octadecyl-coated silica gel, anion exchange chromatography and affinity chromatography on membrane-bound (Na+ + K+)-ATPase. The partially purified substance behaved like the material described by Cloix et al. (1985) Biochem. Biophys. Res. Commun. 131:1234-1240 and competed with digoxin for digoxin antibodies. Ascorbic acid isolated on the search for an inhibitor of (Na+ + K+)-ATPase from beef brain inhibited [3H]ouabain binding due to a decrease of the ouabain binding sites by a reduction of a group within the ATP binding site of the enzyme. Unsaturated fatty acids claimed to be "endogenous digitalis (Tamura et al. [1985] J. Biol. Chem. 260:9672-9677)" also lowered the capacity of the cardiac glycoside binding site but did not compete with ouabain.

Investigation of the endogenous Na+-pump inhibitor in essential hypertension and blood volume expansion.

The digitalis-like activities of plasma extracts from 108 patients and normal subjects were measured by their ability to compete with ouabain for binding to the digitalis sites of the Na+-pump. High levels were found in 18 of 54 untreated patients with moderate hypertension, 10 of 14 patients with end-stage renal failure and six patients with active acromegaly. These levels returned to control values after dialysis in the patients with renal insufficiency and high levels of the inhibitor, and after successful surgery and cobalt therapy in seven acromegalic patients. An increase in circulating Na+, K+-ATPase inhibitor was also found in rats after chronic sodium loading. These results indicate that levels of the circulating compound with digitalis-like properties do not result from high blood pressure but, rather, are related to blood volume and Na+ balance.

Evidence against a circulating ouabain-like transport inhibitor as a cause of increased red cell sodium in essential hypertension.

Red cell sodium and potassium concentrations, and total ouabain-sensitive and ouabain-insensitive first order red constants for sodium efflux, were measured in 15 patients (11 female, 4 male) with essential hypertension (162 +/- 14/99 +/- 7 (s.d. mmHg) and 15 normotensive control subjects (117 +/- 17/64 +/- 10 mmHg) individually matched for age (45.8 +/- 10 versus 45.7 +/- 10 years, respectively, sex, weight (68.6 +/- 16.1 versus 64.7 +/- 11.2 kg) and blood group. To test for possible plasma inhibitors of sodium transport in hypertension, total efflux rate constants were measured in red cells incubated in two plasma samples, from either the same or the complementary (paired) subjects, respectively. Intracellular sodium was significantly increased in patients (11.8 +/- 3.45 versus 8.75 +/- 2.48 mmols/l of red cell water; P = 0.023). Intracellular potassium and total and ouabain-sensitive sodium efflux rate constants, were similar in both groups. Sodium efflux was similar when cells were incubated in the homologous and complementary plasma samples. Ouabain-insensitive rate constants were decreased in the patients (0.16 +/- 0.08 versus 0.20 +/- 0.05 h-1) but the difference was of borderline significance (P = 0.059). These results confirm the presence of abnormal intracellular sodium concentrations and membrane transport in essential hypertension but are not consistent with the suggestion that the abnormalities are due to a circulating sodium transport inhibitor.

Release of an active sodium transport inhibitor (ASTI) from rat hypothalamic cells in culture.

To investigate the hypothesis whether the hypothalamus releases an active (ouabain-sensitive) sodium transport inhibitor, we cultured hypothalamic and cortical cells from day 17 fetal rats. Culture media from hypothalamic cells reduced the total erythrocyte sodium efflux rate constant from 0.487 +/- (SE) 0.014 to 0.408 +/- 0.013 (P less than 0.001), and the ouabain-sensitive rate constant from 0.305 +/- 0.015 to 0.240 +/- 0.016 (P less than 0.01). Hypothalamic media also showed a dose-dependent displacement of [3H]-ouabain-binding to erythrocyte membranes. Neither cortical nor conditioned media (incubated without cells) had any effect. Various well-characterized hormones of hypothalamic origin failed to inhibit sodium efflux rate constant. These studies demonstrate that fetal rat hypothalamic cells contain and release a factor which inhibits sodium transport in human erythrocytes.

Plasma sodium pump inhibitor in essential hypertension and normotensive subjects with hypertensive heredity.

The presence in plasma extracts of a sodium pump inhibitor with digitalis-like properties was investigated by two complementary tests: decrease in the affinity of ouabain binding to human red blood cells and inhibition of Na+,K+-ATPase. The results of the two methods were correlated (r = 0.76, n = 44, p less than 0.01), suggesting that the same factor may be responsible for both effects. All subjects with elevated values were hypertensive or normotensive and had a family history of hypertension. Forty percent of the subjects in these two groups had high inhibition values. The elevation was significant (p less than 0.01) when compared with values in normotensive subjects with no hypertensive heredity. Increased inhibition was observed in patients taking beta-blocking agents; conversely, diuretics normalized the values. No correlation was found between pump inhibition and age, sex, blood pressure, levels of plasma K+ or Na+, or plasma renin activity. These data show the existence of a sodium pump inhibitor in the plasma of some subjects and point to a possible association with hypertension. They also underline the importance of genetic background and the heterogeneity of essential hypertension.

The use of lithium carbonate in the treatment of mood disorders.

The use of lithium ions in the treatment of manic states is discussed. Lithium is possibly the only specific drug treatment presently available for the major psychoses and has met with enthusiasm in England, Scandinavia, Australia and, more recently, in Canada and the United States.A number of the published papers on the subject are not sufficiently comprehensive to provide guidance for even its empiric use; some lack the necessary controls and design to permit comparisons with other studies.Some clinicians with wide experience of lithium therapy do not maintain laboratory control of patients by ordering serum lithium determinations but rely entirely on clinical judgement in establishing drug schedules. This is not advised if one has little experience with lithium therapy because of the possible side effects and toxicity.