Inhibition of CO<Subscript>2</Subscript> production from glucose by arginine in brain slices of rats

In the present study we evaluated the in vivo effect of arginine on CO₂ production from glucose in a medium with physiological and high extracellular K⁺ concentrations. We also tested the influence of the nitric oxide synthase inhibitor, N^ω-nitro-L-arginine methyl ester (L-NAME), on the effects elicited by arginine in order to investigate the possible participation of NO and/or its derivatives on the effects of arginine on CO₂ production from glucose. Sixty-day-old rats were treated with a single intraperitoneal injection of saline (control; group I), arginine (0.8 g/kg; group II), L-NAME (2.0 mg/kg; group III) or arginine (0.8 g/kg) plus L-NAME (2.0 mg/kg; group IV) and were killed 1 h later. Results showed that arginine administration inhibited CO₂ production from glucose at physiological extracellular K⁺ concentration and L-NAME prevented such effect. In contrast, arginine administration had no effect on CO₂ production from glucose at high extracellular K⁺ concentration. Based on these data, we also investigated the in vitro effect of arginine on CO₂ production from glucose in a medium with physiological extracellular K⁺ concentration in hippocampus slices. Results showed that arginine (0.1–1.5 mM) when added to the incubation medium did not alter CO₂ production from glucose in hippocampus slices of untreated rats. In addition, we also demonstrated that arginine inhibits Na⁺, K⁺-ATPase activity. The data indicate that the reduction of CO₂ production by arginine was probably mediated by NO and/or its derivatives, which could act inhibiting the activity of Na⁺, K⁺-ATPase. The results suggest that arginine impairs energy metabolism in hippocampus slices of rats.     

Backflux of ammonia from brain to blood in human subjects with and without hepatic encephalopathy

In patients with hepatic encephalopathy (HE) the blood concentration of ammonia is usually highly elevated. Ammonia readily enters brain cells from the blood, and toxic effects of ammonia on brain metabolism and neurotransmission are believed to play a key role in the pathogenesis of HE. It has, however, been a matter of great controversy whether backflux of unmetabolized ammonia (NH₃ + NH₄ ⁺) from brain cells to the blood occurs in man. In the present analysis of data from a dynamic PET study of brain ¹³N–ammonia metabolism in healthy subjects and cirrhotic patients with and without HE, we provide the first unambiguous evidence for backflux of ammonia from brain cells to the blood in man. The high temporal and spatial resolution of modern PET technology was employed to distinguish between unidirectional blood-brain transport of ammonia and subsequent metabolism of the ammonia in the brain. In all 16 subjects, clearance of the unidirectional transport of ¹³N–ammonia from the blood to brain cells (K₁) was higher than the metabolic clearance of ¹³N–ammonia from the blood (K_met=K₁ k₃/(k₂+k₃). This can only be explained by backflux (k₂) of ammonia from brain cells to the blood. In conclusion, backflux of ammonia from the brain to the blood does indeed occur in both healthy subjects and cirrhotic patients with and without hepatic encephalopathy.     

A Sustainable Technique to Prepare High-Purity Vanadium Pentoxide via Purification with Low Ammonium Consumption

The general preparation method for V₂O₅ is ammonium salt vanadium precipitation, which inevitably produces large amounts of ammonia nitrogen wastewater. In this paper, we propose an environmentally friendly method for preparing high-purity V₂O₅ with low ammonium consumption. The purity of the V₂O₅ product reaches more than 99% while reducing the level of ammonium consumption. The vanadium precipitation efficiency reaches 99.23% and the V₂O₅ purity of the product reaches 99.05% under the following conditions: precipitation time of 1.5 h, precipitation temperature of 98 °C, initial precipitation pH of 2, ammonium addition coefficient of 2, purification time of 5 min with purification performed twice, purification temperature of 65 °C. In this study, compared with the use of ammonia spirit for vanadium precipitation and ammonium salt vanadium precipitation, the ammonia consumption levels are reduced by 79.80% and 80.00%, and the purity levels are increased by 0.70% and 1.01%, respectively. The compositions of the precipitated (NaV₃O₈∙xH₂O) and purified ((NH₄)₂V₆O₁₆·1.5H₂O) hydrolysis products are characterized via XRD. The TGA results show that NaV₃O₈∙xH₂O contains 1.5 times the amount of crystal water. The FTIR results explain that the two V₃O₈⁻ layers are combined end-to-end to form a V₆O₁₆²⁻ layer. The change of the product image indicates that the purification process includes three stages. Firstly, heating and NH₄⁺ attack expand the V₃O₈⁻ layer. NH4⁺ diffuses more easily into the V₃O₈⁻ layer. Secondly, NH₄⁺ destroys the electrostatic interaction between Na⁺ with the V₃O₈⁻ layer and replacing Na⁺. Finally, V₃O₈⁻ is polymerized into V₆O₁₆²⁻ to keep the crystal structure stable.     

Cell Volume Regulation During Hyperosmotic Shrinkage Is Mediated by Na+/K+-ATPase and Na+–K+–2Cl− Cotransporter in Necturus Gastrics Surface Epithelial Cells

Cell volume regulation was investigated in gastric surface epithelial cells during hypertonic conditions. Isolated Necturus antral mucosa was perfused on the serosal side with Ringer's solution (pH 7.25, 95%O₂/5%CO₂) and on the mucosal side successively with 150–500 mM NaCl. Amiloride, ouabain, and bumetanide were used to experimentally inhibit Na⁺/H⁺, Na⁺/K⁺ ATPase or Na⁺–K⁺–2Cl^– ion transporters. Intracellular sodium activity and cell volume changes were measured with liquid sensor microelectrodes. The increase in intracellular sodium activity caused by luminal hyperosmolar exposure was mainly due to cell shrinkage. Inhibition of Na⁺/K⁺ ATPase or Na⁺–K⁺–2Cl^– cotransporter increased hyperosmotic cell shrinkage (−52 ± 5%, −85 ± 19%, and −77 ± 9% for control, ouabain, and bumetanide, respectively). Inhibition of Na⁺/K⁺ ATPase increased intracellular sodium activity (from 18 ± 4 to 52 ± 12 mM). Cell volume regulation in gastric epithelial surface cells during mucosal hyperosmolar exposure is maintained by the basolateral Na⁺–K⁺–2Cl^– cotransporter, while Na⁺/K⁺ ATPase maintains sodium balance, but Na⁺/H⁺ antiport seems to have a less important role. This work was supported by the Research Foundation of Helsinki University Central Hospital (EVO), Academy of Finland, Antti and Jenny Wihuri Foundation, and Biomedicum Research Foundation, Helsinki, Finland. The authors thank Mrs Paula Kokko for excellent technical assistance.     

The Effect of Intragastric Ammonia Production on Titratable Gastric Acid Output in Helicobacter pylori-Infected Patients with Chronic Gastritis

The purpose of this study was to assess whether intragastric neutralization of HCl by ammonia in Helicobacter pylori-infected patients could meaningfully affect the titratable acid output as a measure of gastric acid secretion in a relation to the severity of infection. In 79 patients with different degrees of Helicobacter pylori infection and chronic gastritis, the basal acid output (BAO) and maximal acid output (MAO) after pentagastrin (6 μg/kg s.c.) was estimated. Cl⁻ and NH₄⁺ contents in these fractions were also assayed. H⁺/Cl⁻ ratio in the MAO fraction was diminished in markedly infected patients (68.1 ± 3.9%, vs 84.1 ± 3.3% in noninfected patients; P < 0.005). Ammonium content was maximal in patients with marked infection (0.912 ± 0.086 vs 0.149 ± 0.034 mmol/hr in MAO [P < 0.001] and 0.475 ± 0.063 vs 0.105 ± 0.016 mmol/hr in BAO of noninfected patients [P < 0.001]), with intermediate values in mild and moderate infection. The NH₄⁺/(H⁺ + NH₄⁺) ratio reached 27.01 ± 7.34% in the BAO of moderately infected patients, vs 10.22 ± 3.81% in noninfected patients (P = 0.05), and 7.25 ± 1.06% in the MAO of markedly infected patients, vs 1.14 ± 0.33% in noninfected patients (P < 0.001). The intragastric ammonia production affects the titratable acid output in Helicobacter pylori-infected patients dependent on the severity of infection. Therefore this factor should be taken into consideration in the evaluation of gastric secretory function in Helicobacter pylori-infected patients. This study was supported by Medical University of Bialystok Grant 514 928.     

Dofetilide Enhances the Contractility of Rat Ventricular Myocytes via Augmentation of Na+–Ca2+ Exchange

Purpose  Dofetilide (DOF), a novel Class III antiarrhythmic drug, prolongs the action potential duration (APD) and shows a positive inotropic effect in guinea pig papillary muscle. The present experiments were designed to study the positive inotropic effect of DOF on rat ventricle and explore its possible mechanism(s). Methods  Hearts from male Wistar rats (260–320 g) were divided into five groups and perfused in Langendorff mode. Ventricular myocytes were enzymatically isolated from male Wistar rats. Whole-cell voltage-clamping technique was used to test the Na⁺–Ca²⁺ exchange (NCE) current (I _NCX); Calcium transients and cell shortening provoked by field stimulation or using calcium current command waveform were observed synchronously with an ionic imaging system. Results  DOF (0.03–1.0 μM) increased left ventricular function in isolated rat hearts in a concentration-dependent manner. DOF (0.03–1.0 μM) also concentration-dependently increased both inward and outward I _NCX in isolated rat ventricular cells. The EC₅₀ values of DOF were 0.149 μM for the inward I _NCX and 0.249 μM for outward I _NCX, respectively. DOF 0.2 μM significantly enhanced Ca²⁺ transient and cell shortening in single rat ventricular myocytes driven by field electric stimulation. When the patch clamp system was connected to the ionic imaging system, Ca²⁺ current (I _Ca), Ca²⁺ transient and cell shortening amplitude in a same cell were recorded synchronously. Application of DOF 0.2 μM had no effect on I _Ca, but significantly increased Ca²⁺ transient and cell shortening. NCX inhibitor KB-R7943 0.6 μM significantly depressed the effects of DOF on Ca²⁺ transient and cell shortening. Conclusions  We conclude that DOF enhanced contractility of rat ventricular myocytes. The enhancement of NCE may be involved in the positive inotropic action of DOF.     

Androgen support of lacrimal gland function

The effects of dihydrotestosterone (DHT) (1 mg/kg) on biochemical parameters related to lacrimal secretion, basal tear flow rate, and pilocarpine-stimulated lacrimal gland fluid secretion, in mature ovariectomized rabbits were studied. The effects of the synthetic estrogen, diethylstilbestrol (DES) (100 μg/kg), on lacrimal gland biochemical parameters in normal mature female rabbits was also studied. Ovariectomy decreased the total serum levels of testosterone (T) by 88.5% and androstenedione by 35.9%, without changing the levels of dehydroepiandrosterone (DHEA) or its sulfate. Ovariectomy caused a significant regression of the lacrimal glands, decreasing total DNA by 35%, and total protein by 22%. DHT treatment of ovariectomized animals prevented lacrimal gland regression, increasing total gland DNA (31%) and total protein (18%). DHT treatment also increases Na⁺,K⁺-ATPase activity (29%) and ?-adrenergic receptor binding sites (23%) compared to the ovariectomized group. DHT increased pilocarpine stimulated lacrimal gland fluid secretion (13.26±1.47 μL/min) compared to the ovariectomized group (7.72±0.41 μL/min), but DHT treatment paradoxically decreased basal tear flow rate (1.02±0.04 μL/min) as compared to the ovariectomized rabbits (1.96±0.12 μL/min). DES decreased the total serum T from 59.33±10.54 pg/mL to 21.5±6.06 pg/mL. DES decreased total Na⁺,K⁺-ATPase by 12% and increased ?-adrenergic receptor binding sites by 83.3%. These results suggest that androgens play a major role in supporting lacrimal gland secretory function. Additionally, they suggest that estrogens may influence certain aspects of lacrimal functions, although it is not clear to what extent those actions are elicited directly or indirectly.     

Deleterious effects of digitalis on reperfusion-induced arrhythmias and myocardial injury in ischemic rat hearts: possible involvements of myocardial Na+ and Ca2+ imbalance

Summary Isolated rat hearts were made ischemic for 25 min after an initial recirculating perfusion, followed by 30 min of reperfusion. In some hearts, interventions including administration of ouabain and/or high [K⁺] in the buffer were performed during the first 10 min of reperfusion.During ischemia, intracellular Na⁺ (Na_i) increased from 15 to 64 [mol/g dry weight (dwt). During reperfusion, Na_i declined rapidly (at 10 min of reperfusion: 48 nol/g dwt, at 30 min: 25 mol/g dwt) and regular rhythm was recovered within 10 min in hearts without any intervention during reperfusion.⁴⁵Ca²⁺ uptake increased from 0.8 to 7.5 mol/g dwt after 30 min of reperfusion. Ventricular function recovered by 45 %.A 10-min perfusion with 10 or 50 M of ouabain increased Na_i (17 to 21 or 27 mol/g dwt) with increased left-ventricular (LV) contractile function, but these effects were reversed by combination of high perfusate [K⁺] (20 mM) in non-ischemic hearts.A 10-min reperfusion with ouabain retarded or stopped the decline in Na_i (at 10 min of reperfusion: 54 or 63 mol/g dwt, at 30 min: 32 or 40 mol/g dwt). These amounts of ouabain also increased the incidence of ventricular tachyarrhythmias during reperfusion to 30 % or 50 %, and increased the duration of ventricular fibrillation from 6.5 to 11.5 or 18.0 min.⁴⁵Ca²⁺ uptake reached to 8.8 or 10.0 mol/g dwt, and function recovered only 35 % or 28 %. When high perfusate [K⁺] was combined with ouabain during reperfusion, the retarded decline in Nai, augmented⁴⁵Ca²⁺ uptake, and reduced recovery of function caused by ouabain alone were attenuated. These results suggest that digitalis has toxic effects on reperfused ischemic hearts by inhibition of rapid active outward transport of previously elevated Na_i and potentiation of Ca²⁺ overload.The work was supported in part by grant HL 37936 from the National Heart, Lung and Blood Institute. J. R. Neely was deceased on November 29, 1988     

GDP-mannose pyrophosphorylase is a genetic determinant of ammonium sensitivity in Arabidopsis thaliana

Higher plant species differ widely in their growth responses to ammonium (NH₄⁺). However, the molecular genetic mechanisms underlying NH₄⁺ sensitivity in plants remain unknown. Here, we report that mutations in the Arabidopsis gene encoding GDP-mannose pyrophosphorylase (GMPase) essential for synthesizing GDP-mannose confer hypersensitivity to NH₄⁺. The in planta activities of WT and mutant GMPases all were inhibited by NH₄⁺, but the magnitude of the inhibition was significantly larger in the mutant. Despite the involvement of GDP-mannose in both l-ascorbic acid (AsA) and N-glycoprotein biosynthesis, defective protein glycosylation in the roots, rather than decreased AsA content, was linked to the hypersensitivity of GMPase mutants to NH₄⁺. We conclude that NH₄⁺ inhibits GMPase activity and that the level of GMPase activity regulates Arabidopsis sensitivity to NH₄⁺. Further analysis showed that defective N-glycosylation of proteins, unfolded protein response, and cell death in the roots are likely important downstream molecular events involved in the growth inhibition of Arabidopsis by NH₄⁺.     

Effects of hyper- and hypothyroidism on acetylcholinesterase, (Na<Superscript>+</Superscript>, K<Superscript>+</Superscript>)- and Mg <Superscript><Emphasis Type="Bold">2+</Emphasis></Superscript>-ATPase activities of adult rat hypothalamus and cerebellum

Thyroid hormones (THs) are recognized as key metabolic hormones, and the metabolic rate increases in hyperthyroidism, while it decreases in hypothyroidism. The aim of this work was to investigate how changes in metabolism induced by THs could affect the activities of acetylcholinesterase (AChE), (Na⁺, K⁺)- and Mg²⁺-ATPase in the hypothalamus and the cerebellum of adult rats. Hyperthyroidism was induced by subcutaneous administration of thyroxine (25μg/100 g body weight) once daily for 14 days, while hypothyroidism was induced by oral administration of propylthiouracil (0.05%) for 21 days. All enzyme activities were evaluated spectrophotometrically in the homogenated brain regions of 10 three-animal pools. Neither hyper-, nor hypothyroidism had any effect on the examined hypothalamic enzyme activities. In the cerebellum, hyperthyroidism provoked a significant decrease in both the AChE (−23%, p < 0.001) and the Na⁺, K⁺-ATPase activities (−26%, p < 0.001). Moreover, hypothyroidism had a similar effect on the examined enzyme activities: AChE (−17%, p < 0.001) and Na⁺, K⁺-ATPase (−27%, p < 0.001). Mg²⁺-ATPase activity was found unaltered in both the hyper- and the hypothyroid brain regions. In conclusion: neither hyper-, nor hypothyroidism had any effect on the examined hypothalamic enzyme activities. In the cerebellum, hyperthyroidism provoked a significant decrease in both the AChE and the Na⁺, K⁺-ATPase activities. The decreased (by the THs) Na⁺, K⁺-ATPase activities may increase the synaptic acetylcholine release, and thus, could result in a decrease in the cerebellar AChE activity. Moreover, the above TH-induced changes may affect the monoamine neurotransmitter systems.     

Effect of hypermethioninemia on some parameters of oxidative stress and on Na<Superscript>+</Superscript>,K<Superscript>+</Superscript>-ATPase activity in hippocampus of rats

In the present study we investigated the effect of chronic administration of methionine, a metabolite accumulated in many inherited pathological conditions such as methionine adenosyltransferase deficiency and homocystinuria, on some parameters of oxidative stress, namely thiobarbituric acid reactive substances (TBARS), catalase activity and total thiol content, as well as on Na⁺,K⁺-ATPase activity in rat hippocampus. For chronic treatment, rats received subcutaneous injections of methionine (1.34–2.68 μmol/g of body weight), twice a day, from the 6th to the 28th day of age and controls received saline. Animals were killed 12 h after the last injection. Results showed that chronic hypermethioninemia significantly increased TBARS, decreased Na⁺,K⁺-ATPase activity but did not alter catalase and total thiol content. Since chronic hypermethioninemia altered TBARS and Na⁺,K⁺-ATPase activity at 12 h after methionine administration, we also investigated the effect of acute administration of this amino acid on the same parameters studied after chronic methionine administration. For acute treatment,29-day-old rats received one single injection of methionine (2.68 μmol/g of body weight) or saline and were killed 1, 3 or 12 h later. Results showed that rats subjected to acute hypermethioninemia presented a reduction of Na⁺,K⁺-ATPase activity and an increase in TBARS when the animals were killed at 3 and 12 h, but not at 1 h, after methionine administration. These data indicate that hypermethioninemia increases lipid peroxidation which may, at least partially, explain the effect of methionine on the reduction in Na⁺,K⁺-ATPase activity. If confirmed in human beings, our findings could suggest that the induction of oxidative stress and the inhibition of Na⁺,K⁺-ATPase activity caused by methionine might contribute to the neurophysiopathology observed in patients with severe hypermethioninemia.     

Valproic acid induction of coma in rats: Synergism with NH 4 + and pentobarbital

Dose-response curves for the incidence of coma after intraperitoneal injections of various doses of valproic acid (VP) and octnoic acid (OA) showed that, mole for mole, valproic acid was less toxic than octanoic acid. However, a simultaneous subcoma dose of pentobarbital (PB) enhanced the toxicity of VP more than that of OA. The dose-response curve for NH₄C1 was affected by simultaneous subcoma doses of VP and OA but not by PB. VP enhanced the toxicity of the NH ₄ ⁺ by 52%; OA enhanced the toxicity by 12%. PB added significantly to the toxicity of VP and NH ₄ ⁺ when the three were given simultaneously. Doses of 0.7 mmol NH ₄ ⁺ and 0.5 mmol VP given separately had little or no encephalopathic effect, with blood ammonias of 250–1250g/dl. When given simultaneously they induced a deep coma and raised the blood ammonia threefold, to about 3600g/dl. Similar doses of OA and NH ₄ ⁺ , induced a similar deep coma, but blood levels of ammonia were not as high. Simultaneous injections of 250 mg glucose did not alter the results. Thus VP toxicity is enhanced substantially by its synergistic interactions with PB and the NH ₄ ⁺ .     

Identifying the direct effects of ammonia on the brain

Elevated concentrations of ammonia in the brain as a result of hyperammonemia leads to cerebral dysfunction involving a spectrum of neuropsychiatric and neurological symptoms (impaired memory, shortened attention span, sleep-wake inversions, brain edema, intracranial hypertension, seizures, ataxia and coma). Many studies have demonstrated ammonia as a major player involved in the neuropathophysiology associated with liver failure and inherited urea cycle enzyme disorders. Ammonia in solution is composed of a gas (NH₃) and an ionic (NH₄ ⁺) component which are both capable of crossing plasma membranes through diffusion, channels and transport mechanisms and as a result have a direct effect on pH. Furthermore, NH₄ ⁺ has similar properties as K⁺ and, therefore, competes with K⁺ on K⁺ transporters and channels resulting in a direct effect on membrane potential. Ammonia is also a product as well as a substrate for many different biochemical reactions and consequently, an increase in brain ammonia accompanies disturbances in cerebral metabolism. These direct effects of elevated ammonia concentrations on the brain will lead to a cascade of secondary effects and encephalopathy.

In vitro chemosensitivity of human pancreatic cancer cell lines

Summary Conclusion These results show that eight pancreatic cancer cell lines are broadly sensitive to CDDP, and that chemotherapy for pancreatic cancer may improve the prognosis by more effective drug delivery to cancer cells. Background Chemotherapy for pancreatic cancer does not satisfactorily improve prognosis. The efficacy of chemotherapy depends on choosing sensitive anticancer drugs. Methods The in vitro chemosensitivity of eight human pancreatic cancer cell lines was investigated. Growth inhibition was measured by³H-thymidine incorporation assays for doxorubicin hydrochloride (ADM), mitomycin C (MMC), cisplatin (CDDP), and etoposide (VP-16), and by Alamar Blue^TM assay for (AB assay) 5-fluorouracil (5-FU). The cells were exposed to ADM, MMC, CDDP, and VP-16 for 2h, and 5-FU for 72 h. From the dose-response curves, the 50% growth inhibition (IC₅₀) level for each drug was estimated. Results The IC₅₀ after 2 h of exposure of each of the eight kinds of cell lines to each anticancer drug ranged from 0.12–8.2 μg/mL for ADM, 0.066–25 μg/mL for MMC, 0.57–7 μg/mL for CDDP, 0.68–300 μg/mL for VP-16. IC₅₀ after 72 h of exposure to 5-FU ranged from 1.8–23 μg/mL.     

Changes in Antioxidant Status,Protein Concentration,Acetylcholinesterase, (Na<Superscript>+</Superscript>,K<Superscript>+</Superscript>)-, and Mg<Superscript>2+</Superscript>-ATPase Activities in the Brain of Hyper- and Hypothyroid Adult Rats

It is a common knowledge that metabolic reactions increase in hyperthyroidism and decrease in hypothyroidism. The aim of this work was to investigate how the metabolic reactions could affect the total antioxidant status (TAS), protein concentration (PC) and the activities of acetylcholinesterase (AChE), (Na⁺,K⁺)-ATPase and Mg²⁺-ATPase in the brain of hyper- and hypothyroid adult male rats. Hyperthyroidism was induced in rats by subcutaneous administration of thyroxine (25 g/100 g body weight) once daily for 14 days, while hypothyroidism was induced by oral administration of propylthiouracil (0.05%) for 21 days. TAS, PC, and enzyme activities were evaluated spectrophotometrically in the homogenated brain of each animal. TAS, PC, and Mg²⁺-ATPase activity were found unaffected in hyperthyroidism, while AChE and Na⁺,K⁺-ATPase activities were reduced by 25% (p < 0.01). In contrast, TAS, (Na⁺,K⁺)-ATPase and Mg²⁺-ATPase activities were found to be increased (approx. 23–30%, p < 0.001) in the hypothyroid brain, while AChE activity and PC were shown to be inhibited (approx. 23–30%, p < 0.001). These changes on brain enzyme activities may reflect the different metabolic effects of hyper- and hypothyroidism. Such changes of the enzyme activities may differentially modulate the brain intracellular Mg²⁺, neural excitability, as well as the uptake and release of biogenic amines.     

Castration reduces mRNA levels for calcium regulatory proteins in rat heart

Sex-related differences in the cardiac phenotype have been well established. This study was designed to determine whether androgens regulate myocardial gene expression and play a role in the sex-related differences in the myocardial phenotype. Gonadectomized male rats were treated with testosterone, and myocardial gene expression was examined in whole heart using quantitative real-time PCR. Gonadectomy produced a substantial decrease in mRNA levels for the androgen receptor, Na⁺/Ca²⁺ exchanger, L-type calcium channel, and β₁-adrenergic receptor (β₁AR). Supplementation of testosterone in castrates produced a fivefold increase in androgen receptor mRNA levels. Testosterone treatment of castrates produced almost a sixfold increase in Na⁺/Ca²⁺ exchanger mRNA, a tenfold increase in L-type calcium channel mRNA accumulation, and a fourfold increase in β₁AR mRNA levels. Increased calcium channel expression, β₁AR expression, and Na⁺/Ca²⁺ exchanger expression together may alter cytosolic calcium. These results provide the first evidence that testoster-one regulates expression of myocardial calcium regulating genes and thus may play a role in modulating the cardiac phenotype in males.     

Effect of phenylalanine and p-chlorophenylalanine on Na+, K+-ATPase activity in the synaptic plasma membrane from the cerebral cortex of rats

Na⁺, K⁺-ATPase activity was measured in synaptic plasma membrane from cerebral cortex of Wistar rats subjected to experimental phenylketonuria, i.e., chemical hyperphenylalaninemia induced by subcutaneous administration of 5.2 μmol phenylalanine /g body weight (twice a day) plus 0.9 μmol p-chlorophenylalanine /g body weight (once a day). The treatment was performed from the 6^th to the 14^th postpartum day and rats were killed 12 h after the last injection. Synaptic plasma membrane from cerebral cortex was prepared by a discontinuous density sucrose gradient for Na⁺, K⁺-ATPase activity determination. The results showed that the enzyme activity was decreased by 30% in animals subjected to experimental phenylketonuria when compared to control. Thein vitro effects of the drugs on Na⁺, K⁺-ATPase activity were also investigated. Phenylalanine and p-chlorophenylalanine inhibited the enzyme activity and this inhibition was reversed by alanine. In addition, competition between phenylalanine and p-chlorophenylalanine for binding to the enzyme was observed, suggesting a common binding site for these substances. Our results suggest that reduction of Na⁺, K⁺-ATPase activity may be one of the mechanisms related to the brain dysfunction observed in human PKU.     

Effects of short-term acid and aluminum exposure on the parr-smolt transformation in Atlantic salmon (Salmo salar): disruption of seawater tolerance and endocrine status

Episodic acidification resulting in increased acidity and inorganic aluminum (Al_i) is known to interfere with the parr-smolt transformation of Atlantic salmon (Salmo salar), and has been implicated as a possible cause of population decline. To determine the extent and mechanism(s) by which short-term acid/Al exposure compromises smolt development, Atlantic salmon smolts were exposed to either control (pH 6.7–6.9) or acid/Al (pH 5.4–6.3, 28–64 μg l⁻¹ Al_i) conditions for 2 and 5 days, and impacts on freshwater (FW) ion regulation, seawater (SW) tolerance, plasma hormone levels and stress response were examined. Gill Al concentrations were elevated in all smolts exposed to acid/Al relative to controls confirming exposure to increased Al_i. There was no effect of acid/Al on plasma ion concentrations in FW however, smolts exposed to acid/Al followed by a 24 h SW challenge exhibited greater plasma Cl⁻ levels than controls, indicating reduced SW tolerance. Loss of SW tolerance was accompanied by reductions in gill Na⁺,K⁺-ATPase (NKA) activity and Na⁺,K⁺,2Cl⁻ (NKCC) cotransporter protein abundance. Acid/Al exposure resulted in decreased plasma insulin-like growth factor (IGF-I) and 3,3′,5′-triiodo-l-thyronine (T₃) levels, whereas no effect of treatment was seen on plasma cortisol, growth hormone (GH), or thyroxine (T₄) levels. Acid/Al exposure resulted in increased hematocrit and plasma glucose levels in FW, but both returned to control levels after 24 h in SW. The results indicate that smolt development and SW tolerance are compromised by short-term exposure to acid/Al in the absence of detectable impacts on FW ion regulation. Loss of SW tolerance during short-term acid/Al exposure likely results from reductions in gill NKA and NKCC, possibly mediated by decreases in plasma IGF-I and T₃.     

Hydrogen peroxide-induced vascular relaxation in porcine coronary arteries is mediated by Ca2+-activated K+ channels

Summary Hydrogen peroxide (H₂O₂) elicited concentration-dependent relaxation of endothelium-denuded rings of porcine coronary arteries. The relaxation induced by the H₂O₂ was markedly attenuated by 10μM 1H-[1,2,4]oxadiazolo [4,3,a]quinoxalin-1-one (ODQ), an inhibitor of soluble guanylate cyclase, or by 100nM charybdotoxin, an inhibitor of large-conductance Ca²⁺-activated K⁺ (K_Ca) channels. A combination of the ODQ and charybdotoxin abolished the H₂O₂-induced relaxation. Pretreatment with 25 μM of an Rp stereoisomer of adenosine-3′,5′-cyclic monophosphothioate (Rp-cAMPS), 20μM glibenclamide, or 1mM 4-aminopyridine did not affect the vascular response to H₂O₂. The presence of catalase at 1000U/ml significantly attenuated the H₂O₂-induced relaxation. Exposure of cultured smooth muscle cells to H₂O₂ activated K_Ca channels in a concentration-dependent manner in cell-attached patches. Pretreatment with catalase significantly inhibited the activation of K_Ca channels. Rp-cAMPS did not inhibit the H₂O₂-induced activation of K_Ca channels. The activation of K_Ca channels by H₂O₂ was markedly decreased in the presence of ODQ. However, even in the presence of ODQ, H₂O₂ activated K_Ca channels in a concentration-dependent manner. In inside-out patches, H₂O₂ significantly activated K_Ca channels through a process independent of cyclic guanosine 3′,5′-monophosphate (cGMP). In conclusion, H₂O₂ elicits vascular relaxation due to activation of K_Ca channels, which is mediated partly by a direct action on the channel and partly by activation of soluble guanylate cyclase, resulting in the generation of cGMP.     

Sodium salicylate has no effect on cerebrospinal fluid [H+] in dogs with normal acid-base balance

The experiments described here were designed to investigate the possibility that central stimulation of respiration by salicylates may be due to changes in [H⁺] of cerebral fluids. Two groups (n=6 in each) of anesthetized, paralyzed, and mechanically ventilated dogs were studied for 6 hr. Renal pedicles were ligated to maintain blood salicylate level constant. Group II received 150 mg/kg Na salicylate intravenously at 0 hr after samples had been obtained. Group I (control) received equal volume of half-normal saline. Mean plasma salicylate levels were 18.9, 18.4, and 19.6 mg % at 0.5, 3, and 6 hr after administration of Na salicylate. Respective cisternal cerebrospinal fluid (CSF) levels were 3.2, 4.8, and 5.9 mg %. Salicylate-induced hyperthermia was prevented by peritoneal cold dialysis, and a rise in PaCO₂ was prevented by increasing ventilation. During the 6 hr of relatively normal systemic acid-base balance, cisternal CSF mean PCO₂ values were 45.3, 43.6, and 49.3 mm Hg at 0, 3, and 6 hr in the control group; in group II, respective values were 46.9, 45.7, and 47.7 mm Hg. Cisternal CSF [H⁺] were 44.4, 45.2, and 50.5 nEq/L in group I at 0, 3, and 6 hr. Respective values in group II were 45.0, 47.5, and 50.6 nEq/L. These values were similar and statistically insignificant from those in group I. In both groups cisternal CSF [HCO ₃ ⁻ ] fell about 2 and CSF lactate concentration rose about 1 mEq/L at 6 hr. We conclude that at the dose used in the present study, Na salicylate increases metabolic rate but does not significantly influence cisternal CSF [H⁺] up to 6 hr in dogs with normal systemic acid-base balance.