Evidence of nitric oxide and angiotensin II regulation of circulation and cutaneous drinking in Bufo marinus

The nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester (l-NAME) increased vascular resistance (VR) 10% above baseline of 3.08+/-0.08 (n=11) mmHg/mL/min at 10 mg/kg and 20% above 3.05+/-0.08 (n=9) at 50 mg/kg in anesthetized toads (Bufo marinus). Blood pressure was unaffected by either dose of L-NAME. Blood flow decreased at the higher dose of L-NAME. L-arginine (300 mg/kg) reversed the effects of L-NAME on VR and blood flow in toads treated with 10 mg/kg but not with 50 mg/kg. Injection of 50 mg/kg L-NAME into empty-bladder toads produced a 10% decrease in water uptake, J(v), resulting in a J(v) of 1,267+/-11 cm(3)/cm(2)/s x 10(-7) (n=9) compared to 1,385+/-12 (n=8) for controls. Injection of 10 microg/kg angiotensin II (ANG II) increased J(v) 15% across the pelvic patch (J(v), cm(3)/cm(2)/s x 10(-7)), resulting in a J(v) of 1,723+/-12 cm(3)/cm(2)/s x 10(-7) (n=8) compared to 1,471+/-12 (n=8) for controls. It is hypothesized that during cutaneous drinking blood flow into the capillary bed of the pelvic patch is regulated by nitric oxide and ANG II.     

Angiotensin II alters blood flow distribution in amphibians.

In toads, angiotensin II (ANG II) induces the water absorption response (WR) during which the seat patch (pelvic+inner-thigh skin) is pressed to a wet substrate from which water flows osmotically into the animal. Since ANG II is a potent vasoconstrictor, it has the potential to redistribute blood flow. To determine the regional circulatory effects of ANG II, we used microsphere methods to measure relative changes in blood flow to several skin regions and other organs before and after ANG II administration in terrestrial toads and aquatic bullfrogs. In toads, after ANG II administration, seat patch and bladder blood flow increased by 264.2%+/-197.6% and 287.2%+/-86.7%, respectively (P<0.05), while dorsal and pectoral skin flow decreased by 48.0%+/-19.4% and 21.3%+/-25.4%, respectively (P<0.05). In bullfrogs, ANG II caused no significant changes in blood flow. Our results support our hypothesis that, in toads, ANG II increases and decreases blood flow to regions of the body associated with water gain and water loss, respectively.     

Angiotensin II stimulates nitric oxide production in pulmonary artery endothelium via the type 2 receptor

We previously reported that angiotensin II stimulates an increase in nitric oxide production in pulmonary artery endothelial cells. The aims of this study were to determine which receptor subtype mediates the angiotensin II-dependent increase in nitric oxide production and to investigate the roles of the angiotensin type 1 and type 2 receptors in modulating angiotensin II-dependent vasoconstriction in pulmonary arteries. Pulmonary artery endothelial cells express both angiotensin II type 1 and type 2 receptors as assessed by RT-PCR, Western blot analysis, and flow cytometry. Treatment of the endothelial cells with PD-123319, a type 2 receptor antagonist, prevented the angiotensin II-dependent increase in nitric oxide synthase mRNA, protein levels, and nitric oxide production. In contrast, the type 1 receptor antagonist losartan enhanced nitric oxide synthase mRNA levels, protein expression, and nitric oxide production. Pretreatment of the endothelial cells with either PD-123319 or an anti-angiotensin II antibody prevented this losartan enhancement of nitric oxide production. Angiotensin II-dependent enhanced hypoxic contractions in pulmonary arteries were blocked by the type 1 receptor antagonist candesartan; however, PD-123319 enhanced hypoxic contractions in angiotensin II-treated endothelium-intact vessels. These data demonstrate that angiotensin II stimulates an increase in nitric oxide synthase mRNA, protein expression, and nitric oxide production via the type 2 receptor, whereas signaling via the type 1 receptor negatively regulates nitric oxide production in the pulmonary endothelium. This endothelial, type 2 receptor-dependent increase in nitric oxide may serve to counterbalance the angiotensin II-dependent vasoconstriction in smooth muscle cells, ultimately regulating pulmonary vascular tone.     

Acidification and sodium entry in frog skin epithelium

Acidification of the external medium by isolated frog skin epithelium (Rana catesbeiana, Rana temporaria, and Caudiververa caudiververa) and its relationship to Na⁺ uptake was studied. Acidification was measured by the pH-stat technique under short-circuit or open-circuit conditions. The results of this study demonstrate that (a) acidification by these species of in vitro frog skins is not directly coupled to Na⁺ or anion transport; (b) acidification can be inhibited by the diuretic drug amiloride, but only at high external Na⁺ concentrations; (c) acidification rate in these species of frog skin is controlled in part by the metabolic production of CO₂; and (d) the positive correlation between net Na⁺ absorption and net acidification observed in whole animal studies could not be replicated in the in vitro skin preparation, even when the frogs were first chronically stressed by salt depletion, a physiological state comparable to that used in the in vivo experiments.     

Angiotensin II and nitric oxide in neural control of intrarenal blood flow

We investigated the roles of the renin-angiotensin system and the significance of interactions between angiotensin II and nitric oxide, in responses of regional kidney perfusion to electrical renal nerve stimulation (RNS) in pentobarbital sodium-anesthetized rabbits. Under control conditions, RNS (0.5-8 Hz) reduced total renal blood flow (RBF; -89 +/- 3% at 8 Hz) and cortical perfusion (CBF; -90 +/- 2% at 8 Hz) more than medullary perfusion (MBF; -55 +/- 5% at 8 Hz). Angiotensin II type 1 (AT(1))-receptor antagonism (candesartan) blunted RNS-induced reductions in RBF (P = 0.03), CBF (P = 0.007), and MBF (P = 0.04), particularly at 4 and 8 Hz. Nitric oxide synthase inhibition with N(G)-nitro-L-arginine (L-NNA) enhanced RBF (P = 0.003), CBF (P = 0.001), and MBF (P = 0.03) responses to RNS, particularly at frequencies of 2 Hz and less. After candesartan pretreatment, L-NNA significantly enhanced RNS-induced reductions in RBF (P = 0.04) and CBF (P = 0.007) but not MBF (P = 0.66). Renal arterial infusion of angiotensin II (5 ng.kg(-1).min(-1)) selectively enhanced responses of MBF to RNS in L-NNA-pretreated but not in vehicle-pretreated rabbits. In contrast, greater doses of angiotensin II (5-15 ng.kg(-1).min(-1)) blunted responses of MBF to RNS in rabbits with intact nitric oxide synthase. These results suggest that endogenous angiotensin II enhances, whereas nitric oxide blunts, neurally mediated vasoconstriction in the renal cortical and medullary circulations. In the renal medulla, but not the cortex, angiotensin II also appears to be able to blunt neurally mediated vasoconstriction.     

Angiotensin II induces oxidative stress in prostate cancer

Angiotensin II has been shown to be a cytokine especially acting as a growth factor. A local renin-angiotensin system has been identified in the prostate gland, and the physiologic function of angiotensin II seems to be similar in prostate cancer, as we previously reported. In the present study, we explored the biological role of angiotensin II in oxidative stress of prostate cancer cells. Activated Akt was determined, and the expression of oxidative stress-related proteins (p47phox, manganese superoxide dismutase 2, glutathione peroxidase) was examined by Western blotting in LNCaP cells, which were stimulated with angiotensin II and/or an angiotensin II receptor type 1 blocker, candesartan. To examine DNA damage induced by angiotensin II, 8-hydroxy-2'-deoxyguanosine was determined, and Western blots were analyzed to detect checkpoint proteins including p53, Chk2, and cdc2. Immunocytochemical studies of inducible nitric oxide synthase and superoxide anion radical (O(2)(-)) were done in LNCaP cells stimulated with angiotensin II. The phosphorylation of Akt was induced by angiotensin II treatment and inhibited by candesartan, as well as by LY294002, an inhibitor of phosphoinositide 3-kinase. Oxidative stress-related proteins were up-regulated by angiotensin II and inhibited by pretreatment with candesartan or catalase. The level of 8-hydroxy-2'-deoxyguanosine was increased by angiotensin II and conversely decreased by candesartan. Immunocytochemical studies showed that angiotensin II enhanced an inflammatory marker, inducible nitric oxide synthase, and the production of O(2)(-) radical. The hypothesis that angiotensin II has the potential to induce oxidative stress, which may be implicated in carcinogenesis of the prostate gland through long-term exposure to chronic inflammation is proposed.     

Galactosamine decreases nitric oxide formation in cultured rat hepatocytes: mechanism of suppression

We have shown that nitric oxide production is dramatically decreased in rat primary hepatocyte cultures exposed to galactosamine. Cotreatment of the cells with uridine, which is known to prevent cytotoxicity, was found to also attenuate NO loss. In the present study, two possible mechanisms for the decreased nitric oxide production were examined. First, we examined the possibility that galactosamine could interfere with the uptake of extracellular arginine by the cultured hepatocytes. Cellular uptake of arginine was determined after addition of 14C-arginine at the time of hepatocyte attachment. Uptake of arginine was rapid in control cultures, and both the rate and level of uptake were unchanged by the addition of a cytotoxic concentration of galactosamine (4 mM). In addition, increased concentrations of arginine in the cell culture medium did not ameliorate the galactosamine-induced decrease in production of nitric oxide. Second, we determined whether the synthesis of inducible nitric oxide synthase in the hepatocyte cultures was inhibited by addition of galactosamine. Hepatocyte levels of inducible nitric oxide synthase were determined immunochemically at various times after the addition of galactosamine (4 mM). In control cultures, inducible nitric oxide synthase was detectable at 7 and 24 hours after attachment. In contrast, no nitric oxide synthase protein was detectable at any time in the galactosamine-treated cultures. Furthermore, addition of galactosamine after inducible nitric oxide synthase had already been synthesized (6.5 h after attachment) did not result in suppression of nitric oxide production in the hepatocyte cultures. The present studies suggest that galactosamine suppresses nitric oxide production in hepatocyte cultures by inhibiting synthesis of inducible nitric oxide synthase, rather than by interference in cellular uptake of arginine.     

Presence and activity of nitric oxide synthase isoforms in ischemia-reperfusion-injured flaps

Nitric oxide is produced from the amino acid L-arginine by nitric oxide synthase, which has three known isoforms: (1) endothelial nitric oxide synthase and (2) brain nitric oxide synthase, both of which are constitutive nitric oxide synthase; and (3) inducible nitric oxide synthase. The authors' hypothesis is that after reperfusion injury, endothelial cell dysfunction leads to disruption of nitric oxide synthase-mediated nitric oxide production and that this may in part explain the deleterious effects of ischemia-reperfusion injury on tissue survival and blood reflow in flaps. An experiment was designed to study the effects of ischemia-reperfusion injury on the bioactivity of all three isoforms of nitric oxide synthase. Buttock skin flaps and latissimus dorsi myocutaneous flaps were elevated in eight pigs. Flaps on one side of the animal were randomized to receive 6 hours of arterial ischemia, whereas flaps on the other side served as controls. At 6 hours of ischemia and at 1, 4, and 18 hours after reflow, tissue biopsy specimens were obtained and were processed for both constitutive nitric oxide synthase and inducible nitric oxide synthase enzyme activity on the basis of the L-citrulline assay. In addition, specimens were processed for Western blot analysis of the three isoforms. The authors' results revealed three key findings: first, there was a statistically significant (p < 0.001) decrease in constitutive nitric oxide synthase activity of ischemia-reperfusion-injured flaps as compared with controls in both skin and muscle for all time intervals measured. Second, Western blot analyses of endothelial nitric oxide synthase and brain nitric oxide synthase showed a significant decrease in the signal intensity in ischemic and reperfused tissue as compared with controls. Third, the inducible nitric oxide synthase isoform's activity and protein remained undetectable in both tissue types for all time points measured. The authors' data demonstrated that following ischemia-reperfusion injury in the pig flap model there was a disruption of constitutive nitric oxide synthase expression and activity, which may lead to decreased nitric oxide production. The significant decrease in nitric oxide synthase activity found in the current study may partly explain the mechanism of tissue damage in flaps subjected to ischemia-reperfusion injury. Knowledge of the kinetics of nitric oxide synthase activity under conditions of ischemia-reperfusion injury has important implications for the choice and timing of delivery of therapeutic agents whose goal is to increase the bioavailability of nitric oxide in reperfused tissue.     

NO and cGMP mediate angiotensin AT2 receptor-induced renal renin inhibition in young rats

We hypothesized that angiotensin subtype-2 receptor (AT(2)R) inhibits renal renin biosynthesis in young rats via nitric oxide (NO). We monitored changes in renal NO, cGMP, renal renin content (RRC), and ANG II in 4-wk-old rats in response to low sodium (LNa(+)) intake alone and combined with 8-h direct renal cortical administration of AT(1) receptor blocker valsartan (VAL), AT(2)R blocker PD123319 (PD), NO synthase inhibitor N(G)-nitro-l-arginine methyl ester (l-NAME), NO donor S-nitroso-N-acetyl penicillamine (SNAP), or guanylyl cyclase inhibitor 1H-[1,2,4] oxadiazolo[4,2-alpha] quinoxaline-1-one (ODQ). In addition, we monitored renal endothelial nitric oxide synthase (eNOS) and neuronal nitric oxide synthase (nNOS) in response to VAL or PD. LNa(+), VAL, PD, l-NAME, and ODQ increased RRC, ANG II, and renin mRNA. PD and l-NAME decreased NO and cGMP, while SNAP reduced RRC, ANG II, renin mRNA, and reversed the effects of PD. PD also reduced eNOS and nNOS protein and mRNA. Combined treatment with PD, l-NAME, or ODQ and VAL reversed the effects of VAL and caused further increase in RRC, ANG II, renin mRNA, and protein. ODQ reversed the effects of SNAP. These data demonstrate that the renal AT(2) receptor decreases renal renin biosynthesis and ANG II production in young rats. Reversal of the PD effects by SNAP and SNAP effects by ODQ confirms that NO and cGMP mediate the AT(2) receptor inhibition of renal renin production.     

Water potential receptors in the skin regulate blood perfusion in the ventral pelvic patch of toads

Blood cell flux (BCF) in ventral pelvic skin capillaries was measured in conscious unrestrained Bufo bufo, using a laser Doppler flowcytometer. Hydrated toads responded to water contact with a small but significant increase in BCF. Dehydration alone did not change the BCF in seat patch skin before water contact. However, water contact by dehydrated toads elicited a rapid 600% increase in BCF. The BCF and water uptake of dehydrated toads rehydrating in water declined over 2 h but remained significantly above the low, constant values measured in hydrated toads. Arginine vasotocin injection in hydrated toads did not change skin BCF, but water uptake increased, and urine production decreased. Injection of the beta -adrenergic agonist isoproterenol increased BCF in hydrated toads by 900% and also increased the rate of water uptake. These increases corresponded in magnitude and duration to the response to water contact observed in dehydrated toads. Injection of dehydrated toads with the beta -adrenergic antagonist propranolol significantly reduced both BCF and water uptake. These results are consistent with an autonomic reflex mediated by skin water potential receptors that regulate blood perfusion of ventral pelvic skin.     

牛蛙致病变形菌的鉴定及其敏感药物筛选

【目的】分离鉴定引起牛蛙(Rana catesbeiana)皮肤溃疡病症的致病菌,筛选抗菌药物。【方法】采用无菌解剖组织划线分离法分离致病菌,通过人工感染试验、菌体和菌落形态观察、API20E系统鉴定、16S r RNA基因序列分析,确定分离菌的致病性及分类地位,并对该菌进行药物敏感性试验。【结果】从患病濒死牛蛙体内分离细菌NWG20141026,通过形态特征观察、生理生化试验、API 20E系统鉴定和16S r RNA基因序列相似性分析,认为NWG20141026菌株为普通变形菌(Proteus vulgaris)。人工感染试验显示,NWG20141026菌株不仅可以通过皮肤伤口感染引起蛙体表溃疡溃烂病症,也可通过消化道感染引起蛙肠炎病。药敏实验结果表明,氟苯尼考、四环素、多西环素、萘啶酸、磺胺异噁唑、恩诺沙星、红霉素等7种药物对普通变形菌具有较好的抑杀菌作用。【结论】引起牛蛙(R.catesbeiana)皮肤溃疡病症的致病菌NWG20141026为普通变形菌(P.vulgaris),所患疾病命名为牛蛙变形菌病。普通变形菌对健康牛蛙具有较强致病性,氟苯尼考、四环素、多西环素、萘啶酸、磺胺异噁唑等5种药可作为防治牛蛙变形菌病的候选药物。     

Histone deacetylase inhibitor LMK235 attenuates vascular constriction and aortic remodelling in hypertension

Here, we report that LMK235, a class I and histone deacetylase (HDAC6)‐preferential HDAC inhibitor, reduces hypertension via inhibition of vascular contraction and vessel hypertrophy. Angiotensin II‐infusion mice and spontaneously hypertensive rats (SHRs) were used to test the anti‐hypertensive effect of LMK235. Daily injection of LMK235 lowered angiotensin II‐induced systolic blood pressure (BP). A reduction in systolic BP in SHRs was observed on the second day when SHRs were treated with 3 mg/kg LMK235 every 3 days. However, LMK235 treatment did not affect angiotensin‐converting enzyme 1 and angiotensin II receptor mRNA expression in either hypertensive model. LMK235, acting via the nitric oxide pathway, facilitated the relaxing of vascular contractions induced by a thromboxane A2 agonist in the rat aortic and mesenteric artery ring test. In addition, LMK235 increased nitric oxide production in HUVECs and inhibited the increasing of aortic wall thickness in both animal hypertensive models. LMK235 decreased the enhanced cell cycle‐related genes cyclin D1 and E2F3 in angiotensin II‐infusion mice and restored the decreased p21 expression. In addition, LMK235 suppressed calcium calmodulin‐dependent protein kinase II (CaMKII) α, which is related to vascular smooth muscle cell proliferation. Inhibition or knockdown of HDAC5 blocked the CaMKIIα‐induced cell cycle gene expression. Immunoprecipitation demonstrated that class I HDACs were involved in the inhibition of CaMKII α‐induced HDAC4/5 by LMK235. We suggest that LMK235 should be further investigated for its use in the development of new therapeutic options to treat hypertension via reducing vascular hyperplasia or vasoconstriction.     

Comparison of amphibian whole-cell and cell-free activity towards different transferrins.

1. A cell-free preparation from immature-erythrocytes of the bullfrog (Rana catesbeiana) was about equally active towards iron bound to homologous, rabbit or human transferrin. 2. In contrast with the cell-free system, whole immature erythrocytes exhibited optimum, intermediate or low uptake of iron from the above and from other transferrins.     

Role of superoxide and thromboxane receptors in acute angiotensin II-induced vasoconstriction of rabbit vessels

This study explored the hypothesis that a portion of angiotensin II-induced contractions is dependent on superoxide generation and release of a previously unidentified arachidonic acid metabolite that activates vascular smooth muscle thromboxane receptors. Treatment of rabbit aorta or mesentery artery with the thromboxane receptor antagonist SQ29548 (10 μM) reduced angiotensin II-induced contractions (maximal contraction in aorta; control vs. SQ29548: 134 ± 16 vs. 93 ± 10%). A subset of rabbits deficient in vascular thromboxane receptors also displayed decreased contractions to angiotensin II. The superoxide dismutase mimetic Tiron (30 mM) attenuated angiotensin II-induced contractions only in rabbits with functional vascular thromboxane receptors (maximal contraction in aorta; control vs. Tiron: 105 ± 5 vs. 69 ± 11%). Removal of the endothelium or treatment with a nitric oxide synthase inhibitor, nitro-l-arginine (30 μM) did not alter angiotensin II-induced contractions. Tiron and SQ29548 decreased angiotensin II-induced contractions in the denuded aortas by a similar percentage as that observed in intact vessels. The cyclooxygenase inhibitor indomethacin (10 μM) or thromboxane synthase inhibitor dazoxiben (10 μM) had no effect on angiotensin II-induced contractions indicating that the vasoconstrictor was not thromboxane. Angiotensin II increased the formation of a 15-series isoprostane. Isoprostanes are free radical-derived products of arachidonic acid. The unidentified isoprostane increased when vessels were incubated with the superoxide-generating system xanthine/xanthine oxidase. Pretreatment of rabbit aorta with the isoprostane isolated from aortic incubations enhanced angiotensin II-induced contractions. Results suggest the factor activating thromboxane receptors and contributing to angiotensin II vasoconstriction involves the superoxide-mediated generation of a 15-series isoprostane.     

Evaluation of nitric oxide synthase activity and inhibition kinetics by chemiluminescence.

The binding affinity (K(I)) and inactivation rate (k(inact)) parameters of nitric oxide synthase (NOS) inhibitors are typically estimated by kinetic activity studies. Methods currently used in the estimation of these parameters frequently employ radiolabeled materials and require intensive sample preparation. We have devised a simple, reproducible, and sensitive method for the kinetic analysis of NOS activity and inhibition kinetics using chemiluminescence. We have used this method to characterize enzyme activity for purified murine macrophage nitric oxide synthase (NOS II). Using this method, we have also estimated the inhibitory parameters for a series of competitive antagonists and mechanism-based inactivators of NOS II. The estimated parameters are in agreement with those reported using other methods. We conclude that the chemiluminescence method can be used for kinetic studies of NOS activity and inhibition. This method represents a more efficient means for conducting kinetic studies of NOS inhibition.     

The potential role of nitric oxide in the hypertrophic growth of the left ventricle

Left ventricular hypertrophy (LVH) is the result of interaction between a chronic hemodynamic overload and non-hemodynamic factors. There are several lines of evidence presented in this work suggesting that nitric oxide (NO) may participate in the hypertrophic growth of the myocardium. First, endothelial NO production was shown to be decreased in several types of hemodynamically overloaded circulation both in animals and humans. Second, compounds stimulating NO production were able to diminish the extent or modify the nature of LVH in some models of myocardial hypertrophic growth. Third, arterial hypertension can be induced by inhibition of nitric oxide synthase activity. This NO-deficient hypertension is associated with the development of concentric LVH, myocardial fibrosis and protein remodeling of the left ventricle. The mechanism of LVH development in NO-deficient hypertension is complex and involves decreased NO production and increased activation of the renin-angiotensin-aldosterone system. Cardiovascular protection via ACE inhibition in NO-deficient hypertension may be induced by mechanisms not involving an improvement of NO production. In conclusion, the hypertrophic growth of the LV appears to be the result of interaction of vasoconstrictive and growth stimulating effects of angiotensin II on the one hand and of vasodilating and antiproliferative effects of nitric oxide on the other.     

Modulation of brain mitochondrial function by deprenyl

The present study shows that deprenyl, a known inhibitor of monoamine oxidase B (MAO B), may generate changes in mitochondrial function. Brain submitochondrial membranes (SMP), synaptosomes and cytosolic fractions were incubated with different deprenyl concentrations and nitric oxide synthase (NOS) activity was measured. The effect of deprenyl on oxygen consumption, calcium-induced permeability transition and hydrogen peroxide (H(2)O(2)) production rates was studied in intact mitochondria. Respiratory complexes and monoamine oxidase activities were also measured in submitochondrial membranes. Incubation of brain submitochondrial membranes with deprenyl 10, 25 and 50 microM inhibited nitric oxide synthase activity in a concentration-dependent manner. The same effect was observed in cytosolic fractions and synaptosomes. Monoamine oxidase activity was inhibited at lower deprenyl concentrations (from 0.5 microM). Cytochrome oxidase (complex IV) activity was found 42% increased in the presence of 25 microM deprenyl in a condition of maximal nitric oxide synthase activity. Incubation of brain mitochondria with deprenyl 25 microM produced a 60% increase in oxygen uptake in state 3, but no significant changes were observed in state 4. Pre-incubation of brain mitochondria with deprenyl 0.5 and 1 microM inhibited calcium-induced mitochondrial permeability transition and decreased hydrogen peroxide production rates. Our results suggest that in vitro effects of deprenyl on mitochondrial function can occur through two different mechanisms, involving nitric oxide synthase inhibition and decreased hydrogen peroxide production.     

Indomethacin prevents the induction of inducible nitric oxide synthase in murine peritoneal macrophages and decreases their nitric oxide production

Indomethacin (0.14-.5 mM concentration) inhibits nitric oxide production in murine peritoneal macrophages. This was evidenced by measuring both nitrite production or 14C-L-citrulline formation. The inhibition was caused by the diminution of de novo inducible nitric oxide synthase production as demonstrated by Western blotting experiment. The effect of indomethacin after 4 h treatment was irreversible. NO synthase and arginase activities and the uptake of arginine were not directly affected by the drug. Indomethacin also decreased uridine incorporation in macrophages. The effect of indomethacin on the induction of other enzymes (i.e. arginase) was weaker.     

Interrelationships among epidermal Na-K ATPase, developmental stage and length of Rana catesbeiana tadpoles

Sodium and potassium activated adenosine triphosphatase (Na-K ATPase) was extracted from the skin of Rana catesbeiana tadpoles and adults. Using carefully staged tadpoles, it was noted that the enzyme level increases two or three stages before there is a perceptible potential difference generated across the skin. The level of non-ouabain-inhibitable ATPase remains constant throughout the life cycle. It was also concluded that Rana catesbeiana tadpoles cannot be reliably staged using length as a key trait.     

Distinct effects of N-acetylcysteine and nitric oxide on angiotensin II-induced epidermal growth factor receptor phosphorylation and intracellular Ca(2+) levels

These studies describe inhibitory effects of N-acetylcysteine on several biochemical events associated with the activation of extracellular signal-regulated kinases (ERK) by angiotensin II in the cardiac fibroblast and compare these effects with those of the nitric oxide donor, S-nitroso-N-acetylpenicillamine, an agent we showed previously to inhibit angiotensin II-induced ERK activation and the concomitant phosphorylation of proline-rich tyrosine kinase 2 (Wang, D., Yu, X., and Brecher, P. (1999) J. Biol. Chem. 274, 24342-24348). The transactivation of the epidermal growth factor receptor by angiotensin II, a process required for the activation of ERK, was inhibited by N-acetylcysteine but not by nitric oxide. The transactivation of the epidermal growth factor receptor by angiotensin II was shown to be independent of intracellular calcium increases. Nitric oxide, but not N-acetylcysteine, inhibited the angiotensin II-induced increase in intracellular Ca(2+). Neither nitric oxide nor N-acetylcysteine inhibited either phospholipase C activation or inositol triphosphate generation in response to angiotensin II. N-Acetylcysteine did inhibit the phosphorylation of the calcium sensitive tyrosine kinases PYK2 and Src, effects that also occurred using nitric oxide. These studies describe a novel effect of N-acetylcysteine on cross-talk between a G protein-linked receptor and a tyrosine kinase receptor and offer additional molecular insight to explain how N-acetylcysteine and nitric oxide act at different sites and might have an additive effect on specific hormonal responses.