Site of action of endogenous nitric oxide on pulmonary vasculature in rats

The effect of endogenous nitric oxide (NO) on the pulmonary hypoxic vasoconstriction was studied in isolated and blood perfused rat lungs. By applying the occlusion technique we partitioned the total pulmonary vascular resistance (PVR) into four segments: (1) large arteries (R _a), (2) small arteries (R _a′), (3) small veins (R _v′), and (4) large veins (R _v). The resistances were evaluated under baseline (BL) conditions and during; hypoxic vasoconstriction and acetylcholine (Ach) which was injected during hypoxic vasoconstriction. After recovery from hypoxia and Ach, N ^ω-nitro-L-arginine (L-NA) was added to the reservoir and the responses to hypoxia and Ach were reevaluated. Before L-NA, hypoxia caused significant increase in the resistances of all segments (P < 0.05), with the largest being in R _a and R _a′. Ach-induced relaxation during hypoxia occurred in R _a, R _a′ and R _v′ (P < 0.05). L-NA did not change the basal tone of the pulmonary vasculature significantly. However, after L-NA, hypoxic vasoconstriction was markedly enhanced in R _a, R _a′, and R _v′ (P < 0.01) compared with the hypoxic response before L-NA. Ach-induced relaxation was abolished after L-NA. We conclude that, in rat lungs, inhibition of NO production during hypoxia enhances the response in the small arteries and veins as well as in the large arteries. The results suggest that hypoxic vasoconstriction in the large pulmonary arteries and small vessels is attenuated by NO release. Received: 22 May 1995/received after revision: 2 October 1995/Accepted: 5 March 1996     

Buffering airway acid decreases exhaled nitric oxide in asthma

BACKGROUND: The human airway is believed to be acidified in asthma. In an acidic environment nitrite is converted to nitric oxide (NO). OBJECTIVE: We hypothesized that buffering airway lining fluid acid would decrease the fraction of exhaled NO (F(ENO)). METHODS: We treated 28 adult nonsmoking subjects (9 healthy control subjects, 11 subjects with mild intermittent asthma, and 8 subjects with persistent asthma) with 3 mL of 10 mmol/L phosphate buffered saline (PBS) through a nebulizer and then serially measured F(ENO) levels. Six subjects also received PBS mouthwash alone. RESULTS: F(ENO) levels decreased after buffer inhalation. The maximal decrease occurred between 15 and 30 minutes after treatment; F(ENO) levels returned to pretreatment levels by 60 minutes. The decrease was greatest in subjects with persistent asthma (-7.1 +/- 1.0 ppb); this was more than in those with either mild asthma (-2.9 +/- 0.3 ppb) or healthy control subjects (-1.7 +/- 0.3 ppb, P < .001). Levels did not decrease in subjects who used PBS mouthwash. CONCLUSION: Neutralizing airway acid decreases F(ENO) levels. The magnitude of this change is greatest in persistent asthma. These data suggest that airway pH is a determinant of F(ENO) levels downstream from NO synthase activation. CLINICAL IMPLICATIONS: Airway biochemistry modulates F(ENO) levels. For example, nitrite is converted to NO in the airway, particularly the inflamed airway, by means of acid-based chemistry. Thus airway pH should be considered in interpreting clinical F(ENO) values. In fact, PBS challenge testing integrates airway pH and F(ENO) analysis, potentially improving the utility of F(ENO) as a noninvasive test for the type and severity of asthmatic airway inflammation.     

内源性一氧化氮预防大鼠乙醇性胃粘膜损伤的研究

目的：探讨内源性一氧化氮在大鼠乙醇性胃粘膜损伤时保护胃粘膜作用及机制。方法：大鼠７２ 只,随机分为４ 组,正常对照组,单纯乙醇灌胃（Ｅｔｈ） 组,Ｌ－ 精氨酸＋ 乙醇（Ｌ－ ａｒｇ ＋ Ｅｔｈ） 组,Ｌ－ 硝基精氨酸＋ 乙醇（Ｌ－ ＮＮＡ＋Ｅｔｈ） 组,观察其血浆ＮＯ 含量对胃粘膜的大体及组织学损伤程度、胃粘膜血流量（ｇａｓｔｒｉｃ ｍｕｃｏｓａｌ ｂｌｏｏｄ ｆｌｏｗ ,ＧＭＢＦ） 和胃粘液分泌量的影响。结果：Ｅｔｈ 组血浆ＮＯ 含量,ＧＭＢＦ 和粘液分泌量明显少于对照组（ Ｐ 均＜ ０－０５） ,Ｌ－ ａｒｇ ＋ Ｅｔｈ 组血浆ＮＯ 含量,ＧＭＢＦ 和粘液分泌量明显多于Ｅｔｈ 组（ Ｐ 均＜ ０ ．０５） ,而乙醇对胃粘膜损伤明显减轻（ Ｐ＜ ０－０５） ,Ｌ－ ＮＮＡ ＋ Ｅｔｈ组血浆ＮＯ 含量ＧＭＢＦ 和粘液分泌量明显少于前３ 组（ Ｐ 均＜ ０－０５） ,而胃粘膜损伤程度明显较重（ Ｐ＜ ０－０５） 。结论：内源性ＮＯ 保护胃粘膜对抗乙醇引起的损伤,是通过增加胃粘膜血流量和促进粘液分泌而实现的。     

The effect of endogenous nitric oxide on mechanical ciliostimulation of human nasal mucosa

BACKGROUND: Endogenous nitric oxide (NO) production by the inducible NO-synthase is enhanced in the nasal respiratory epithelium of patients with allergic rhinitis. Recent experimental data suggest endogenous NO to be strongly involved in the regulation of ciliary activity, the driving force of the mucociliary transport system. OBJECTIVE: In this study, we investigated the effect of endogenous NO on mechanical stimulation of ciliary activity in a nasal mucosa explant model. METHODS: Cultures of nasal mucosa explants were incubated with TNF-alpha and bacterial lipopolysaccharides (LPS) to enhance endogenous NO production. Direct in vitro NO imaging was performed by the fluorescent NO-indicator DAF-2 DA and laser scanning confocal microscopy. Ciliary beat frequency (CBF) was determined using a photoelectric technique. Mechanical stimulation was performed by two consecutive flow increments in a closed perfusion chamber. Endogenous NO-synthesis was blocked by l-NAME before the second flow stimulation. RESULTS: Under control conditions the mean rise of CBF relative to baseline was 30.2% during the first flow increment and 30.7% during the second flow increment. Blocking of the endogenous NO synthesis in TNF-alpha/LPS-stimulated cultures reduced baseline CBF by 10.6+/-2.1% (P<0.05) but the effect of mechanical ciliostimulation on CBF remained unchanged (36.0% vs. 38.2%). CONCLUSION: In conclusion, endogenous NO- and Ca(2+)-dependent mechanical stimulation of ciliary activity probably use independent intracellular signalling pathways. The combination of both effects on ciliary activity is likely to improve the local defence against inhaled allergens in patients with nasal allergies.     

Signaling pathways involved in the A and B receptor-mediated cortisol secretagogue effect of endothelins in the human adrenal cortex

Endothelins (ETs) are a family of 21-amino acid hypertensive peptides, which together with their receptors ETA and ETB are expressed in human adrenal cortex. Evidence has been provided that ETs exert a potent secretagogue effect on human adrenocortical cells, acting through both ETA and ETB receptors. Therefore, it seemed worthwhile to study the signaling cascades mediating the cortisol secretagogue effect of the two receptor subtypes. Normal adrenal glands were obtained from consenting patients undergoing unilateral nephrectomy with ipsilateral adrenalectomy for renal cancer. Dispersed zona fasciculata-reticularis (ZF/R) cells were obtained by collagenase digestion and mechanical disaggregation. The selective activation of ETA and ETB receptors was obtained by exposing dispersed cells to ET-1 plus the ETB receptor antagonist BQ-788 and to the selective ETB receptor agonist BQ-3020, respectively. ETA and ETB receptors about equally contributed to the cortisol response of dispersed ZF/R cells to ETs. The phospholipase (PL) C inhibitor U-73122 abolished ETA-mediated secretory response, but only partially prevented the ETB-mediated one. The phosphatidylinositol 3-kinase inhibitor wortmannin and the protein kinase (PK) C inhibitor calphostin-C significantly blunted the secretory responses ensuing from the activation of both receptor subtypes, while the Ca(2+)-channel blocker nifedipine was ineffective. The ETB receptor-, but not the ETA receptor-mediated cortisol response was partially reversed by the cyclooxygenase (COX) inhibitor indomethacin, which when added together with U-73122 abolished it. The inhibitors of adenylate cyclase, PKA, tyrosine kinase and lipoxygenase did not affect the secretory response to the activation of either receptor subtype. ETA-receptor activation raised inositol triphosphate (IP3) production from dispersed ZF/R cells, while ETB-receptor stimulation enhanced both IP3 and prostaglandin-E(2) production. Collectively, our findings indicate that ETs stimulate cortisol secretion from human ZF/R cells, acting through ETA receptors exclusively coupled with PLC/PKC-dependent pathway and ETB receptors coupled with both PLC/PKC- and COX-dependent cascades.     

Regulatory effect of regucalcin on nitric oxide synthase activity in rat kidney cortex cytosol: Role of endogenous regucalcin in transgenic rats

The effect of regucalcin, a regulatory protein in Ca2+ signaling, on nitric oxid (NO) synthase activity in the cytosol of kidney cortex of rats was investigated. The presence of calcium chloride (10 micro M) in the enzyme reaction mixture caused a significant increase in NO synthase activity. This increase was significantly prevented by the addition of trifluoperazine (TFP; 20 or 50 micro M), an antagonist of calmodulin, supporting the existence of Ca2+/calmodulin-dependent NO synthase in rat kidney cortex cytosol. NO synthase activity was significantly decreased by the addition of regucalcin (10(-10)-10(-8) M) in the reaction mixture in the absence or presence of calcium chloride (10 micro M). The regucalcin (10(-8) M) effect was not seen in the presence of Nw-nitro-L-argine metylester (NAME; 10(-6) or 10(-5) M), an inhibitor of NO synthase. Regucalcin significantly reduced NO synthase activity in the presence of TFP (50 micro micro M) or EGTA (1 mM) which has a significant inhibitory effect on the enzyme activity. The presence of anti-regucalcin monoclonal antibody (25 or 50 ng/ml) in the reaction mixture caused a significant increase in NO synthase activity. This increase was completely abolished by the addition of regucalcin (10(-7) M). NO synthase activity was not significantly changed in the kidney cortex cytosol of regucalcin transgenic rats overexpressing endogenous regucalcin as compared with that of wild-type rats. However, the effect of calcium chloride (10 micro M) in increasing NO synthase activity in the kidney cortex cytosol of wild-type rats was significantly weakened in regucalcin transgenic rats. The present study demonstrates that endogenous regucalcin has a suppressive effect on NO synthase activity in the kidney cortex cytosol of rats.     

Mode of nitric oxide action on the renal vasculature

Our study aimed to characterize the essential cellular pathways along which nitric oxide (NO) exerts its well-known vasodilatatory properties in the kidney. Using the isolated perfused rat kidney model we examined the roles of potassium channels, cGMP-protein kinase activity and cAMP-phosphodiesterases (PDE) in the effect of NO on renovascular resistance. We found that neither potassium channel activity nor G-kinase activity was essential for the vasodilatatory effect of NO. The effect of NO, however, was essentially mimicked by pharmacological inhibition of PDE-3, which is a cGMP-inhibitable PDE. As PDE-3 is strongly expressed in renal preglomerular vessels and NO stimulates cGMP formation in renal vessels, it appears likely that inhibition of cAMP degradation and consequently the cAMP pathway are crucially involved in mediating the effects of NO on renal vascular resistance.     

Influence of endogenous nitric oxide on sympathetic vasoconstriction in normoxia, acute and chronic systemic hypoxia in the rat

We studied the role of nitric oxide (NO) in blunting sympathetically evoked muscle vasoconstriction during acute and chronic systemic hypoxia. Experiments were performed on anaesthetized normoxic (N) and chronically hypoxic (CH) rats that had been acclimated to 12% O₂ for 3–4 weeks. The lumbar sympathetic chain was stimulated for 1 min with bursts at 20 or 40 Hz and continuously at 2 Hz. In N rats, acute hypoxia (breathing 8% O₂) reduced baseline femoral vascular resistance (FVR) and depressed increases in FVR evoked by all three patterns of stimulation, but infusion of the NO donor sodium nitroprusside (SNP), so as to similarly reduce baseline FVR, did not affect sympathetically evoked responses. Blockade of NO synthase (NOS) with l -NAME increased baseline FVR and facilitated the sympathetically evoked increases in FVR, but when baseline FVR was restored by SNP infusion, these evoked responses were restored. Acute hypoxia after l -NAME still reduced baseline FVR and depressed evoked responses. In CH rats breathing 12% O₂, baseline FVR was lower than in N rats breathing air, but l -NAME had qualitatively similar effects on baseline FVR and sympathetically evoked increases in FVR. SNP similarly restored baseline FVR and evoked responses. Inhibition of neuronal NOS or inducible NOS did not affect baselines, or evoked responses. We propose that in N and CH rats sympathetically evoked muscle vasoconstriction is modulated by tonically released NO, but not depressed by additional NO released on sympathetic activation. The present results suggest that hypoxia-induced blunting of sympathetic vasoconstriction in skeletal muscle is not mediated by NO.     

Acute effects of endothelins on endogenous adrenomedullin system in the rat heart: Immunocytochemical and autoradiographic studies

Many lines of evidence indicate that adrenomedullin (AM) through its coronary vasodilatory and inotropic effects, exerts a major cardiac protective action. Conversely, endothelins (ETs) exert a cardiac detrimental action, which seems to be mainly mediated by the ETA receptor, whose activation promotes coronary constriction and decreases cardiac blood flow. Hence, we have investigated by immunocytochemistry (ICC) and autoradiography the acute effects of ET-1 on endogenous AM system of adult rat heart. Isolated hearts were perfused for 20 min, according to the Langendorff technique, with 2x10(-9) M ET-1[1-21] or ET-1[1-31], which are mixed ETA/ETB and selective ETA receptor agonists, respectively. ICC demonstrated AM-immunoreactivity (IR) in cardiomyocytes, endocardium and especially in the wall of coronary vessels. Quantitative densitometry showed that ET-1[1-31], but not ET1[1-21], significantly decreased AM-IR in coronary vessels, thereby suggesting that ET-1 elicits AM release in the heart through the activation of ETA receptors. Autoradiography demonstrated [125I]AM-binding sites in cardiomyocytes and especially in the wall of coronary venules, and treatment with both ET-1s did not apparently affect them. This location af AM receptors suggests that AM raises cardiac blood flow by evoking coronary artery dilation indirectly, probably via its stimulating effect on NO production, and by decreasing postcapillary resistance via cardiac vein dilation. Collectively, our findings indicate that important functional interrelationships occur between ET and AM systems in the rat heart, where ETs may at least partly hamper their own ETA receptor-mediated decrease in blood flow by increasing the local release of endogenous AM.     

内源性一氧化氮在急性高原病中的研究进展

1一氧化氮(nitric oxide,NO)与内源性一氧化氮(endogenous NO,eNO) NO是具有调节血管张力、血流等众多生物学作用的脂溶性气体信号分子,eNO是由L-精氨酸(L-arginine,L-Arg)经过体内一氧化氮合酶[nitric oxide synthase,NOS;主要有神经源型(neuronal NOS,nNOS)、诱导型(inducible NOS,iNOS)和内皮源型(endothelial NOS,eNOS)3种]催化合成的,广泛存在于全身组织[1].NO通过一些生物分子和细胞间相互作用参与机体的保护、调节及逆转等生理过程[2].近年来的研究发现NO在调节血管张力、抑制炎症以及抗动脉粥样硬化中发挥关键作用[1-2].     

慢性低氧性肺动脉高压大鼠内源性一氧化氮代谢及其外源性吸入的研究

目的:研究一氧化氮(NO)在大鼠慢性低氧性肺动脉高压发生发展中的代谢变化,观察外源性吸入NO对肺血流动力学的疗效。方法:67只SD雄性成年大鼠,随机分为7组:常氧对照组(n=9),慢性间断性低氧(6h/d,7d/周)1周组(n=7),低氧2周组(n=11),低氧3周组(n=11),L-NAME(NO合成酶抑制剂,30mg/kg,灌饲)处理组(n=10),L-Arg(NO合成前质,10mg/kg,灌饲)处理组(n=9),NO吸入(0.0004%,20min)组(n=10)。插管测肺动脉平均压(MPAP),分离右心室(R)、室间膈+左心室(S+L),计算R/(S+L)(g/g)和R/Wt(Wt:体重,g/kg)。结果:①低氧1周组MPAP显著高于对照组,低氧2周时更高,R/(S+L)和R/Wt也显著高于对照组;②血浆NO₂^-/NO₃^-含量在低氧2周组显著高于对照组,而在低氧3周组显著低于低氧2周组和低氧1周组;低氧1周,血浆ET-1含量显著高于对照组,血浆ET-1含量与MPAP和R/(S+L)均呈显著正相关,r分别为0.43和0.46,P值均＜0.01;③L-NAME组大鼠血浆NO₂^-/NO₃^-含量降低33.2%,R/(S+L)显著增加15.2%,P＜0.05;④L-Arg组大鼠血浆NO₂^-/NO₃^-含量和PAPM无显著改变,但R/(S+L)降低8.7%,P＜0.05;⑤吸入NO,MPAP降低17.8%,P＜0.01。结论:内源性NO在慢性低氧早期(1-2周)继发性增加,但进一步低氧时则减少,血浆ET-1含量的增加在低氧性心肺血管重建中可能具有重要意义,吸入低浓度NO对以肺血管增生和右心室肥大为主要基础的肺动脉高压仍具有明显降低作用。     

Influence of nitric oxide agents in the rat amygdala on anxiogenic-like effect induced by histamine

Background/aims: Both histamine and nitric oxide (NO) may play a role in anxiety-like behavior. Within the brain, the amygdala is an important area involved in processing emotional responses such as anxiety. The aim of the present study was to assess whether the NO system in the basolateral amygdala (BLA) influences histamine-induced anxiety-like behavior in rats. Methods: Male Wistar rats weighing 200–220 g were used. Bilateral cannulae were implanted in the BLA place for microinjections of drugs and the elevated plus maze apparatus has been used to test parameters (%OAT, %OAE, locomotor activity) of anxiety-like behavior. Results: Intra-BLA administration of histamine (2.5 and 5 μg/rat) decreased %OAT [P < 0.001]. Histamine (5 μg/rat) also reduced %OAE [P < 0.05] but not locomotor activity. The results obtained may indicate an anxiolytic response for histamine. Furthermore, bilateral intra-BLA microinjections of different doses of l-arginine (l-arg), an NO precursor (0.5 and 1 μg/rat) increased %OAT [P < 0.01], %OAE [P < 0.01] and locomotor activity [P < 0.001] while NG-nitro-l-arg methylester (l-NAME), a potent inhibitor of NO-synthase (NOS; 0.025, 0.05 and 0.1 μg/rat) decreased %OAT [P < 0.05] and locomotor activity [P < 0.001] but not %OAE. The combination of l-arg (0.5 μg/rat) with histamine increased %OAE [P < 0.001] but had no effect on %OAT and locomotor activity. Finally, the combination of l-NAME (0.025 μg/rat) with histamine decreased %OAT [P < 0.001] and locomotor activity [P < 0.05] but increased %OAE. Conclusion: The results indicate a modulatory role for NO in BLA in the anxiogenic response of histamine in rats.     

Inhibitory effect of nitric oxide on neuronal activity in the periaqueductal grey matter of the rat

Experiments were carried out in urethane-anaesthetized rats to examine the effect of nitric oxide (NO) on neuronal activity within the dorsolateral sector of the midbrain periaqueductal grey matter (PAG), an area which is rich in NO synthesizing neurones. NADPH dependent diaphorase histochemistry revealed small NO synthase containing perikarya, 15.4±3.1 m (mean±SEM) in diameter, in a longitudinal column in the dorsolateral sector of the PAG. The labelled cell bodies were surrounded by a dense meshwork of stained fibres and processes in which unlabelled neurones were embedded. In order to establish whether NO was generated when NO donors were ejected iontophoretically from micropipettes, a chemiluminescence method was used to estimate the output of NO in vitro after iontophoresis of two chemically different classes of NO donor: the sydnonimine 3 morpholino sydnonimin hydrochloride (SIN 1) and the nitrosothiol S nitroso glutathione (SNOG). Iontophoresis of both NO donors into 200 l aliquots of 165 mM NaCl using ejection currents between 6000 and 18000 nA·min produced a current related increase in the concentration of NO. Iontophoresis of SIN 1 in vivo produced a reproducible, current related inhibition of firing in 40 of 59 neurones in the dorsolateral PAG. In 8 of 10 neurones the effect of SIN 1 was significantly reduced after iontophoresis of methylene blue (10–30 nA for 2.7–5 min). The inhibition took up to 7 min to develop and lasted for up to 13 min. Inhibitory responses to GABA were not affected by methylene blue. Iontophoresis of SNOG also inhibited ongoing activity of 18 of 24 neurones tested in the PAG. The experiments demonstrate firstly that NO donors can be used in vivo to deliver NO in the vicinity of neurones by iontophoresis from micropipettes. Secondly, NO appears to inhibit neuronal activity within the PAG.     

高糖对腹膜巨噬细胞诱导型一氧化氮合酶的作用

目的：探讨在体外葡萄糖对SD大鼠腹膜巨噬细胞诱导型一氧化氮合酶（iNOS）mRNA的表达及蛋白的合成。方法:取雄性SD大鼠腹腔巨噬细胞培养，培养的巨噬细胞分为5组：(1)正常对照组；(2)30 mmol/L甘露醇组；(3)90 mmol/L甘露醇组；(4)30 mmol/L葡萄糖组；(5)90 mmol/L葡萄糖组。每组6个培养孔，24 h后提取细胞RNA及蛋白，分别做RT-PCR及Western blotting以检测腹膜巨噬细胞iNOS mRNA的表达及蛋白的合成。结果:30 mmol/L及90 mmol/L甘露醇刺激腹膜巨噬细胞iNOS mRNA的表达及蛋白的合成均不明显， 30 mmol/L及90 mmol/L葡萄糖能明显刺激腹膜巨噬细胞iNOS mRNA的表达及蛋白的合成，90 mmol/L葡萄糖与30 mmol/L的葡萄糖相比更能刺激巨噬细胞iNOS mRNA的表达及蛋白的合成。结果:葡萄糖能明显刺激SD大鼠腹膜巨噬细胞iNOS mRNA的表达及蛋白的合成。     

Cytotoxic effect of autocrine and macrophage-derived nitric oxide on cultured rat mesangial cells

Expression of the inducible form of nitric oxide synthase (iNOS) has been found to be up-regulated in cytokine-stimulated mesangial cells (MC) and in experimental glomerulonephritis. Since direct toxicity of nitric oxide (NO) has been implicated in damage of bacteria, neoplastic and intact pancreatic cells, we investigated whether NO is cytotoxic to cultured MC, which may be relevant to pathogenesis of glomerular injury. MC isolated from rat glomeruli generated substantial amounts of nitrite, the stable NO end-product, when cells were stimulated with IL-1β and tumour necrosis factor-alpha (TNF-α). Total DNA synthesis was significantly reduced in the presence of IL-1β and TNF-α, and this effect was completely reversed by N^G-monomethyl-l-arginine (l-NMMA), an inhibitor of iNOS. Stimulation of MC with IL-1β and TNF-α caused remarkable toxicity to these cells, measured by the MTT test (3-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide cleavage, specific cytotoxicity 41.5 ± 20.3%), and much less prominent MC lysis (³H-thymidine release, specific cytolysis 11.5 ± 5.3%). Toxic effects of cytokines were fully reversible by the iNOS inhibitor. Lipopolysaccharide (LPS) and interferon-gamma (IFN-γ), but not IL-1β and TNF-α, induced rat peritoneal macrophages to produce large amounts of nitrite. In co-culture, such prestimulated macrophages had significantly cytotoxic (MTT test 62.9 ± 19.9%) and cytolytic (³H-thymidine release 57.9 ± 13.8%) effects on MC. Again, this toxicity was totally inhibited in the presence of l-NMMA. We conclude from these results that cytokine-stimulated generation of NO by MC or macrophages is directly toxic to MC, and may play a role in pathogenesis of glomerular injury involving mesangiolysis.     

The effect of pinealectomy on adrenocortical rhytmhm in the blinded rat

The effect of the pineal on circadian periodicity of plasma corticosterone in rats was studied. Male rats were pinealectomized, blinded, or pinealectomized and blinded, and exposed to a lighting cycle of 12 hr of light and 12 hr of darkness. The diurnal variation in plasma corticosterone in pinealectomized animals was not different from that found in intact controls. Blinded rats did not show normal steroid cyclicity. In contrast, rats that were both blinded and pinealectomized had a diurnal variation similar to that of the controls. The pineal gland, then, appears to contribute to the disturbed plasma corticosterone rhythm found in blinded rats.     

On the secretagogue effect of dibutyryl cyclic AMP in the rat exocrine pancreas

Summary DbcAMP0.1 mM induces the discharge of exportable enzymes from rat pancreas fragments incubated in vitro. This effect is qualitatively similar to the action of physiological secretagogues acting via hormone receptors: 1) it is accompanied by the appearance of exocytotic images at the acinar cell apex; 2) it is energy dependent but energy supply is low while that required for the carbamylcholine or caerulein response is high and can only be afforded by oxidative phosphorylation; 3) it is calcium dependent, but no alteration of inward or outward calcium movement can be observed; 4) it is altered by agents known to disrupt the microfilamentous microtubular system [41]. However, the secretory response to DbcAMP is quantitatively less than that obtained with hormonal stimuli. A damaging effect of DbcAMP on pancreatic acinar cells is ruled out on histological and biochemical grounds: there is no significant leakage of LDH; protein synthesis, 2-deoxy-d-glucose andl-leucine uptake are unaltered. The secretagogue effect of DbcAMP is reversible, dose-related and specific. It is not mediated by neuro-transmitter release or by interaction with their receptors. The evidence presented points to a direct interaction of DbcAMP on the pancreatic acinar cell and suggests the last step of the secretory cycle as the most probable site of action of the nucleotide derivative.Abbreviations cAMP Adenosine-3,5-monophosphate - cyclic DbcAMP: N⁶-2-O-Dibutyryl-adenosine-3,5-monophosphate, cyclic - DbcGMP N²-2-O-Dibutyryl-guanosine-3,5-monophosphate, cyclic - 5-AMP Adenosine-5-monophosphate - TCA Trichloracetic acid - ATP Adenosine triphosphate - NADH Nicotinamide-adenine dinucleotide - Tris Tris-(hydroxy-methyl) amino-methane - EGTA Ethylene glycol-bis ( aminoethylether)-NN-tetraacetic acid - LDH Lactic dehydrogenase Part of this work has been presented in abstract from at the VIIIth Symposium of the European Pancreatic Club, Toulouse, France, October, 1975, 23rd–25th and at the Biochemical Society of Belgium [40]This work was partially carried out under contracts from the Ministère de la Politique Scientifique within the framework of the Association Euratom—University of Brussels—University of Pisa, and the Institut National de la Santé et de la Recherche Médicale (France)     

Kv1.1 expression in microglia regulates production and release of proinflammatory cytokines,endothelins and nitric oxide

Potassium channels play an important role in microglial activation but their involvement in main functions of microglia including secretion of proinflammatory cytokines has remained uncertain. This study has revealed the specific expression of Kv1.1 in microglia both in vivo and in vitro. Kv1.1 immunoreactivity was localized in the amoeboid microglia in the rat brain between postnatal (P) day 1 (P1) and day 10 (P10); it was, however, progressively reduced with age and was hardly detected at P14 and P21 in ramified microglia, a derivative cell of amoeboid microglia. Following hypoxic exposure, Kv1.1 expression in amoeboid microglia was enhanced or induced in ramified microglia in more mature brain at P21 when compared with their matching controls. RT-PCR and Western blot analysis confirmed Kv1.1 mRNA and protein expression in murine BV-2 cells which was up-regulated by hypoxia or lipopolysaccharide (LPS) treatment; it was reduced significantly by dexamethasone. Neutralization with Kv1.1 antibody suppressed the expression and release of tumor necrosis factor-α, interleukin-1β, endothelins and nitric oxide (NO) in LPS-activated BV-2 cells. It is concluded that Kv1.1, constitutively expressed by microglia, is elicited by hypoxia and LPS and this may be linked to production of proinflammatory cytokines, endothelins and NO.     

Roles of endogenous nitric oxide synthase inhibitors and endothelin-1 for regulating myometrial contractions during gestation in the rat

This study investigated the roles of endogenous nitric oxide synthase (NOS) inhibitors and endothelin-1 (ET-1) for regulating myometrial contractions during gestation in the rat. Basal and stimulated cyclic GMP production with L-arginine as a NOS substrate or sodium nitroprusside (SNP) as a NO donor were significantly enhanced at the middle of gestation (14th day), while these were greatly decreased at term (22nd day), suggesting the accelerated NO production and/or up-regulation of guanylate cyclase at the middle of gestation. NOS within the myometrium was mainly Ca(2+)dependent and partly Ca(2 + )independent and remained unaffected by aminoguanidine as an inhibitor of inducible NOS in non-pregnant and gestational myometrium. NOS activity per se and endothelial NOS (eNOS) protein expression remained unchanged at the middle and term gestation. Neuronal NOS (nNOS) and inducible NOS (iNOS) proteins were undetectable. SNP at a high concentration of 100 micromol/l failed to modify the spontaneous and ET-1-induced rhythmic contractions in non-pregnant and gestational myometrium. Contents of N(G)-monomethyl-L-arginine (L-NMMA) plus asymmetric N(G),N(G)-dimethyl-L-arginine (ADMA) as endogenous NOS inhibitors and ET-1 within the myometrium were significantly decreased at 14th and 20th days of gestation, whereas these were significantly increased at term gestation (22nd day) and after delivery. There was a significant and positive correlation between endogenous NOS inhibitor content and ET-1. ET-1 within the myometrium was significantly increased with a concomitant decrease in cyclic GMP production after the intraperitoneal application of authentic L-NMMA for 2 weeks, suggesting that the impaired NO production with endogenous NOS inhibitors would result in increased ET-1 content. These results suggest that endogenous NOS inhibitors such as L-NMMA and ADMA play an important role for regulating NO production in rat myometrium. The impaired NO production due to accumulated endogenous NOS inhibitors possibly results in increased ET-1 content within the myometrium, thereby increasing myometrial contractions at term gestation and after delivery.