'The effect of different genres of music on the stress levels of kennelled dogs'.

Classical music has been shown to reduce stress in kennelled dogs; however, rapid habituation of dogs to this form of auditory enrichment has also been demonstrated. The current study investigated the physiological and behavioural response of kennelled dogs (n=38) to medium-term (5days) auditory enrichment with five different genres of music including Soft Rock, Motown, Pop, Reggae and Classical, to determine whether increasing the variety of auditory stimulation reduces the level of habituation to auditory enrichment. Dogs were found to spend significantly more time lying and significantly less time standing when music was played, regardless of genre. There was no observable effect of music on barking, however, dogs were significantly (z=2.2, P<0.05) more likely to bark following cessation of auditory enrichment. Heart Rate Variability (HRV) was significantly higher, indicative of decreased stress, when dogs were played Soft Rock and Reggae, with a lesser effect observed when Motown, Pop and Classical genres were played. Relative to the silent period prior to auditory enrichment, urinary cortisol:creatanine (UCCR) values were significantly higher during Soft Rock (t=2.781, P<0.01) and the second silent control period following auditory enrichment (t=2.46, P<0.05). Despite the mixed response to different genres, the physiological and behavioural changes observed remained constant over the 5d of enrichment suggesting that the effect of habituation may be reduced by increasing the variety of auditory enrichment provided.

Impacts of Fungal Stalk Rot Pathogens on Physicochemical Properties of Sorghum Grain.

Stalk rot diseases are among the most ubiquitous and damaging fungal diseases of sorghum (Sorghum bicolor (L.) Moench) worldwide. Although reports of quantitative yield losses to stalk rots are available, the impact of stalk rot on grain quality attributes is unknown. This study was conducted to test whether stalk rot diseases could affect grain mineral (N, P, K; Ca, Mg, Cu, Fe, Mn, and Zn) and macronutrient (protein, fat, and starch) content, ash content, and physical traits (unit grain weight, hardness, and diameter). A field experiment was conducted in 2013 and 2014 with four sorghum genotypes (two hybrids and two lines). Plants from each genotype were inoculated with four stalk rot pathogens (Fusarium andiyazi, F. proliferatum, F. thapsinum, and Macrophomina phaseolina) and mock-inoculated with phosphate-buffered saline (control). Grains collected from infected and control plants were analyzed for macronutrient and ash content using near-infrared reflectance spectroscopy, grain hardness and diameter using the single-kernel characterization system, and mineral content using the Rapid Flow Analyzer (Model RFA-300 for N) and inductively coupled plasma spectrometer (for P, K, Ca, Mg, Cu, Fe, Mn, and Zn). Although stalk rot pathogens significantly reduced unit grain weight, they did not significantly affect grain hardness and diameter and, therefore, may not affect milling quality. Pathogens significantly reduced all macronutrient and most mineral contents across genotypes and environments on a per-unit-grain basis, except N and Mg, which were affected in a genotype- and environment-specific manner, and Fe, which was not significantly affected. Most minerals tested were significantly and negatively correlated with disease severity (lesion length) and total grain weight per panicle. The hybrid tested (Pioneer 84G62) exhibited reduced mineral and macronutritional changes after stalk rot infection, providing insights into the possibility of producing high-yielding, nutritionally stable hybrids under stalk rot disease pressure through dedicated breeding efforts.

'Four Seasons' in an animal rescue centre; classical music reduces environmental stress in kennelled dogs.

On admission to rescue and rehoming centres dogs are faced with a variety of short- and long-term stressors including novelty, spatial/social restriction and increased noise levels. Animate and inanimate environmental enrichment techniques have been employed within the kennel environment in an attempt to minimise stress experienced by dogs. Previous studies have shown the potential physiological and psychological benefits of auditory stimulation, particularly classical music, within the kennel environment. This study determined the physiological/psychological changes that occur when kennelled dogs are exposed to long-term (7 days) auditory stimulation in the form of classical music through assessment of effects on heart rate variability (HRV), salivary cortisol and behaviour. The study utilised a cross over design in which two groups were exposed to two consecutive 7 day treatments; silence (control) and classical music (test). Group A was studied under silent conditions followed by 7 days of test conditions during which a fixed classical music playlist was played from 10:00-16:30 h. Group B received treatment in the reverse order. Results showed that auditory stimulation induced changes in HRV and behavioural data indicative of reduced stress levels in dogs in both groups (salivary cortisol data did not show any consistent patterns of change throughout the study). Specifically, there was a significant increase in HRV parameters such as µRR, STDRR, RMSSD, pNN50, RRTI, SD1 and SD2 and a significant decrease in µHR and LF/HF from the first day of silence (S1) to the first day of music (M1). Similarly, examination of behavioural data showed that dogs in both groups spent significantly more time sitting/lying and silent and less time standing and barking during auditory stimulation. General Regression Analysis (GRA) of the change in HRV parameters from S1 to M1 revealed that male dogs responded better to auditory stimulation relative to female. Interestingly, HRV and behavioural data collected on the seventh day of music (M2) was similar to that collected on S1 suggesting that the calming effects of music are lost within the 7 days of exposure. A small '9-Day' study was conducted in attempt to determine the time-scale in which dogs become habituated to classical music and examination of the results suggests that this occurs within as soon as the second day of exposure. The results of this study show the potential of auditory stimulation as a highly effective environmental enrichment technique for kennelled dogs. However, the results also indicate the requirement for further investigations into the way in which auditory stimulation should be incorporated within the daily kennel management regime in order to harness the full physiological and psychological benefits of music.

Investigation of the effects of a polymerised bovine haemoglobin solution on tension in isolated canine saphenous artery.

OBJECTIVES: To investigate the vasoconstriction induced by a polymerised bovine haemoglobin solution, Hb-200, in isolated canine arteries. METHODS: Rings of canine saphenous artery, from euthanatized dogs, were mounted between stainless steel wires in Krebs' solution (95% O2 , 5% CO2 , 37°C) for isometric tension recording. Following incubation with Hb-200, cumulative concentration response curves to phenylephrine (vasoconstrictor) and acetylcholine (vasodilator) were investigated. Responses to acute addition of Hb-200 were also examined in pre-constricted or pre-dilated arteries. Responses were further studied in the presence or absence of the endothelium, inhibitors of endothelium-dependent vasodilation (L-NAME, charybdotoxin and apamin), an endothelin antagonist (BQ-788) and the antioxidant superoxide dismutase. RESULTS: Incubation with Hb-200 (0·2 or 2 g/L) significantly enhanced phenylephrine-induced contraction (decreasing half maximal effective concentration, EC50 , P=0·0035) and inhibited acetylcholine-induced relaxation (increasing EC50 , P<0·0001). Acute addition of Hb-200 (0·2 or 2 g/L) significantly increased tension in pre-constricted arteries (P=0·0059) and reversed relaxation in pre-dilated arteries (P=0·0005). These acute responses were abolished in endothelium-denuded arteries and arteries incubated with L-NAME. Responses to Hb-200 were unaffected by incubation with charybdotoxin and apamin, BQ-788, or superoxide dismutase. CLINICAL SIGNIFICANCE: Low concentrations of Hb-200 enhance vasoconstriction in isolated canine saphenous artery, primarily by antagonism of nitric oxide. This effect may be detrimental in some dogs (e.g. those at risk of volume overload) but beneficial in others (e.g. those in septic shock).

Differential vasoactive effects of oestrogen, oestrogen receptor agonists and selective oestrogen receptor modulators in rat middle cerebral artery.

Cerebrovascular disorders are less common in pre-menopausal than post-menopausal women and in females than males. This protection may be due, in part at least, to direct effects of oestrogens on blood vessels. Oestrogen's vasodilatory mechanisms have been reported to be via the endothelium, vascular smooth muscle and extracellular matrix, depending on the vascular bed studied. Herein we investigated the vasoactive effects of oestrogen, oestrogen receptor (ER) and GPR30 agonists and selective ER modulators (SERMs) in the rat middle cerebral artery(MCA), an artery affected in focal ischaemia. MCAs isolated from male Sprague Dawley rats were mounted on a wire myograph. Concentration response curves were constructed to 17ß-oestradiol, ERα agonist-PPT, ERß agonist-DPN, GPR30 agonist-G1 and novel SERMs (LY362321 and LY2120310) in pre-constricted vessels, in the presence and absence of endothelium, blocking agents for nitric oxide synthase (L-NAME), classic ER antagonist (ICI182,780) or plasma membrane specific ERα (ERα-36) antibody. 17ß-oestradiol induced rapid vasorelaxation of the MCA which was not affected by endothelium removal, L-NAME or ICI182,780. Vasorelaxation was mimicked by PPT, DPN and G1 but not by the SERMs. Using ERα-36 antibody, effects of oestrogen were partially blocked. PPT had a greater vasorelaxation, while DPN and G1 had a lesser effect than 17ß-oestradiol. These findings indicate that activation of plasma membrane bound ERα, ß and GPR30 elicits rapid, endothelial-nitric oxide-independent relaxation of the rat MCA.

Age-grading the biting midge Culicoides sonorensis using near-infrared spectroscopy.

Age-grading of insects is important in the control and monitoring of both insect populations and vector-borne diseases. Microscopy and morphological techniques exist to age-grade most blood-feeding flies, but these techniques are laborious, often destructive to the insects, and slow. Near-infrared spectroscopy (NIRS) can be automated and is a non-destructive technique for age-grading. We applied NIRS techniques to age-grade females of the biting midge, Culicoides sonorensis Wirth & Jones (Diptera: Ceratopogonidae), the vector of bluetongue and other arboviruses in North America. Female flies of five known age cohorts (1, 3, 6, 9 and 12 days post-emergence) from three laboratory colonies were used. The data indicate that NIRS can be used to differentiate age groups of C. sonorensis.

Neutering affects mRNA expression levels for the LH- and GnRH-receptors in the canine urinary bladder.

Neutering a bitch increases the incidence of acquired urinary incontinence (AUI) 20-fold. Mechanistically this effect is thought to be related to altered steroid/reproductive hormone concentrations and a recent study showed that gonadotrophin releasing hormone (GnRH) analogue treatment improved continence in bitches with AUI. The aim of this study was to examine mRNA expression levels for luteinizing hormone (LH)- and GnRH-receptors in the canine bladder and the correlation between these and in vitro contractility of the bladder using age matched entire and neutered, male and female canines and canines with AUI. Biopsies from the dome of the bladder were dissected post mortem with informed owner consent. mRNA expression for LH- and GnRH-receptor was quantified by rtPCR (relative to beta-actin). Contractility was assessed (cumulative concentration response curve for carbachol) in strips of bladder muscle using standard protocols. Analysis of variance (Tukey post-test) demonstrated thet neutering was associated with significantly increased levels of expression of LH- and GnRH-receptor mRNA in both sexes (P<0.01). mRNA expression for both receptors was significantly higher in female versus male canines. Neither effect was affected by animals' age and/or weight. A significant inverse correlation (Spearman's test) was found between bladder contractility and mRNA expression for both receptors. This effect was most pronounced in canines with AUI which demonstrated the highest mRNA expression levels yet had the lowest contractility of all animals studied. This suggests that increased LH- and GnRH-receptor mRNA expression is associated with changes in bladder function that increase an animal's predisposition to develop AUI.

Flow-induced enhancement of vasoconstriction and blockade of endothelium-derived hyperpolarizing factor (EDHF) by ascorbate in the rat mesentery.

BACKGROUND AND PURPOSE: We previously reported that ascorbate inhibits flow- and agonist-induced, EDHF-mediated vasodilatation in the bovine ciliary circulation. This study examined whether ascorbate had similar actions in the rat mesenteric vasculature. EXPERIMENTAL APPROACH: The effects of ascorbate were examined both in rat second order mesenteric arterial rings suspended in a static wire myograph and the rat mesentery perfused at different rates of flow. KEY RESULTS: Ascorbate (50 microM) had no effect on U46619-induced tone or acetylcholine-induced, EDHF-mediated vasodilatation in either rings of mesenteric artery or the perfused mesentery at rates of flow below 10 ml min(-1). At higher rates of flow, ascorbate produced two distinct effects in the rat mesentery: a rapid and maintained enhancement of vasoconstrictor tone and a slow (max at 3 h) inhibition of acetylcholine-induced, EDHF-mediated vasodilatation. The enhancement of vasoconstrictor tone appeared to be due to inhibition of flow-induced EDHF-like activity, since it was endothelium-dependent, but could be elicited during blockade of nitric oxide synthase and cyclooxygenase. Despite this, the classical inhibitors of EDHF, apamin and charybdotoxin, failed to affect the ascorbate-induced enhancement of tone, although they inhibited acetylcholine-induced vasodilatation. CONCLUSIONS AND IMPLICATIONS: Ascorbate inhibits both flow- and agonist-induced EDHF in the rat mesentery. The strikingly different timecourses of these two effects, together with their differential sensitivity to apamin and charybdotoxin, suggest that the flow- and agonist-induced EDHFs in the rat mesenteric vasculature may either be different entities or operate by different mechanisms.

Differential effects of glucose on agonist-induced relaxations in human mesenteric and subcutaneous arteries.

BACKGROUND AND PURPOSE: Acute periods of hyperglycaemia are strongly associated with vascular disorder, yet the specific effects of high glucose on human blood vessel function are not fully understood. In this study we (1) characterized the endothelial-dependent relaxation of two similarly sized but anatomically distinct human arteries to two different agonists and (2) determined how these responses are modified by acute exposure to high glucose. EXPERIMENTAL APPROACH: Ring segments of human mesenteric and subcutaneous arteries were mounted in a wire myograph. Relaxations to acetylcholine and bradykinin were determined in a control (5 mM) and high glucose (20 mM) environment over a 2 and 6 h incubation period. KEY RESULTS: Bradykinin-induced relaxation in both sets of vessels was mediated entirely by EDHF whilst that generated by acetylcholine, though principally generated by EDHF, also had contribution from prostacyclin and possibly nitric oxide in mesenteric and subcutaneous vessels, respectively. A 2-h incubation of high glucose impaired bradykinin-induced relaxation of subcutaneous vessels whilst, in contrast, the relaxation generated by bradykinin in mesenteric vessels was enhanced at the same time point. High glucose significantly augmented the relaxation generated by acetylcholine in mesenteric and subcutaneous vessels at a 2 and 6 h incubation point, respectively. CONCLUSIONS AND IMPLICATIONS: Short periods of high glucose exert a variable influence on endothelial function in human isolated blood vessels that is dependent on factors of time, agonist-used and vessel studied. This has implications for how we view the effects of acute hyperglycaemia found in patients with diabetes mellitus as well as other conditions.

The effect of the polymerized bovine haemoglobin solution, Hb-200, on endothelial function in isolated arterial rings from rats.

Haemoglobin-based oxygen carriers have been developed as 'blood substitutes'. Early cross-linked haemoglobins caused marked vasoconstriction, however, polymerized haemoglobin solutions, such as the bovine product haemoglobin glutamer-200 (bovine) (Hb-200), are said to have lesser vascular effects. The aim of this study was to quantify the effect of Hb-200 on isolated rat arterial rings. The actions of Hb-200 were investigated using rings of aorta and second-order mesenteric arteries from female Wistar rats. The arterial rings were mounted for isometric tension recording. Addition of Hb-200 (1 microM) to arterial rings, precontracted with phenylephrine (50-70% maximum phenylephrine-induced tone), resulted in vasoconstriction. Addition of Hb-200 to arteries precontracted with phenylephrine and relaxed with acetylcholine (1 microM) also caused contraction. Furthermore preincubation with Hb-200 (1 microM) enhanced the response to phenylephrine (1 nm to 10 microM) and impaired acetylcholine-induced relaxation (10 nm to 10 microM). Responses to Hb-200 were similar to those in response to the nitric oxide (NO) synthase inhibitor, NG-nitro-l-arginine methyl ester (L-NAME, 100 microM), alone or in combination with Hb-200. These data demonstrate that Hb-200 has a significant vasoconstrictor effect similar to L-NAME. Thus the vascular actions of Hb-200 are consistent with activity as a NO scavenger. This may have implications for the clinical use of Hb-200.

Evidence for the expression and enzymatic activity of haem oxygenase-1 in the lungs of horses.

The aim of this study was to characterise the expression of haem oxygenase-1 (HO-1) in healthy lung tissue from horses and to measure its activity. Samples of lung tissue were collected from six horses euthanased for reasons other than respiratory disease. HO-1 expression and activity were detected in type II alveolar epithelial cells, macrophages and neutrophils in all the samples examined. The activity was dependent on the presence of NADPH and inhibited quantitatively by the addition of increasing concentrations of a competitive inhibitor of HO-1, tin mesoporphyrin IX.

Sex separation of tsetse fly pupae using near-infrared spectroscopy.

Implementation of the sterile insect technique for tsetse (Glossina spp.) requires that only sterile male insects be released; thus, at some stage of the fly production process the females have to be removed. A further constraint in the use of the sterile insect technique for tsetse is that the females are needed for colony production and hence, a non-destructive method of sex separation is required. In most tsetse sterile insect technique programmes thus far, females have been eliminated from the released material by hand-separation of chilled adults. Using near-infrared (NIR) spectroscopy, significant differences have been found between the spectra for the pupae of male and female G. pallidipes Austen. Significantly, the differences appear to be maximized 4-5 days before emergence of the adults. Tsetse fly pupae up to five days before emergence can be sexed with accuracies that generally range from 80 to 100%. This system, when refined, will enable effective separation of male and female pupae to be carried out, with emerged females being returned to the colony and males being irradiated and released. If separation can be achieved five days before emergence, this will also enable irradiated male pupae to be shipped to other destinations as required. Other Diptera were evaluated using this system but had lower classification accuracies of 50-74%. This may be due to the difference in reproductive physiology between these different fly groups.

Transgenic avidin maize is resistant to storage insect pests.

Avidin is a glycoprotein found in chicken egg white, that sequesters the vitamin biotin. Here we show that when present in maize at levels of > or =100 p.p.m., avidin is toxic to and prevents development of insects that damage grains during storage. Insect toxicity is caused by a biotin deficiency, as shown by prevention of toxicity with biotin supplementation. The avidin maize is not, however, toxic to mice when administered as the sole component of their diet for 21 days. These dates suggest that avidin expression in food or feed grain crops can be used as a biopesticide against a spectrum of stored-produce insect pests.

Decreased basal despite enhanced agonist-stimulated effects of nitric oxide in 12-week-old stroke-prone spontaneously hypertensive rat.

This study examined both basal and agonist-stimulated effects of nitric oxide in rings of thoracic aorta and carotid artery from 12-week-old stroke-prone spontaneously hypertensive rats (SHRSP) and compared them to those found in rings from normotensive Wistar Kyoto (WKY) controls. Acetylcholine-induced endothelium-dependent relaxation was found to be five-fold more sensitive in both male and female SHRSP when compared with those from age- and sex-matched WKY rats. In contrast, we found a reduction in the effects of basal nitric oxide in the SHRSP rat. Specifically, the ability of basal nitric oxide to depress contractile responses to phenylephrine was found to be reduced in vessels from SHRSP when compared with those from WKY rats. In addition, the endothelium-dependent depression of vasodilator responses to the nitric oxide donor, glyceryl trinitrate, was reduced in vessels from SHRSP when compared to those from WKY rats. Thus, we have shown that the effects of basal nitric oxide are impaired in the SHRSP rat at an age when the effects of agonist-stimulated nitric oxide are actually enhanced. This impairment may be related to the greater susceptibility of basal nitric oxide to destruction by superoxide anion which is known to be produced in excess in this model of hypertension.

Gene transfer of endothelial nitric oxide synthase improves nitric oxide-dependent endothelial function in a hypertensive rat model.

OBJECTIVE: We have shown previously that there is a relative nitric oxide deficiency at the level of vascular endothelium in the stroke-prone spontaneously hypertensive rat (SHRSP), a model of human essential hypertension, as compared to its normotensive reference strain Wistar Kyoto (WKY) rat. The aim of the current study was to investigate whether adenoviral-mediated gene transfer of an endothelial nitric oxide synthase (eNOS) cDNA (AdCMVeNOS) into carotid arteries of the SHRSP may improve endothelial function. METHODS: Enzyme activity of the recombinant eNOS protein encoded by AdCMVeNOS was tested using a Griess assay in endothelial cells in culture. Left carotid arteries of SHRSP were surgically isolated and exposed to either the AdCMVeNOS or control beta-galactosidase-containing virus, (2 x 10(9) pfu/ml) ex vivo and in vivo. The vessels were harvested 24 h after surgery and analysed by Western blotting, immunohistochemistry and by examining endothelial function ex vivo. RESULTS: Cultured endothelial cells showed almost 100% transduction with both viruses and a dose response of eNOS expression showed a five-fold increase in nitrite production for AdCMVeNOS with no change for beta-galactosidase-containing virus. Western blotting demonstrated a significant increase of eNOS expression in vessels infused with AdCMVeNOS when compared to controls. Immunohistochemistry showed highly positive staining with monoclonal antibodies against eNOS in the intact endothelial cells of the AdCMVeNOS infused vessels. The areas under the curve of the concentration responses to phenylephrine (10(-9) to 3 x 10(-6) M) in the absence and presence of NG-nitroarginine methyl ester (100 microM) showed increased basal nitric oxide bioavailability in the carotid arteries infused with AdCMVeNOS compared to the control (n = 6 for each; P = 0.0069; 95% CI, 0.864 to 3.277). CONCLUSIONS: Our results show that AdCMVeNOS is an effective tool for vascular gene transfer and that it can improve endothelial NO availability in the SHRSP, a genetic model of essential hypertension and endothelial dysfunction.

Interaction between peroxynitrite and L-cysteine: effects on rat aorta.

In rings of rat aorta previously exposed to peroxynitrite (1 mM), L-cysteine and its analogues containing, but not those lacking, a thiol group produced a powerful transient relaxation. This relaxation is likely to result from the release of nitric oxide from a nitrated/nitrosated compound formed following reaction of peroxynitrite with a component of the tissue or bathing medium. Furthermore, when peroxynitrite was pre-mixed with L-cysteine a new relaxant species was formed. Analogues of L-cysteine with a free thiol reacted with peroxynitrite to form species with similar relaxant potencies. Analogues lacking a thiol formed products with relaxant activity, but less than with L-cysteine. Analogues with a free amino but no thiol or carboxylic functions formed products with potencies similar to those lacking only the thiol. If the amino is substituted and the thiol removed, no relaxant activity was generated. Thus, peroxynitrite reacts with L-cysteine to form a novel relaxant whose activity derives mainly from formation of its S-nitrosothiol, with a lesser component perhaps from an N-nitroso derivative.

Estimating aflatoxin in farmers' stock peanut lots by measuring aflatoxin in various peanut-grade components.

Five, 2 kg test samples were taken from each of 120 farmers' stock peanut lots contaminated with aflatoxin. Kernels from each 2 kg sample were divided into the following grade components: sound mature kernels plus sound splits (SMKSS), other kernels (OK), loose shelled kernels (LSK), and damaged kernels (DAM). Kernel mass, aflatoxin mass, and aflatoxin concentration were measured for each of the 2400 component samples. For 120 lots tested, average aflatoxin concentrations in SMKSS, OK, LSK, and DAM components were 235, 2543, 11,775, and 69,775 ng/g, respectively. Aflatoxins in SMKSS, OK, LSK, and DAM components represented 6.9, 7.9, 33.3, and 51.9% of the total aflatoxin mass, respectively. Cumulatively, 3 aflatoxin risk components--OK, LSK, and DAM--accounted for 93.1% of total aflatoxin, but only 18.4% percent of test sample mass. Correlation analysis suggests that the most accurate predictor of aflatoxin concentration in the lot is the cumulative aflatoxin mass in the high 3 risk components OK + LSK + DAM (correlation coefficient, r = 0.996). If the aflatoxin in the combined OK + LSK + DAM components is expressed in concentration units, r decreases to 0.939. Linear regression equations relating aflatoxin in OK + LSK + DAM to aflatoxin concentration in the lot were developed. The cumulative aflatoxin in the OK + LSK + DAM components was not an accurate predictor (r = 0.539) of aflatoxin in the SMKSS component. Statistical analyses of 3 other data sets published previously yielded similar results.

Effects of chronic losartan treatment on vascular reactivity in normotensive rats.

OBJECTIVE: To investigate the vasoactive properties of large (aorta) and small (mesenteric) arteries in vitro after chronic losartan treatment of normotensive rats, hence providing information on the role played by angiotensin II in vascular tone. METHODS: Wistar rats were treated with 10 mg/kg per day losartan for 3 weeks. Ring segments of thoracic aorta and mesenteric resistance arteries (200 microns diameter) were mounted in myographs and wall force measured isometrically. RESULTS: The mean carotid blood pressure was reduced significantly after chronic losartan treatment (108 +/- 3 mmHg, n = 17 versus 116 +/- 2 mmHg, n = 16 in control rats, P < 0.05). In the mesenteric resistance artery the contractile response to 125 mmol/l K+, phenylephrine and angiotensin II was not affected significantly by losartan treatment. A subcontractile concentration of angiotensin II (0.1 nmol/l) induced a significant potentiation of the response to 0.03-100 mumol/l) phenylephrine (450 +/- 180 to 150 +/- 20% of the previous response to phenylephrine in control rats). This potentiation was attenuated significantly in the losartan group (240 +/- 80 to 100 +/- 15% of the previous response, P < 0.01 versus control rats). In the aorta, the response to 125 mmol/l K+ was not affected by chronic losartan treatment. The concentration required for the half-maximal effect for phenylephrine was increased significantly in the losartan group (0.51 +/- 0.11 mumol/l versus 0.17 +/- 0.03 mumol/l in controls rats; no change in maximum response) and the maximum response to angiotensin II was reduced significantly (0.7 +/- 0.08 mN/mg tissue versus 1.9 +/- 0.2 mN/mg tissue in control rats; the concentration for the half-maximal effect was not affected). Potentiation of phenylephrine-induced tone by 0.1 nmol/l angiotensin II (273 +/- 55 to 122 +/- 12% of the previous response in control rats) was attenuated significantly by losartan treatment (91 +/- 46 to 95 +/0 23% of the previous response, P < 0.01 versus control) CONCLUSIONS: Chronic administration of losartan could act on resistance arteries in normotensive rats by blocking the potentiation by angiotensin II of the agonist-induced tone.

Alteration of flow-induced dilatation in mesenteric resistance arteries of L-NAME treated rats and its partial association with induction of cyclo-oxygenase-2.

1. We investigated the response to pressure (myogenic tone) and flow of rat mesenteric resistance arteries cannulated in an arteriograph which allowed the measurement of intraluminal diameter for controlled pressures and flows. Rats were treated for 3 weeks with NG-nitro-L-arginine methyl ester (L-NAME, 50 mg kg-1 day-1) or L-NAME plus the angiotensin I converting enzyme inhibitor (ACEI) quinapril (10 mg kg-1 day-1). 2. Mean blood pressure increased significantly in chronic L-NAME-treated rats (155 +/- 4 mmHg, n = 8, vs control 121 +/- 6 mmHg, n = 10; P < 0.05). L-NAME-treated rats excreted significantly more dinor-6-keto prostaglandin F1 alpha (dinor-6-keto PGF1 alpha), the stable urinary metabolite of prostacyclin, than control rats. The ACEI prevented the rise in blood pressure and the rise in urinary dinor-6-keto PGF1 alpha due to L-NAME. 3. Isolated mesenteric resistance arteries, developed myogenic tone in response to stepwise increases in pressure (42 +/- 6 to 847 +/- 10 mN mm-1, from 25 to 150 mmHg, n = 9). Myogenic tone was not significantly affected by the chronic treatment with L-NAME or L-NAME + ACEI. 4. Flow (100 microliters min-1) significantly attenuated myogenic tone by 50 +/- 6% at 150 mmHg (n = 10). Flow-induced dilatation was significantly attenuated by chronic L-NAME to 22 +/- 6% at 150 mmHg (n = 10, p = 0.0001) and was not affected in the L-NAME + ACEI group. 5. Acute in vitro NG-nitro-L-arginine (L-NOARG, 10 microM) significantly decreased flow-induced dilation in control but not in L-NAME or L-NAME + ACEI rats. Both acute indomethacin (10 microM) and acute NS 398 (cyclo-oxygenase-2 (COX-2) inhibitor, 1 microM) did not change significantly flow-induced dilatation in controls but they both decreased flow-induced dilatation in the L-NAME and L-NAME + ACEI groups. Acute Hoe 140 (bradykinin receptor inhibitor, 1 microM) induced a significant contraction of the isolated mesenteric arteries which was the same in the 3 groups. 6. Immunofluorescence analysis of COX-2 showed that the enzyme was expressed in resistance mesenteric arteries in L-NAME and L-NAME + ACEI groups but not in control. COX-1 expression was identical in all 3 groups. 7. We conclude that chronic inhibition of nitric oxide synthesis is associated with a decreased flow-induced dilatation in resistance mesenteric arteries which was compensated by an overproduction of vasodilator prostaglandins resulting in part from COX-2 expression. The decrease in flow-induced dilatation was prevented by the ACEI, quinapril.

The effects of peroxynitrite on rat aorta: interaction with glucose and related substances.

Peroxynitrite (1-100 microM) induced a concentration-dependent relaxation of rat aortic rings; the logEC50 and maximum relaxation on endothelium-denuded rings were -5.31 +/- 0.03 and 105 +/- 5%, n = 6, respectively. The presence of the endothelium significantly impaired this relaxation (logEC50, -4.41 +/- 0.04; maximum relaxation, 71 +/- 4%; n = 6); an effect which was reversed by the inhibitor of nitric oxide synthase, N(G)-nitro-L-arginine methyl ester (L-NAME; 100 microM). Incubation with a high concentration of peroxynitrite (1 mM, 10 min followed by washout) had no effect on subsequent relaxation to acetylcholine (0.01-1 microM). It did, however, significantly depress subsequent contraction to phenylephrine (1-300 nM). This depression was dependent upon the presence of D-glucose in the Krebs solution, could be reversed by the inhibitor of soluble guanylate cyclase, methylene blue (10 microM) and reversed spontaneously after 2 h. When peroxynitrite (1 mM) was mixed with D-glucose (11 mM) and subsequently neutralised to remove unreacted peroxynitrite, a new more potent relaxant was formed. Despite this, the ability of peroxynitrite (1-100 microM) to produce relaxation of endothelium-denuded rings was similar in normal and glucose-free Krebs. Glycerol (22 mM), which like D-glucose is membrane permeant, also reacted with peroxynitrite (1 mM) to form a new more potent relaxant. L-cysteine (1 mM) had no effect by itself on the tone of aortic rings and when present in the tissue bath had no effect on the ability of peroxynitrite or neutralised peroxynitrite (1-100 microM) to produce relaxation. It did, however, potentiate the relaxant actions of the products formed from the reaction of peroxynitrite with D-glucose or glycerol. The membrane impermeant sugars, mannitol and sorbitol (each 11 mM) also reacted with peroxynitrite (1 mM), but expression of the vasorelaxant properties of their respective derivatives was seen only in the presence of L-cysteine (1 mM). Membrane permeance cannot, however, explain why peroxynitrite reacts with D-glucose and glycerol, but not mannitol or sorbitol to form products with intrinsic relaxant activity, as the product formed from the impermeant sugar, L-glucose (11 mM), also has intrinsic activity. The relaxant potency of this product was equipotent to that formed from D-glucose and was also potentiated by L-cysteine (1 mM). These result confirm that peroxynitrite can react with glucose and other compounds with alcohol functional groups to form vasorelaxant species. The relaxation induced when peroxynitrite is added to rat aortic rings is not, however, dependent upon this reaction since it occurs in glucose-free Krebs.