Transport of cisplatin by the copper efflux transporter ATP7B

ATP7B is a P-type ATPase that mediates the efflux of copper. Recent studies have demonstrated that ATP7B regulates the cellular efflux of cisplatin (DDP) and controls sensitivity to the cytotoxic effects of this drug. To determine whether DDP is a substrate for ATP7B, DDP transport was assayed in vesicles isolated from Sf9 cells infected with a baculovirus that expressed either the wild-type ATP7B or a mutant ATP7B that was unable to transport copper as a result of conversion of the transmembrane metal binding CPC motif to CPA. Only the wild-type ATP7B-expressing vesicles exhibited copper-dependent ATPase activity, copper-induced acyl-phosphate formation, and ATP-dependent transport of copper. The amount of DDP that became bound was higher for vesicles expressing either type of ATP7B than for those not expressing either form of ATP7B, but only the vesicles expressing wild-type ATP7B mediated ATP-dependent accumulation of the drug. At pH 4.6, the vesicles expressing the wild-type ATP7B exhibited ATP-dependent accumulation of DDP with an apparent K(m) of 1.2 +/- 0.5 (S.E.M.) muM and V(max) of 0.03 +/- 0.002 (S.E.M.) nmol/mg of protein/min. DDP also induced the acyl-phosphorylation of ATP7B but at a much slower rate than copper. Copper and DDP each inhibited the ATP-dependent transport of the other. These results establish that DDP is a substrate for ATP7B but is transported at a much slower rate than copper.     

Displacement of nortriptyline and uptake of 14C-lidocaine in the lung after administration of 14C-lidocaine to nortriptyline intoxicated pigs

Six anaesthetized Swedish land-race pigs were intoxicated by an intravenous infusion of nortriptyline-HCl (NT) up to a concentration of 4.58 +/- 0.58 (mean +/- S.E.M.) microM in arterial whole blood. A rapid injection of 2 mg/kg b.wt. lidocaine-HCl in the right atrium was followed by a rise in arterial whole blood concentration of NT up to a maximum of 7.32 +/- 0.28 microM NT. Amount displaced NT from the cardio-pulmonary circulation after the 14C-lidocaine bolus, was calculated to be 0.66 +/- 0.03 mumol. Lung uptake of 14C-lidocaine during first-pass through the lung was not influenced to any statistically signficant degree compared to a control group. Thus, first pass uptake (FPU) was 30 +/- 8 (mean +/- S.E.M.)% and 39 +/- 5% respectively. The duration of the QRS-complex of the ECG was increased (P less than 0.01) during the infusion of NT from 0.07 +/- 0.01 (mean +/- S.E.M.) sec. to 0.14 +/- 0.02 sec. when 250 mg NT-HCl had been administered. The QRS-duration was decreased (P less than 0.01) after the injection of the 14C-lidocaine bolus to 0.09 +/- 0.01 sec. Mean arterial blood pressure and heartrate decreased slightly during the infusion of NT, but did not change immediately after the 14C-lidocaine bolus.     

Possible role of histamine (H1- and H2-) receptors in the regulation of meningeal blood flow

1. Vasodilatation in the dura mater has been suggested to play an important role in the pathophysiology of vascular headaches. Histamine may contribute to these vascular changes. The aim of the present study was to examine the role of different histamine receptors in histamine-induced meningeal hyperperfusion using laser Doppler flowmetry. 2. The blood flow in the medial meningeal artery was monitored in the exposed parietal dura mater encephali of barbiturate anaesthetized rats. Local application of histamine (10(-5) and 10(-4) M) onto the dura caused increases in flow to 114.2+/-9.6 and 135.1+/-19.1%, respectively, of the basal flow. 3. Flow increases induced by topical application of histamine (10(-4) M) were reduced by local pretreatment with the H(2)-receptor antagonist cimetidine (0.4 and 4 mM) to 63.4+/-17 and 37.8+/-18.8%, respectively. Systemic pre-administration of cimetidine (5 mg kg(-1) i.v.) did not change histamine-induced flow increases. 4. Local pretreatment with the H(1)-receptor antagonist cetirizine (2 micro M) further increased the flow evoked by topical histamine administration (10(-4) M) to 123.5+/-14.7% of the histamine control. 5. Increases in blood flow induced by i.v. administration of histamine (10 micro g kg(-1)) were reduced by i.v. pre-injection of cetirizine (50 micro g kg(-1)) to 31.9+/-9% but not by i.v. cimetidine (5 mg kg(-1)). 6. We conclude that histamine-induced relaxation of dural arterial vessels is mediated by H(2)-receptors, most likely located on vascular smooth muscle cells, and by endothelial H(1)-receptors. In addition, H(1)-receptors on smooth muscle cells may mediate vasoconstriction.     

The effects of nisoldipine (Bay K 5552) on cardiovascular performance and regional blood flow in pentobarbital-anaesthetized pigs with or without beta-adrenoceptor blockade.

The effects of the 1,4-dihydropyridine derivative nisoldipine, infused intravenously (i.v.) at 3 different rates (0.25, 0.5 and 1.0 microgram kg-1 min-1), were studied in anaesthetized pigs on cardiovascular performance with or without beta-adrenoceptor blockade produced by propranolol. Nisoldipine caused dose-dependent decreases in arterial blood pressure (30%), systemic vascular resistance (30%) and left ventricular filling pressure (15%), but raised heart rate (25%) and LV dP/dt max (20%). Cardiac output was not significantly affected. Transmural myocardial blood flow and vascular conductances increased dose-dependently after nisoldipine. The elevation in blood flow to the left ventricle favoured epicardial layers. Endocardial blood flow showed small increases as the changes in conductance of the endocardial layer more than compensated for the loss in perfusion pressure. The endo-epi blood flow ratio decreased from 1.16 +/- 0.05 to 0.70 +/- 0.01. Myocardial O2-consumption was unaltered as the decrease in arterial-coronary venous O2-content difference (30%) was balanced by the increase in transmural blood flow. Nisoldipine increased blood flow to skeletal muscle (500%), stomach (50%) and adrenals (25%), but decreased that to the liver (50%), spleen (25%) and kidneys (25%). No changes were noticed in the small intestine, skin and brain. In spite of differential effects on blood flow, vascular conductance in all organs and tissues, with the exception of the liver, increased. After beta-adrenoceptor blockade the responses of mean arterial blood pressure, cardiac output and systemic vascular resistance to nisoldipine remained virtually unchanged, but the elevations in heart rate and LV dP/dt max were abolished, as was the decrease in left ventricular filling pressure. A higher dose of nisoldipine was required after beta-adrenoceptor blockade to elicit significant vasodilatation in the epi- and endocardial layers. However, the reduction in endo-epi blood flow ratio by nisoldipine was not affected by propranolol. Myocardial O2-consumption tended to decrease as the diminution in the arterial-coronary venous O2-content difference (30%) slightly exceeded the increase of left ventricular blood flow (30%). Except for the brain and liver, effects of nisoldipine on regional vascular conductances were attenuated after beta-adrenoceptor blockade.     

Effects of S9977 and dihydroergotamine in an animal experimental model for migraine.

The present study concerns the effects of S9977, a trimethylxanthine derivative with potential antimigraine characteristics, on the distribution of carotid blood flow in the anaesthetized pig. Furthermore, the effects of dihydroergotamine have been analysed for comparison. Dihydroergotamine (3, 10, 30 and 100 micrograms/kg, i.v.) elicited dose-dependent pressor and bradycardic responses which were probably mediated by its partial agonist action on alpha-adrenoceptors and dopamine2 receptors. In contrast, S9977 (0.3, 1, 3 and 10 mg/kg, i.v.) caused a moderate hypotension and bradycardia. The carotid haemodynamic effects of dihydroergotamine (3, 10, 30 and 100 micrograms/kg, i.v.) consisted of a dose-dependent reduction of arteriovenous anastomotic blood flow and conductance and an increase in nutrient (tissue) blood flow and conductance. As a consequence, jugular venous PO2 decreased. These findings, demonstrating an active constriction of arteriovenous anastomoses, are in agreement with earlier findings in cats. Though S9977 (0.3, 1, 3 and 10 mg/kg, i.v.) decreased carotid (two highest doses) and arteriovenous anastomotic (highest dose) blood flow, there was no concomitant decrease in the vascular conductances. Therefore, the effects of S9977 seem to be related to a decrease in arterial blood pressure and not to an active vasoconstriction of arteriovenous anastomoses. These results are discussed in terms of the potential therapeutical usefulness of S9977 in the treatment of migraine.     

Effects of digoxin in isolated human pulmonary vessels.

In isoalted human pulmonary arteries and veins the contractile response to noradrenaline (1.8 X 10(-5)M) was 33 +/- 7.4% and 20 +/- 4.5% (Mean +/- S.E.M.) of that induced by potassium (127 mM). A variable degree of spontaneous contractile activity was recorded in the vein preparations. This activity was abolished by nifedipine (2.9 X 10(-6)M). Digoxin (10(-6)M) induced contractions in pulmonary vessels. In the arteries, the digoxin contraction developed slowly and reached a maximum amplitude of 90 +/- 4% (Mean +/- S.E.M.) of the potassium evoked contraction. In the veins, the amplitude of the digoxin contraction was 32 +/- 5% of that induced by potassium. Digoxin (10(-6)M) also increased the maximum response to noradrenaline and potassium in both arteries and veins. In the arteries, the noradrenaline and potassium contractions increased to 211 +/- 6.8% and 145 +/- 8.9 of control, and in the veins to 169 +/- 13.5% and 163 +/- 9.9%, respectively. Nifedipine in concentrations which completely relaxes arterial and venous preparations contracted by potassium, had only a slight relaxing effect on digoxin induced contraction. It is concluded that digoxin increases the tension in pulmonary arteries and veins, and increases the maximum response to noradrenaline and potassium in both types of vessels. The digoxin induced contraction is highly resistant to blockade of extracellular calcium influx.     

Investigation of the role of 5-HT1B and 5-HT1D receptors in the sumatriptan-induced constriction of porcine carotid arteriovenous anastomoses.

1. It has previously been shown that the antimigraine drug sumatriptan constricts porcine carotid arteriovenous anastomoses via 5-HT1-like receptors, identical to 5-H1B/1D receptors. The recent availability of silent antagonists selective for the 5-HT1B (SB224289) and 5-HT1D (BRL15572) receptor led us to further analyse the nature of receptors involved. 2. In pentobarbitone-anaesthetized, bilaterally vagosympathectomized pigs, sumatriptan (30, 100 and 300 microg kg(-1), i.v.) dose-dependently decreased carotid arteriovenous anastomotic conductance by up to 70+/-5%. 3. The dose-related decreases in carotid arteriovenous anastomotic conductance by sumatriptan (30, 100 and 300 microg kg(-1), i.v.) remained unchanged in animals treated (i.v.) with 1 mg kg(-1) of BRL15572 (maximum decrease: 72+/-3%), but were significantly attenuated by 1 mg kg(-1) (maximum decrease: 30+/-11%) and abolished by 3 mg kg(-1) (maximum decrease: 3+/-7%) of SB224289. The highest dose of SB224289 did not attenuate the hypertension, tachycardia or increases in carotid blood flow induced by bolus injections of noradrenaline (0.1-3 microg kg(-1), i.v.). 4. The results indicate that sumatriptan constricts porcine carotid arteriovenous anastomoses primarily via 5-HT1B, but not via 5-HT1D receptors.     

Determination of corticosteroids in tissue samples by liquid chromatography-tandem mass spectrometry

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for determination of corticosterone and 11-dehydrocorticosterone (11-DHC) levels in KKA(y) mouse liver and adipose tissue, and hydrocortisone and cortisone levels in human adipose tissue has been developed. The corticosteroids were extracted from liver tissue with methanol/water and ethyl acetate for adipose tissue samples. Corticosterone and 11-DHC were separated with a methanol gradient and hydrocortisone and cortisone with an acetonitrile gradient containing trifluoroacetic acid on a reversed-phase column within 15 min. The corticosteroids were detected after electrospray ionization in positive mode with multiple reaction monitoring (MRM). The limits of quantification (LOQ) were estimated to be 15 nmol/kg liver and 1.6 nmol/kg adipose tissue for corticosterone and 5.4 nmol/kg liver and 0.92 nmol/kg adipose tissue for 11-DHC. The LOQ was estimated to be 0.2 nmol/kg adipose tissue for hydrocortisone and 0.4 nmol/kg adipose tissue for cortisone. The limits of detection (LOD) at 3 times S/N were estimated to be 0.07 nmol/kg adipose tissue for hydrocortisone 0.1 nmol/kg adipose tissue for cortisone. The variation of endogenous levels in KKA(y) mouse from different animals (CV%) was high with mean liver tissue levels of 117+/-25 (S.D.)nmol/kg for corticosterone and 62+/-19 (S.D.)nmol/kg for 11-DHC (n=5) and adipose tissue levels of 39+/-20 (S.D.)nmol/kg for corticosterone and 2.4+/-0.9 (S.D.)nmol/kg for 11-DHC (n=9). Endogenous levels in human biopsy samples from adipose tissue were 12+/-7.0 (S.D.)nmol/kg for hydrocortisone and 3.0+/-1.6 (S.D.)nmol/kg for cortisone (n=16). The new LC-MS/MS methods showed sufficient sensitivity and selectivity for determination of endogenous levels of corticosteroids in both KKA(y) mouse liver and adipose tissue samples and human adipose tissue samples. The selectivity of the methods was verified by analysis of two different productions from each analyte.     

The quantification of endogenous steroids in bovine aqueous humour and vitreous humour using isotope dilution GC-NCI-MS

Pentafluorobenzyloxime-trimethylsilyl derivatives of androgens, progestogens and corticosteroids were prepared and used for the analysis of these steroids in bovine aqueous humour and vitreous humour by GC-MS method. Appropriate deuteriated isotopomers of the parent steroids were labelled with deuterium via simple synthetic procedure and used as internal standards. The concentration (ng ml(-1), +/- S.E.M.) of these steroids in bovine aqueous humour and vitreous humour were found to be as follow: (1) aqueous humour (n = 17); hydrocortisone (n = 17; 2.40 +/- 0.54), progesterone (n = 15; 0.06 +/- 0.01), 4-androstene-3,17-dione (n = 8; 0.15 +/- 0.07) and testosterone (n = 4; 0.14 +/- 0.04); and (2) bovine vitreous humour (n = 19); hydrocortisone (n = 19; 1.78 +/- 0.25), progesterone (n = 18; 0.09 +/- 0.01), 4-androstene-3,17-dione (n = 19; 0.11 +/- 0.02), 11-deoxycorticosterone (n = 12; 29.27 +/- 6.42), 17alpha-hydroxyprogesterone (n = 6; 5.55 +/- 3.12). The concentration of corticosterone, 11-deoxycorticosterone and 17alpha-hydroxyprogesterone and testosterone and corticosterone were below the limit of detection in aqueous humour and vitreous humour, respectively.     

Effects of BIBN4096BS on cardiac output distribution and on CGRP-induced carotid haemodynamic responses in the pig

Calcitonin gene related peptide (CGRP) seems to be involved in the pathogenesis of migraine, since plasma CGRP levels increase during the headache phase. In the present study, we investigated the effects of a novel CGRP receptor antagonist, BIBN4096BS (1-piperidinecarboxamide, N-[2-[[5-amino-1-[[4-(4-pyridinyl)-1-piperazinyl]carbonyl] pentyl] amino]-1-[(3,5-dibromo-4-hydroxyphenyl) methyl]-2-oxoethyl]-4-(1,4-dihydro-2-oxo-3(2H)-quinazolinyl)-, [R-(R*,S*)]-), on the regional cardiac output distribution and on the carotid haemodynamic changes induced by alpha-CGRP in anaesthetised pigs. Treatment with BIBN4096BS (100, 300 and 1000 microg kg(-1), i.v.) did not affect the heart rate, mean arterial blood pressure or systemic vascular conductance, but a small decrease in cardiac output was noticed; the latter was, however, not significantly different from that in vehicle-treated animals. The highest dose of BIBN4096BS moderately decreased vascular conductance in the lungs, kidneys, spleen and adrenals. Vascular conductance in other tissues including the brain, heart, gastrointestinal system, skin and skeletal muscles remained unchanged. Intracarotid artery infusions of alpha-CGRP (10, 30 and 100 pmol kg(-1) min(-1) during 3 min) increased the total carotid blood flow and conductance, but decreased the arterial blood pressure. These responses were dose-dependently blocked by BIBN4096BS. The above results show that BIBN4096BS is a CGRP receptor antagonist in the porcine carotid and systemic circulations, but the endogenous CGRP does not seem to play an important physiological role in regulating basal vascular tone. These findings suggest that BIBN4096BS may have therapeutic usefulness in migraine.     

Regional blood flow responses to acute ANG II infusion: effects of nitric oxide synthase inhibition.

We hypothesized that nitric oxide (NO) opposes regional vasoconstriction caused by acute angiotensin II (ANG II) infusion in conscious rats. Mean arterial pressure (MAP), blood flow, and vascular conductance (regional blood flow/ MAP; ml/min/100 g/mm Hg) were measured and/or calculated before and at 2 min of ANG II infusion (0.05 or 1 microg/kg/min, i.a.) in the absence and presence of NO synthase (NOS) inhibition [N(G)-nitro-L-arginine methyl ester (L-NAME), 0.25 or 1 mg/kg, i.a.]. ANG II reduced stomach and hindlimb conductance only after NOS inhibition. For example, whereas 0.05 microg/kg/min ANG II did not attenuate conductance in the stomach (i.e., 1.04+/-0.08 to 0.93+/-0.12 ml/min/100 g/mm Hg), this variable was reduced (i.e., 0.57+/-0.14 to 0.34-/+0.05 ml/min/100 g/mm Hg; p < 0.05) when ANG II was infused after 0.25 mg/kg L-NAME. In addition, whereas hindlimb conductance was similar before and after administering 1 microg/kg/min ANG II (i.e., 0.13+/-0.01 and 0.09+/-0.02, respectively), this variable was reduced (i.e., 0.07+/-0.01 and 0.02+/-0.00, respectively; p < 0.05) when ANG II was infused after 1 mg/kg L-NAME. These findings indicate that NO opposes ANG II-induced vasoconstriction in the stomach and hindlimb. In contrast, whereas both doses of ANG II decreased (p < 0.05) vascular conductance in the kidneys and small and large intestine regardless of whether NOS inhibition was present, absolute vascular conductance was lower (p < 0.05) after L-NAME. For example, 1 microg/kg ANG II reduced renal conductance from 3.34+/-0.31 to 1.22+/-0.14 (p < 0.05). After 1 mg/kg L-NAME, renal conductance decreased from 1.39+/-0.18 to 0.72+/-0.16 (p < 0.05) during ANG II administration. Therefore the constrictor effects of NOS inhibition and ANG II are additive in these circulations. Taken together, our results indicate that the ability of NO to oppose ANG II-induced constriction is not homogeneous among regional circulations.     

The pharmacological selectivity of three NMDA antagonists

Three N-methyl-D-aspartate (NMDA) antagonists (+/-)2-amino-5-phosphonopentanoate (AP5), 3-((+/-)-2-carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP) and ((+/-)-5-methyl-10,11-dihydro-5H-dibenzo(a.d.)cyclohepten-5,10- imin e maleate) (MK-801) have been tested for selectivity against depolarization of motoneurones induced by carbachol, 5-hydroxytryptamine, noradrenaline and substance P in isolated immature rat spinal cord preparations. AP5 (400 microM) and CPP (50 microM) gave mean dose-ratios, for antagonism against NMDA, of 103 +/- 14.9 S.E.M. (eight preparations) and 34.1 +/- 1.9 S.E.M. (14 preparations). MK-801 (1 and 10 microM) was the most potent of the three antagonists yielding dose ratios greater than 100 after 120 min treatment. MK-801 potentiated responses induced by 5-hydroxytryptamine and noradrenaline given dose-ratios of 0.22 +/- 0.16 S.E.M. and 0.20 +/- 0.06 S.E.M., respectively (four preparations). The three antagonists produced no significant antagonism of the non-amino acid agonists (four preparations for each agonist) when dose-ratios against NMDA were at least 40. The observations support the use of these antagonists as tools to identify sites of excitatory amino acid-mediated transmission.     

Degradation of busulfan in aqueous solution

The influence of pH, phosphate buffer components and temperature on the degradation rate of busulfan was studied. The analysis was performed using gas chromatography with electron capture detection and reversed-phase liquid chromatography with radioactivity monitoring. The degradation rate of busulfan showed no pH dependence in the range pH 1.5-11 and increased at higher pH values. The degradation rate constant was 0.034 +/- 0.001 h(-1) (S.E.M.) for the degradation of busulfan in pure water and 0.45 +/- 0.01 h(-1)M(-1) (S.E.M.) for the reaction of busulfan with the hydroxide ion at 37 degrees C. The reactivity of HPO(4)(-2) was six times higher than the reactivity of H(2)PO(4)(-1) towards busulfan. The hydrolysis products were identified as tetrahydrofuran and methanesulphonic acid by nuclear magnetic resonance spectroscopy.     

Ouabain at nanomolar concentration promotes synthesis and release of angiotensin II from the endothelium of the tail vascular bed of spontaneously hypertensive rats

The effects of 1 nM ouabain (OUA) on the contractile actions of phenylephrine (PHE, 0.001-100 microg) and functional activity of the sodium pump (NKA) in isolated-perfused tail vascular beds from WKY and SHR were investigated. In preparations from SHR, perfusion with OUA in the presence of endothelium (E+) increased the sensitivity (pED50) of PHE (before: 2.14 +/- 0.06 versus after: 2.47 +/- 0.07; P < 0.05) without altering the maximal response (Emax). After endothelial damage, OUA reduced the Emax of PHE in SHR (before: 350 +/- 29 versus after: 293 +/- 25 mm Hg; P < 0.05). In SHR/E+, pretreatment with losartan (10 microM) or enalaprilat (1 microM) prevented the increased sensitivity to PHE induced by OUA. OUA increased NKA activity in SHR/E+ (before: 45 +/- 6 versus after: 58 +/- 5%, P < 0.05). Losartan (10 mg/Kg, i.v.) also abolished the increment in systolic and diastolic blood pressure induced by OUA (0.18 microg/Kg, i.v.) in anesthetized SHR. OUA did not alter the actions of PHE in either anesthetized WKY rats or vascular preparations. Results suggest that 1 nM OUA increased the vascular reactivity to PHE only in SHR/E+. This effect is mediated by OUA-induced activation of endothelial angiotensin converting enzyme that promotes the local formation of angiotensin II, which sensitizes the vascular smooth muscle to the actions of PHE.     

Inhibition and reversal of endotoxin-, aggregated IgG- and paf-induced hypotension in the rat by SRI 63-072, a paf receptor antagonist

Platelet activating factor (paf) given intravenously produces systemic hypotension in the rat. Similar effects can be induced using endotoxin or heat-aggregated IgG challenges, which are thought to involve endogenous paf release. Extending this concept, we have examined the ability of the paf antagonist SRI 63-072 to inhibit or reverse systemic hypotension induced with paf, heat-aggregated IgG or endotoxin 0111-B4 in rats. At 100 ng kg-1 paf, there occurred a 38.6 +/- 5.1% decrease in carotid mean arterial pressure (MAP) followed by a 3.2 +/- 0.7 min recovery period (RP) to return to normal pressure values. The ED50 of SRI 63-072 was 0.16 mg kg-1 i.v. (MAP) and 0.25 mg kg-1 (RP) when given 1-5 min before the paf challenge. Endotoxin (15 mg kg-1 i.v.) produced a hypotensive response (54 +/- 8% decrease in MAP) and a corresponding 80% decrease in mesenteric artery blood flow. When given 2-8 min after endotoxin, 1.0 mg kg-1 i.v. SRI 63-072 totally restored blood pressure and artery blood flow. SRI 63-072 similarly reversed heat-aggregated IgG (10 mg kg-1) induced reduction of MAP, with an ED50 of 0.05 mg kg-1 i.v. The observations that SRI 63-072 can inhibit or reverse systemic vascular effects produced from paf and other provocators of endogenous paf release strongly implicates paf as a common final mediator of hypotension and shock. As SRI 63-072 is a competitive receptor antagonist, the hypotensive effects of these provocators appear to be mediated by vascular receptors for paf.     

Effects of palytoxin on Na, K and ATP contents of vascular smooth muscle of rabbit aorta

Palytoxin (PTX) at concentrations higher than 10(-9) M increased tissue Na and decreased tissue K contents in the smooth muscle of rabbit aorta. The decrease in the tissue K content induced by PTX (10(-8)M) was complete within 1 hr. Saponin (1 mg/ml) and Triton X-100 (0.1% wt./vol.) also rapidly decreased the tissue K content. On the other hand, a high concentration of ouabain (10(-3)M) did not change the tissue K content within 1 hr of application, and the maximum loss of K was obtained after 6 hr. Loss of tissue Na into Na- and K-free solution from Na loaded muscle was accelerated by PTX (10(-8)M). The PTX-induced increase in loss of Na was inhibited in proportion to the decrease in the temperature from 37 to 10 degrees C, while the loss of Na in the absence of PTX was almost completely inhibited at 24 degrees C. Decrease in the wet weight of the muscle induced by hyperosmotic solution was inhibited by pretreatment with PTX (10(-8)M) or saponin (1 mg/ml) for 1 hr. PTX (10(-9) and 10(-8)M) had no effect on the ATP content of the muscle. However, PTX at concentrations above 10(-7)M reduced the ATP content, and a significant amount of ATP was detected in the incubation medium. Saponin (1 mg/ml) and Triton X-100 (0.2% wt./vol.) induced a release of Na from liposomes prepared with synthesized lecithin or total lipids of rabbit red blood cells. However, no Na leak was induced by PTX (10(-8)-10(-6)M) in these liposomes. These results suggest that PTX at low concentrations (10(-9)-10(-8)M) increases the membrane permeability of vascular smooth muscle cells to Na and K ions. At higher concentrations (10(-7)-10(-6)M). PTX seems to form pores which are permeable to a larger molecule like ATP. The results further suggest that the mode of action of PTX is different from that of saponin or detergent.     

Effects of BY-1949 on evoked responses of cortical blood flow and Meynert neurons.

The effects of BY-1949, a novel dibenzoxazepine derivative, injected into the cerebral ventricle on noxious stimulus-induced responses of regional blood flow in the cortex and neuronal activity in the nucleus basalis of Meynert were studied in male rats. These induced responses were markedly enhanced by administration of BY-1949 (16.4 +/- 0.5 ng/100 g, S.E.M.). These data indicate that BY-1949 acts on the central nervous system to modulate the response to a noxious stimulus of regional cerebral blood flow and neuronal activity in the nucleus basalis of Meynert.     

Effect of Larrea divaricata Cav. extract and nordihydroguaiaretic acid upon peroxidase secretion in rat submandibulary glands.

Free radicals are involved in several diseases, including cancer, central nervous system alterations and inflammatory pathologies. Peroxidase is an oral enzyme implicated in the defence of oral cavity. It has been determined that flavonoids and lignans possess antioxidant and free radical scavenging either directly or indirectly, usually by means of increasing the secretion of free radicals scavenger enzymes. Larrea divaricata Cav. is a plant used in folk Argentine medicine for the treatment of cancer and inflammatory ailments. In this study, we have determined the effect and mechanism of action of an aqueous extract of the leaves of L. divaricata and NDGA on peroxidase secretion in female rat submandibular glands. The extract significantly increased the secretion and total peroxidase. % of secreted peroxidase (X +/- S.E.M.): extract maximum response: 150 +/- 10; % of total peroxidase (X +/- S.E.M.): extract maximum response: 1000 +/- 90. The effect of the extract on peroxidase secretion was mediated by beta1 adrenoceptors (% of secreted peroxidase: extract + atenol maximum response: 50 +/- 4 ). Meanwhile, NDGA produced a decrease in peroxidase secretion (peroxidase secreted: basal: 0.44 +/- 0.03; NDGA 2.5 x 10(-6) M: 0.20 +/- 0.02; prostaglandins E2 (PGE2) 10(-7)M: 1.32 +/- 0.5; NDGA + PGE2: 0.46 +/- 0.035), an effect that was exerted by the inhibition on prostaglandins synthesis.     

Effects of trapidil and nitroglycerin on coronary circulation in conscious dogs

Effects of trapidil and nitroglycerin (glyceryl trinitrate) on coronary blood flow or epicardial coronary diameter were studied in conscious dogs, instrumented with a Doppler flow probe or a pair of ultrasonic dimension crystals on the left circumflex coronary artery. Bolus intravenous injections of trapidil (0.5, 1.0 and 2.0 mg/kg i.v.) increased coronary blood flow, dose-dependently, such being comparable at the peak value seen with nitroglycerin (5, 10 and 20 micrograms/kg i.v.). Coronary blood flow following the intravenous administration of trapidil or nitroglycerin increased biphasically and returned to pre-drug levels in 1.4 +/- 0.2 min (trapidil 1 mg/kg i.v.) or 1.0 +/- 0.1 min (nitroglycerin 10 micrograms/kg i.v.), while the mean coronary diameter increased monophasically and approached the control level 5 min after drug administration. The first peak was observed before the maximal decrease in aortic pressure and the second peak was associated with concomitant increases in heart rate and myocardial contractility induced by a sudden hypotension. Propranolol, a beta-blocker, did not modify the initial peak but attenuated markedly the second peak (P less than 0.05) in case of trapidil (1.0 mg/kg i.v.) or nitroglycerin (10 micrograms/kg i.v.), which corresponded with reduced changes in reflex tachycardia and positive inotropism. Therefore, the direct effects of trapidil and nitroglycerin on coronary circulation of conscious dogs are an initial transient dilatation of the resistance vessels followed by a continuation of the dilatation of the conductance coronary vessels.     

Endogenous prostaglandins limit angiotensin-II induced regional vasoconstriction in conscious rats

In conscious rats, we tested the hypothesis that prostaglandins attenuate regional vasoconstriction caused by acute infusion of angiotensin II. Mean arterial pressure, regional blood flow, and vascular conductance in response to 2-minute infusions of 0.05 or 1 microg/kg/min Ang II were assessed before and during indomethacin treatment (5 mg/kg). Effects of the lower dose of Ang II (n=8) on regional blood flow were not altered by indomethacin, but conductance in the kidney (2.98+/-0.35 vs. 2.19+/-0.32), stomach (1.15+/-0.13 vs. 0.83+/-0.13), and white gastrocnemius muscle (0.11+/-0.02 vs. 0.07+/-0.01 mL/min/100g/mm Hg) were reduced. Changes in conductance were not seen in the pancreas or spleen. In response to the higher dose of Ang II (n=7), indomethacin reduced blood flow in the kidney, red and white gastrocnemius, and soleus muscles. Reductions in conductance were found in the kidney, stomach and small intestine, and in the red and white gastrocnemius, and soleus muscles (2.27+/-0.9 vs. 1.79+/-0.14, 0.44+/-0.07 vs. 0.27+/-0.03, 0.68+/-0.11 vs. 0.60+/-0.07, 0.43+/-0.08 vs. 0.16+/-0.03, 0.10+/-0.02 vs. 0.05+/-0.01, and 0.26+/-0.03 vs. 0.15+/-0.02 mL/min/100g, respectively). No changes occurred in the pancreas and spleen. Indomethacin had no effect on baseline blood flow or conductance in any of these organs. These results suggest that prostaglandins attenuate vasoconstriction caused by Ang II in a manner that is organ-specific and dependent on the dose of Ang II. Consequently, prostaglandins may limit vasoconstriction and potential ischemia caused by elevated levels of this hormone.