Responsiveness of microcirculation and local cold vasodilation to capsaicin in the intact and chronically denervated canine tongue

Lingual blood flow and its distribution were determined at rest and in response to local cooling of the tongue (32 degrees C) in 6 anaesthetized, paralyzed and artificially ventilated dogs before and after two intraarterial (i.a.) injections of capsaicin (2.5 mg) at an interval of about 40 min. In 3 dogs, the same protocol was performed after degeneration of the chorda-lingual and glossopharyngeal nerves due to prior transection. In general the first i.a. injection of capsaicin resulted in a marked and the second injection in a smaller decrease of lingual blood flow. Local cooling of the tongue induced significant increases in lingual blood flow before as well as after capsaicin treatment, regardless of whether sensory innervation was intact or degenerated. In both the untreated and capsaicin treated dogs the increase in lingual blood flow during local cooling of the tongue was solely due to an increase in blood flow through the arteriovenous anastomoses, while blood flow through the capillaries of the mucosa and muscles even decreased. The findings suggest that capsaicin-induced vasoconstriction of the tongue vessels is due to a direct effect on vascular receptors. It is further suggested that cold vasodilatation of the canine tongue is not mediated by axon collaterals releasing substance P. Direct thermal effects on the intramural ganglia and the postganglionic vasomotor efferents innervating the AVAs, or on AVAs basal tone itself are suggested as the underlying mechanism.     

Mechanism of adenosine-induced vasodilation in rat diaphragm microcirculation

The mechanism of adenosine-induced vasodilation in rat diaphragm microcirculation was investigated using laser Doppler flowmetry. Adenosine (10(-5), 3.2 x 10(-5), and 10(-4) M), the nonselective adenosine agonist 5'-N-ethylcarboxamido-adenosine (NECA) (10(-8)-10(-7) M), the specific A(2A) agonist 2-p-(2-carboxyethyl)phenyl-amino-5'-N-ethyl carboxamidoadenosine (CGS-21680) (10(-8)-10(-7) M), and the adenosine agonist with higher A(1)-receptor affinity, R-N(6)-phenylisopropyladenosine (R-PIA) (10(-7), 3.2 x 10(-7), and 10(-6) M) elicited a similar degree of incremental increase of microcirculatory flow in a dose-dependent manner. The ATP-dependent potassium (K(ATP)) channel blocker glibenclamide (3.2 x 10(-6) M) significantly attenuated the vasodilation effects of these agonists. Adenosine-induced vasodilation could be significantly attenuated by the nonselective adenosine antagonist 8-(p-sulfophenyl)-theophylline (3 x 10(-5) M) or the selective A(2A) antagonist 4-(2-[7-amino-2-(2-furyl)[1,2, 4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl) phenol (ZM-241385, 10(-6) M), but not by the selective A(1) antagonist 8-cyclopentyl-1, 3-dipropylxanthine (5 x 10(-8) M). Adenylate cyclase inhibitor N-(cis-2-phenyl-cyclopentyl) azacyclotridecan-2-imine-hydrochloride (MDL-12330A, 10(-5)M) effectively suppressed the vasodilator response of adenosine and forskolin. These results suggest that adenosine-induced vasodilation in rat diaphragm microcirculation is mediated through the stimulation of A(2A) receptors, which are coupled to adenylate cyclase activation and opening of the K(ATP) channel.     

Lack of renal vasodilation during intrarenal infusion of synthetic atriopeptin II in conscious intact SHR

Synthetic atriopeptin II (APII) was infused directly into the right renal artery of intact conscious SHR at rates of 0.25-1 microgram/kg/min, while simultaneously measuring blood pressure (MAP) and selected regional blood flows. The latter were measured using chronically implanted miniaturized Doppler flowprobes that were placed on the right and left renal artery, superior mesenteric artery and abdominal aorta. The effects of intrarenally (i.r.) infused APII on these vascular beds were compared to the effects of the same amounts of APII given intravenously (i.v.) in the same SHR. I.r. and i.v. infusions caused similar reductions of MAP and all four blood flows. Also effects on calculated resistances were comparable, implying that resistance increased most in the mesentery and least in the two kidneys. The increase in right renal resistance during i.v. infusions of APII was not different from the effect during i.r. infusions. Also, during i.r. infusions into the right kidney, effects on the left and right kidney were not different. Our observations suggest that synthetic APII has no direct effects on the renal vasculature of intact conscious SHR.     

Vascular and neural mechanisms of ACh-mediated vasodilation in the forearm cutaneous microcirculation.

The relative contribution of endothelial vasodilating factors to acetylcholine (ACh)-mediated vasodilation in the forearm cutaneous microcirculation is unclear. The aims of this study were to investigate the contributions of prostanoids and cutaneous C fibers to basal cutaneous blood flow (CuBF) and ACh-mediated vasodilation. ACh was iontophoresed into the forearm, and cutaneous perfusion was measured by laser-Doppler flowmetry. To inhibit the production of prostanoids, four doses of acetylsalicylic acid (ASA; 81, 648, 972, and 1,944 mg) were administered orally. Cutaneous nerve fibers were blocked with topical anesthesia. Cyclooxygenase inhibition did not change basal CuBF or endothelium-mediated vasodilation to ACh. In contrast, ASA (972 and 1,944 mg) significantly reduced the C-fiber-mediated axon reflex in a dose-dependent fashion. Blockade of C-fiber function significantly reduced axon reflex-mediated vasodilation but did not affect basal CuBF or endothelium-dependent vasodilation. The findings suggest that prostanoids do not contribute significantly to basal CuBF or endothelium-dependent vasodilation in the forearm microcirculation. In contrast, prostanoids are mediators of the ACh-provoked axon reflex.     

Relationship between dietary copper concentration and acetylcholine-induced vasodilation in the microcirculation of rats

Dietary copper deficiency has been shown to significantly reduce acetylcholine (Ach)-induced vascular smooth muscle relaxation. The current study was designed to examine the relative relationship between dietary copper and the vasodilator response to Ach in the microcirculation of the rat. Male weanling rats were fed a purified basal diet supplemented with 6.0, 3.0, 1.5 or 0.0 microg Cu/g diet for 4 weeks to provide an adequate, two marginal, and deficient intakes of dietary copper. Arteriole dilation in response to increasing concentrations of acetylcholine (10(-7) to 10(-4) M) was measured in the in vivo cremaster muscle microcirculation for each dietary group. Liver copper and both aortic and erythrocyte Cu,Zn-SOD activity were used as indices of systemic copper status. Dilation to the increasing concentrations of Ach was only different in the 0 microg Cu supplemented group compared to the copper-adequate control values. However, the combined results showed an exponential increase in 10(-5) M Ach-induced vasodilation as liver copper concentration increases from 0 microg Cu/g dry wt. This relationship suggests that dilation is attenuated at liver Cu concentrations below 5 microg/g dry wt. The results indicate that Ach-induced vasodilation is copper-dependent but that the pathway is not very sensitive to short-term marginal restriction of copper intake.     

Kinetic measurements of gas exchange in the intact pulmonary microcirculation.

The kinetics of gas exchange are monitored in an isolated perfused lung preparation contained within a plethysmograph. The lungs are perfused with buffer, and there is no gas exchange until a 2.0-ml bolus of reactant is injected into the perfusion system. Subsequent gas exchange produces a pressure transient that is related to the corresponding volume of exchanged gas. The observed rate of volume change is the result of two separate processes: 1) the rate of gas exchange during transit through the capillary bed and 2) the distribution of vascular transit times between the point of injection and the capillary bed. The latter is assessed by a control injection containing a dissolved inert gas that is liberated in the alveoli as the bolus enters the capillary bed. Analysis of the experimental curves permits the separation of these two processes. A model of exchange kinetics indicates that this method has the capability of measuring kinetic events occurring during gas exchange in the microcirculation under physiological conditions.     

Oxygen transport and peripheral microcirculation in long-term diabetes

The purpose of this investigation was to evaluate the impact of long-term diabetes on muscle blood flow (MBF) and oxygen transport (vO2) during exercise. Twelve male patients (58 +/- 8 years, mean +/- SD), with at least a 10-year history of diabetes controlled by insulin, and seven age-matched controls (56 +/- 5 years, mean +/- SD) participated in this study. No patient had been clinically diagnosed as having peripheral vascular disease, and on the average resting ankle/arm systolic blood pressure ratios were normal. Following a baseline period, 5 min of cycle ergometer exercises at 75 W were performed in the upright position and, after 1-hr recovery, in the supine position. Continuous vO2 was determined via breath-by-breath analysis. MBF was measured in the vastus lateralis (VL) and tibialis anterior (TA) by 133Xe clearance. In the erect position, the diabetic group (compared with the control group, respectively) exhibited significantly (P less than 0.05) lower exercise MBF [ml. (100 g.min)-1] in both VL (19 +/- 2.5 vs 30.9 +/- 2) and TA (13.7 +/- 2 vs 22.0 +/- 4), a lower steady-state VO2 (1.3 +/- 0.3 vs 1.7 +/- 0.2 liters.min-1) during exercise including the values in the last 15 sec of exercise, and greater accumulation of blood lactate (35 +/- 2 vs 22.0 +/- 2 mg/100 ml). The same trends in the data were observed during supine exercise; however, the blood pressure of the diabetics was significantly elevated during exercise when compared with that of controls. The reduced exercise MBF in the TA and VL demonstrated that impaired microvascular flow, without clinically overt peripheral vascular disease, in long-term diabetics leads to reduced oxygen delivery and exercise tolerance.     

Role of pertussis toxin-sensitive G protein in metabolic vasodilation of coronary microcirculation

We have previously demonstrated that pertussis toxin (PTX)-sensitive G protein (G(PTX)) plays a major role in coronary microvascular vasomotion during hypoperfusion. We aimed to elucidate the role of G(PTX) during increasing metabolic demand. In 18 mongrel dogs, coronary arteriolar diameters were measured by fluorescence microangiography using a floating objective. Myocardial oxygen consumption (MVO(2)) was increased by rapid left atrial pacing. In six dogs, PTX (300 ng/ml) was superfused onto the heart surface for 2 h to locally block G(PTX). In eight dogs, the vehicle (Krebs solution) was superfused in the same way. Before and after each treatment, the diameters were measured during control (130 beats/min) and rapid pacing (260 beats/min) in each group. Metabolic stimulation before and after the vehicle treatment caused 8.6 +/- 1. 8 and 16.1 +/- 3.6% dilation of coronary arterioles <100 microm in diameter (57 +/- 8 microm at control, n = 10), respectively. PTX treatment clearly abolished the dilation of arterioles (12.8 +/- 2. 5% before and 0.9 +/- 1.6% after the treatment, P < 0.001 vs. vehicle; 66 +/- 8 microm at control, n = 11) in response to metabolic stimulation. The increases in MVO(2) and coronary flow velocity were comparable between the vehicle and PTX groups. In four dogs, 8-phenyltheophylline (10 microM, superfusion for 30 min) did not affect the metabolic dilation of arterioles (15.3 +/- 2.0% before and 16.4 +/- 3.8% after treatment; 84.3 +/- 11.0 microm at control, n = 8). Thus we conclude that G(PTX) plays a major role in regulating the coronary microvascular tone during active hyperemia, and adenosine does not contribute to metabolic vasodilation via G(PTX) activation.     

Effects of Pseudomonas aeruginosa endotoxin on vasodilation in the intact spinotrapezius muscle.

The purpose of this study was to determine whether short-term exposure to clinically relevant concentrations of Pseudomonas aeruginosa lipopolysaccharide (LPS) impairs vasoreactivity of resistance arterioles in the intact spinotrapezius muscle microcirculation and, if so, to determine the mechanisms mediating this response. Using intravital microscopy, we found that 60-min suffusion of P. aeruginosa LPS (0.03-3.0 microg/ml) on the in situ hamster spinotrapezius muscle elicited an immediate, profound, and prolonged concentration-dependent vasodilation (P < 0.05). This response was reversible once suffusion of P. aeruginosa LPS was stopped. Pretreatment with N(G)-nitro-L-arginine methyl ester (10.0 microM), a nonselective nitric oxide (NO) synthase inhibitor, but not N(G)-nitro-D-arginine methyl ester, abrogated P. aeruginosa LPS-induced vasodilation and elicited a small, albeit significant, vasoconstriction. Indomethacin had no significant effects on P. aeruginosa LPS-induced responses. P. aeruginosa LPS had no significant effects on acetylcholine- and nitroglycerin-induced vasodilation in the spinotrapezius muscle. Collectively, these data indicate that short-term exposure to clinically relevant concentrations of P. aeruginosa LPS evokes an immediate, potent, prolonged, and reversible NO-dependent, prostaglandin-independent vasodilation in skeletal muscles in vivo. We suggest this response could play an important role in the pathophysiology of the profound vasomotor dysfunction observed in the peripheral circulation of patients with P. aeruginosa sepsis syndrome.     

Estimation of the light distribution between photosystems I and II in intact wheat leaves by fluorescence and photoacoustic measurements

Usisng intact leaves, the extent of the decrease in chlorophyll a fluorescenece caused by the addition of continuous 710 nm light superimposed on modulated (20 Hz) 550 nm light was used to determine the distribution of this absorbed light between photosystems I () and II (). The F_o and F_m levels, which defined the total variable fluorescenece, were taken as equal to those obtained with excess 710 nm light and with saturating blue-green light, respectively.An analogous procedure was used with a photoacoustic detector, saturating white light defining a base line for oxygen yield, the levels with an without 710 nm light being used to define and respectively.The two methods gave similar values for the distribution of light between the two photosystems for the experimental conditions used, averaging 0.55 for a range of Triticum genotypes and Brachypodium sylvaticum grown in high or low light.     

Chiral separation based on immobilized intact and fragmented cellobiohydrolase II (CBH II): A comparison with cellobiohydrolase I (CBH I)

Intact and fragmented cellobiohydrolase II (CBH II) were immobilized to silica and used as chiral stationary phases (CSPs) for liquid chromatographic separations of enantiomers. Both acidic and basic chiral compounds could be resolved into their enantiomers on these phases. The enantioselectivities obtained on intact CBH II and its core were almost equivalent. Comparisons were also made with CBH I silica. It was found that the new materials show quite different chiral and chromatographic properties. The enzymatic activity of the CBH II in free solution was influenced by alprenolol and mexiletine, both separated on the corresponding CSP. It indicates that the sites for catalysis and for chiral recognition overlap. © 1995 Wiley-Liss, Inc.     

Resolution of the Photosystem I and Photosystem II contributions to chlorophyll fluorescence of intact leaves at room temperature

Green leaves illuminated with photosynthetically active light emit red fluorescence, whose time-dependent intensity variations reflect photosynthetic electron transport (the Kautsky effect). Usually, fluorescence variations are discussed by considering only the contribution of PSII-associated chlorophyll a, although it is known that the fluorescence of PSI-associated chlorophyll a also contributes to the total fluorescence [Aust. J. Plant Physiol. 22 (1995) 131]. Because the fluorescence emitted by each photosystem cannot be measured separately by selecting the emission wavelength in in vivo conditions, the contribution of PSI to total fluorescence at room temperature is still in ambiguity. By using a diode array detector, we measured fluorescence emission spectra corresponding to the minimal (F(O)) and maximal (F(M)) fluorescence states. We showed that the different shapes of these spectra were mainly due to a higher contribution of PSI chlorophylls in the F(O) spectrum. By exciting PSI preferentially, we recorded a reference PSI emission spectrum in the near far-red region. From the F(O) and F(M) spectra and from this PSI reference spectrum, we derived specific PSI and PSII emission spectra in both the F(O) and F(M) states. This enables to estimate true value of the relative variable fluorescence of PSII, which was underestimated in previous works. Accurate separation of PSI-PSII fluorescence emission spectra will also enable further investigations of the distribution of excitation energy between PSI and PSII under in vivo conditions.     

Photosynthetic energy storage of Photosystems I and II in the spectral range of photosynthetically active radiation in intact sugar maple leaves

The relative activity of Photosystems (PS) I and II in the spectral range between 400 and 720 nm was studied by measuring photosynthetic energy storage (ES) of an intact sugar maple leaf using photoacoustic spectroscopy. ES, determined with a modulated (80 Hz) monochromatic light beam in the presence of saturating intensity of background non-modulated white light, indicated the total energy stored by both photosystems (ES_T). Using background far-red light, ES of PS I (ES_{PS I}) was quantified. ES_{PS II} was derived from ES_T-ES_{PS I}. ES_T dependence on intensity and wavelength of modulated light was studied at 470, 560, 640 and 680 nm. ES_T was maximum in red light and minimum in blue light. It decreased with an increase in modulated light intensity. The ratio ES_{PS II}/ES_{PS I}, measured at 640 nm, remained nearly constant with an increase in modulated light intensity. The relative quantum yield of ES_T spectrum showed two peaks around 610 and 660 nm, and declined sharply after 680 nm, revealing a clear red drop. ES_{PS I} spectrum presented peaks around 610 and 670 nm, and a minimum between 440 and 470 nm. ES_{PS I} was observed beyond 700 nm up to 720 nm, indicating the energy stored by cyclic electron transport. ES_{PS II} spectrum showed broad peaks, around 460, 490, 600 and 660 nm, and a shoulder between 530 and 560 nm. ES_{PS II} was always higher than ES_{PS I} between 400 and 690 nm and reached zero around 700 nm.Abbreviations ES energy storage - ES_{PS I} energy storage of PS I - ES_{PS II} energy storage of PS II - ES_T energy storage of PS I and PS II - PA photoacoustic - PS I Photosystem I - PS II Photosystem II - Q_m PA signal in the absence of any background light - Q_ma PA signal in the presence of background white light - Q_mfrl PA signal in the presence of background far-red light - S/N signal to noise     

Comparative characteristics of microcirculation disorders in the serous membrane of the uterus in intact and pregnant rats treated with oxytocin

The studies were performed on intact and pregnant rats after the administration of 10(-2) U of oxytocin. Oxytocin-induced increase in spasm duration and microvascular dilatation was noted both in intact and pregnant animals. However, in the former case spasm duration was markedly increased, while in the latter case there was an equal increase both in spasm duration and dilatation. Dilatation phase was characterized by an increase in microvessel diameter, as compared to the initial one. Maximal spasm phase after oxytocin administration in intact rats was characterized by hemostasis. In pregnant animals blood flow was delayed, with no marked spasm present. The initial level of microcirculation was restored 38-40 minutes later.     

Altered peripheral noradrenergic activity in intact and sinoaortic denervated Dahl rats

Development of salt-induced hypertension in Dahl salt-sensitive (S) rats is dependent on sympathetic overactivity which may be partially related to arterial baroreflex dysfunction and, therefore, is regionally selective. Our first experiment was designed to determine which regions have elevated sympathetic activity in Dahl S compared with Dahl salt-resistant (R) rats. Weanling (4-week-old) female Dahl R and S rats were fed low or high salt diets (0.13% and 8% NaCl) until 10 weeks of age. Norepinephrine (NE) synthesis was blocked with alpha-methyl-p-tyrosine, and the fractional decline of NE concentration was measured in various tissues. Dahl S rats with increases in both arterial pressure and left ventricular weight demonstrated increased NE turnover in the sinoatrial node, the atrial appendages, the cardiac ventricles, and the renal cortex. In all of these tissues except the cardiac ventricle, increases were associated with high salt intake. Our second experiment was designed to test if arterial baroreflex dysfunction could account for regional increases in sympathetic activity. Separate groups of Dahl R and S rats fed high salt were subjected to either sham surgery or sinoaortic baroreceptor denervation 1 week prior to turnover determinations. Sinoaortic baroreceptor denervation abolished differences in NE turnover between salt-fed Dahl R and S rats in the cardiac sinoatrial node and the atrial appendages, but not in the cardiac ventricles and the renal cortex. Sinoaortic baroreceptor denervation also abolished differences between salt-fed Dahl S and R rats in the spleen but not the duodenum.(ABSTRACT TRUNCATED AT 250 WORDS)     

Insulin action in intact mouse diaphragm I.

Summary The incubation of intact mouse diaphragms with insulin caused a dose and time dependent increase in the independent activity of glycogen synthase in tissue extracts. 2-deoxyglucose (2–10 mm) alone markedly stimulated the conversion of glycogen synthase to the independent activity under conditions in which tissue ATP concentrations were not affected. The incubation of diaphragms with both insulin and 2-deoxyglucose resulted in a greater than additive effect. Insulin stimulated the uptake of 2-deoxyglucose into mouse diaphragms, accumulating as 2-deoxyglucose-6-phosphate. The accumulation of 2-deoxyglucose-6-phosphate correlated well with the increase in the independent activity of glycogen synthase and with the activation of glycogen synthase phosphatase in tissue extracts. The uptake of 3-0-methyl glucose was also markedly stimulated by insulin, without affecting the activity of glycogen synthase. Both glucose-6-phosphate and 2-deoxyglucose-6-phosphate stimulated the activation of endogenous glycogen synthase phosphatase activity in muscle homogenates. We conclude that insulin, in addition to its effects in the absence of exogenous sugars, increases the independent activity of glycogen synthase through increased sugar transport resulting in increased concentrations of sugar-phosphates which promote the activity of glycogen synthase phosphatase.Abbreviations GS Glycogen synthase - GS-I Glycogen synthase activity independent of G6P - GS-D Glycogen synthase activity dependent on G6P - G6P Glucose-6-phosphate - ATP Adenosine triphosphate - EDTA Ethylene diamine tetracetic acid - Mops Morpholinopropane sulfonic acid - 2DG 2-Deoxy glucose - 3-0-MG 3-0-Methyl glucose - tricine N-tris(Hydroxymethyl)methyl glycine Enzymes: Glycogen Synthase — UDPGlucose — Glycogen Glucosyl — Transferase (EC 2.4.1.11) J. Larner is an established investigator of the American Diabetes Association.     

Insulin action in intact mouse diaphragm II.

Summary Incubation of intact mouse diaphragms with insulin in the absence of glucose resulted in a rapid inhibition of the subsequent cell-free phosphorylation of endogenous protein substrates in tissue extracts. The phosphorylation of added histone was inhibited to a lesser extent. The inhibition was observed both in the absence and in the presence of added cyclic 35 adenosine monophosphate. Acrylamide gel electrophoresis of phosphorylation products revealed a number of major phosphorylated polypeptides. The phosphorylation of several polypeptides was inhibited following short treatment with insulin. These results represent a novel experimental approach to the elucidation of the mechanism of the action of insulin and are consistent with our hypothesis that the inhibition of protein kinase activities in the tissue may be the first step in this mechanism.Abbreviations Tricine N-tris(hydroxymethyl)methyl glycine - EDTA ethylenediamine tetraacetic acid - EGTA ethyleneglycol-bis-(-amino ethyl ether) N,N¹-tetraacetic acid - Mes 2(N-morpholino)ethane sulfonic acid J. Larner is an established investigator of the American Diabetes Association.     

Effect of T-activin on peripheral immunity organs in intact and thymectomized mice

Morphometry of the spleen, axillary lymph nodes and cytological assay of the bone marrow and peripheral blood were performed in (CBA X C57BL)F1 mice 1, 5, 10 and 15 days after subcutaneous injection of 0.5 microgram T-activin to intact and thymectomized (when adult) mice 2 months after operation. It was demonstrated that in intact animals, injection of T-activin stimulated the whole system of immunogenesis. The time course of plasmatization and the response of the germinative centers differing from that seen during antigen administration suggests that T-activin is not immunogenous, acting as a stimulant of the previous immune responses. The permanent amount of the degenerating cells attests to the lack of the toxic drug effect.