bluuebunny/crossref_metadata_2025 huggingface dataset:
Tyrosine kinase inhibitors and the contractile action of epidermal growth factor – urogastrone and other agonists in gastric smooth muscle
Authors: Yang Song-Gui , Saifeddine Mahmoud , Hollenberg Morley D.
Published: April 2011

We examined the effects of the tyrosine kinase (TK) inhibitors, genistein, and tyrphostin (RG-50864) on the contractile action of epidermal growth factor – urogastrone (EGF–URO), transforming growth factor-α (TGF-α), and other agonists in two smooth muscle bioassay systems (guinea pig gastric longitudinal muscle, LM, and circular muscle, CM). We also studied the inhibition by tyrphostin of EGF–URO stimulated protein phosphorylation in identical smooth muscle strips. The selective inhibition by genistein and tyrphostin of EGF–URO and TGF-α induced contraction, but not of carbachol- and bradykinin-mediated contraction, occurred at much lower concentrations (genistein, <7.4 μM (2 μg/mL); tyrphostin, <20 μM (4 μg/mL)) than those used in previously published studies with these TK inhibitors. In LM tissue, the IC50 values were for genistein 1.1 ± 0.1 μM (0.30 μg/mL; mean ± SEM) and 3.6 ± 0.5 μM (0.74 μg/mL) for tyrphostin, yielding a molar potency ratio (GS:TP) of 1:3 in the longitudinal preparation. In CM tissue, the IC50 values were 3.0 ± 0.3 μM (0.81 μg/mL) for genistein and 2.4 ± 0.2 μM (0.49 μg/mL) for tyrphostin, yielding a molar potency ratio (GS:TP) of 1.0:0.8 in the circular strips. The inhibition by genistein and tyrphostin of EGF–URO and TGF-α mediated contraction was rapid (beginning within minutes) and was reversible upon washing the preparations free from the enzyme inhibitors. In intact tissue strips studied under bioassay conditions, tyrphostin (40 μM) also blocked EGF–URO triggered phosphorylation of substrates detected on Western blots using monoclonal antiphosphotyrosine antibodies. In contrast with the results with EGF–URO, genistein at 7.4 μM (2 μg/mL) and tyrphostin at 20 μM (4 μg/mL) had no effect on either bradykinin (1 μM) or carbachol (1 μM) stimulated contraction in both LM and CM preparations. However, the contractile action of prostaglandin F2α (1 μM) was partially inhibited by genistein (7.4 μM), up to 10 ± 5% in the LM preparation and 35 ± 3% in the CM preparation (p < 0.05, LM vs. CM preparations) but not by tyrphostin (20 μM). Additionally, at the same concentrations, these tyrosine kinase inhibitors completely blocked angiotensin II mediated contraction in the LM preparation and partially inhibited contraction (43 ± 3%, mean ± SEM) in the CM preparation. The data suggest that tyrosine kinase activity plays an important role in EGF–URO induced contraction of gastric smooth muscle. Additionally, our results suggest that tyrosine kinase activity may also play a role in the signal transduction pathways by which smooth muscle contraction is induced by agonists such as angiotensin II and prostaglandin F2α which are known to act via a G-protein receptor coupled pathway.Key words: smooth muscle, tyrosine kinase, growth factor, angiotensin.
Dicarboxylate transport in renal basolateral and brush-border membrane vesicles
Authors: Kim Yong Keun , Jung Jin Sup , Lee Sang Ho
Published: April 2011

Characteristics of succinate transport were determined in basolateral and brush-border membrane vesicles (BLMV and BBMV, respectively) isolated in parallel from rabbit renal cortex. The uptake of succinate was markedly stimulated by the imposition of an inwardly directed Na+ gradient, showing an "overshoot" phenomenon in both membrane preparations. The stimulation of succinate uptake by an inwardly directed Na+ gradient was not significantly affected by pH clamp or inhibition of Na+–H+ exchange. The Na+-dependent and -independent succinate uptakes were not stimulated by an outwardly directed pH gradient. The Na dependence of succinate uptake exhibited sigmoidal kinetics, with Hill coefficients of 2.17 and 2.38 in BLMV and BBMV, respectively. The Na+-dependent succinate uptake by BLMV and BBMV was stimulated by a valinomycin-induced inside-negative potential. The Na+-dependent succinate uptake by BLMV and BBMV followed a simple Michaelis–Menten kinetics, with an apparent Km of 22.20 ± 4.08 and 71.52 ± 0.14 μM and a Vmax of 39.0 ± 3.72 and 70.20 ± 0.96 nmol/(mg·min), respectively. The substrate specificity and the inhibitor sensitivity of the succinate transport system appeared to be very similar in both membranes. These results indicate that both the renal brush-border and basolateral membranes possess the Na+-dependent dicarboxylate transport system with very similar properties but with different substrate affinity and transport capacity.Key words: dicarboxylate transport, brush border membrane, basolateral membrane, inhibitors, rabbit kidney.
Muscle triglyceride metabolism during exercise
Authors: Gorski Jan
Published: April 2011

Skeletal muscle cell contains a considerable amount of triglycerides. The amount stored depends on the animal species as well as on muscle fiber composition. It is well documented that triglycerides in the fast-twitch red muscle and to a lesser extent in the slow-twitch muscle, but not those in the fast-twitch white muscle, are mobilized during prolonged exercise. Yet, little is known about the regulation of the metabolism of muscle triglycerides either at rest or during exercise. This is well reflected by the fact that an enzyme responsible for the hydrolysis of muscle triglycerides has not been identified. Mobilization of muscle triglycerides during exercise seems to be under both adrenergic and noradrenergic control. Accumulation of lactic acid and reduction in muscle pH are likely to be strong inhibitors of muscle triglyceride lipolysis. Reduction of carbohydrate availability accelerates mobilization of muscle triglycerides during exercise. The relationship between the plasma free fatty acids and muscle triglyceride metabolism seems to be complex. It has been proposed that most free fatty acids entering the muscle cell is esterified before being oxidized, but this is arguable for contracting skeletal muscles. It is suggested that most free fatty acids entering contracting high oxidative myocytes are transported directly to the mitochondria. A much lesser portion is likely esterified. It is proposed that triglycerides stored in the contracting muscle cell are mobilized when the delivery of the blood-borne-free fatty acids to the mitochondria is insufficient.Key words: muscle triglycerides, free fatty acids, exercise.
Regulation of skeletal muscle metabolism by enzyme binding
Authors: Parkhouse Wade S.
Published: April 2011

The random diffusion mechanism is usually assumed in analyzing the energetics of specific pathways despite the findings that enzymes associate with each other and (or) with various membranous and contractile elements of the cell. Successive glycolytic enzymes have been shown to associate in the cytosol as enzyme complexes or bind to the thin filaments. Furthermore, the degree of glycolytic enzyme interactions have been shown to change with altered rates of carbon flux through the pathway. In particular, the proportions of aldolase, phosphofructokinase, and glyceraldehyde phosphate dehydrogenase bound to the contractile proteins have been found to increase with increased rates of glycolysis. In addition, decreasing pH and ionic strength are also associated with an increase in glycolytic enzyme interactions. The kinetics displayed by interacting enzymes generally serve to enhance their catalytic efficiencies. The associations of the glycolytic enzymes serve to enhance metabolite transfer rates, increase the local concentrations of intermediates, and provide for regulation of activity via effectors. Therefore these interactions provide an additional mechanism for regulating glycolytic flux in skeletal muscle.Key words: glycolysis, skeletal muscle, enzymes, binding.
Anaerobic metabolism in human skeletal muscle during short-term, intense activity
Authors: Spriet Lawrence L.
Published: April 2011

The ability of human skeletal muscle to provide anaerobically derived ATP during short-term, intense activity is examined. The paper emphasizes the information obtained from direct measurements of substrates, intermediates, and products of the pathways in muscle that provide anaerobically derived ATP. The capacity of muscle to provide ATP via anaerobic pathways is ~370 mmol/kg dry muscle (dm) during dynamic exercise lasting ~3 min. Anaerobic glycolysis provided ~80%, phosphocreatine (PCr) degradation ~16%, and depletion of the ATP store ~4% of the total ATP provided. When the blood flow to the working muscles is reduced or occluded, the anaerobic capacity decreases to ~300 mmol/kg dm. This reduction is due to a lower glycolytic capacity associated with an inability to remove lactate from the muscles. Directly measured maximal rates of anaerobically derived ATP provision from PCr degradation and glycolysis during intense muscular activity are each ~9–10 mmol∙kg−1 dm∙s−1. Evidence suggests that both of these pathways are activated instantaneously at the onset of maximal activity. Spring training does little to the capacity or rates of the pathways, although a 10–20% increase in glycolytic ATP provision has been reported. The only study comparing direct and indirect estimates of the anaerobic capacity in humans suggests that O2 deficit measured at the mouth accurately predicts the anaerobic capacity of a single muscle group and that O2 debt does not. There are many unresolved issues regarding the capacity of the PCr and glycogenolytic–glycolytic systems to provide ATP during short-term intense muscular activity in humans. Considerable effort is now being directed to understanding the in vivo regulation of the regulatory and flux-generating glycogenolytic enzyme, phosphorylase.Key words: glycogenosis, glycolysis, phosphocreatine, ATP, sprinting.
Is there a seasonal variation in mucus transport and nutrient absorption in the leopard frog?
Authors: Rubin Bruce K. , Cheeseman Chris I. , Gourishankar Sita , King Malcolm
Published: April 2011

We postulated that as a hibernating species, frogs might have variable demands for nutrients at different seasons of the year and that this must be reflected in seasonal variations of physiologic processes related to nutrient transport and absorption. We examined the rate of mucus transport on the ciliated palate and the movement of nutrients across the intestinal lumen of leopard frogs, Rana pipiens. Mucus transport on the frog palate was strongly influenced by season, with maximal transport occurring in late June (Julian day 178, p = 0.0001; r = 0.58). This increased transport rate was associated with a summertime increase in mucus recoil (lower tangent δ) and a decrease in mucus hydration (increase in percent solids composition). Intestinal transport of leucine, lysine, and galactose did not appear to exhibit seasonal variability. These data suggest that different mechanisms may operate in determining seasonal variability in physiologic responses.Key words: mucociliary clearance, mucus viscoelasticity, intestinal absorption, Anura, seasonal variation.
Fast Na+ channels in smooth muscle from pregnant rat uterus
Authors: Sperelakis Nicholas , Inoue Yoshihito , Ohya Yusuke
Published: April 2011

Smooth muscle cells normally do not possess fast Na+ channels, but inward current is carried through two types of Ca2+ channels: slow (L type) Ca2+ channels and fast (T type) Ca2+ channels. Whole-cell voltage clamp was done on single smooth muscle cells isolated from the longitudinal layer of the 18-day pregnant rat uterus. Depolarizing pulses, applied from a holding potential of −90 mV, evoked two types of inward current, fast and slow. The fast inward current decayed within 30 ms, depended on [Na]o, and was inhibited by tetrodotoxin (TTX) (K0.5 = 27 nM). The slow inward current decayed slowly, was dependent on [Ca]o (or Ba2+), and was inhibited by nifedipine. These results suggest that the fast inward current is a fast Na+ channel current and that the slow inward current is a Ca2+ slow channel current. A fast-inactivating Ca2+ channel current was not evident. We conclude that the ion channels that generate inward currents in pregnant rat uterine cells are TTX-sensitive fast Na+ channels and dihydropyridine-sensitive slow Ca2+ channels. The number of fast Na+ channels increased during gestation. The averaged current density increased from 0 on day 5, to 0.19 on day 9, to 0.56 on day 14, to 0.90 on day 18, and to 0.86 pA/pF on day 21. This almost linear increase occurs because of an increase in the fraction of cells that possess fast Na+ channels. The Ca2+ channel current density was also higher during the latter half of gestation. These results indicate that the fast Na+ channels and Ca2+ slow channels in myometrium become more numerous as term approaches, and we suggest that the fast Na+ current may be involved in spread of excitation. Isoproterenol (β-agonist) did not affect either ICa(s) or INa(f), whereas Mg2+ (K0.5 = 12 mM) and nifedipine (K0.5 = 3.3 nM) depressed ICa(s). Oxytocin had no effect on INa(f) and actually depressed ICa(s) to a small extent. Therefore, the tocolytic action of β-agonists cannot be explained by an inhibition of ICa(s), whereas that of Mg2+ can be so explained. The stimulating action of oxytocin on uterine contractions cannot be explained by a stimulation of ICa(s).Key words: sodium current, fast sodium current, calcium currents, myometrial smooth muscle cells, pregnant uterine muscle.
The biophysics and biochemistry of smooth muscle contraction
Authors: Stephens N. L. , Seow C. Y. , Halayko A. J. , Jiang H.
Published: April 2011

In this review the biophysics and biochemistry of smooth muscle contraction are dealt with. We describe a new model for the study of bronchial smooth muscle, which facilitates study of cellular contractile mechanisms. A new concept emerging is that study of steady-state mechanical parameters such as maximal isometric force (Po) velocity is inadequate because two types of crossbridges (normally cycling (NBR) and latch) seem to be sequentially active during smooth muscle contraction. Thus quick-release techniques are required to characterize the force–velocity properties of the two types of bridges. Pathophysiological processes that affect the muscle's shortening ability seem to affect the early NBRs only. With respect to maximal shortening capacity of the smooth muscle, the role of loading is very important. The differences between isotonic, elastic, and viscous loading are considerable. Ultimately, the time course and magnitude of loading should exactly resemble that operative in vivo. Once again, it is the characteristic of loading in the early phase of contraction that is crucial, as most of the shortening in smooth muscle occurs early in the contraction. While the maximum force developed by smooth muscle per unit cross-sectional area is the same as for striated muscle, the velocity is 50 times less. The properties of the series and parallel elastic elements of smooth muscle are described. The latter, when in compression mode, acts as an internal resistance to shortening and probably limits it. Isotonic relaxation has therefore not been studied in smooth muscle. We have developed a shortening parameter that is independent of the load on the muscle and of the initial length of the muscle's contractile element. We report the novel observation that isotonically relaxing smooth muscle reactivates itself, resulting in terminal slowing of the relaxation process. With respect to the biochemistry of smooth muscle contraction, contractile (actin isoforms, myosin heavy and light chains and their isoforms), regulatory (calmodulin–4 Ca2+, myosin light chain kinase, myosin light chain and its phosphorylation, tropomyosin, caldesmon, and calponin), and cytoskeletal (chiefly desmin and vimentin) proteins are discussed. While the kinase activates the contractile system, caldesmon and calponin modulate the activity downward. The cytoskeletal proteins desmin, vimentin, and α-actinin could constitute the muscle cell's internal resistor.Key words: smooth muscle mechanics, force–velocity, smooth muscle, internal resistor, smooth muscle retardation, contractile proteins, regulatory proteins.
Cerebral vascular changes associated with hemorrhagic stroke in hypertension
Authors: Smeda John S.
Published: April 2011

There are a number of alterations that protect the cerebrovasculature from hemorrhagic stroke development during hypertension. The upper limit of cerebral blood flow autoregulation is shifted to higher blood pressure levels; this allows a constant blood flow to be maintained during hypertension. Studies we have performed have indicated that the middle cerebral arteries (MCA) of Wistar–Kyoto stroke-prone spontaneously hypertensive rats (spSHR) lose their ability to constrict in response to elevations in transmural pressure. The decline in such function precedes stroke development and totally disappears at an age where there is a 100% mortality from stroke. Prior to stroke development, spSHR also develop uremic conditions and signs of renal failure. The induction of uremia in stroke-resistant SHR (srSHR) via nephrectomy induces these animals to develop stroke. Like prestroke spSHR, prestroke uremic srSHR also have MCA with attenuated pressure-dependent myogenic function. It is hypothesized that the inability to increase vascular resistance in response to elevations in pressure might promote overperfusion of the more distal vasculature leading to cerebral hemorrhage formation. Since uremia promotes bleeding tendencies, such alterations along with the loss of cerebrovascular myogenic function could initiate or aggrevate hemorrhage formation.Key words: stroke-prone SHR, stroke, cerebral vasculature, myogenic response, autoregulation.
Airway smooth muscle contraction at birth: in vivo versus in vitro comparisons to the adult
Authors: Fisher John T.
Published: April 2011

It is clear from the literature that considerable postnatal development occurs in the contractile properties of skeletal and cardiac muscle. Nevertheless, few studies have focused on developmental changes in airway smooth muscle or on the functional capabilities of airway innervation in the newborn. Conclusions about force generation, based on measurements of pulmonary mechanics during stimulation of the vagus nerves, suggest that the newborn possesses a reduced capability to narrow airway diameter relative to the adult. This reduced in vivo response is accompanied by a reduction in maximal force generating capabilities when compared on the basis of force per unit tissue cross-sectional area (stress) in vitro. However, studies of porcine airways suggest that such a finding may simply reflect a reduction in the relative amount of contractile protein (myosin heavy chain) as seen in fetal or preterm smooth muscle. Thus, comparisons based on force normalized per cross-sectional area of myosin alter conclusions from one in which fetal tracheal smooth muscle generates less maximal force than the adult, to one in which the fetal trachea has greater contractile capabilities. Interestingly, comparisons of maximal isometric force in bronchial smooth muscle between different age groups remain unaffected when myosin heavy chain normalization is applied. Finally, there appears to be an age at which maximal force is significantly greater than at any other age, independent of the amount of smooth muscle (determined morphologically), smooth muscle myosin content, or myosin isoform. Whether this enhanced in vitro response is reflected in vivo, or is counteracted by other physiological mechanisms, remains to be seen.Key words: development, airway smooth muscle, lung resistance, force generation, normalization, myosin.
Normal blood flow but lower oxygen tension in diabetes of young rats: microenvironment and the influence of sympathectomy
Authors: Zochodne Douglas W. , Ho Lam T.
Published: April 2011

Studies of rats with experimental streptozotocin (STZ)-induced diabetes at 4 months have identified sciatic nerve trunk oligemia and hypoxia, but it is uncertain how early these abnormalities develop or which develops first. We studied young (4-week-old) rats after 6 or 16 weeks of STZ-induced diabetes (or after citrate buffer injection in controls) by recording multi-fiber conduction in three different nerve territories and by measuring sciatic endoneurial blood flow (NBF) and oxygen tension [Formula: see text] at end point. To evaluate the impact of sympathectomy on this diabetic model, separate animal groups were treated for 5 weeks with guanethidine monosulfate given at the onset of diabetes (group 1, end point 6 weeks) or after 6 weeks of diabetes (group 2, end point 16 weeks). Diabetes was associated with deficits in sensory and motor caudal conduction and increased resistance to ischemic conduction failure (RICF). NBF was comparable to control animals at both time points and was within the published normal range of NBF. In contrast, oxygen tensions were shifted to lower values in diabetic animals. Sympathectomy was associated with blunting of the RICF increase in group 2 but worsened caudal sensory conduction despite evidence of modest improvement in sciatic nerve oxygenation. Our findings support the concept that neuropathy occurs early in diabetes and that hypoxia develops before oligemia. Sympathectomy did not benefit this diabetic model.Key words: diabetic neuropathy, hydrogen clearance, sympathectomy, nerve conduction.
Effect of omega-3 fatty acids, docosahexaenoic and eicosapentaenoic, on norepinephrine-induced contractions
Authors: Engler Mary B.
Published: April 2011

The relaxant responses of the rat thoracic aorta to omega-3 fatty acids, docosahexaenoic and eicosapentaenoic, on norepinephrine- and potassium-induced contractions were investigated. Relaxation was enhanced in vessels contracted with norepinephrine. Docosahexaenoic acid at concentrations as low as 1, 3, and 10 μM evoked significant relaxant responses (15, 23, 30%) in norepinephrine-contracted vessels as compared with responses (5, 9, 12%) in potassium-contracted vessels. Results for eicosapentaenoic acid under similar conditions were 3, 8, and 19% in norepinephrine-contracted vessels and 3, 3, and 8% in potassium-contracted vessels. Pretreatment with eicosapentaenoic (10 μM) or docosahexaenoic acids (1–10 μM) decreased the contractile response to physiologic concentrations of norepinephrine. In the presence of calcium-free medium, the omega-3 fatty acids (1–30 μM) significantly abolished sustained norepinephrine contractions but did not reduce the phasic contractions when incubated prior to norepinephrine contraction. Comparatively, the effects of docosahexaenoic acid were greater than eicosapentaenoic acid. These findings suggest that the relaxant effects of the omega-3 fatty acids are specific to the mode of contraction, i.e., α-adrenoceptor stimuli. This effect may be related to intracellular calcium mechanisms, since both fatty acids reversed norepinephrine-induced sustained contractions in the absence of extracellular calcium.Key words: omega-3 polyunsaturated fatty acids, eicosapentaenoic acid, docosahexaenoic acid, vascular responses, fish oils.
Studies of the inhibitory activity of MK-0591 (3-[1-(4-chlorobenzyl)-3-(t-butylthio)-5-(quinolin-2-yl-methoxy)-indol-2-yl]-2,2-dimethyl propanoic acid) on arachidonic acid metabolism in human phagocytes
Authors: Ménard Luc , Laviolette Michel , Borgeat Pierre
Published: April 2011

We have investigated the inhibitory activity of compound MK-0591 (3-[1-(4-chlorobenzyl)-3-(t-butylthio)-5-(quinolin-2-yl-methoxy)-indol-2-yl]-2,2-dimethyl propanoic acid) on 5-lipoxygenase (5-LO) product synthesis in various human phagocytes stimulated with either the ionophore A23187, opsonized zymosan (OPZ), platelet-activating factor (PAF), or forrnyl-methionyl-leucyl-phenylalanine (fMLP). The lipoxygenase products were analyzed by reversed-phase HPLC. MK-0591 inhibited the formation of 5-hydroxyeicosatetraenoic acid, leukotriene (LT) B4, its Ω-oxidation products, and 6-trans-isomers with IC50 values of 2.8–4.8 nM in A23187-stimulated neutrophils. In these conditions, arachidonic acid at a concentration of 10 μM had no effect on MK-0591 inhibitory activity. In neutrophils stimulated with OPZ, the synthesis of LTB4, its Ω-oxidation products, and 6-trans-isomers was inhibited with IC50 values of 9.5–11.0 nM. MK-0591 inhibited 5-LO product synthesis in A23187-stimulated blood monocytes, eosinophils, and alveolar macrophages with IC50 values of 0.3–0.9, 3.7–5.3, and 8.5–17.3 nM, respectively. In neutrophils primed with granulocyte – macrophage colony-stimulating factor and stimulated with PAF, lipoxygenase product synthesis was inhibited with IC50 values of 7.7–8.7 nM. At the concentration of 1 μM, MK-0591 had no inhibitory effect on 15-lipoxygenase activity in human polymorphonuclear leukocytes, nor on human platelet 12-lipoxygenase and cyclooxygenase. In conclusion, MK-0591 is a very potent and specific inhibitor of 5-LO product synthesis in various types of human phagocytes.Key words: 5-lipoxygenase inhibition, colony-stimulating factor, leukotriene, neutrophil, eosinophil, monocyte, macrophage, platelet-activating factor.
Trophic regulation of action potential in bullfrog sympathetic neurones
Authors: Traynor Philip , Dryden William F. , Smith Peter A.
Published: April 2011

These experiments tested the hypothesis that the normal electrophysiological properties of mature bullfrog sympathetic ganglion (BFSG) neurones are maintained by the retrograde supply of nerve growth factor-like molecules from peripheral target tissues. Maintenance of these cells in explant culture in the absence of nerve growth factor (NGF) for up to 30 days produced electrophysiological changes that resemble those previously shown to accompany axotomy in vivo. These included (i) an increase in action potential (ap) duration (spike width), (ii) a decrease in the amplitude of the afterhyperpolarization (ahp), which follows the ap, and (iii) a rapidly developing decrease in ahp duration. When murine NGF (2.5 s; 50 ng/mL) was included in the culture medium there was less attentuation of ahp amplitude. Inclusion of affinity-isolated sheep lgG antibodies (0.5 μg/mL; raised against murine 2.5 s NGF) in the culture medium promoted a greater reduction in ahp amplitude than was seen in the "control" explants that were maintained in the absence of NGF. By contrast, the decrease in ahp duration that occurred in control explants was neither attenuated by exposure to NGF nor was it enhanced by NGF antibodies. Also, the increase in spike width that was seen in control explants was enhanced both by murine NGF and by NGF antibodies. Although some of the data support the hypothesis that factor(s) with some similarity to NGF may be synthesized by BFSG in vitro, loss of the retrograde transport of such factors does not explain all aspects of the electrophysiological response to target deprivation and (or) axotomy.Key words: sympathetic ganglion, axotomy, tissue culture, action potential afterhyperpolarization, nerve growth factor.
Conserved charge of glomerular and mesangial cell proteoglycans: possible role of amino acid-derived sulphate
Authors: Templeton Douglas M. , Wang Aimin
Published: April 2011

Sulphation of proteoglycans was studied in isolated glomeruli and cultured rat mesangial cells. Both preparations produced heparan, dermatan, and chondroitin sulphates, recoverable both from the tissue layers and the conditioned media. The proportion of heparan sulphate made by mesangial cells was independent of the age of the culture, but declined in later passage. These preparations differed from several other nontransformed cell types studied to date in that the degree of proteoglycan sulphation was independent of the concentration of inorganic sulphate provided. Even when no exogenous sulphate was added, sulphation-dependent charge density of newly synthesized proteoglycans was conserved. Both isolated glomeruli and cultured mesangial cells produced proteoglycans with 35S-labelled sulphate esters when [35S]methionine was provided as the sole source of labelled sulphate. Conversion of methionine to cysteine and subsequent oxidation of organic sulphate via the sulphinyl pyruvate pathway is the only mechanism known in eukaryotic cells that can account for this observation. We conclude that facile oxidation of sulphur-containing amino acids can contribute to sulphation of glomerular proteoglycans and may serve to sustain the charge density of these multifunctional molecules when the supply of inorganic sulphate is otherwise compromised.Key words: renal glomerulus, proteoglycans, sulphation, heparan sulphate, basement membrane.
Attempts to alter the heterogeneous response to ANP in sodium-retaining caval dogs
Authors: Legault Louis , Cernacek Peter , Levy Mortimer
Published: April 2011

Chronic caval dogs with ascites were identified as being natriuretic "responders" or "nonresponders" (ΔUNaV < 20 μequiv./min) following an infusion of atrial natriuretic peptide (ANP) (100 ng∙kg−1∙min−1). To learn more about the factors modulating tubular resistance to ANP, we attempted to convert responders into nonresponders and vice versa by manipulating the physiological environment. To responding dogs, we readministered ANP in the presence of noradrenaline (n = 5), angiotensin (n = 5), indomethacin (n = 4), and adenosine receptor blockage with theophylline (n = 4), and with purposeful reduction of blood pressure (n = 5). To nonresponding dogs, we readministered the ANP in the face of α-adrenergic blockade (n = 4), saralasin (n = 4), dipyridamole to block adenosine cellular uptake (n = 5), and elevation of blood pressure (n = 4). In no case were we able to alter the initial natriuretic response to ANP. Binding parameters of ANP receptors in suspensions of renal papillary cells were equivalent in responding caval dogs (n = 6), nonresponding dogs (n = 7), and normal controls (n = 7), as was cGMP generation. We conclude that the tubular resistance to ANP in caval dogs unresponsive to this natriuretic peptide is not due to antagonism from catecholamines or angiotensin but may be due to a post-cGMP problem in signal transduction, or a reduction in the delivery of ANP to the distal nephron.Key words: edema, ascites, natriuretic peptide, sodium excretion.
Exercise training has a heparin-like effect on lipoprotein lipase activity in muscle
Authors: Oscai Lawrence B. , Tsika Richard W. , Essig David A.
Published: April 2011

Lipoprotein lipase (LPL) is anchored with high affinity to heparan sulphate proteoglycans on the luminal surface of the capillary endothelium. The levels of pre-heparin perfusate LPL activity increased from 16 ± 1 to 145 ± 6 U/hindlimb (ninefold increase) in hindlimb muscle of exercise-trained rats measured immediately after the last bout of work. At the same time, post-heparin perfusate LPL activity decreased from 63 ± 2 to 13 ± 1 U/hindlimb (p < 0.001). These results provide evidence that exercise-training has a heparin-like effect on capillary-bound LPL. The total amount of LPL (i.e., pre-heparin perfusate plus post-heparin perfusate) was twofold greater in the hindlimb of the trained animals versus the controls. The effect of exercise on muscle LPL activity appears to last for as long as 5 days after cessation of exercise. Serum triglycerides were reduced 38% and plasma free fatty acids increased fourfold. These results provide evidence that training increases the capacity to remove triglycerides from circulation.Key words: lipoprotein lipase, skeletal muscle, exercise, serum triglycerides, heparin.
γ-Aminobutyric acid activity in the olfactory bulb of the rat during the sexual cycle and response to olfactory stimuli
Authors: Becerra N. Navarro , Munaro N. I.
Published: April 2011

Glutamic acid decarboxylase activity in the main and accessory olfactory bulbs throughout the sexual cycle of the rat was studied. The effect of male pheromonal secretion on enzyme activity during proestrus and estrus day was also tested. The enzyme activity showed circadian rhythm during the estrous cycle. This rhythm was disrupted during diestrus-2 afternoon in the main bulb and came back during proestrus afternoon. A different pattern of enzyme activity was present in the accessory bulb, since the circadian rhythm was altered during proestrus morning, returning during estrus afternoon. Male odor exposition did not change enzyme profile activity during proestrus day and during estrus morning in the main bulb. In contrast, in the accessory bulb the olfactory stimuli induced opposite changes to that found in rats from the vivarium during proestrus. Comparison of enzyme activity in olfactory stimuli-deprived rats with that of pheromone-stimulated rats during proestrus showed that male odor exposure specifically affects accessory bulb enzyme activity. It is concluded that the changes of the olfactory bulb GABAergic system during proestrus and estrus day, or that evoked by odor stimuli, demonstrate the discriminative response of this system between the accessory olfactory bulb and the main olfactory bulb.Key words: glutamic acid decarboxylase, pheromone, olfactory bulb, GABA, sexual cycle.
Increased microalbuminuria in diabetic rats is independent of angiotensin II or glomerular proteoglycan synthesis
Authors: Whiteside C. , Templeton D. M.
Published: April 2011

Increased microalbuminuria is seen early in rats with both streptozotocin-induced and genetic (Bio-Breeding) diabetes. This study examines the roles of angiotensin II-dependent mechanism(s) and sulfation of glomerular proteoglycans in this phenomenon, as both processes have been implicated by several lines of circumstantial evidence. Anionic sites in the glomerular basement membrane, attributed to the presence of heparan sulfate, were quantitated by polyethylenimine staining at 15, 21, and 70 days of diabetes in rats treated with streptozotocin, with or without insulin, and at 70 days in the Bio-Breeding rats. All diabetic rats developed increased microalbuminuria: control, 0.08 ± 0.03 μg/mL glomerular filtration rate, [Formula: see text]; streptozotocin without insulin at 15 days, 0.92 ± 0.06 μg/mL (p < 0.05); streptozotocin with insulin at 21 days, 0.61 ± 0.37 μg/mL (p < 0.05 vs. control). At 70 days, both the Bio-Breeding and the streptozotocin rats sustained their microalbuminuria to the same degree (p < 0.05 vs. control). Enalapril (250 mg/L) in the drinking water of diabetic animals did not reduce the microalbuminuria. Although the polyethylenimine-stained heparan sulfate sites decreased significantly in the streptozotocin rats, they remained unchanged in the Bio-Breeding rats. To determine the cause of reduced heparan sulfate staining, the in vitro synthesis and degree of sulfation of proteoglycans by glomeruli isolated from control and streptozotocin diabetic rat kidneys were compared. The amount of heparan sulfate synthesis and degree of sulfation were unchanged in diabetic glomeruli, although lower incorporation into the extracellular matrix and greater secretion into the medium were noted. These effects were partially normalized by insulin therapy. Neither glomerular cell response to angiotensin II nor altered synthesis of heparan sulfate can account for increased microalbuminuria in the diabetic rat.Key words: microalbuminuria, heparan sulfate, enalapril, glomerulus, polyethylenimine.
Effects of gaseous anesthetics and ultrashort and short-acting barbiturates on human blood platelet free cytosolic calcium: relevance to their effects on platelet aggregation
Authors: Philp R. B. , Arora P. , McIver D. J.
Published: April 2011

The effects of elevated pressures (to 6 atmospheres absolute (ATA)) of nitrous oxide (N2O) and of xenon (Xe), and barbiturates on platelet free cytosolic calcium ([Ca2+]i) and platelet aggregation were studied. N2O inhibited the ADP-induced rise in [Ca2+]i whereas Xe had no effect. Neither affected basal levels. Pentobarbital and methohexital had little effect on basal or stimulated levels in the presence or "absence" of extracellular Ca2+; but both, at concentrations > 10−4 M, inhibited platelet aggregation induced by adenosine diphosphate. Thiopental increased basal and stimulated [Ca2+]i when extracellular Ca2+ was present, but not when it was absent, and displayed a bimodal effect with low and high doses being more active than intermediate ones. It also potentiated aggregation. Methitural displayed similar, but nonsignificant, effects. These patterns held for all agents whether or not acetylsalicylic acid was present. Pentobarbital and methohexital inhibited phorbol myristate acetate aggregation in low extracellular calcium and no potentiation was seen with thiopental. In the absence of extracellular Ca2+, no potentiation was observed in stimulated platelets. Potentiation of aggregation previously reported for Xe does not involve increased Ca2+ uptake and did not occur in the absence of extracellular Ca2+. A common mechanism of action for these agents cannot be inferred from their effects on platelet aggregation or [Ca2+]i, as their pharmacological profiles differ markedly. It is evident that their inhibitory properties in this cell are not dependent on extracellular Ca2+, whereas the potentiation observed with pentobarbital, and formerly with Xe, is so dependent.Key words: platelets, aggregation, calcium, anesthetics, narcotic gases.
Regulation of glucose transport in isolated adipocytes by levamisole
Authors: Basi Nirmal S. , Thomaskutty K. G. , Pointer Richard H.
Published: April 2011

When isolated rat adipocytes were incubated with increasing concentrations of levamisole (0.5–5 mM), basal glucose oxidation decreased by almost 50% and insulin-stimulated glucose oxidation decreased by 90%. The decrease in glucose oxidation correlated with an inhibition of glucose transport, since levamisole at 5.0 mM decreased basal 3-O-methylglucose transport by 60% and insulin-stimulated transport by 80%. Diamide-stimulated glucose transport was also inhibited approximately 80% by 5.0 mM levamisole. Levamisole at concentrations up to 5.0 mM had no effect on phosphofructokinase activity. The present results suggest that levamisole inhibits glucose utilization by inhibiting glucose transport in a concentration-dependent manner.Key words: insulin, levamisole, glucose transport, adipocytes.
Effect of synthetic enantiomeric cannabinoids on platelet aggregation
Authors: Nathan I. , Agam G. , Mechoulam R. , Dvilansky A. , Livne A. A.
Published: April 2011

The effect of a synthetic pair of enantiomeric cannabinoids on platelet function was evaluated. The nonpsychotropic enantiomer, the 1,1-dimethylheptyl homolog of (+)-(3S,4S)-7-hydroxy-Δ-6-tetrahydrocannabinol (HU-211), was found to be more active in inhibiting ADP-induced platelet aggregation than the highly psychotropic (−)-enantiomer (HU-210). The related (+)-(3R,4R) cannabinoid, HU-213, which lacks the 7-hydroxy moiety, exerted its inhibitory effect within a wider range of concentrations. The results indicate a differentiation between psychotropic activity and inhibition of platelet aggregation in the cannabinoid group of compounds.Key words: enantiomeric cannabinoids, platelet aggregation.
Role of environmental temperature and photoperiod in regulation of seasonal testicular activity in the frog, Rana perezi
Authors: Delgado M. J. , Alonso-Gómez A. L. , Alonso-Bedate M.
Published: April 2011

To analyze the role of environmental temperature and photoperiod in the regulation of the annual testicular cycle in Rana perezi, we performed experiments combining high (25 ± 1 °C) or low (6 ± 1 °C) temperature and different photoperiod regimens (18L:6D, 12L:12D, and 6L:18D (hours light: hours dark)) during three phases of the reproductive cycle: winter stage (December) and prebreeding (February) and postbreeding (May, June) periods. Low temperature and short photoperiod in winter induced the arrest of the maturation phase of spermatogenesis and the activation of primary spermatogonia proliferation and spermiohistogenesis. Rana perezi testis responded to long days stimulus in winter, even at low temperature, with induction of the maturation phase of the cycle. Exposure of male frogs to either high temperature or long photoperiod induced a decrease in testosterone levels in winter. During the prebreeding period, an increase in environmental temperature caused a reduction in testosterone, and a lengthening in photoperiod produced the opposite effect. Photoperiod had no effect on testosterone levels during the postbreeding period, but low temperature increased testosterone plasma levels. These results suggest that both temperature and photoperiod effects can vary seasonally, depending on the phase of the annual reproductive cycle in R. perezi.Key words: environmental factors, spermatogenesis, steroidogenesis, Amphibia.
Moderate increases in peripheral blood estradiol concentration in the adult ram do not directly inhibit testosterone secretion
Authors: Melnyk Peter M. , Sanford Lee M. , Robaire Bernard
Published: April 2011

Two experiments were conducted in July with adult Dorset × Leicester × Suffolk rams to determine whether increases of 150 or 300% in estradiol (E2) concentration in peripheral blood (from 6.3 ± 0.8 pg/mL in control rams) would affect testosterone secretion directly as well as indirectly via the hypothalamic–pituitary axis. After 4 days of estradiol treatment (experiment 1) provided with subcutaneous polydimethylsiloxane implants filled with crystalline estradiol, luteinizing hormone (LH) and testosterone secretions were reduced by 50% (p < 0.05) in both groups of rams because of subtle decreases in pulse frequencies and amplitudes. Estradiol treatments were also associated with decreases in mean follicle-stimulating hormone (FSH) concentration (30–50% in both groups, p < 0.05) and increases in mean prolactin concentration (35% in low-E2 group; 105% in high-E2 group, p < 0.05), but testicular responsiveness to an LH challenge (single intravenous dose, 10 μg NIH-LH-S25) remained normal. When along with estradiol treatment, 10-μg doses LH were given every 80 min (experiment 2), testosterone secretion increased by 265% (p < 0.05) in both treated and control rams. Relative to day −1, secretion on day 4 was characterized by higher (p < 0.05) pulse frequencies and baseline concentrations and lower (p < 0.05) pulse amplitudes; values for all characteristics were similar to those for Dorset × Leicester × Suffolk rams in the breeding season. Interestingly, the decreases in mean FSH concentration brought about by estradiol and (or) LH treatments were not any greater than in experiment 1, and estradiol's ability to elevate mean prolactin concentration was blocked completely. We conclude that moderate increases in estradiol concentration in peripheral blood of rams suppress testosterone secretion only by impairing LH release.Key words: estradiol, hypothalamic–pituitary axis, testes, ram.
Inositol phosphate production in response to [Arg8]vasopressin, endothelin 1, and prostaglandin F2α in rat aorta and mesenteric arteries
Authors: Parent Angèle , Nguyen Paul V. , Yang Xiao Ping , Schiffrin Ernesto L.
Published: April 2011

Vascular tissues such as rat aorta and mesenteric arteries are extensively used experimentally for the study of cardiovascular diseases. To further our understanding of the signal transduction mechanisms involved in responses to several potent vasoconstrictors, such as [Arg8]vasopressin (AVP), endothelin 1 (ET-1), and prostaglandin F2α (PGF2α), we have investigated the time course for production of inositol monophosphate (InsP1), bisphosphate (InsP2), and trisphosphate (InsP3) in response to these agonists as well as their relative potency for phosphatidylinositol hydrolysis. Time-course studies of production of the different inositol phosphates in response to AVP and PGF2α showed an early increase after 15–30 s of stimulation. Thereafter InsP3 declined towards baseline, with a secondary increase towards steady state after 5–10 min. Rapid turnover of InsP3 was reflected by accumulation of InsP1 and InsP2 in the presence of LiCl (20 mM) to inhibit monophosphatases. After 15–30 min of stimulation, there was accumulation of the Ins(1,3,4)P3 isomer. All three agonists induced greater accumulation of InsP2 in mesenteric arteries than in thoracic aorta, suggesting that turnover of Ins(1,4,5)P3 may be faster in the former than in the latter. The accumulation of total inositol phosphates induced by maximum concentrations of ET-1 was greater than in response to AVP or PGF2α. Dose–response curves showed that the rank order of potency for stimulation of production of inositol phosphates was AVP > ET-1 > PGF2α, similar to the sensitivity of blood vessels to these agents. Comparison of responses to ET-1 and ET-3 showed that the receptors stimulated by endothelins were of the isopeptide selective ETA subtype. In conclusion, all three agonists (AVP, ET-1, PGF2α) stimulate phospholipase C activity in rat aorta and in mesenteric arteries, although with different potencies. This study demonstrates that intact blood vessels allow a detailed investigation of inositol phosphate responses to different agonists of interest in cardiovascular research.Key words: phosphoinositide metabolism, phospholipase C, inositol trisphosphate, vasoconstrictors, blood vessels.
Mechanoreceptor afferent activity compared with receptor field dimensions and pressure changes in feline urinary bladder
Authors: Downie John W. , Armour J. Andrew
Published: April 2011

The relationship between vesical mechanoreceptor field dimensions and afferent nerve activity recorded in pelvic plexus nerve filaments was examined in chloralose-anesthetized cats. Orthogonal receptor field dimensions were monitored with piezoelectric ultrasonic crystals. Reflexly generated bladder contractile activity made measurements difficult, therefore data were collected from cats subjected to actual sacral rhizotomy. Afferent activity was episodic and was initiated at different pressure and receptor field dimension thresholds. Maximum afferent activity did not correlate with maximum volume or pressure. Furthermore, activity was not linearly related to intravesical pressure, receptor field dimensions, or calculated wall tension. Pressure–length hysteresis of the receptor fields occurred. The responses of identified afferent units and their associated receptor field dimensions to brief contractions elicited by the ganglion stimulant 1,1-dimethyl-4-phenylpiperazinium iodide (2.5–20 μg i.a.), studied under constant volume or constant pressure conditions, are compatible with bladder mechanoreceptors behaving as tension receptors. Because activity generated by bladder mechanoreceptors did not correlate in a simple fashion with intravesical pressure or receptor field dimensions, it is concluded that such receptors are influenced by the viscoelastic properties of the bladder wall. Furthermore, as a result of the heterogeneity of the bladder wall, receptor field tension appears to offer a more precise relationship with the activity of bladder wall mechanoreceptors than does intravesical pressure.Key words: bladder distension, intravesical pressure, sacral rhizotomy, viscoelasticity.