Ferrokinetics after high-dose methotrexate therapy

Summary After high-dose methotrexate with doses ranging from 2–10 g/m², ferrokinetics were performed in 11 patients with different tumors. Iron-III-citrate-⁵⁹Fe was injected at methotrexate serum concentrations ranging between 2.7×10^–7–1.3×10^–8 M. With MTX levels 4.2×10^–8 M the plasma iron clearance was always retarded, and the plasma iron turnover was diminished in 5 of 6 patients with levels of 3.1–8.9×10^–8 M at the time of the injection. In all cases with MTX concentrations 5×10^–8 M the⁵⁹Fe-utilization as the measure of effective erythropoiesis was reduced pathologically below normal range. These results show that the erythropoiesis resumes its normal extent even in case of normal methotrexate clearance only when the methotrexate serum concentrations have fallen down below 5×10^–8 and that erythropoiesis is more sensitive against methotrexate toxicity than the granulocytopoiesis.     

Ion-induced release of LHRH,TRH and α-MSH from hypothalamic synaptosomes and its inhibition by vinblastine

Homogenates of rat hypothalamic tissue were fractionated by means of discontinuous sucrose density gradient centrifugation. Immunoreactive luteinizing hormone releasing hormone (LHRH), thyrotropin releasing hormone (TRH), and -melanocyte stimulating hormone (-MSH) were found to be concentrated in the synaptosome-enriched fraction. This fraction was suspended in 0.32 M sucrose and the release of the three peptides was investigated. After incubation, the synaptosomes were re-isolated by ultrafiltration, and the concentration of each peptide in the ultrafiltrate was determined by radioimmunoassay. When the synaptosomal fraction was incubated at 30° C in 0.32 M sucrose containing either 60 mM K⁺-2 mM Ca²⁺ or 140 mM Na⁺ alone a release of LHRH, TRH, and -MSH occurred. Of the total content 30–50% of LHRH but only about 10% of TRH and -MSH was releasable. When the synaptosome preparation was preincubated for 30 min at 30°C with 10^–4 M vinblastine. K⁺- as well as Na⁺-induced release of LHRH, THR, and -MSH was inhibited, and the stimulatory effect of each cation was almost totally blocked by preincubation with 5×10^–4 and 10^–3 M vinblastine. The inhibitory action of vinblastine (5×10^–4 M) did not affect the oxidation of glucose to CO₂ by the synaptosomes. The results of the present investigation demonstrate that synaptosome-enriched fractions of hypothalamic origin are more stable with respect to LHRH, TRH, and -MSH release during incubation in isotonic sucrose than they are in ionic solutions, and that the peptides are released by a vinblastine-sensitive mechanism.Supported by grants from the National Institute of Arthritis, Metabolism, and Digestive Diseases (AMO 1237), the National Institute of Child Health and Human Development (HDO8672), and the National Institutes of Health contract (5-P17-HL1487-06)Supported by Grant No. 512-6951 from the Danish Medical Research Council     

Non-quantal release of transmitter at mouse neuromuscular junction and its dependence on the activity of Na+−K+ ATP-ase

Summary The effect of val⁵-angiotensin II on steady-state sodium concentration gradients (c_Na) was studied in rat proximal tubules by stationary micro-perfusion combined with perfusion of the peritubular capillaries. Angiotensin added to the peritubular perfusion fluid had a biphasic action with stimulation of sodium reabsorption at low doses (10^–12–10^–10 M) and inhibition at high doses (3×10^–7–3×10^–6 M). Stimulation of transport was also observed with intraluminal angiotensin but only at a dose of 10^–9M. Transepithelial potential difference was calculated from the steady-state chloride distribution; no significant change was observed at low (10^–11M) or high (10^–6M) concentrations and a direct action on sodium transport is postulated. This biphasic effect is discussed in relation to the responses of the intact kidney to intra-renal infusion of angiotensin, and to the control of tubulo-glomerular feed-back.     

The effects of pinacidil and tolbutamide in feline pial arteries in situ

The vasomotor effect of the K⁺ channel opener pinacidil was investigated in feline pial arteries of the parietal cortex. Perivascular microapplication (5l in 40 sec) and an image splitting method for the measurement of vascular diameter were employed. Pinacidil (10^–11 –10^–7 M) induced concentration-dependent dilatations at 10^–9 M and higher concentrations. A maximal dilatation of about 42% was achieved at 10^–8 M, the dilatation at 10^–7 M was reduced to 22%. The sulphonylurea tolbutamide exerted per se no effect in pial arteries but it blocked concentration-dependently the pinacidil induced dilatation. This is consistent with the presence of ATP-sensitive K⁺ channels in pial arteries which are closed under resting conditions.     

Effects of gold compounds on function of phagocytic cells

The effect of sodium aurothiomalate and auranofin on the generation of superoxide anions (O ₂ ^– ) by polymorphonuclear leukocytes (PMNLs) and adherent mononuclear phagocytic cells (AMNCs) has been investigated. Sodium aurothiomalate at final concentrations of 1, 10, and 100 g Au/ml and auranofin ranging from 0.1 to 2.0 g Au/ml were used in the reactions involving all celt types. Results have been compared between cells drawn from normal controls and patients with active rheumatoid disease. The effect of gold compounds on both cell types was assessed following activation by phorbyl myristate acetate (1×10^–8 M) and N-formyl-methionylleucyl-phenylalanine (1×10^–4 M) using a cytochromec reduction method. Sodium aurothiomalate at the maximum concentration modestly inhibited O ₂ ^– generation by PMNLs but not AMNCs. Auranofin inhibits O ₂ ^– generation by both cell types. Inhibition of cells from patients with rheumatoid arthritis was greater than that seen with cells from normal controls.     

The inotropic effects of dopamine and its precursor levodopa on isolated human ventricular myocardium

Summary The direct positive inotropic effects of dopamine and its precursor, levodopa, were measured using isolated, contracting human papillary muscle strips taken from patients during mitral valve replacement. Levodopa did not produce any positive inotropic effect at concentrations up to 3×10^–3 M. The positive inotropic effects of dopamine were observed at concentrations above 1×10^–5 M with the maximal effect at 3×10^–3 M — concentrations higher than those observed in therapy. This inotropic effect was reduced by the ₁ antagonist, 1-practolol (1×10^–6 M); the ₂ antagonist, ICI 118,551 HCl (1×10^–6 M); the dopamine antagonist, haloperidol (3×10^–6 M); the neuronal uptake inhibitor, cocaine (3×10^–5 M), but not by the ₁, prazosin (1×10^–7 M). This indicates that dopamine exerts its positive inotropic effects on human heart muscle mainly through release of noradrenaline, together with possible interactions at -and dopamine-receptors. The maximal inotropic effect of dopamine was about 50% that of calcium (15 mM, 6.2±0.7 mN) or ouabain (1×10^–7 M, 5.0±0.8 mN) when measured in the same muscle strips, possibly due to the reduced cardiac noradrenaline content together with the reduced -receptor number in congestive heart failure. This concentration of ouabain (1×10^–7 M) gave almost maximal inotropy without marked toxicity; when dopamine was then added, only toxicity developed without any further increases in force of contraction. Any haemodynamic benefits of dopamine therapy in optimally digitalis-treated patients are probably due to other cardiovascular effects such as vasodilatation.This study was carried out using human heart samples provided by: Prof. E. Kreuzer, Dr. B. Kemkes, Dr. C. Weinhold, Herzchirurgische Klinik der Universität, Klinikum Großhadern, and Dr. K. Holper, Dr. W. Klövekorn, Herzchirurgische Klinik der Deutschen Herzzentrums, MünchenSupported by the Deutsche Forschungsgemeinschaft (Er 65/4-4)     

General features of electrical and mechanical properties of smooth muscle cells in the guinea-pig abdominal aorta

The membrane potential in smooth muscle cells of the guinea-pig's abdominal aorta has a mean value of –57.2 mV. These cells are electrically connected and the space and time constant are 0.66 mm and 180 ms respectively. An increased K-concentration elicited a contraction at 20 mM and the maximum was reached at 77mM. The maximum depolarization produced by a tenfold increase of [K]₀ was 45 mV. Tetraethylammonium at concentrations exceeding 2 mM, depolarized the membrane, increased the membrane resistance and reduced the rectifying properties of the membrane. Only at 20 mM a small active response could be induced by outward current pulses. Low concentrations of noradrenaline (<10^–8 M) hyperpolarized the membrane, while higher concentrations (10^–7 M) depolarized. Isoprenaline at concentrations below 10^–7 M also hyperpolarized, but it depolarized from 10^–5 M onward. Acetylcholine at concentrations over 10^–8 M hyperpolarized the cells without exerting an effect on the resting tension, but it reduced a noradrenaline induced contraction. Low concentrations of caffeine (2 mM) hyperpolarize the membrane, while higher concentration (5 mM) depolarize. Caffeine is found to be a more efficient releaser of cellular Ca than noradrenaline. This might be due to the weak -agonist action of noradrenaline appearing at high noradrenaline concentrations. The hypothesis is supported by the finding that a -stimulation increases the Ca-uptake in the intracellular store. The study of the electrophysiological effects of different stimuli do not suggest an important role for electromechanical coupling in this tissue.     

In vitro effects of milbemycin oxime: mechanism of action againstAngiostrongylus cantonensis andDirofilaria immitis

The neuropharmacological mechanisms underlying the action of milbemycin oxime on the motility ofAngiostrongylus cantonensis andDirofilaria immitis were examined in vitro. InA. cantonensis, milbemycin oxime caused inhibitory effects at low concentrations of 10^–9 g/ml, and paralysis was elicited at 10^–8–10^–6 g/ml. The paralysis was antagonized by picrotoxin and bicuculline but not by dibenamine. In addition, stimulatory effects were observed when the antibiotic was used at higher concentrations of 3–5×10^–6 g/ml, and the action was antagonized by strychnine. Both effects were also observed in the preparation contracted by eserine or pyrantel. WhenD. immitis was treated with milbemycin oxime at concentrations of 10^–7 and 3–5×10^–6 g/ml, only slight inhibitory and stimulatory effects, respectively, were observed. These effects were partially antagonized by picrotoxin and strychnine, respectively. These results suggest that the inhibitory and stimulatory actions of milbemycin oxime are caused through gabergic and cholinergic mechanisms inA. cantonensis andD. immitis.Studies on chemotherapy of parasitic helminths XXIX     

Effect of atrial natriuretic factor and cyclic guanosine monophosphate on water and urea transport in the inner medullary collecting duct

We examined the action of high (2×10^–8M) and low (6×10^–9M) concentrations of atrial natriuretic factor (ANF) on water and urea transport in the rat inner medullary collecting duct (IMCD) using the in vitro microperfusion technique. We measured the hydraulic conductivity (Lp ×10^–6 cm/atm per second) and both lumen-to-bath (P _u(lb)) and bath-to-lumen (P _u(bl)) ¹⁴C-urea permeabilities (P _u× 10^–5 cm/s) in the absence and in the presence of vasopressin (VP). High concentrations of ANF were able to inhibit the maximum activity of (50 U/ml) VP-stimulated L _p but physiological concentration of ANF inhibit only submaximum activity (10 U/ml) of VP-stimulated L _p. The hydrosmotic effect of dibutyryl-cyclic 3,5 adenosine monophosphate (cAMP) (10^–4M) was unchanged by high concentrations of ANF (2×10^–8M). Also we found that high (10^–4M) and low (10^–6M) concentrations of exogenous cyclic 3,5-guanosine monophosphate (GMP) while unable to change the Lp in the absence of VP, decreased the maximum activity of VP-stimulated Lp significantly. We also found that ANF inhibits partially and in a reversible manner the VP-stimulated P _u(lb) but not the VP-stimulated P _u(bl). These results demonstrated that plasma concentrations of ANF observed during volume expansion (10^–10M) are able to inhibit submaximum activity of VP-stimulated (10 U/ml) L _p in the rat IMCD, this effect seems to occur before cAMP formation and it appears to be mediated by cGMP. ANF (6× 10^–9M) also reduced the VP-stimulated urea outflux. Therefore, the increase in water excretion produced by ANF could be explained, at least in part, by the inhibition by ANF of vasopressin effects on water and urea transport in the IMCD.This study was presented in part at the VI Latin American Congress of Nephrology, Brazil, October 1985 and at the X^th International Congress of Nephrology, London, July 1987.     

A study of the interaction of catecholamines and antidiuretic hormone on water permeability and the cyclic AMP system in isolated papillae of the rat

The diffusional water permeability of collecting ducts in vitro and the cyclic A.M.P. content of isolated papillae were measured after exposure to different concentrations of antidiuretic hormone, isoproterenol and noradrenalin. Antidiuretic hormone 25 units/ml. caused a 25% increase in diffusional water permeability. This response was not affected by isoproterenol (10^–6 M) or noradrenalin (2 ×10^–6 M). Antidiuretic hormone 100 unit ml^–1 caused a 50% increase in diffusional water permeability which likewise was not altered by isoproterenol or noradrenalin. Isoproterenol (10^–6 M) and noradrenalin (2×10^–6 M) had no significant effect on basal levels of diffusional water permeability.Isoproterenol had no significant effect on the tissue concentration of cyclic A.M.P. or on the increase in cyclic A.M.P. concentration induced by antidiuretic hormone. Noradrenalin (2×10^–6 and 10^–4) had no significant effect on basal cyclic A.M.P. concentration. However, noradrenalin inhibited the stimulation of cyclic A.M.P. induced by antidiuretic hormone. This effect was inhibited by phentolamine.This study suggests that catecholamines do not alter water handling by a direct action on the water permeability of the kidney but probably exert their action through an effect of A.D.H. release.Supported by N.H.M.R.C. and the Repatriation Department     

Inhibitory effects of palmitoylcarnitine and lysophosphatidylcholine on the sodium current of cardiac ventricular cells

We investigated the effects of ischemia-related amphipathic compounds, palmitoylcarnitine (PamCar, 0.5–50 M) and lysophosphatidylcholine (lysoPtdCho, 5–50 M) on sodium current (I _Na) of guinea-pig ventricular myocytes. The cells were perfused with low-Na⁺ (60 mM) Tyrode's solution, and Ca²⁺ and K⁺ currents were blocked by external Co²⁺ (3 mM) and internal Cs⁺ (140 mM), respectively. I _Na was elicited by depolarizing voltage steps from a holding potential of –100mV at a temperature of 33 °C. PamCar (5 M) decreased the peak I _Na (attained at –20mV or –30mV) from 6.1±2.1 nA to 3.9±1.4 nA (n=11), or by 36.1% within 2 min, and shifted the curve of steady-state I _Na inactivation by 5.4 mV in the positive direction (from –76.3±4.6 mV, control to –70.9±4.0 mV, in PamCar, n=4). Partial restoration of the amplitude and the shift of the steady-state inactivation curve of I _Na was attained after washout of PamCar. In contrast, lysoPtdCho at concentrations over 10 M irreversibly depressed the I _Na within 0.5–3 min and the reduction of IinNa was followed by cell contracture or cell death (n=9). The survival time, defined as a period from the start of lysoPtdCho application to the time of the last successful recording of the I _Na (before evolution of sudden changes in the holding current), depended on the concentrations of lysoPtdCho. Both PamCar and lysoPtdCho retarded the time course of activation and inactivation of I _Na. These findings are compatible with the idea that PamCar and lysoPtdCho decrease the maximum Na⁺ conductance and alter the surface negative charge of the membrane, perhaps via amphiphilic intervention in the phospholipid bilayers. However, PamCar had an additional effect that indicates more direct but reversible incorporation of this agent with the Na⁺ channels or integral membrane proteins.     

Cyclic AMP and enzyme secretion by the isolated rabbit pancreas

Summary The isolated rabbit pancreas secreted protein and -amylase at a relatively constant rate during an eight-hour period. Both pancreozymin and acetylcholine caused a massive release of protein and -amylase into pancreatic fluid. Theophylline (10^–2 M) stimulated protein secretion in vitro maximally by 155%, while for -amylase secretion a maximal average stimulation of 80% was observed. Theobromine (10^–2 M) exerted a slight increase on enzyme secretion, equal to that given by 10^–3 M theophylline. Cyclic AMP (10^–3M) also increased enzyme secretion. For protein secretion a maximal average stimulation of 64% was observed, while -amylase secretion was maximally stimulated by 31%. Theophylline (10^–2 M) potentiated the stimulatory effect of pancreozymin (8.5 U/l) on pancreatic enzyme secretion. These results indicate that cyclic AMP is a mediator in enzyme secretion.     

Cardiac glycoside tolerance in cultured chicken heart muscle cells — A dose-dependent phenomenon

Summary In cultured heart muscle cells from 10–13 day-old chicken embryos, the effects of acute (4 h) and chronic (3 days) exposure of the cells to varying concentrations of ouabain have been studied. In these cells, the cardiac glycoside ouabain binds to a specific cardiac glycoside receptor (K_D=4 × 10^–7 M; 750,000 receptors/cell). Binding to this receptor results in inhibition of active Na⁺/K⁺-transport [EC₅₀ for active (⁸⁶Rb⁺ + K⁺)-influx=4 × 10^–6 M], and in an increase in beating velocity (positive inotropic effect;; EC₅₀=4 × 10^–7 M); toxic signs (arrhythmias) appear at concentrations 6 × 10^–7 M. During exposure of the cells to 3 × 10^–6 M ouabain for 3 days, tolerance develops with respect to both the positive inotropic and the toxic effect. The mechanism underlying this tolerance is identified as an increase in the number of active sodium pump molecules per cell, while the binding properties of the cardiac glycoside receptor remain unchanged. The development of cardiac glycoside tolerance is only observed in the presence of severe impairment of Na⁺/K⁺-homeostasis, due to cardiac glycoside-induced inhibition of active Na⁺/K⁺-transport. This, however, only occurs in the presence of toxic (receptor occupation 60%), but not in the presence of positive inotropic, non-toxic (receptor occupation 20–60%), ouabain concentrations. We conclude that the development of cardiac glycoside tolerance during long-term treatment in patients with heart failure should not occur with submaximal dose regimens, when toxic signs (arrhythmias) are absent.Abbreviations CGP-12177 (4-3-tert-butylamino-2-hydroxy-propoxy)-benzimidazol-2-one hydrochloride) - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulphonic acid Some of the results were presented at the IXth European Congress of Cardiology, July 1984, Düsseldorf, Germany [49]     

On the mechanism of histamine induced enhancement of the cardiac Ca2+ current

In guinea pig ventricular myocytes, the effect of histamine on the slow Ca²⁺ current (I_Ca) was studied and the following results were obtained: (1) Superfusion of cells with histamine resulted in a dose-dependent enhancement of the amplitude of I_Ca. The threshold concentration of histamine was 10^–8 M, half maximal increase occurred at 3×10^–7 M and maximal enhancement (about 3–4-fold) at 5×10^–6 M. (2) The histamine effect was greatly reduced by the H₂ antagonist cimetidine (10^–5 M) but only slightly by the H₁ antagonist diphenhydramine (10^–5M). (3) Effects of isoprenaline (ISP) and histamine at maximal effective concentrations on I_Ca were not additive, suggesting that both agents use the same intracellular pathway. Intracellular infusion of a blocker of the cAMP-dependent protein kinase, R_p-cAMPS (10^–4 M), prevented the histamine effect. (4) The involvement of GTP-dependent transducer proteins was studied by cell dialysis with several GTP derivatives. Intracellular application of the stable GDP-analogue, GDP--S, reduced the histamine effect on I_Ca, whereas the stable GTP analogue, GTP--S, mimicked the histamine effect.     

α1-Adrenergic regulation of Cl− and Ca2+ movements in rat parotid acinar cells

In rat parotid acinar cells, maximal ₁-adrenergic receptor stimulation (10^–5 M epinephrine +10^–5 M propranolol) leads to a rapid (<10 s), 4–5-fold elevation in cytosolic Ca²⁺ ( 800 nM at peak) which decreases to 50% of peak Ca²⁺ by 3–4 min. Similarly, cells preloaded with³⁶Cl^– show a rapid (<10 s) 35–50% loss of³⁶Cl^– which returns to 80% of resting values in 3–4 min. Both responses are dependent on epinephrine, with half-maximal effects achieved at 2×10^–7 M and 2×10^–6 M agonist for Cl^– and Ca²⁺, respectively. In the presence of low extracellular Ca²⁺ (i.e. with EGTA), the initial rapid changes in cellular Ca²⁺ and Cl^– are unaltered. However, cellular Ca²⁺ and Cl^– levels return to basal values sooner than when extracellular Ca²⁺ is present (within 2 and 3 min, respectively). Maximal epinephrine-induced Ca²⁺ and Cl^– responses are unaffected by the ₂-adrenergic antagonist, yohimbine, are completely blocked by the ₁-adrenergic antagonist, SZL-49, and are similar to ion fluxes induced by maximal muscarinic-cholinergic receptor stimulation (10^–5 M carbachol). The data suggest that a close association exists between mobilization of intracellular Ca²⁺ and Cl^– content in rat parotid acinar cells after ₁-adrenocetor stimulation.     

Role of Na/K-ATPase in Regulation of Neurite Growth in Sensory Neurons

We studied the effects of acetylcholine and norepinephrine on the growth of neurites in organotypic culture of 10–12-day-old chick embryo sensory neurons. Acetylcholine (10^–8 M) and norepinephrine (10^–13 M) stimulated the growth on sensory neuron neurites. Experiments with combined application of acetylcholine and norepinephrine against the background of ouabain showed that the nonspecific action of the test neurotransmitters is related to modulation of Na/K-ATPase activity.__________Translated from Byulleten Eksperimentalnoi Biologii i Meditsiny, Vol. 139, No. 2, pp. 157–159, February, 2005     

Basolateral membrane chloride permeability of A6 cells: implication in cell volume regulation

The permeability to Cl^– of the basolateral membrane (blm) was investigated in renal (A6) epithelial cells, assessing their role in transepithelial ion transport under steady-state conditions (isoosmotic) and following a hypoosmotic shock (i.e. in a regulatory volume decrease, RVD). Three different complementary studies were made by measuring: (1) the Cl^– transport rates (F/F _o · s^–1 (× 10^–3)), where F is the fluorescence of N-(6-methoxyquinoyl) acetoethyl ester, MQAE, and F _o the maximal fluorescence (×10^–3) of both membranes by following the intracellular Cl^–3 activities (a _iCl^–, measured with MQAE) after extracellular Cl^– substitution (2) the blm ⁸⁶Rb and ³⁶Cl uptakes and (3) the cellular potential and Cl^– current using the wholecell patch-clamp technique to differentiate between the different Cl^– transport mechanisms. The permeability of the blm to Cl^– was found to be much greater than that of the apical membranes under resting conditions: a _iCl^– changes were 5.3±0.7 mM and 25.5±1.05 mM (n=79) when Cl^– was substituted by NO₃ ^– in the media bathing apical and basolateral membranes. The Cl^– transport rate of the blm was blocked by bumetanide (100 M) and 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB, 50 M) but not by N-phenylanthranilic acid (DPC, 100 M). ⁸⁶Rb and ³⁶C1 uptake experiments confirmed the presence of a bumetanide- and a NPPB-sensitive Cl^– pathway, the latter being approximately three times more important than the former (Na/K/2Cl cotransporter). Application of a hypoosmotic medium to the serosal side of the cell increased F/F _o · s^–1 (×10^–3) after extracellular Cl^–3 substitution (1.03±0.10 and 2.45±0.17 arbitrary fluorescent units·s^–1 for isoosmotic and hypoosmotic conditions respectively, n=11); this F/F _o·s^–1 (×10^–3) increase was totally blocked by serosal NPPB application; on the other hand, cotransporter activity was decreased by the hypoosmotic shock. Cellular Ca²⁺ depletion had no effect on F/F _o·s^–1 (×10^–3) under isoosmotic conditions, but blocked the F/F _o·s^–1 (×10^–3) increase induced by a hypoosmotic stress. Under isotonic conditions the measured cellular potential at rest was –37.2±4.0 mV but reached a maximal and transient depolarization of –25.1±3.7 mV (n=9) under hypoosmotic conditions. The cellular current at a patch-clamping cellular potential of –85 mV (close to the Nernst equilibrium potential for K⁺) was blocked by NPPB and transiently increased by hypoosmotic shock ( 50% maximum increase). This study demonstrates that the major component of Cl^– transport through the blm of the A6 monolayer is a conductive pathway (NPPB-sensitive Cl^– channels) and not a Na/K/2Cl cotransporter. These channels could play a role in transepithelial Cl^– absorption and cell volume regulation. The increase in the blm Cl^– conductance, inducing a depolarization of these membranes, is proposed as one of the early events responsible for the stimulation of the ⁸⁶Rb efflux involved in cell volume regulation.     

The effect of antisecretory factor on the permeability of nerve cell membrane to chloride ion

The antisecretory factor (ASF) is a hormone-like protein (m.w. 60,000) that most effectively counteracts hypersecretion in vivo in the small intestine of pigs and rats. The present report demonstrate that 10^–13 moles of ASF inhibits significantly the³⁶Cl^– permeation through the isolated neuronal plasma membrane of Deiters' cells in rabbits. This effect was enhanced by 0.2 mM -aminobutyric acid (GABA), and quenched by the addition of anti-ASF immunoglobulins; pretreatment of the neuronal membrane with nipecotic acid (10^–6 M) or with bicuculline (10^–3 M) abolished the ASF action whilst picrotoxin (10^–4 M) pretreatment left the inhibitory effect of ASF unaffected. The results suggest that ASF blocks chloride channels in neuronal membranes, including those channels activated by GABA.Abbreviations ASF antisecretory factor - GABA -aminobutyric acid - SEM standard error of the mean     

Adenosine as inhibitor of myocardial effects of catecholamines

Summary Infusion of adenosine into the coronary arteries of isolated guinea pig hearts produced a dosedependent inhibition of dP/dt_max caused by bolus injections of isoproterenol (4×10^–11 moles). Threshold concentration of adenosine was 10^–7 M and maximal inhibition (90%) occurred at 10^–5 M. Coronary dilation induced by, papaverine did not influence the contractile response to catecholamines. In addition to its influence on cardiac performance, adenosine (10^–5 M) effectively inhibited the isoproterenol (10^–7 M) induced initial rise in myocardial levels of cyclic 35-AMP, glucose-1-phosphate and glucose-6-phosphate. Adenosine also antagonized the effect of isoproterenol on adenylate cyclase activity in a crude membrane preparation from guinea pig ventricles; it was without effect on the activity of the membrane phosphodiesterase. Theophylline inhibited the actions of adenosine both on adenylate cyclase activity and on contractile force development.-Upon infusion of isoproterenol (3×10^–7 M) into the coronary arteries of the isolated heart (perfusion at constant pressure), the adenosine concentration in the effluent perfusate increased within 45 s from 10^–8 M to about 10^–6 M. It thus appears conceivable that in ventricular myocardium endogenously formed adenosine may serve 2 functions: dilation of the coronary arteries and limitation of the inotropic and metabolic effects of catecholamines.A preliminary report of these studies was presented at the 47th Meeting of the German Physiological Society in Regensburg, Germany [Pflügers Arch.365, R4 (1976)]     

Effects of two vasodilatory phosphodiesterase inhibitors on bradykinin-induced permeability increase in the hamster cheek pouch

Two inhibitors with selective effect on cyclic nucleotide phosphodiesterases (PDEs, preferentially hydrolyzing cAMP), milrinone (cGMP-inhibited PDE) and rolipram (cAMP-specific PDE) were studied for their effects on bradykinin-induced plasma leakage in comparison with the ₂-receptor stimulant terbutaline. The dilation of arterioles induced by milrinone and rolipram was studied in the concentration range 10^–7–10^–4 M. Maximal arteriolar dilation was 53% for milrinone at 10^–4 M and 28% for rolipram at 10^–4 M. The hamster cheek pouch preparation was used as prepared for intravital microscopy of fluorescein-labelled dextran, FITC-dextran. Bradykinin was applied topically to the cheek pouch at a final concentration of 4×10^–7 M and caused rapid and reversible increase in plasma leakage (number of leakage sites) from postcapillary venules. Milrinone (M), rolipram (R) and terbutaline (T) were also applied topically starting 5 min prior to bradykinin application and at final concentration of 10^–4 and 10^–5 M (M), 10^–5 and 10^–6 M (R) and 10^–7 M (T). These local concentrations resulted in significant (p<0.05) and reversible inhibition of the bradykinin-induced response by 44% and 33% (M), 77% and 67% (R) and 46% (T). Combining M and R individually with T resulted in a significantly larger inhibition of the bradykinin response than with each of the drugs given separately.It is concluded that selective inhibitors of PDEs, preferentially hydrolyzing cAMP, can result in a reduced response to bradykinin as seen with ₂-receptor agonists and that the potency of these two PDE inhibitors to counteract plasma leakage was not correlated to their potency as vasodilators.