Electropharmacological effects of berberine on canine cardiac Purkinje fibres and ventricular muscle and atrial muscle of the rabbit.

1. Conventional microelectrode techniques were used for intracellular recordings of the transmembrane electrical potentials, the effects of berberine were studied on canine cardiac Purkinje and ventricular muscle fibres and on rabbit atrial fibres. 2. Berberine (3-30 microM) increased in a concentration-dependent manner, the action potential duration (APD) in canine Purkinje and ventricular muscle without affecting other parameters of the action potential. 3. The berberine-induced enlargement of the APD showed reverse use-dependence, so that the effect was greater at lower rates of stimulation. 4. Preparations perfused with berberine (30 microM) and driven at rates below 0.5 Hz exhibited early after depolarizations which persisted 3-4 h after washing. 5. The early afterdepolarizations were reversibly abolished by perfusion with lignocaine (3 microM) or by the increase in the rate of stimulation. 6. The effective refractory period (ERP) of Purkinje fibres was greatly increased by berberine (30 microM); however, the ratio ERP/APD was not significantly affected. 7. Berberine (10-100 microM) decreased in a concentration-dependent manner the spontaneous frequency of rabbit sinoatrial cells. The decrease in frequency was accompanied by a depression of the phase 4 depolarization, without significant changes in other parameters of the nodal action potential. 8. Atropine (2.5 microM) did not affect the bradycardic effect of berberine. On the other hand, berberine (30 microM) did not alter the chronotropic effect of isoprenaline. 9. Berberine (30 microM) also increased the duration of slow responses in K-depolarized rabbit atrial muscle fibres, other parameters being unaffected. 10. It is suggested that berberine exerts Class III antiarrhythmic and proarrhythmic actions in cardiac muscle of the dog in vitro.     

In vitro cardiac models of dog Purkinje fibre triggered and spontaneous electrical activity: effects of nicorandil.

1. The effects of nicorandil (30 microM and 100 microM) on two models of triggered activity [early afterdepolarizations (EADs) and delayed afterdepolarizations (DADs)] and on spontaneous automaticity occurring from both normal and depolarized levels of membrane potential were examined in isolated cardiac Purkinje fibres of the dog. Standard intracellular microelectrode techniques were used. 2. Nicorandil (30 microM) abolished EADs provoked by superfusion with Tyrode solution containing 2.7 mM K+ and 3 mM Cs. 3. DADs were induced by 0.2 microM acetylstrophanthidin in Tyrode solution containing 5.4 mM K+. Nicorandil (30 microM) significantly reduced the amplitude of these DADs from 12.5 +/- 2.5 mV to 5.5 +/- 0.2 mV (P less than 0.02, n = 6), while DADs were fully abolished by 100 microM nicorandil. 4. In unstimulated Purkinje strands, superfused with 2.7 mM K+ containing Tyrode solution having a pH of either 7.4 or 6.8, spontaneous depolarizations developed with a mean maximum diastolic potential (MDP) of -84.6 +/- 1.6 mV (n = 9) or -54.0 +/- 1.2 mV (n = 9), respectively. Nicorandil significantly reduced the frequency of this automatic activity and caused its cessation, at either level of MDP. Nicorandil, however, produced significant hyperpolarization only when automaticity occurred from the depolarized level of potential. 5. These results suggest that nicorandil may exert significant antiarrhythmic actions in vivo by abolishing both spontaneous and triggered electrical activity.     

Effects of cyproheptadine on electrophysiological properties of isolated cardiac muscle of dogs and rabbits

The effects of cyproheptadine were studied on cardiac Purkinje and ventricular muscle fibres of the dog and on cells of the sinoatrial (SA) node region of rabbit hearts, by means of electrophysiological techniques. Cyproheptadine (2-8 microM) decreased, in a dose-dependent manner, the plateau amplitude and the action potential duration to 50% repolarization of Purkinje and ventricular muscle cells. Higher concentrations also depressed the action potential amplitude, the overshoot and the maximum rate of rise of the upstroke. These effects were only partially reversed on washing. A four fold increase in Ca concentration of the standard Tyrode solution antagonized the effects of cyproheptadine on the action potential characteristics. The 'slow response' obtained in K-depolarized isoprenaline-treated fibres was blocked by cyproheptadine (4 microM). Cyproheptadine (10 microM) depolarized and suppressed the automaticity of spontaneously beating Purkinje fibres. The frequency of discharge of the SA node cells was slowed or abolished by cyproheptadine (2-4 microM). Atropine (2.6 microM) did not affect the negative chronotropic effect, whereas adrenaline (5 microM) reversed it. It is suggested that cyproheptadine depresses the slow inward current in all types of myocardial fibres studied. Higher concentrations might also affect the fast inward sodium current system.     

Relaxant actions of isoprenaline on guinea-pig isolated tracheal smooth muscle.

1. The effects of isoprenaline on membrane potential and intracellular Ca2+ concentration ([Ca2+]i) in guinea-pig isolated tracheal muscle were studied by use of intracellular micro-electrodes and fura-2 signals respectively. Measurements of membrane potential were carried out in the presence of spontaneously-generated muscle tone, whereas fura-2 signals were measured during contraction produced by exogenous prostaglandin E2 (100 nM). The potency of isoprenaline in causing relaxation was the same in these two different situations. 2. Isoprenaline (0.01 microM) produced relaxation accompanied by 5 mV hyperpolarization. A combination of tetraethylammonium (TEA, 10 mM) and verapamil (3 microM) did not alter the effects of isoprenaline. Removal of external K+ did not increase the degree of hyperpolarization produced by isoprenaline. 3. In the presence of TEA (10 mM) and verapamil (3 microM), isoprenaline (0.03-1 microM) reduced [Ca2+]i concentration-dependently. A similar degree of inhibition was observed when isoprenaline was applied during the maintained contraction induced by prostaglandin E2 and against the contraction evoked by the addition of Ca2+ to tissues bathed in a Ca(2+)-free medium and pretreated with both isoprenaline and prostaglandin E2. 4. It is concluded that activation of TEA-sensitive Ca(2+)-dependent K+ channels does not play a significant role in isoprenaline-induced relaxation. We propose that, in the guinea-pig tracheal muscle, isoprenaline may produce relaxation mainly by inhibiting a receptor-operated pathway for Ca2+ influx across the plasma membrane which is normally activated by prostaglandins.     

Effects of nitric oxide (NO) and NO donors on the membrane conductance of circular smooth muscle cells of the guinea-pig proximal colon.

1. The membrane conductance changes underlying the membrane hyperpolarizations induced by nitric oxide (NO), S-nitroso-L-cysteine (NC) and sodium nitroprusside (SNP) were investigated in the circular smooth muscle cells of the guinea-pig proximal colon, by use of standard intracellular microelectrode recording techniques. 2. NO (1%), NC (2.5-25 microM) and SNP (1-1000 microM) induced membrane hyperpolarization in a concentration-dependent manner, the hyperpolarizations to NO and NC developing more rapidly than those to SNP. The slower-developing responses to SNP were mimicked by the membrane permeable analogue of guanosine 3':5' cyclic-monophosphate (cyclic GMP), 8-bromo-cyclic GMP (500 microM), and by isoprenaline (10 microM). 3. The hyperpolarizations to NC and SNP were reduced in a low Ca2+ (0.25 mM) saline and upon the addition of haemoglobin (20 microM), but were not effected by NG-nitro-L-arginine (L-NOARG) (100 microM) or omega-conotoxin GVIA (100 nM). the hyperpolarizations to SNP were also significantly reduced by methylene blue (50 microM). 4. Apamin (250 nM) depolarized the membrane potential approximately 10 mV and reduced the initial transient component of the hyperpolarization to NO (1%) and NC (25 microM), but had no effects on the hyperpolarizations to SNP and cyclic GMP. Tetraethylammonium (TEA) (5-15 mM), had little effect on the membrane responses to NO(1%), NC(2.5-25 microM), SNP(100(-1000) microM) or cyclic GMP(500 microM). However, TEA (5-15 mM) reduced the membrane hyperpolarizations to SNP (10 microM) and isoprenaline (10 microM) in a concentration-dependent manner. The hyperpolarization to isoprenaline (10 microM) remaining in the presence of 15 mM TEA was blocked by ouabain (10 microM). 5. The amplitude of electronic potentials (1 s duration) elicited during NO donor hyperpolarizations were little changed or only slightly reduced (5-25%). However, the amplitude of the electrotonic potentials elicited during maintained electrically-induced hyperpolarizations of similar amplitude were significantly increased (30-150%), suggesting that the non-linear membrane properties of the proximal colon partially mask an increase in membrane conductance elicited during the NO donor hyperpolarizations. 6. Membrane hyperpolarization in the presence of an NO donor, 8-bromo-cyclic GMP, isoprenaline, or upon application of a maintained hyperpolarizing electrical current, often evoked oscillations of the membrane potential. These oscillations were prevented by Cs+ (1 mM). 7. These results indicate that NO and NC hyperpolarize the circular muscle of the proximal colon by activating at least two TEA-resistant membrane K+ conductances, one of which is sensitive to apamin blockade. The K+ conductance increases activated by SNP or 8-bromo-cyclic GMP were little effected by apamin, perhaps suggesting a common mechanism. In contrast, the hyperpolarization to isoprenaline appears to involve the activation of TEA-sensitive Ca2(+)-activated K+ ('BK') channels, as well as a Na:K ATPase. Finally, the 'background' membrane conductance of the circular muscle cells of the proximal colon decreased upon membrane hyperpolarization to reveal oscillations of the membrane potential which may well represent 'pacemaker' or 'slow wave' activity.     

Modulation of the isoprenaline-induced membrane hyperpolarization of mouse skeletal muscle cells.

1. The hyperpolarization of the resting membrane potential, Vm, induced by isoprenaline in the lumbrical muscle fibres of the mouse, was investigated by use of intracellular microelectrodes. 2. In normal Krebs-Henseleit solution (potassium concentration: K+o = 5.7 mM, 'control'), Vm was -7.40 +/- 0.2 mV; lowering K+o to 0.76 mM ('low K+o') resulted in either a hyperpolarization (Vm = -95.7 +/- 2.9 mV), or a depolarization (Vm = -52.0 +/- 0.3 mV). 3. Isoprenaline (> or = 200 nM) induced a hyperpolarization of Vm by delta Vm = -5.6 +/- 0.4 mV in control solution. 4. When Vm hyperpolarized after switching to low K+o, the addition of isoprenaline resulted in increased hyperpolarization Vm: delta Vm = -16.3 +/- 3.2 mV to a final Vm = -110.1 +/- 3.4 mV. Adding iso-prenaline when Vm depolarized in low K+o, leads to a hyperpolarization of either by -11.6 +/- 0.5 mV to -63.6 +/- 0.8 mV or by -51.7 +/- 2.7 mV to -106.9 +/- 3.9 mV. 5. Ouabain (0.1 to 1 mM) did not suppress the hyperpolarization by isoprenaline in 5.7 mM K+o (delta Vm = -6.7 +/- 0.4 mV) or the hyperpolarization of the depolarized cells in low K+- (delta Vm = -9.7 +/- 1.5 mV). 6. The hyperpolarization is a logarithmically decreasing function of K+o in the range between 2 and 20 mM (12 mV/decade). 7.IBMX and 8Br-cyclic AMP mimicked the response to isoprenaline whereas forskolin (FSK) induced in low K+o a hyperpolarization of -7.0 +/- 0.7 mV that could be augmented by addition of isoprenaline (delta Vm = -8.2 +/- 1.8 mV). 8. In control and low K+o, Ba2+ (0.6 mM) inhibited the hyperpolarization induced by isoprenaline, IBMX or 8Br-cyclic AMP. Other blockers of the potassium conductance such as TEA (5 mM) and apamin (0.4 microM) had no effect. 9. We conclude that in the lumbrical muscle of the mouse the isoprenaline-induced hyperpolarization is primarily due to an increase in potassium permeability.     

Electrophysiological effects of labetalol on canine atrial, cardiac Purkinje fibres and ventricular muscle.

1. Using conventional microelectrode techniques for the intracellular recordings of the membrane potential, the effects of labetalol were studied on cardiac Purkinje, atrial and ventricular muscle fibres of the dog. 2. Labetalol (1-10 microM) reduced, in a concentration-dependent manner, the action potential amplitude (APA) and the maximum rate of rise of the action potential (Vmax) in Purkinje fibres. 3. The action potential duration (APD) was decreased in Purkinje fibres but significantly increased in ventricular fibres after small concentrations of labetalol (1-3 microM). The atrial fibres were not very sensitive to labetalol. 4. Depolarization of the cardiac Purkinje fibres by increasing the external potassium concentration (8-12 mM), potentiated the labetalol effects on APA and Vmax but blocked its effects on the APD. 5. The effects of labetalol on Vmax of Purkinje fibres were dependent on the frequency of stimulation. 6. The ratio of the effective refractory period to the APD was increased both in normally polarized and depolarized Purkinje fibres after treatment with labetalol (10 microM). 7. Labetalol (10 microM) shifted the membrane responsiveness curve of Purkinje fibres by about 10 mV in the hyperpolarizing direction. 8. The slow response obtained in K-depolarized, Ba-treated Purkinje fibres was not significantly affected by labetalol (10-100 microM). 9. It is suggested that labetalol can exert Class I and Class III antiarrhythmic actions in cardiac muscle of the dog in vitro.     

Modulation of agonist-induced phosphoinositide metabolism, Ca2+ signalling and contraction of airway smooth muscle by cyclic AMP-dependent mechanisms.

1. The effects of increased cellular cyclic AMP levels induced by isoprenaline, forskolin and 8-bromoadenosine 3':5'-cyclic monophosphate (8-Br-cyclic AMP) on phosphoinositide metabolism and changes in intracellular Ca2+ elicited by methacholine and histamine were examined in bovine isolated tracheal smooth muscle (BTSM) cells. 2. Isoprenaline (pD2 (-log10 EC50) = 6.32 +/- 0.24) and forskolin (pD2 = 5.6 +/- 0.05) enhanced cyclic AMP levels in a concentration-dependent fashion in these cells, while methacholine (pD2 = 5.64 +/- 0.12) and histamine (pD2 = 4.90 +/- 0.04) caused a concentration-related increase in [3H]-inositol phosphates (IP) accumulation in the presence of 10 mM LiCl. 3. Preincubation of the cells (5 min, 37 degrees C) with isoprenaline (1 microM), forskolin (10 microM) and 8-Br-cyclic AMP (1 mM) did not affect the IP accumulation induced by methacholine, but significantly reduced the maximal IP production by histamine (1 mM). However, the effect of isoprenaline was small (15.0 +/- 0.6% inhibition) and insignificant at histamine concentrations between 0.1 and 100 microM. 4. Both methacholine and histamine induced a fast (max. in 0.5-2 s) and transient increase of intracellular Ca2+ concentration ([Ca2+]i) followed by a sustained phase lasting several minutes. EGTA (5 mM) attenuated the sustained phase, indicating that this phase depends on extracellular Ca2+. 5. Preincubation of the cells (5 min, 37 degrees C) with isoprenaline (1 microM), forskolin (10 microM) and 8-Br-cyclic AMP (1 microM) significantly attenuated both the Ca(2+)-transient and the sustained phase generated at equipotent IP producing concentrations of 1 microM methacholine and 100 microM histamine (approx. 40% of maximal methacholine-induced IP response), but did not affect changes in [Ca2+]i induced by 100 microM methacholine (95.2 +/- 3.5% of maximal methacholine-induced IP response). 6. Significant correlations were found between the isoprenaline-induced inhibition of BTSM contraction and inhibition of Ca2+ mobilization or influx induced by methacholine and histamine, that were similar for each contractile agonist. 7. These data indicate that (a) cyclic AMP-dependent inhibition of Ca2+ mobilization in BTSM cells is not primarily caused by attenuation of IP production, suggesting that cyclic AMP induced protein kinase A (PKA) activation is effective at a different level in the [Ca2+]i homeostasis, (b) that attenuation of intracellular Ca2+ concentration plays a major role in beta-adrenoceptor-mediated relaxation of methacholine- and histamine-induced airway smooth muscle contraction, and (c) that the relative resistance of the muscarinic agonist-induced contraction to beta-adrenoceptor agonists, especially at (supra) maximal contractile concentrations is largely determined by its higher potency in inducing intracellular Ca2+ changes.     

Inhibitory responses to nicotine and transmural stimulation in hyoscine-treated guinea-pig isolated trachealis: an electrical and mechanical study.

Guinea-pig isolated trachealis muscle treated with hyoscine (1 microM) exhibited mechanical tone which could be suppressed by transmural stimulation and, in a concentration-dependent manner, by nicotine (10-1000 microM). Hexamethonium (500 microM) did not itself cause tone changes, antagonized effects of nicotine but did not antagonize those of isoprenaline. Tetrodotoxin (0.3 microM) did not itself cause tone changes, did not modify the action of isoprenaline but antagonized the effects of nicotine and very markedly reduced responses to transmural electrical stimulation. Guanethidine (50 microM) did not itself cause tone changes, potentiated the action of isoprenaline, antagonized effects of nicotine and reduced responses to transmural electrical stimulation. Propranolol (1 microM) did not itself cause tone changes, antagonized effects of both isoprenaline and nicotine and reduced responses to transmural electrical stimulation. Propranolol (10 microM) caused greater antagonism of isoprenaline but did not further antagonize nicotine or further reduce responses to electrical stimulation. Intracellular electrophysiological recording from hyoscine-treated trachealis showed that 10 microM nicotine caused little or no mechanical or electrical change. Higher concentrations (100 microM and 1 mM) evoked relaxation which was often though not invariably accompanied by transient hyperpolarization and transient inhibition of electrical slow waves in the impaled cell. Hexamethonium (500 microM), tetrodotoxin (0.3 microM), guanethidine (50 microM) and propranolol (1 microM) each suppressed the electrical or mechanical changes evoked by nicotine (100 microM). However, nicotine (1 mM) tested in the presence of propranolol (1 microM), caused relaxation which could be accompanied by slow wave suppression but not by change in resting membrane potential. Transmural stimulation of hyoscine-treated trachea with single pulses of supramaximal voltage and 0.5 ms duration evoked neither relaxation nor membrane potential changes. Stimulation with similar pulses in trains of 5 s duration evoked relaxation which was dependent on pulse frequency. In many cells this relaxation was not accompanied by membrane potential change. In other cells suppression of slow waves occurred. At high pulse frequencies (greater than 16 Hz) this was generally accompanied by membrane hyperpolarization. In tissue treated with hyoscine and propranolol (both 1 microM), transmural stimulation with pulse trains as described above always evoked relaxation but no membrane potential changes were observed.(ABSTRACT TRUNCATED AT 400 WORDS)     

Augmentation by external Mg ions of beta-adrenoceptor-mediated actions in the longitudinal muscle of rat uterus.

1. The longitudinal muscle isolated from the uterus of oestrogen-treated rats was not spontaneously active in Locke solution, and electrical stimulation evoked phasic contraction. Isoprenaline (3 x 10(-11) - 10(-8) M) and dibutyryl cyclic AMP (db cyclic AMP, 0.1-0.8 mM) depressed the phasic contraction; the depression was enhanced in the presence of 0.6 mM Mg. 2. The contracture generated by 40 mM K was partially relaxed by isoprenaline (10(-11) - 10(-8) M) and db cyclic AMP (0.1-0.8 mM). Mg (0.6 mM) enhanced the isoprenaline-induced relaxation, but not that induced by db cyclic AMP. 3. The membrane potential of the muscle was -61 mV, and electrical stimulation induced an action potential which consisted of spike and plateau components. Application of isoprenaline and db cyclic AMP mainly reduced the duration of the plateau potential. The effect was potentiated by 0.6 mM Mg. 4. The membrane was hyperpolarized, accompanied by a decrease in membrane resistance, when 10(-8) M isoprenaline or 0.8 mM db cyclic AMP was applied. The effects of isoprenaline were prominently augmented in the presence of 1.2 mM Mg, while those of db cyclic AMP were slightly potentiated. 5. Forskolin (0.1 microM) or papaverine (10 microM) inhibited the phasic contraction and the K-contracture. The effect on the phasic contraction was potentiated by 0.6 mM Mg, while that on the K-contracture was not affected. 6. Forskolin shortened the action potential at 0.3 microM, and hyperpolarized the membrane with a decrease in membrane resistance at 3.0 microM. The membrane effects were augmented by 0.6 and 1.2 mM Mg, respectively. 7. It was hypothesized that external Mg ions could affect at least two processes involved in actions at beta-adrenoceptors on rat myometrium; receptor-agonist interaction and cyclic AMP-mediated inhibition of membrane excitability.     

Electrophysiological and other aspects of the relaxant action of isoprenaline in guinea-pig isolated trachealis.

In guinea-pig isolated trachealis isoprenaline (0.001-0.1 mumol l-1) caused concentration-dependent relaxation. Propranolol (1 mumol l-1) antagonized the effects of isoprenaline by more than 100 fold but did not modify the relaxant action of sodium nitrite. The tracheal relaxant actions of isoprenaline and ATP were unaffected by apamin (0.1 mumol l-1) but apamin profoundly antagonized the effects of noradrenaline and ATP on guinea-pig isolated taenia caeci. Tetraethylammonium (TEA; 8 mmol l-1) and procaine (5 mmol l-1) each evoked tracheal spasm but neither agent antagonized the isoprenaline-evoked relaxation of the trachealis. Trachealis exposed to K+-rich (120 mmol l-1) Krebs solution developed near-maximal tension. Both isoprenaline and sodium nitrite relaxed the K+-depolarized tissue though concentration-effect curves for both relaxants were moved to the right compared to those obtained in non-depolarized tissues. The maximal effect of sodium nitrite was markedly reduced. Intracellular electrophysiological recording showed that isoprenaline (0.01-1 mumol l-1) caused hyperpolarization and reduced or abolished slow wave discharge in trachealis muscle. These effects were accompanied by relaxation. Propranolol (1 mumol l-1) virtually abolished both the electrical and mechanical responses to isoprenaline (0.1 mumol l-1). Apamin (0.1 mumol l-1) did not alter the spontaneous electrical activity of trachealis cells or their electrical and mechanical responses to isoprenaline (0.1 mumol l-1). TEA (8 mmol l-1) caused depolarization and often increased slow wave amplitude and induced spike discharge. Isoprenaline (0.01 mumol l-1) failed to hyperpolarize TEA-treated trachealis cells. Higher concentrations of isoprenaline suppressed TEA-induced spasm, caused hyperpolarization and thereby increased slow wave or spike amplitude. Slow wave or spike frequency decreased as the hyperpolarization progressed but abolition of slow waves or spikes sometimes required more than 4 min exposure to isoprenaline. Procaine (5 mmol l-1) increased the amplitude of slow waves and induced spike discharge. Procaine markedly reduced the hyperpolarization induced by isoprenaline (0.1 and 1 mumol l-1) but had little effect on isoprenaline-induced relaxation. It is concluded that isoprenaline activates beta-adrenoceptors in guinea-pig trachealis and thereby evokes relaxation and hyperpolarization of the smooth muscle. The hyperpolarization does not involve the opening of apamin-sensitive K+-channels and it probably plays a supportive rather than a crucial role in the process by which isoprenaline-induced relaxation is achieved.     

Electrophysiological effects of the salicylates on isolated atrial muscle of the rabbit

1 Intracellular recordings were made from cells of the sinoatrial (S-A) node region and from atrial muscle fibres of rabbit hearts. The effects of sodium salicylate and 5-bromo salicylate on various parameters of the membrane action potential were studied.

2 5-Bromo salicylate (30-100 μM) and sodium salicylate (300-500 μM) caused a dose-dependent decrease in the frequency of discharge of the SA node cells. Applications of atropine (2.6 μM) with propranolol (3.3 μM) did not affect the negative chronotropic effect, whereas adrenaline (5 μM) reversed it.

3 Depolarization and shortening of the action potential duration were found in atrial muscle fibres after the application of 5-bromo salicylate (60-100 μM). The reduction of the action potential duration (APD) was not affected by atropine (2.6 μM).

4 Higher concentrations of 5-bromo salicylate (> 100 μM) also caused a dose-dependent reduction in the action potential amplitude (APA), in the overshoot (OS) of the action potential and in the maximum rate of rise of the action potential (V_max). All these effects were completely reversed on washing.

5 Substitution of the NaCl of the bathing Tyrode solution by an equimolar concentration of Na isethionate did not affect the plateau depression induced by the salicylates in atrial muscle fibres.

6 After increasing the K concentration to 27 mM in the presence of isoprenaline (1 μM), `slow responses' were obtained upon stimulation. 5-Bromo salicylate (20-60 μM) and sodium salicylate (100 μM) decreased reversibly the amplitude and the rate of rise of the `slow response'.

7 A four fold increase in Ca concentration of the standard Tyrode solution did not antagonize the plateau depression of atrial muscle fibres or the negative chronotropism induced by salicylates.

8 Addition of CsCl (10 mM) to the Tyrode solution did not affect the shortening of the APD induced by the salicylates in atrial muscle fibres.

9 When the K concentration in the Tyrode solution was increased from 2.7 mM to 5.4 mM, the effects of 5-bromo salicylate on the APA, OS and V_max were potentiated. However, a significant reduction in the shortening of the APD produced by the salicylate was observed.

10 It is suggested that the salicylates possibly depress the slow inward current in both S-A node cells and atrial muscle fibres of the rabbit heart. In atrial muscle fibres, a concomitant increase in the outward potassium current is probably involved.

     

The effects of alpha- and beta-adrenoceptor activation on tension and membrane properties of the longitudinal smooth muscle of the chicken rectum.

1. Isolated longitudinal muscle strips from the chicken rectum responded to isoprenaline, adrenaline and noradrenaline with a prolonged relaxation. The concentrations required to produce 50% of the maximum relaxation were 1.3 x 10(-8) M for isoprenaline, 1.7 x 10(-8) M for adrenaline and 10(-6) M for noradrenaline. The relaxing potency of isoprenaline is about equal to that of adrenaline, but more than 50 times that of noradrenaline. 2. Propranolol, 3.4 x 10(-6) M, blocked the isoprenaline-induced relaxation, and in the presence of this drug the responses to adrenaline and noradrenaline were converted into small, transient relaxations. The residual relaxation was blocked by phentolamine, 2.6 x 10(-6) M. 3. These catecholamines suppressed spontaneous spike discharge and produced membrane hyperpolarization. Propranolol, 3.4 x 10(-6) M, prevented the inhibitory effects of isoprenaline, and reduced but did not completely abolish those of adrenaline and noradrenaline. 4. Adrenaline and noradrenaline, but not isoprenaline, reduced membrane resistance in some preparations. 5. In the rectal muscle of the chicken, the beta-adrenoceptor mediates a prolonged relaxation and the alpha-adrenoceptor a fast and short-lasting relaxation which is usually obscured by the beta-response and unmasked only after blockade of the beta-adrenoceptors. The alpha- and beta-mediated relaxations are each associated with the suppression of spontaneous spike activity.     

Leukotriene receptor antagonism and augmentation of beta-receptor-mediated events by LY171883.

LY171883, (1-[2-hydroxy-3-propyl-4-[4(1H-tetrazol-5-yl)butoxy)phenyl]etha none), a leukotriene (LT) D4/E4 receptor antagonist, was assessed in comparison with two well known phosphodiesterase inhibitors, isobutylmethyl-xanthine (IBMX) and theophylline, for its ability to augment beta-receptor-mediated responses. Relaxation of carbachol-contracted guinea-pig trachea by isoprenaline was enhanced by the three agents in a dose-dependent manner. A two-fold enhancement of isoprenaline-induced smooth muscle relaxation was produced by 2.5 microM IBMX, 28 microM LY171883, or 140 microM theophylline. Similar concentrations of IBMX or theophylline did not antagonize LTE4-induced tracheal contractions; LY171883 totally inhibited the response and had significant LTE4 receptor antagonist activity even at 10-fold lower concentrations. Antigen-induced release of histamine and LTC4 from guinea-pig lung was reduced by isoprenaline. Prior treatment with LY171883, IBMX, or theophylline did not enhance this action. Isoprenaline reduced histamine-induced bronchospasm in anaesthetized guinea-pigs. LY171883, 30 mg kg-1, or IBMX, 1 mg kg-1, did not affect the isoprenaline-induced decrease in the histamine response. IBMX, 3 mg kg-1, and theophylline, 30 mg kg-1, augmented the isoprenaline-induced bronchodilation. LTE4-induced bronchoconstriction was not affected by IBMX or theophylline whereas LY171883 antagonized this response at doses as low as 3 mg kg-1. Therefore, in both in-vitro and in-vivo test systems, LY171883 functioned primarily as a leukotriene receptor antagonist with minimal pharmacological activity attributable to its ability to potentiate isoprenaline.     

Effects of isoprenaline and dibutyryl-C-AMP on the electrical and mechanical activity of the rabbit myometrium.

Mechanical and electrical responses of spontaneously contracting muscle strips from oestrogen dominated rabbit uterus were investigated by the sucrose gap method. The effects of isoprenaline and dibutyryl-c-AMP in the presence and absence of propranolol were tested. It has been shown previously that the beta-adrenoceptor agonist isoprenaline inhibits the spontaneously contracting uterus even in the presence of the beta-blocker propranolol. The rise in c-amp which is caused by isoprenaline, is, however, blocked by propranolol. The aim of this investigation was to study further the role of c-AMP in the isoprenaline-induced relaxation of the uterus. Isoprenaline (10(-6) M) consistently abolished spikes and mechanical contractions, usually accompanied by hyperpolarization. Propranolol (3 X 10(-6) M) markedly reduced the incidence of hyperpolarization, but did not affect the other actions of isoprenaline. Hyperpolarization does not seem to be a prerequisite for inhibition of uterine contraction. Dibutyryl-c-AMP mimicked the actions of isoprenaline. No positive evidence to support a hypothesis of a c-AMP-independent mechanism was found. Therefore a possible explanation of the results could be that isoprenaline increases c-AMP in a small intracellular pool, mediating electrical and mechanical responses but not influenced by propranolol.     

Phaclofen-insensitive presynaptic inhibitory action of (+/-)-baclofen in neonatal rat motoneurones in vitro.

1. Intracellular recordings were made from antidromically identified motoneurones in transverse spinal cord slices from neonatal (12-16 day) rats. 2. Superfusion of (+/-)-baclofen (0.5-50 microM) reduced the excitatory postsynaptic potentials (e.p.s.ps) and inhibitory postsynaptic potentials (i.p.s.ps) evoked by dorsal root or dorsal root entry zone stimulation in a concentration-dependent manner; the calculated EC50 was 2.4 microM. Baclofen in comparable concentrations also reversibly eliminated spontaneously occurring e.p.s.ps and i.p.s.ps. 3. (-)-Baclofen was more effective as compared to baclofen in reducing the synaptic responses, whereas (+)-baclofen at concentrations as high as 50 microM was ineffective. 4. Baclofen (less than 5 microM) attenuated the synaptic responses without causing a significant change of passive membrane properties and depolarizations induced by exogenously applied glutamate. In addition to synaptic depression, baclofen (greater than 5 microM) caused a hyperpolarization associated with decreased membrane resistance in some of the motoneurones; the glutamate responses were also attenuated. 5. Baclofen reversibly depressed the spike after-hyperpolarization of the motoneurones. 6. GABA (1-10 mM) depressed synaptic transmission and depolarized or hyperpolarized motoneurones. While potentiated by the uptake inhibitor nipecotic acid, the synaptic depressant effect of GABA was not antagonized by bicuculline. 7. The synaptic depressant effect of baclofen was neither blocked by GABAA antagonists bicuculline and picrotoxin (10-50 microM) nor by the GABAB antagonist phaclofen (0.1-1 mM).(ABSTRACT TRUNCATED AT 250 WORDS)     

The effects of phorbol 12,13-diacetate on responses of guinea-pig isolated trachea to methylxanthines, isoprenaline and ryanodine.

1. Using guinea-pig isolated trachea, we have studied how phorbol 12,13-diacetate (PDA) modulates mechanical responses of the tissue to methylxanthines, isoprenaline and ryanodine. 2. Caffeine (10 microM-5 mM), theophylline (10 microM-5 mM) and isoprenaline (1 nM-1 microM), each inhibited the spontaneous tone of the trachea. Pretreatment with PDA (0.1-10 microM) converted relaxant responses to high concentrations of the methylxanthines into contractions. PDA produced no equivalent effect against isoprenaline. Pretreatment with verapamil (1 or 10 microM), nifedipine (0.1 microM) or incubation with Ca(2+)-free, EGTA (0.1 mM)-containing physiological salt solution (PSS) suppressed the contraction produced by caffeine or theophylline in PDA (5 microM)-treated tissues. 3. The ability of PDA (5 microM) to convert caffeine-induced relaxation into caffeine-induced contraction was retained in tissues pretreated with a combination of atropine (1 microM) and mepyramine (1 microM) and in tissues denuded of the airway epithelium. 4. Caffeine (10 microM-5 mM), theophylline (10 microM-5 mM) and isoprenaline (1 nM-1 microM), each relaxed trachea contracted with histamine (0.1 mM). The relaxation induced by caffeine, theophylline and isoprenaline was markedly reduced in the presence of PDA (5 microM) and the responses to high concentrations of caffeine and theophylline, but not those to isoprenaline, were reversed to contractions. Verapamil (10 microM) prevented the effects of PDA against caffeine- or theophylline-induced relaxation. 5. PDA (1 microM) enhanced the tracheal spasm produced by caffeine (10 mM) and theophylline (10 mM) in indomethacin (2.8 microM)-treated trachea maintained at 20 degrees C. This enhancement was reduced in the presence of verapamil (10 microM).(ABSTRACT TRUNCATED AT 250 WORDS)     

Control of cyclic AMP levels in primary cultures of human tracheal smooth muscle cells.

1. [3H]-adenosine 3':5'-cyclic monophosphate ([3H]-cyclic AMP) responses were studied in primary cultures of human tracheal smooth muscle cells derived from explants of human trachealis muscle and in short term cultures of acutely dissociated trachealis cells. 2. Isoprenaline induced concentration-dependent [3H]-cyclic AMP formation with an EC50 of 0.2 microM. The response to 10 microM isoprenaline reached a maximum after 5-10 min stimulation and remained stable for periods of up to 1 h. After 10 min stimulation, 1 microM isoprenaline produced a 9.5 fold increase over basal [3H]-cyclic AMP levels. The response to isoprenaline was inhibited by ICI 118551 (10 nM), (apparent KA 1.9 x 10(9) M-1) indicating the probable involvement of a beta 2-adrenoceptor in this response in human cultured tracheal smooth muscle cells. However, with 50 nM ICI 118551 there was a reduction in the maximum response to isoprenaline. Prostaglandin E2 also produced concentration-dependent [3H]-cyclic AMP formation (EC50 0.7 microM, response to 1 microM PGE2 6.4 fold over basal). 3. Forskolin (1 nM - 100 microM) induced concentration-dependent [3H]-cyclic AMP formation in these cells. A 1.6 fold (over basal) response was also observed following stimulation with NaF (10 mM). 4. The nonselective phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX) (0.1 mM) and the type IV, cyclic AMP selective, phosphodiesterase inhibitor rolipram (0.1 mM) both elevated basal [3H]-cyclic AMP levels by 1.8 and 1.5 fold respectively. IBMX (1-100 microM) and low concentrations of rolipram (< 10 microM), also potentiated the response to 1 microM isoprenaline.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cromakalim-induced relaxation of guinea-pig isolated trachealis: antagonism by glibenclamide and by phentolamine.

1. Tested against the spontaneous tone of guinea-pig isolated trachealis, cromakalim (0.1-100 microM), isoprenaline (1 nM-1 microM) and theophylline (1 microM-1 mM) each produced concentration-dependent relaxation. 2. Glibenclamide (0.1-10 microM) did not itself alter the spontaneous tone of the trachea nor did it modify the relaxant actions of isoprenaline or theophylline. In contrast, glibenclamide (0.1 and 1 microM) caused a concentration-dependent rightward shift of the log concentration-effect curve of cromakalim. Glibenclamide (10 microM) reduced the slope of the log concentration-effect curve of cromakalim and moved the foot of the curve back towards the control position. 3. Phentolamine (1, 10 and 100 microm) did not itself alter the spontaneous tone of the trachea nor did it modify the relaxant actions of isoprenaline or theophylline. In contrast phentolamine caused concentration-dependent depression of the log concentration-effect curve of cromakalim. 4. Neither prazosin (1 microM) nor yohimbine (10 microM) modified the spontaneous tone of the trachea. Prazosin and yohimbine each failed to antagonise the effects of cromakalim, isoprenaline and theophylline. 5. Intracellular electrophysiological recording showed that glibenclamide (1 microM) and phentolamine (100 microM) caused minor change in the resting membrane potential of trachealis cells. Slow wave activity was slightly depressed by these agents. In contrast tetraethylammonium (TEA; 8 mM) caused marked depolarisation, and promoted the conversion of slow waves into regenerative action potentials. These electrical changes were accompanied by tonic tension development. 6. Phentolamine (100 microM) and glibenclamide (1 microM) reduced and reversed both the relaxation and the hyperpolarisation induced by cromakalim (10 microM). 7. It is concluded that glibenclamide and phentolamine each provide selective antagonism of the relaxant action of cromakalim in guinea-pig trachealis. These agents also inhibit the plasmalemmal hyperpolarisation induced by cromakalim. The effect of phentolamine is unrelated to the blockade of alpha 1- or alpha 2-adrenoceptors. If either glibenclamide or phentolamine act to block the K+ channels opened by cromakalim, then such channels are not identical to those which endow the trachealis plasmalemma with its powerful rectifying behaviour.     

Effects of Isoprenaline and Dibutyryl-C-AMP on the Electrical and Mechanical Activity of the Rabbit Myometrium

Abstract Mechanical and electrical responses of spontaneously contracting muscle strips from oestrogen dominated rabbit uterus were investigated by the sucrose gap method. The effects of isoprenaline and dibutyryl-c-AMP in the presence and absence of propranolol were tested. It has been shown previously that the β-adrenoceptor agonist isoprenaline inhibits the spontaneously contracting uterus even in the presence of the β-blocker propranolol. The rise in c-AMP which is caused by isoprenaline, is, however, blocked by propranolol. The aim of this investigation was to study further the role of c-AMP in the isoprenaline-induced relaxation of the uterus. Isoprenaline (10^-6 M) consistently abolished spikes and mechanical contractions, usually accompanied by hyperpolarization. Propranolol (3 × 10^-6 M) markedly reduced the incidence of hyperpolarization, but did not affect the other actions of isoprenaline. Hyperpolarization does not seem to be a prerequisite for inhibition of uterine contraction. Dibutyryl-c-AMP mimicked the actions of isoprenaline. No positive evidence to support a hypothesis of a c-AMP-independent mechanism was found. Therefore a possible explanation of the results could be that isoprenaline increases c-AMP in a small intracellular pool, mediating electrical and mechanical responses but not influenced by propranolol.