The consequences of loss followed by recovery of noradrenergic nerve function on muscarinic receptors in the chick expansor secundariorum muscle.

The effects of chemical sympathectomy with 6-hydroxydopamine on the response of the expansor secundariorum muscle to noradrenergic nerve stimulation, noradrenaline and acetylcholine have been investigated. Expansor muscles from 60 day old chicks were sensitive to noradrenergic nerve stimulation and exogenous noradrenaline but virtually unresponsive to acetylcholine. Chemical sympathectomy with 6-hydroxydopamine caused loss of function of noradrenergic nerves of the expansor muscle, induced supersensitivity to exogenous noradrenaline and gradually increased the response of the expansor muscle to acetylcholine. As the patency of noradrenergic nerves reappeared there was a decline in the extent of supersensitivity to noradrenaline and the response to acetylcholine gradually declined. The time courses of these changes differed, indicating that the mechanisms responsible for changes in response to noradrenaline and acetycholine are different.     

Effect of potassium on the contractile response of the rat detrusor muscle to field stimulation at increasing calcium concentrations

• 1.1. At 0.6 and 1.8 mM calcium, increasing the potassium concentration from 2.5 to both 7.5 and 12.5 mM significantly increased the contractile response to field stimulation at 0.5, 1, 2 and 4 Hz. There were no effects on the contractile responses to 8, 16, 32 and 64 Hz. 22.5 mM potassium significantly reduced the contractile response to all frequencies of stimulation.
• 2.2. At 5.4 mM calcium, the contractile responses to all frequencies in the presence of 2.5, 7.5 and 12.5 mM potassium were similar. The responses to all frequencies of stimulation in the presence of 22.5 mM potassium were significantly reduced.
• 3.3. In general, the relative responses to low frequencies of stimulation were significantly greater in the presence of 5.4 mM calcium than in the presence of 0.6 or 1.8 mM calcium.
• 4.4. The magnitude of the inhibition in the presence of 22.5 mM potassium was inversely proportional to the extracellular calcium concentration, i.e. the lower the calcium concentration, the greater the inhibition.
• 5.5. The bladder base responded to alterations in the potassium and calcium concentrations similarly to the bladder body.
• 6.6. In conclusion, increasing the potassium concentration from 2.5 mM (normal bath concentration) to 7.5 and 12.5 mM significantly enhanced the contractile response to low frequency stimulation at both 0.6 and 1.8 mM (normal bath concentration) calcium.

     

The influence of prostaglandins on the contractile response to electrical field stimulation in rabbit myometrial and cervical smooth muscles

Indomethacin (10 μM) significantly reduced the contractile responses to electrical field stimulation (EFS) in the non-pregnant myometrium (5–50 Hz by an average of 25.7%) and the pregnant myometrium (30–50 Hz by an average of 16.5%). On the other hand, it significantly increased the contractile responses of the cervix in non-pregnant (2.5-30 Hz by an average of 21.5%) and pregnant (2.5-10 Hz by an average of 12.5%) muscular strips. Prostaglandin (PG) E ₂ (0.1-1000 nM) strongly and PGI₂ (0.1-1000 nM) slightly and dose-dependently inhibited the contractile response to EFS of the non-pregnant cervical muscle, but not of the myometrium. In the pregnant uterus, the contractile responses of the cervical muscle were similar to those of the non-pregnant cervix, while the response of the myometrium was increased significantly and dose-dependently by the appliaction of PGE₂ or PGI₂. PGF_2θ (1000 nM) significantly increased the contractile response of the myometrium but not cervical muscle to EFS in the pregnant rabbits. In non-pregnant muscles, the contractile responses of the myometrium and cervical smooth muscle to EFS were not influenced by the treatment with PGF₂. PGE₂ (0.1-1000 nM) also inhibited the contractile response to direct muscle stimulation, but the inhibition was significantly less than that of the response to indirect stimulation. PGI₂ slightly inhibited the contractile responses to both indirect and direct stimulation. These findings indicate the PGE₂, PGI₂ and PGF₂ increase the contractile response of pregnant myometrium to EFS. PGE₂ has both a pre- and a post-junctional inhibitory effect on excitatory transmission in the cervix, but PGI₂ acts only post-junctionally. It is suggested that the physiological action PGE₂ and of PGI₂ is exerted differently on the myometrium and cervical smooth muscles.     

The role of hydrogen peroxide in the contractile response to angiotensin II

In the last years, reactive oxygen species (ROS) have been proposed as mediators of proliferative/hypertrophic responses to angiotensin II (Ang II), both in vivo and in vitro. However, the hypothesis that the Ang II-dependent cell contraction could be mediated by ROS, particularly H2O2, has not been tested. Present experiments were devoted to test this hypothesis and to analyze the possible mechanisms involved. Catalase (CAT) prevented the increased myosin light chain phosphorylation and the decreased planar cell surface area (PCSA) induced by 1 microM Ang II in cultured rat vascular smooth muscle cells (VSMC). This preventive effect of CAT was also detected when 1 microM platelet-activating factor (PAF) was used as a contractile agonist instead of Ang II. Similar results were found when using horseradish peroxidase as an H2O2 scavenger or cultured rat mesangial cells. In vascular smooth muscle cells, CAT modified neither the binding of labeled Ang II nor the Ang II-induced inositol 1,4,5-trisphosphate (IP3) synthesis. However, it completely abolished the Ang II-dependent calcium peak, in a dose-dependent fashion. CAT-loaded cells (increased intracellular CAT concentration over 3-fold) did not show either a decreased PCSA or an increased intracellular calcium concentration after Ang II treatment. Ang II stimulated the H2O2 synthesis by cultured cells, and the presence of CAT in the extracellular compartment significantly diminished the Ang II-dependent increased intracellular H2O2 concentration. The physiological importance of these findings was tested in rat thoracic aortic rings: CAT prevented the contraction elicited by Ang II. In summary, present experiments point to H2O2 as a critical intracellular metabolite in the regulation of cell contraction.     

Inhibition by propranolol of the contractile response of the rat diaphragm to tetanic field stimulation in vitro.

Contraction of the rat isolated diaphragm in response to maximal tetanic stimulation was examined before and after isoprenaline or propranolol. Isoprenaline (10(-4)M) did not affect maximum isometric force, whereas propranolol depressed maximum force in a concentration-dependent manner (10(-6)-10(-4)M). Inhibition due to propranolol (10(-4)M) could not be overcome by increasing the intensity or duration of electrical stimulation, and was only partially reversed (mean 73% +/- 10 s.e. mean) after washing. Pretreatment with isoprenaline did not alter the response of the muscle to propranolol, nor did neuromuscular blockade with (+)-tubocurarine. The response to either stereoisomer of propranolol was similar to that obtained with the racemate. Atenolol, a beta-adrenoceptor blocking agent without membrane stabilizing activity, had minimal (less than 10%) depressant effects on diaphragmatic force development. Lignocaine (8.5 X 10(-6)-8.5 X 10(-5)M) produced a concentration-related decrease in isometric force, similar to that with propranolol. It is concluded that propranolol decreases the contractile force of the rat isolated diaphragm by a mechanism related to stabilization of excitable membranes.     

Endotoxin impairs the response of rat anococcygeus muscle to electrical field stimulation.

Rat anococcygeus muscles were isolated 4 h after treatment with either E. coli endotoxin (20 mg kg-1 i.p.) or saline. The contractile responses of the muscle to electrical field stimulation, and to the alpha 1-adrenoceptor agonist cirazoline were impaired by endotoxin treatment. Inhibition of the L-arginine pathway with L-NG-nitro arginine methylester (3 x 10(-5) M) failed to restore responsiveness to either electrical field stimulation or to cirazoline. Endotoxin impaired inhibitory responses to electrical field stimulation in preparations precontracted with cirazoline. Responsiveness to sodium nitroprusside was also impaired by endotoxin under these conditions. These results show an impairment by endotoxin of both contractile and inhibitory responses of the rat anococcygeus muscle to electrical field stimulation. The impairment appears to be post-junctional. In contrast to the findings in vascular smooth muscle, the endotoxin-induced impairment of contractile responses does not appear to involve the L-arginine pathway.     

The comparative effects of aminoglycoside antibiotics and muscle relaxants on electrical field stimulation response in rat bladder smooth muscle

It has been reported that several aminoglycoside antibiotics have a potential of prolonging the action of non-depolarizing muscle relaxants by drug interactions acting pre-synaptically to inhibit acetylcholine release, but antibiotics itself also have a strong effect on relaxing the smooth muscle. In this study, four antibiotics of aminoglycosides such as gentamicin, streptomycin, kanamycin and neomycin were compared with skeletal muscle relaxants baclofen, tubocurarine, pancuronium and succinylcholine, and a smooth muscle relaxant, papaverine. The muscle strips isolated from the rat bladder were stimulated with pulse trains of 40 V in amplitude and 10 s in duration, with pulse duration of 1 ms at the frequency of 1–8 Hz, at 1, 2, 4, 6, 8 Hz respectively. To test the effect of four antibiotics on bladder smooth muscle relaxation, each of them was treated cumulatively from 1 μM to 0.1 mM with an interval of 5 min. Among the four antibiotics, gentamicin and neomycin inhibited the EFS response. The skeletal muscle relaxants (baclofen, tubocurarine, pancuronium and succinylcholine) and inhibitory neurotransmitters (GABA and glycine) did not show any significant effect. However, papaverine, had a significant effect in the relaxation of the smooth muscle. It was suggested that the aminoglycoside antibiotics have inhibitory effect on the bladder smooth muscle.     

The signaling of amitriptyline-induced inhibitory effect on electrical field stimulation response in colon smooth muscle

Amitriptyline, a well-known antidepressant, exerts inhibitory effect on electrically stimulated rat colon smooth muscle contraction. In this study, we investigated the signaling pathway of amitriptyline-induced inhibitory effect. Changes in isometric force of colon muscle were recorded on polygraph, and data were analyzed by measuring the inhibitory extent induced by amitriptyline. Firstly, muscles were contracted by stimulation with electric field stimulation (EFS), and then, amitriptyline was added cumulatively to determine its influence effect on EFS. Amitriptyline significantly inhibited EFS-induced contraction dose dependently. Then, the mechanism of inhibitory effect of amitriptyline was evaluated by pretreating with various antagonists such as L-NAME, methylene blue, atropine, 5-HT receptors blockers, guanethidine, prazosin, guanabenz, isoprenaline, Y27632 (Rho-kinase inhibitor), ML9 (myosin light chain kinase (MLCK) inhibitor), U73122 (PLC inhibitor), and chelerythrine (PKC inhibitor). Then, Ca²⁺ channel blocker (nifedipine) and K⁺channel blockers, tetraethylammonium (TEA), 4-aminopyridine (4-AP), and glybenclamide, were used to determine the involvement of ion channels. L-NAME, guanabenz, 5HT₄ receptor blocker, ML9, and Y27632 enhanced the effect of amitriptyline. Meanwhile, methylene blue, atropine, guanethidine, prazosin, methylsergide, ondansetron, U73122, and chelerythrine blocked its effect. It was also shown that nifedipine enhanced but TEA and glybenclamide blocked amitriptyline-induced inhibitory effect on EFS. Our results indicated that amitriptyline may exert inhibitory effect in response to EFS by inhibiting muscarinic receptors and then PLC-mediated PKC pathway leading to opening of ATP-sensitive potassium channel.     

Effects of pyroxamidine and guanethidine on contractile responses to field stimulation and to noradrenaline in the anococcygeus muscle and vas deferens of the rat

The effects of pyroxamidine (EMD 21192) and guanethidine on contractile responses were studied in the anococcygeus muscle and vas deferens of the rat. Pyroxamidine (10^?6 and 10^?5 M) and guanethidine (6 times 10^?6 and 10^?5 M) potentiated the responses to low concentrations of acetylcholine in the rat anococcygeus muscle. Following incubation of the muscle with 6-hydroxydopamine (10^?3M for 3 h), pyroxamidine (10^?5 M) and guanethidine (10^?5 M) had no effect on responses to acetylcholine. This suggests that the potentiating effect of pyroxamidine and guanethidine on responses to acetylcholine is due to the release of subthreshold concentrations of noradrenaline. In the anococcygeus, pyroxamidine (10^?6 and 10^?5 M) and guanethidine (10^?6 and 10^?5 M) inhibited responses to field stimulation and potentiated responses to exogenously applied (?)-noradrenaline. The responses to field stimulation in the vas deferens were also inhibited by 10^?5M pyroxamidine and by 10^?5 M guanethidine. 10^?6 M guanethidine, but not 10^?5 M pyroxamidine, potentiated responses to (?)-noradrenaline in the vas deferens. In the presence of nortriptyline (10^?6 M), a potent inhibitor of neuronal uptake, the inhibitory effects of pyroxamidine and guanethidine on responses to field stimulation were reduced or reversed and these drugs had no effect on responses to (?)-noradrenaline. This suggests that pyroxamidine is a noradrenergic neuron blocker and that its action is dependent on continued neuronal uptake. Following 6-hydroxydopamine incubation, 10^?5 M pyroxamidine and 10^?5 M guanethidine inhibited the responses to (?)-noradrenaline in the rat anococcygeus muscle. Thus it seems likely, at high concentrations, that these compounds have postsynaptic blocking activity.     

Effects of acetaldehyde on contractile response to nerve stimulation in guinea-pig vas deferens

Twitch contractions of the isolated guinea-pig vas deferens induced by sympathetic nerve stimulation were augmented by acetaldehyde (0.1-10 mM). With high concentrations (5-10 mM), acetaldehyde produced a biphasic response consisting of an initial brief depression and a subsequent potentiation of the contraction. The late effect was associated with repetitive contractions that were not prevented by tetrodotoxin. A low concentration of phentolamine (27 microM) increased and a high concentration (1.3 mM) suppressed the potentiating action of acetaldehyde. Acetaldehyde did not induce contractions in surgically sympathectomized vasa or vasa pretreated with reserpine. Acetaldehyde caused a dose-dependent increase in noradrenaline release into the bathing fluid. The study shows that acetaldehyde has a dual effect on sympathetic neuroeffector transmission, and that an increase in noradrenaline secretion appears to contribute to the late facilitatory effect in the isolated vas deferens.     

Adenosine and dipyridamole: actions and interactions on the contractile response of guinea-pig ileum to high frequency electrical field stimulation.

The action of adenosine on the myenteric plexus-longitudinal muscle strips from guinea-pig ileum to high frequency electrical field stimulation (10 Hz) was investigated. Electrically induced contractions were reduced markedly by tetrodotoxin (0.2 microM) and atropine (1 microM), and partially by noradrenaline (3 microM) and morphine (3 microM). Adenosine, adenosine 5'-monophosphate (AMP) and adenosine triphosphate (ATP) produced a concentration-dependent inhibition of the high frequency contractions over the range of 0.1-100 microM, the most potent being adenosine. The concentration-response curve for adenosine was significantly shifted to the left by dipyridamole (10 nM), while dipyridamole at higher concentrations (30 nM-10 microM), depressed the contraction markedly by itself. Dipyridamole decreased [3H]-adenosine uptake into strips of ileum in a concentration-dependent manner. There was a significant correlationship between the reduction of adenosine uptake and the inhibition of the contraction induced by dipyridamole (r = 0.970). In strips desensitized to adenosine or treated with adenosine deaminase, the inhibitory effect of dipyridamole was significantly reduced. The present investigation revealed that adenosine depressed responses of guinea-pig ileum to high frequency electrical stimulation and suggested that the inhibitory effect of dipyridamole may be closely associated with the behaviour of endogenous adenosine or related compounds.     

Adrenergic mechanisms of the contractile response of rabbit oviduct to electrical stimulation

1. In rabbit oviduct electrical field stimulation evoked contraction in mature animals and contraction followed by relaxation in juvenile animals. 2. Phentolamine (1 microM) inhibited the contraction. The relaxation was partly suppressed by propranolol (1-10 microM) and completely inhibited by guanethidine (5 microM). 3. Cocaine (1 microM) increased the duration or the amplitude of contractions and abolished or did not change the relaxation. The contraction-increasing effect of cocaine was more pronounced in estradiol-treated mature rabbit. 4. It is concluded that electrically evoked responses in rabbit oviduct are mediated by transmitters released from adrenergic terminals and that estradiol increases neuronal uptake of noradrenaline.     

Superoxide anion mediates angiotensin II-induced potentiation of contractile response to sympathetic stimulation

Angiotensin II is known to potentiate vasoconstriction induced by electrical field stimulation (EFS), but the underlying mechanisms for this potentiation are not fully understood. This study was designed to investigate the role of superoxide anion in the potentiation effects of angiotensin II. Contraction of rat mesenteric arterial segments was induced by perivascular nerve stimulation with EFS, and superoxide production was measured with lucigenin-enhanced chemiluminescence. Extracellular signal-regulated kinase (ERK) phosphorylation was determined in cultured smooth muscle cells with Western blot. Angiotensin II concentration dependently potentiated the contraction of rat mesenteric arteries to EFS, which is frequency-dependent. This potentiation was blunted by an angiotensin AT(1) receptor antagonist (2-ethoxy-1-[[2'-(1H-tetrazol-5-yl)biphenyl-4-yl]methyl]-1H-benzimidazole-7-carboxylic acid, CV-11974), NAD(P)H oxidase inhibitor (apocynin), superoxide dismutase (SOD) and its mimetic tiron, but not affected by angiotensin AT(2) receptor antagonist and inhibitors of xanthine oxidase, cytochrome P450, and cyclooxygenase. Angiotensin II increased superoxide production by mesenteric arteries, which was blunted by angiotensin AT(1) receptor antagonist CV-11974, and NAD(P)H oxidase inhibitor apocynin. Superoxide generating compound pyrogallol mimicked the effects of angiotensin II. Tyrosine kinase inhibitor (tyrphostin A25) and mitogen-activated protein kinase (MAPK)/ERK inhibitors (1,4-diamino-2,3-dicyano-1,4-bis [2-aminophenylthio]butadiene (U 0126)) inhibited angiotensin II- and pyrogallol-induced potentiation of EFS-induced contraction, while inactive forms of these inhibitors did not show any inhibitory effects. In cultured smooth muscle cells from mesenteric arteries, angiotensin II and superoxide similarly induced ERK phosphorylation. These results showed that superoxide mediated angiotensin II-induced potentiation of contractile response to EFS and tyrosine kinase-MAPK/ERK activation was involved.     

The rat anococcygeus muscle and its response to nerve stimulation and to some drugs

1. A new smooth muscle preparation, the rat anococcygeus muscle, is described. The muscle is paired, thin, consists of smooth muscle only and the muscle cells are organized in parallel bundles. It has a dense adrenergic innervation distributed throughout the muscle but apparently no cholinergic innervation. The muscles are easily isolated.2. The muscle contracts to noradrenaline, acetylcholine, furmethide, 5-hydroxytryptamine, but not to histamine. Isoprenaline produces contraction at high concentrations. The effects of noradrenaline and acetylcholine are blocked by phentolamine and atropine respectively. The response to isoprenaline is little affected by propranolol.3. The muscle contracts in response to field stimulation or stimulation of extrinsic nerves. This response is completely blocked by phentolamine but unaffected by hexamethonium or atropine.4. Guanethidine 10(-6)-5 x 10(-6)M blocks the motor response to nerve stimulation and potentiates that to noradrenaline. Higher concentrations of guanethidine raise tone. In the presence of raised tone, field stimulation produces an inhibitory response insensitive to hexamethonium but abolished by tetrodotoxin 2 x 10(-7) g/ml. This inhibitory response to stimulation can also be shown after other drugs which raise tone.5. The inhibitory response to nerve stimulation is not mimicked by acetylcholine, isoprenaline or ATP, nor blocked by atropine, phentolamine, phenoxybenzamine, propranolol, hexamethonium or lysergic acid diethylamide.     

Type of electrical stimulation influences diaphragm response to adrenoceptor and calcium channel modulators: the role of extracellular and intracellular calcium events

The influence of direct single pulse and subtetanic electrical stimulation (ES) on isolated rat hemidiaphragm response to adrenoceptor antagonists and calcium channel blockers was investigated. Muscle contractility was stimulated by cumulative micromolar noradrenaline. Noradrenaline effects on developed tension (Td) in the presence of various alpha- and beta-adrenoceptor antagonists (e.g., prazosin and ICI 118551) were qualitatively different during different types of ES. Also, intact L-voltage calcium channels were necessary for noradrenaline-induced potentiation of Td during both types of ES, while the balance between ryanodine receptors- and inositol triphosphate (IP3)-related calcium events in the muscle was influenced by the pattern of the ES.     

The inhibitory effect of furosemide on the contractile response of equine trachealis to cholinergic nerve stimulation

The effects of furosemide on the responses of equine trachealis muscle with and without epithelium to electrical field stimulation (EFS) and exogenous acetylcholine (ACh) were investigated in organ baths. Tissues were pretreated with guanethidine and the parameters used for EFS were those previously demonstrated to activate postganglionic cholinergic neurons. In tissues with intact epithelium, furosemide (100 μm) shifted the frequency-response curve to the right. In the preparations without epithelium, furosemide did not affect the response to EFS. Neither in epithelium-on nor in epithelium-off tissues was the ACh dose-response curve affected by the administration of furosemide. We conclude that furosemide (100 μm) decreases the equine tracheal smooth muscle responses to EFS through inhibition of cholinergic neurotransmission, and that this effect is dependent on epithelium.