Contractile effects of endothelin-1 and localization of endothelin binding sites in rabbit lower urinary tract smooth muscle

In isolated rabbit bladder and urethral smooth muscle, endothelin-1 caused concentration-related, slowly developing contractions that were difficult to wash out. Relative to contractions induced by K+ (124 mM), contractions in bladder preparations reached a higher amplitude than in urethral preparations. There was a marked tachyphylaxis to the effects of the peptide. The endothelin-1-induced contractions were not significantly affected by phentolamine or indomethacin in the urethra, or by scopolamine or indomethacin in the bladder. Incubation for 30 min in a Ca2(+)-free solution abolished the endothelin-1-induced contractions. Nifedipine did not affect the actions of endothelin-1 in the urethra but had a marked inhibitory action on its effects in the bladder. In the presence of endothelin-1, Ca2(+)-induced contractions were significantly blocked by nifedipine in the bladder but not in the urethra. Urethral preparations at resting tension responded to electrical stimulation by tetrodotoxin-sensitive, frequency-dependent contractions sensitive to alpha-adrenoceptor blockade. Pretreatment with endothelin-1 (10(-9) M) produced a significant increase in the nerve-induced contractions but had no significant effect on contractions induced by exogenous noradrenaline. Endothelin-1 did not affect spontaneous or stimulation-induced efflux of 3H-labelled noradrenaline in urethral smooth muscle. Preparations contracted by endothelin-1 were frequency-dependently relaxed by electrical stimulation. The peptide had no significant effect on the responses induced by electrical stimulation in the bladder preparations. In both bladder and urethra, [125]endothelin-1 binding sites were found mainly in the outer longitudinal muscle layer, in vessels and in the submucosa. The highest density of binding sites appeared to be in vessels and the outer muscle layer in both types of muscle. The results suggest that in the rabbit both bladder and urethral smooth muscle contain binding sites for endothelin. The peptide has contractant effects dependent on extracellular calcium in both types of tissue, but voltage-operated calcium channels seem to involved in activation only of bladder smooth muscle. The functional importance of endothelin-1 in the rabbit lower urinary tract remains to be elucidated.     

Different effects of neuropeptide Y on electrically induced contractions in the longitudinal and circular smooth muscle layers of the female rabbit urethra

The effects of neuropeptide Y (NPY) on preparations of isolated longitudinal and circular smooth muscle from rabbit urethra were studied. In both types of muscle, electrically induced contractions and relaxations could be abolished by tetrodotoxin, (TTX). In the longitudinal muscle preparations the contraction was slightly reduced by prazosin, but markedly reduced by scopolamine and NPY. The NPY effect was not influenced by pretreatment with rauwolscine. Pretreatment with NPY had no effect on contractions induced by noradrenaline (NA) or carbachol and the peptide did not relax preparations contracted by these agents. In circular muscle an initial, fast response, not sensitive to prazosin or scopolamine was occasionally observed following electrical stimulation. A slow contraction component was regularly seen; this response was abolished by prazosin. Neuropeptide Y did not influence any of these responses. The preparations were concentration-dependently contracted by NA, whereas carbachol had no effect. Pretreatment with NPY did not affect contractions induced by NA, nor did the peptide relax NA-contracted preparations. In neither longitudinal nor circular muscle strips did NPY affect the electrically induced TTX sensitive relaxation of NA-contracted preparations. The results suggest that in the rabbit urethra NPY reduces contractions in the longitudinal muscle layer by selectively inhibiting the release of acetylcholine from cholinergic nerves. Neuropeptide Y did not appear to have any significant postjunctional effects nor to interfere with the release, or effects of NA or other transmitter agents. The physiological importance of the urethral effects of NPY remains to be established.     

Difference in the actions of calcitonin gene-related peptide on pig detrusor and vesical arterial smooth muscle

Calcitonin gene-related peptide has been demonstrated in urinary bladder nerves, and suggested to play a role in local control of bladder motility. In isolated strips of pig detrusor muscle, calcitonin gene-related peptide did not affect spontaneous contractile activity, or contractions induced by high K+, carbachol, substance P, and electrical field stimulation. In contrast, calcitonin gene-related peptide elicited a concentration-dependent and pronounced (78-99%) relaxation of vesical arteries precontracted with endothelin-1, noradrenaline or prostaglandin F2 alpha. As a vasodilator, CGRP was approximately 50 times more potent than acetylcholine. Removal of the endothelium abolished acetylcholine-induced relaxation, but did not affect the relaxation produced by calcitonin gene-related peptide. Pretreatment with methylene blue, glibenclamide or indomethacin had no influence on CGRP's ability to relax the vessels. The inhibitor of NO-synthesis, NG-nitro-L-arginine, had no effect on the maximum vascular relaxation induced by calcitonin gene-relate peptide. It is concluded that in the pig, calcitonin gene-related peptide has no functionally important mechanical effects on isolated detrusor muscle strips, but is a potent dilator of vesical arteries. The vascular effects of the peptide are endothelium-independent, and seem to be exerted directly on the vascular smooth muscle.     

Functional responses of different muscle types of the female rat urethra in vitro

The influence of muscle type on functional responses of the female rat urethra was investigated using morphological and functional in-vitro techniques. The urethral submucosa was found to contain longitudinally or obliquely oriented smooth muscle cells. The muscularis layer is composed of circularly oriented muscle cells. Near the bladder orifice smooth muscle fibres dominate, but in the mid-urethra the vast majority is circularly oriented striated muscle cells. Circular preparations responded to electrical field stimulation in vitro with a rapid contraction. Stimulation with single impulses resulted in a twitch response; frequencies exceeding 5-10 Hz induced a summation and tetanus. The response was unaffected by phenoxybenzamine, propranolol and scopolamine and had a low sensitivity to calcium-free solution but was sensitive to suxamethonium and tetrodotoxin. Using longer impulse trains stimulation evoked also a slow contraction, sensitive to calcium-free solution. In longitudinal preparations stimulation induced a relaxation followed by a contraction, responses much smaller than those seen in the circular preparations. Both preparations relaxed on addition of calcitonin gene-related peptide or capsaicin. The relaxation to calcitonin gene-related peptide was larger than that to capsaicin in longitudinal preparations but equally large in the circular ones. Substance P and 5-hydroxytryptamine contracted both preparations. The longitudinal urethra showed a larger contraction to 5-hydroxytryptamine than to substance P, whereas both substances induced similar responses in the circular preparations. The study shows a similar muscle arrangement in the female rat urethra as described in humans and further points to a functional differentiation between the different types of muscle.     

Effects of Prostaglandins on the Isolated Human Bladder and Urethra

The effects of prostaglandins F_2α (PGF_2α), E₂ (PGE₁) and E₂ (PGE₂) on the human lower urinary tract were studied in vitro in preparations obtained from patients undergoing total cystourethrectomy because of bladder malignancy. Tissue specimens were taken from different parts of the urethra, the urethrovesical junction, and the bladder. From these specimens, smooth muscle preparations were dissected and mounted in organ baths, that were filled with Krebs solution (37°C) and bubbled with carbogen. Isometric tension was recorded. Preparations from the bladder and all parts of the urethra were contracted by PGF_2α. This effect was not affected by tetrodotoxin, phenoxybenzamine, or atropine; isoprenaline relaxed the PGF_2α induced contractions. PGE₁ and PGE₂ both contracted strips from the bladder. However, urethral preparations contracted by PGF_2α or noradrenaline were relaxed by these agents. This relaxing effect was at least as pronounced as that produced by isoprenaline; it was not affected by propranolol.     

Tetrodotoxin-resistant contractions induced by electrical stimulation of bladder muscle from man, rabbit and rat

Isolated detrusor preparations from man, rabbit and rat were suspended in an organ bath and isometric tension was recorded. The preparations were stimulated electrically in the presence of Bay K8644 and nifedipine before and after neuronal blockade with tetrodotoxin. Transmural electrical stimulation produced frequency-dependent contractions in all preparations. Bay K8644 significantly increased and nifedipine decreased these contractions. TTX effectively suppressed the response to electrical field stimulation in all species. When Bay K8644 was added to TTX blocked preparations, the responses to electrical stimulation were partly restored in bladder strips from man and rat. No increase in response was seen in the rabbit preparations. However, if the extracellular K+-concentration was increased to 10 mM (which per se did not affect the response) Bay K8644 significantly increased the contractions. All responses elicited by electrical stimulation in the presence of TTX were abolished by nifedipine. It is concluded that if the bladder smooth muscle is exposed to factors that can increase its sensitivity to contractile agents, this may result in uncontrolled (unstable) bladder contractions. Such contractions may use the 'normal' transmitter substances, but may be triggered at a lower stimulus intensity than normal. As a non-specific increase in membrane excitability seems to be associated with an influx of calcium through voltage-sensitive calcium channels, calcium antagonists, together with agents specifically blocking relevant transmitter substances, would offer an effective therapy against the unstable bladder.     

Influence of antimuscarinics on alpha-adrenoceptors in the female rabbit urethra

The effects of the tertiary amine atropine and its structural analogues homatropine and scopolamine, as well as the quarternary amine emeprone, were evaluated on noradrenaline (NA)-induced contractions of isolated female rabbit urethral ring preparations. In addition, the abilities of these antimuscarinics to inhibit ³H—dihydro—alpha—ergocryptine (³H—DHE) binding to alpha—adrenoceptors were studied on a crude membrane preparation from the female rabbit bladder base and urethra. Atropine and homatropine depressed the NA- induced contractions in a concentration-dependent way, whereas this was not seen with scopolamine. Emeprone 10^-5 10^-4 M augmented the contractions, an effect possibly attributable to a NA-uptake blocking effect. All antimuscarinics displaced specific ³H— DHE binding, the order of potency being atropine>homatropine>emeprone>scopolamine. In general a good correlation was seen between the binding and mechanical activity studies for atropine, homatropine and scopolamine, while this was not found for emeprone. It is concluded that alpha-adrenoceptor blockade by atropine can be observed at concentrations exceeding 10^-7 M. Scopolamine, showing alpha-adrenoceptor blocking properties only in high concentrations, may be used as an alternative for blockade of muscarinic cholinoceptors.     

Calcitonin gene-related peptide augments parasympathetic contraction of rabbit tracheal smooth musclein vitro

The effect of calcitonin gene-related peptide (CGRP) on parasympathetic contraction of rabbit airway smooth muscle was studied under isometric conditionsin vitro. CGRP (10^–7 M) did not contract tracheal smooth muscle, but it potentiated the contractile response to electrical field stimulation (EFS) in a dose-dependent manner, the maximal increase from the baseline response being 14.4±6.2%. In contrast, the contractile responses to exogenously administered acetylcholine were not altered by this peptide. The CGRP-induced potentiation of the contractile responses to EFS was further augmented by physostigmine (123.1%), but not influenced by phentolamine, propranolol, indomethacin, or thiorphan, an enkephalinase inhibitor. These results suggest that CGRP augments the neurally mediated contraction of rabbit airway smooth muscle probably through a prejunctional mechanism and that this effect may not be modulated by tissue enkephalinase.     

Prostaglandins and neurotransmission at the guinea pig and rabbit urinary bladder

High concentrations of prostaglandins (PGE₁, PGE₂, or PGE₂) (2×10^–6 M) produced a slow contraction of longitudinal strips of detrusor muscle taken from the bladders of guinea pigs and rabbits. At a lower concentration (10^–6 M) prostaglandins enhanced contractions produced by field stimulation of nerves in guinea pig but not rabbit strips. The contractions were not affected by indomethacin. Contractions of guinea pig strips in response to acetylcholine at 10^–4 M were enhanced by prostaglandins and unaffected by indomethacin. Membrane potentials of smooth muscle cells recorded with micro electrodes, were unchanged up to 10^–6 M PGE₂. Above this the cells were depolarized with an increase in frequency of spontaneous action potentials. Synchronous recording of electrical and mechanical activity with the double sucrose gap indicated a decrease in amplitude of the evoked excitatory junction potential and action potential even when the contraction was enhanced in the presence of PGE₂. Responses to repeated stimulation at 10 Hz for 1 min were progressively depressed. This trend was slightly reduced by PGE₂ but unaffected by indomethacin. It is concluded that prostaglandins are not normally released by the nerves to the urinary bladder but are able to facilitate contraction in the guinea pig. This effect is probably on the excitatory-contraction coupling, possibly by mobilizing Ca²⁺. Some modification of transmitter release by the nerves may also occur.     

Spinal micturition reflex mediated by afferents in the deep perineal nerve

Reflexes mediated by urethral sensory pathways are integral to urinary function. This study investigated the changes in bladder pressure and urethral sphincter activity resulting from electrical stimulation of afferents in the deep perineal nerve (DP), which innervates the urethra and surrounding muscles, before and after acute spinal cord transection (SCT) in cats anesthetized with alpha-chloralose monitored by blood pressure and heart rate. DP stimulation elicited bladder contractions before and after SCT but only if the bladder contained a sufficient volume of fluid (78% of the volume needed to cause distention-evoked reflex contractions). The volume dependency was mediated by a neuronal mechanism in the lumbosacral spinal cord and was not attributable to length-tension properties of the detrusor muscle. Stimulation at 2-40 Hz initiated bladder contractions, but 20-40 Hz was more effective than lower frequencies in evoking and sustaining bladder contractions for the duration of the stimulus train. Decreases in urethral sphincter activity occurred during sustained bladder contractions evoked by 20- to 40-Hz stimulation before and within 16 h after SCT. After SCT, average bladder pressure increases evoked by DP stimulation were smaller than those evoked before SCT, but in some animals, bladder pressures elicited by DP stimulation continued to increase as time after SCT increased and reached pretransection amplitudes at 8-16 h posttransection. These data confirm the presence of a spinal circuit that can mediate coordinated bladder-sphincter responses and show that afferents from the DP can activate this circuit under appropriate conditions.     

Sympathetic innervation of the urinary bladder and urethral muscle in the pig

Larsen , J.-J., J. Nordling and B. Christensen , Sympathetic innervation of the urinary bladder and urethral muscle in the pig. Acta physiol. scand. 1978. 104. 485–490. The sympathetic innervation of the lower urinary tract was studied in the female and male pig using a specific histochemical technique for visualization of noradrenaline containing nerves. In smooth muscle from the detrusor few evenly distributed adrenergic nerve terminals were found. A greater number of terminals were observed in the trigone, bladder neck and urethra. There was no sex difference. In smooth muscle specimens from pig detrusor and trigone sympathetic nerve terminals were more abundant than in corresponding tissue from humans.     

Relaxation of sheep urethral muscle induced by electrical stimulation of nerves: involvement of nitric oxide

Isolated smooth muscle preparations from the sheep urethra responded to electrical field stimulation with contraction when basal tension was low (5-6 mN), but with relaxation when the preparations were contracted with noradrenaline (NA), clonidine, or prostaglandin F2a. No relaxant response could be elicited in high K+ (124 mM) contracted preparations. Electrically induced relaxations had a threshold of less than 1 Hz and a maximum at 8 Hz. Both contractant and relaxant responses were abolished by tetrodotoxin, indicating that they were caused by transmitters released from nerves. The amplitude of the relaxant responses showed a highly significant correlation to the tension induced by noradrenaline. A coefficient (R/T) was calculated relating relaxation to noradrenaline-induced tension. In this way it is possible to separate the effect of drugs on muscle tension (non-specific effect) from their action on the electrically induced relaxation (specific effect). Chemical sympathectomy with 6-OHDA did not significantly modify the relaxant response to 6 Hz in noradrenaline contracted strips, as evaluated by the R/T coefficient. The electrically induced relaxation was not affected by hexamethonium, propranolol, phentolamine, muscarinic receptor blockade, cocaine, indomethacin, or methysergide. Both nifedipine and Bay K 8644 inhibited significantly the response induced by electrical stimulation, decreasing its maximum. Nifedipine, but not Bay K 8644, significantly reduced the level of tension induced by noradrenaline, and its effect, evaluated by the R/T coefficient, was an increase in the electrically induced relaxation, whereas Bay K 8644 had a significant inhibitory effect.(ABSTRACT TRUNCATED AT 250 WORDS)     

Energy turnover and lactate dehydrogenase activity in detrusor smooth muscle from rats with streptozotocin-induced diabetes

Force generation and tissue glucose metabolism were measured in the urinary bladder smooth muscle from rats with streptozotocin-induced diabetes (7–8 wk duration). Bladder wet wt was almost 4–fold higher in the diabetic animals compared with the untreated controls. Morphological analysis showed that the growth was associated with hypertrophy of the smooth muscle component in the bladder wall. Force generation of isolated bladder strip preparations was measured in vitro at different ambient oxygen tensions. Activation of intramural nerves, with electrical field stimulation, induced contractions that were unaffected by reduction of oxygen tension down to Po₂ 100 mmHg for both control and diabetic muscle strips. At zero Po₂ force was reduced by approximately 10–20% in both groups. High-K⁺ solution induced ‘tonic’ contractions that were slightly more inhibited by lowering Po₂. At intermediate Po₂ (between 100 and 20 mmHg) the diabetic muscle gave slightly higher force. At zero Po₂ no significant difference could be detected between strips from control and diabetic animals. Oxygen consumption and lactate production in the preparations were determined at a Po₂ of 290 mmHg and related to the volume of smooth muscle. At zero Po₂ lactate formation increased 3- to 4-fold. The metabolic tension cost was lower at zero Po₂ No differences in basal and contraction related metabolic rates could be detected between the two groups under normoxic and anoxic conditions. The maximal activity of lactate dehydrogenase (LDH) determined in tissue sampIes was about 2-fold higher in the diabetic bladder muscle. This increased enzymatic activity could thus not be correlated with any altered metabolic properties of the smooth muscle in the urinary bladder from diabetic rats.     

Actions of prostaglandins and indomethacin on the electrical and mechanical properties of smooth muscle cells of the guinea-pig ileocecal junction

The effects of prostaglandins (PGs) and indomethacin on the mechanical and membrane properties of the smooth muscle cells of the guinea-pig ileocecal junction were studied using microelectrode and tension recording techniques and radioimmunoassay to determine levels of PGs. In the guinea-pig ileocecal junction, we found two distinct cell populations-cells with, and without spontaneous electrical and mechanical activities. PGs (PGE₁, PGE₂, PGF₂) in low concentrations suppressed spontaneous mechanical activity. Correspondingly, PGs (>10^–7 M) suppressed both spontaneously generated spikes and evoked spikes, presumably due to an increase in the threshold for generation of action potential. On the contrary, indomethacin evoked rhythmic spontaneous contractions in the mechanically quiescent muscle preparations and reduced the PGE and PGF content of the muscle. Spontaneous spike discharges occurred during the indomethacin-induced contractions. The spontaneous electrical and mechanical activities induced by indomethacin were suppressed by PGs, in low concentrations. These results indicate that under physiological conditions, the endogenous PGs in the muscle may play an important role in regulating muscle tone as well as the membrane properties, thereby contributing to the regulation of motility of the intestine and possibly the sphincter.     

Localization of Adrenergic Receptors in Guinea Pig Ileum and Rabbit Jejunum to Cholinergic Neurons and to Smooth Muscle Cells

The localization of adrenergic receptors mediating a relaxing action was investigated in innervated and denervated longituclinal muscle strips from guinea pig ileum and rabbit jejunum. Denervated preparations were contracted by drugs that had a direct effect on smooth muscle cells, such as acetylcholine and histamine, but not by stimuli acting on cholinergic neurons, such as electrical field stimulation or nicotine. After blockade of β-adrenoceptors, norepinephrine relaxed the innervated guinea pig ileum contracted by electrical field stimulation, by stimulating α-adrenoceptors. Norepinephrine in low concentrations did not relax denervated preparations contracted by agents acting directly on smooth muscle. In high concentrations, it relaxed denervated preparations by a nonadrenergic mechanism, resistant to α- and/or β-receptor blockade, but which was also activated by l-(3,4-dihydroxyphenyl) ethanol. Phenylephrine only had a weak agonistic effect on the electrically stimulated innervated preparation and did not relax the denervated one. The denervated rabbit intestine contracted by acetylcholine was relaxed by norepinephrine and phenylephrine by stimulation of α-adrenoceptors. In the innervated preparations both drugs were more effective in inhibiting contractions induced by electrical field stimulation or eserine than those induced by exogenous acetylcholine. Both the denervated guinea pig and rabbit intestine were relaxed by stimulation of β-adreno-ceptors. It is suggested that in the guinea pig ileum α-adrenoceptors mediating relaxation are located only in cholinergic neurons, whereas in rabbit jejunum they are located both in these neurons and in the smooth muscle cells. Beta-adrenoceptors are located in the smooth muscle cells of both organs.     

What is the hair cell transduction channel?

Rhythmic electrical activity is a feature of most smooth muscles but the mechanical consequences can vary from regular rapid phasic contractions to sustained contracture. For many years it was thought that spontaneous electrical activity originated in smooth muscle cells but recently it has become apparent that there are specialized pacemaker cells in many organs that are morphologically and functionally distinct from smooth muscle and that the former cells are the source of spontaneous electrical activity. Such a pacemaker function is well documented for the ICC of the gastrointestinal tract but evidence is accumulating that ICC-like cells play a similar role in other types of smooth muscle. We have recently shown that there are specialized pacemaking cells in the rabbit urethra which are spontaneously active when freshly isolated, readily distinguishable from smooth muscle cells under bright field illumination and relatively easy to study using patch-clamp and confocal imaging techniques. Recent results suggest that calcium oscillations in isolated rabbit urethral interstitial cells are initiated by calcium release from ryanodine sensitive intracellular stores, that oscillation frequency is very sensitive to the external calcium concentration and that conversion of the primary oscillation to a propagated calcium wave depends upon IP₃-induced calcium release.     

Neuromuscular actions of endothelin on smooth, cardiac and skeletal muscle from guinea-pig, rat and rabbit

The peptide endothelin (human, porcine) was investigated for effects on basal muscle tone and on responses to transmural nerve stimulation in a series of smooth muscle preparations, as well as in guinea-pig atrium and rat and guinea-pig diaphragm. Endothelin lacked effect on basal tone or on spontaneous and electrically driven contractions in skeletal and atrial muscle. It contracted guinea-pig ileum, pulmonary and femoral arteries, rat anococcygeus, vas deferens and urinary bladder and rabbit taenia coli, whereas guinea-pig taenia was relaxed. Guinea-pig urinary bladder and vas deferens and rabbit iris sphincter were unaffected up to 3 x 10(-8) M. Endothelin thus has a unique pattern of smooth muscle effects, exhibiting mostly contractile but also relaxing effects. Endothelin modified contractile responses to transmural nerve stimulation, yielding marked and persistent enhancement, in guinea-pig and rat vas deferens, and enhancement also in guinea-pig pulmonary artery. In guinea-pig and rat vas deferens the response to exogenous ATP was increased by endothelin, thus suggesting a strong post-junctional enhancement of neurotransmission. In guinea-pig ileum nerve-induced responses were inhibited by endothelin, whereas exogeneous acetylcholine was enhanced, an effect suggesting a simultaneous pre-junctional inhibition and post-junctional enhancement. The Ca2+ channel blocker felodipine counteracted the stimulatory effects of endothelin on tone and transmurally induced contractions. Tachyphylaxis to endothelin action was sometimes evident, but the anococcygeus being less prone to this might be useful for studies on endothelin antagonism. Endothelin thus has prominent post-junctional, and also probably pre-junctional, effects, lending further support for a distinct biological role of this peptide.     

Interaction between interstitial cells and smooth muscles in the lower urinary tract and penis

Smooth muscles in the lower urinary tract and corporal tissue exhibit spontaneous contractile activity which depends on L-type Ca²⁺ channels. The mechanism underlying this activity is spontaneous electrical activity which shows varied form and property between these tissues. Recent studies revealed that interstitial cells (ICs) are widely distributed in the genitourinary system, and suggested their involvement in spontaneous muscle activity. ICs in the system are not a simple analogy of interstitial cells of Cajal (ICC) in the gut, which act as electrical pacemaker, but represent variability amongst tissues which may account for individual characteristics of each organ. In the bladder and corporal tissue, where smooth muscle cells are capable of generating spontaneous electrical activity, ICs may modulate smooth muscle activity. ICs in corporal tissue release prostaglandins via cyclooxygenase-2 (COX-2) activity and reinforce not only spontaneous but also nerve-mediated α-adrenergic contractions. In the bladder, their fundamental role in the integration of signals between populations of cells has been proposed, and thus changes in ICs may contribute to an overactive bladder, a pathological condition which results from increased excitability in detrusor smooth muscles. In the urethra, ICs may act as electrical pacemakers as do ICC. However, overall contractility of urethral smooth muscles does not necessarily rely on pacemaking of ICs, and thus some population of smooth muscles may also have their own excitability.     

Mechanical and electrical responses modulated by excitation of inhibitory nerves during stimulation with high-potassium solutions in circular smooth muscle of the rabbit rectum.

The properties of the mechanical responses produced by solutions containing high concentrations of potassium ion (high-K solution, [K(+)](o) = 9-27 mM) were investigated in circular smooth muscle preparations isolated from the rabbit rectum. Isometric recording of mechanical responses of the muscle revealed spontaneous contractions, which successively decreased and finally disappeared in most preparations. Stimulation of the smooth muscle with high-K solutions elicited an increase in both amplitude and frequency of twitch contractions (sustained component), with about a 2 min delay in the beginning (initial inhibition), and a transient large contraction shortly after the cessation of stimulation (after contraction). Transmural nerve stimulation (TNS) with electrical pulses for 1 min at 1 Hz frequency produced a sustained inhibition, but a transient contraction followed after termination of TNS. In the presence of tetrodotoxin (TTX), the TNS-induced responses were abolished, while a high-K solution elicited increased twitch contractions with a short delay and abolished the after contraction. Suramin produced effects similar to TTX on the responses produced by high-K solutions or TNS, but this was not the case for atropine, guanethidine or N(omega)-nitro-L-arginine (L-NA). Recording membrane potentials with microelectrodes revealed that TNS evoked an inhibitory junction potential (i.j.p.) which was non-adrenergic, non-cholinergic and non-nitrergic in nature. High-K solutions elicited a tri-phasic change in the membrane potential; an initial hyperpolarization, followed by a sustained depolarization and finally a transient depolarization on cessation of high-K stimulation. TTX or suramin inhibited the i.j.p.s and altered the tri-phasic change in the membrane potential produced by a high-K solution to a mono-phasic depolarization. No significant modulation of electrical responses of the membrane induced by TNS or high-K solution was elicited by atropine, guanethidine or L-NA. The results indicated that the circular smooth muscle of the rabbit rectum is innervated by inhibitory nerves, and that stimulation with high-K solutions caused inhibitory neuronal modulation of both electrical and mechanical responses of the smooth muscle, in a suramin-sensitive way.     

Adrenergic and cholinergic induced contractions of tracheal smooth muscle in the rabbit as demonstrated by a new in vivo method

An in vivo tracheal muscle preparation in the rabbit was developed which enabled us to measure changes in the isometric tension of the trachealis muscle in response to electrical stimulation of autonomic nerves and to i.v. administration of autonomic agonists and antagonists. The preparation was very sensitive to injections of carbachol, and showed graded contractions to stimulation of the caudal end of the cut cervical vagus as frequency and strength of stimulation were increased. Stimulation of the rostral end of the cut cervical sympathetic nerve fibers produced contractions in all preparations. This effect was mimicked by the alpha-adrenoceptor agonist, phenylephrine. The effects of both sympathetic stimulation and phenylephrine were blocked by phentolamine and not inhibited by pretreatment with atropine or propranolol. Sympathetic stimulation produced contractions of the trachealis muscle whether the initial tone was normal or actively increased by carbachol, while adrenaline produced relaxation when the initial tone was high. Using this new in vivo trachealis muscle preparation in the rabbit, we could show that sympathetic stimulation produced contractions of the trachealis muscle. This effect is consistent with the existence of smooth muscle activating alpha adrenoceptors.