Effects of Ca-antagonists on neuromuscular transmission in the rabbit ear artery

1. The effects of three Ca-antagonists: diltiazem, nicardipine and flunarizine have been studied on excitatory junction potentials (e.j.p.s), force development and efflux of transmitter during stimulation of perivascular nerves in the rabbit ear artery. 2. Stimulation of these perivascular nerves produces excitatory junction potentials and repetitive stimulation causes facilitation. Increasing the frequency or the number of stimuli initiates an action potential and a large contraction. Both phenomena are completely suppressed by 3 X 10(-7) M TTX. 3. Ca-antagonists at 10(-5) M do not affect the resting membrane potential, but flunarizine and nicardipine at concentrations exceeding 10(-5) M reduce the amplitude of e.j.p.s and of the action potentials and also the concomitant contraction induced by nerve stimulation. Diltiazem at concentrations below 3 X 10(-5) M has no effect on e.j.p.s and action potentials while at 10(-4) M, it largely suppresses e.j.p.s, spikes and contraction. This inhibitory effect of the Ca-antagonists increases with prolonged exposure. 4. All these Ca-antagonists induce an increased release of 3H-DOPEG from the nerve terminals, but in our experiments in vitro they do not reduce the efflux of 3H-noradrenaline induced by nerve stimulation. 5. The results indicate that Ca-antagonists might affect the excitation-contraction coupling in vivo by inhibiting the Ca influx activated by endogenous noradrenaline. They do not exert an acute effect on the noradrenaline release induced by stimulation. The increased DOPEG release by Ca-antagonists remains unexplained.     

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.     

Inhibitory actions of indomethacin on electrical and mechanical responses produced by nerve stimulation in circular smooth muscle of the guinea-pig gastric fundus.

The effects of indomethacin on electrical and mechanical responses produced by transmural nerve stimulation (TNS) were investigated in isolated circular smooth muscle of the guinea-pig gastric fundus. TNS evoked a cholinergic excitatory junction potential (e.j.p.). The e.j.p.s were inhibited by 1-10 microM indomethacin, in a concentration-dependent manner, with no marked alteration of the resting membrane potential. Exogenously applied acetylcholine caused a depolarization of the membrane that was not altered by indomethacin. TNS evoked a cholinergic twitch contraction at low frequencies (0.1 Hz). A train of TNS's at high frequency (1 Hz) produced a transient contraction with a subsequent sustained relaxation. Indomethacin reduced the resting tension and inhibited these TNS-induced contractions. Application of Nomega-nitro-L-arginine (NOLA), an inhibitor of nitric oxide (NO) synthesis, increased the amplitude of twitch contractions, and altered transient contractions to tetanic contractions during TNS at a frequency of 1 Hz, also with an increased amplitude. In the presence of NOLA, indomethacin (5 microM) again reduced the resting tension and inhibited TNS-induced contractions. This inhibition was greater for twitch contractions than for tetanic contractions. Nifedipine reduced the TNS-induced contractions, while addition of indomethacin further reduced the amplitude of contractions. Contractions produced by low concentrations of acetylcholine (0.1 microM) were inhibited by indomethacin, while those produced by 1 microM were not. These results indicate that the inhibitory actions of indomethacin on TNS-induced contractions do not involve enhanced production of NO or selective inhibition of voltage-gated Ca-channels. Prejunctional autoregulatory mechanisms may also not be altered by indomethacin. As indomethacin inhibits the enzyme cyclooxygenase, it is speculated that endogenously produced prostaglandins exert excitatory actions on gastric smooth muscle, and act mainly postjunctionally to facilitate spontaneous and neurogenic electrical and mechanical activity.     

Neural regulation of electrical and mechanical activities in the rat tail artery

Stimulation of the perivascular nerves elicited two types of electrical responses in the rat tail artery—excitatory junction potentials (e.j.p.s) and slow depolarization—and two types of mechanical responses—fast and slow contractions. Fast phasic contractions were triggered whenever action potentials were generated from either the e.j.p. or the slow depolarization reaching threshold. Slow tonic contractions and slow depolarizations were sensitive to -adrenergic blockade. However the slow contraction always preceded the slow depolarization. Bolus doses of exogenous noradrenaline also induced slow contraction and slow depolarization and the development of tension also preceded the membrane potential change. Increasing the external KCl also induced membrane depolarization however, contractions were not observed until the membrane was depolarized positive of –49 mV. In contrast, tension developed readily with membrane potential more negative than –49 mV with exogenous noradrenaline and neural stimulation, suggesting that the action of noradrenaline was not mediated by electromechanical coupling. It was concluded that vascular activity in the rat tail artery could be regulated by the e.j.p., the slow depolarization and also by pharmacomechanical coupling.     

Electrophysiological events during neuroeffector transmission in the spleen of guinea-pigs and rats.

Intracellular recordings were made from smooth muscle cells of arterioles and the capsule of the spleen of guinea-pig and rat, and the responses to periarterial or subcapsular nerve stimulation were recorded. The innervation of the spleen was studied using fluorescence and immunohistochemical techniques. Catecholamine-containing axons were associated with smooth muscle of the splenic capsule, trabeculae, arterioles and amongst cells of the periarteriolar lymphoid sheath. Axons immunoreactive for neuropeptide Y (NPY) and tyrosine hydroxylase were distributed in an identical manner to catecholamine-containing axons, whereas axons immunoreactive for substance P or calcitonin gene-related peptide were present at a very low density in spleens from both species. In segments of arterioles, single transmural stimuli evoked excitatory junction potentials (EJPs) of 1-10 mV amplitude. EJPs facilitated during short trains of stimuli (1-10 Hz) and summated at 10 Hz, often initiating a muscle action potential. EJPs persisted in the presence of prazosin (1 microM) and idazoxan (1 microM), but were abolished by the P2x-purinoceptor antagonist suramin (1 mM). Spontaneous depolarizations were observed in smooth muscle cells of arterioles and capsule. Some events in arterioles were observed in the presence of suramin and so may originate postjunctionally independently of transmitter release. As single transmural stimuli failed to evoke a depolarization in capsular smooth muscle, spontaneous depolarizations in this tissue probably also arise postjunctionally. Short trains of high frequency stimuli (10-35 Hz) evoked biphasic depolarizations of capsular smooth muscle cells. The initial component peaked 2.5 s following the onset of stimulation; the second component peaked 15 s following the onset and decayed exponentially with a time constant of 15 s. By fitting a product of exponentials to the second component, it was possible to define the initial component, which decayed with a time constant of around 1.5 s. Neurally evoked depolarizations of capsular smooth muscle were abolished by 1 microM TTX. Blockade of alpha 1-adrenoceptors with prazosin reduced the initial component of the depolarization, whereas alpha 2-adrenoceptor blockade with idazoxan virtually abolished the second component. In some cells a small, faster depolarization persisted after alpha-adrenoceptor blockade. The slow alpha 2-adrenoceptor-mediated depolarization was identical to that recorded in the rat tail artery and in the guinea-pig mesenteric vein. The data indicate that sympathetic neuroeffector transmission from noradrenergic axons containing NPY to splenic arterial and capsular smooth muscle occur by different mechanisms.     

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.     

Direct and indirect contractile responses of the human vas deferens and actions of noradrenaline and of calcium antagonists

Contractile responses of the isolated human vas deferens, obtained from vasectomy operations, were measured. Large single electrical shocks gave a twitch response with short latency (0.36 s) which was insensitive to prazosin (5 microM) or TTX (0.2 microM) and was thus identified as due to direct muscle stimulation. A train of 100 low intensity shocks gave a response with a longer latency (1.9 s) which was substantially sensitive to both prazosin and TTX; we assume this response is dominated by an indirect nerve-induced contraction. Relaxations, presumably caused by activation of circular muscle, were recorded from regions of some preparations both by direct and indirect stimulation. Noradrenaline (10-20 microM) induced a tonic contracture, spontaneous contractions and a large potentiation of the response to direct stimulation--but not to indirect stimulation implying a strong presynaptic inhibition. Noradrenaline also speeded the relaxation from contractions. Verapamil (1-100 microM) and nifedipine had no effect on the direct responses but verapamil (10 microM) inhibited the indirect response. Calcium removal prevented most, and 5 mM-EDTA all, of the direct response. However, even with EDTA, noradrenaline was able to support spontaneous and stimulus-induced contractions. Thus contraction of the vas, though sustained by external calcium, does not appear to directly depend on it.     

Nitric oxide inhibits smooth muscle responses evoked by cholinergic nerve stimulation in the guinea pig gastric fundus

In circular smooth muscle tissues of the guinea pig gastric fundus, transmural nerve stimulation (TNS) evoked an atropine-sensitive cholinergic excitatory junction potential (e.j.p.) and, after inhibiting the e.j.p. with atropine, an apamin-sensitive nonadrenergic noncholinergic (NANC) inhibitory junction potential (i.j.p.). The amplitude of e.j.p.s was similar when the frequency of TNS was low (<0.5 Hz), but it decreased successively (depression phenomenon) when the frequency was high (>1 Hz). The depression phenomenon was attenuated after inhibiting the production of nitric oxide (NO) with N(omega)-nitro-L-arginine (NOLA), but was not altered by inhibiting the i.j.p. with apamin. The e.j.p.s were increased in amplitude by the inhibition of cholinesterase activity, but they were decreased by NO produced from SNP with no alteration of their depression phenomenon. Isometric twitch contractions were depressed during high-frequency TNS. NOLA caused an increase in the amplitude of twitch contractions and the attenuation of their depression that changed the transient contraction produced by high-frequency TNS (1 Hz) to a tetanic one. SNP reduced the amplitude of twitch contractions, with no alteration of the depression phenomena. Contractions produced by low concentrations of acetylcholine, but not by high concentrations, were attenuated by SNP, with no alteration of the membrane depolarization. The results suggest that NO produced during TNS has inhibitory actions on cholinergic transmission; the depression of e.j.p.s is mainly prejunctional events, and the depression of mechanical responses is mainly postjunctional events.     

Some electrical properties of the rabbit anococcygeus muscle and a comparison of the effects of inhibitory nerve stimulation in the rat and rabbit

1. Simultaneous recordings of mechanical activity and membrane potential of individual smooth muscle cells have been made in the rabbit anococcygeus muscle and the effect of field stimulation on these examined.2. In the absence of tone the mean resting membrane potential was - 48 mV. In the stretched muscle spontaneous tone and rhythmic activity quite frequently appeared and this was associated with depolarization of the muscle cells.3. The response to field stimulation depended on the frequency of stimulation, the level of membrane potential and the presence of myogenic tone. The usual response to single pulses or low frequency stimulation was a hyperpolarization of up to 30 mV (mean 14+/-6.8 mV) after a latency of 185 msec and accompanied by muscle relaxation. Higher frequencies (over 8 Hz) produced an initial depolarization often with a spike potential and followed by hyperpolarization. The mechanical response in these instances was contraction or contraction followed by relaxation. At all frequencies rebound depolarization and an associated contraction followed the end of stimulation).4. Phentolamine (5x10(-6)M) and guanethidine (10(-6)M) blocked the initial depolarization and contraction but had no effect on hyperpolarization, muscle relaxation or rebound depolarization and contraction.5. The effect of field stimulation in the presence of guanethidine (4x10(-5)M) was re-examined in the rat anococcygeus. Single pulses were ineffective, repetitive stimulation produced muscle relaxation but no hyperpolarization comparable to the rabbit. Any oscillations in membrane potential were damped during field stimulation and sometimes a small hyperpolarization was produced with a maximum amplitude of 13 mV and a mean of 1.9+/-1.2 mV.6. The transmembrane potential at the peak of hyperpolarization in the rabbit was rarely more than -70 mV. Passive displacement of the membrane potential by current pulses altered the amplitude of the hyperpolarization and suggested that there was a reversal potential at between -80 and -90 mV.7. No change in input resistance could be measured during inhibitory nerve stimulation in either the rabbit or the rat but measurements based on electrotonic potentials indicated a reducation in membrane resistance, small in the rat but greater in the rabbit.8. These experiments suggest that in both species muscle relaxation is associated with an increase in ionic permeability and a move, at least in the rabbit muscle, towards an equilibrium potential of -80 to -90 mV. In view of the much smaller effect in the rat it is not clear whether this is the cause or at least the sole cause of the muscle relaxation.     

Possible mechanism of adrenergic and nonadrenergic inhibition in intestinal smooth muscle cells

Nonadrenergic synaptic transmission in circular and longitudinal smooth muscles of caecum preexposed to K-free solution for 4–5 h has been studied by means of sucrose gap technique. In addition, the effects of noradrenaline (NA) and ATP on these muscles were investigated under these conditions. The action of the above substances was accompanied by depolarization and contraction. NA induced a decrease in the membrane resistance. Addition of 0.5 mM Ba²⁺ to K-free solution intensified the depolarization. 1 mM of Mn²⁺ blocked depolarization and contraction. Intramural stimulation produced noncholinergic e.j.p.s blocked by TTX. Addition of 0.5 mM Ba²⁺ increased their amplitude. A reversal potential of both NA-induced depolarization and e.j.p. was in the range of +10 to +20 mV. It is supposed that e.j.p.s and depolarization observed in response to ATP and NA action are due to an increase in calcium permeability of the membrane.     

Non-neural electrical responses of smooth muscle cells of the rabbit basilar artery to electrical field stimulation

In smooth muscle cells of the rabbit basilar artery, field stimulation evoked a depolarizing response which consisted of a fast (1-3 s duration) and a following slow (1-4 min duration) component. The amplitude of these responses increased in an intensity-dependent manner and, when exceeding 10-15 mV, a spike potential was generated. During generation of the slow depolarization, ionic conductances of the membrane were increased. When outward current pulses with long duration (2-3 s) were applied to the smooth muscle using the partition stimulating method, electrotonic potentials and spike potentials were generated. The cessation of the current pulse caused repolarization of the membrane with time constant of 250-350 ms. The depolarizing responses were resistant to tetrodotoxin, sympathetic transmission blocking agents (guanethidine, bretylium, or 6-hydroxydopamine treatment), receptor antagonists for 5-hydroxytryptamine (methysergide), dopamine (haloperidol), ACh (atropine), noradrenaline (phentolamine), ATP (alpha,beta-mATP) or histamine (mepyramine), blockade of synthesis of prostaglandins or thromboxane A2 (indomethacin) or high Mg2+, low Ca2+ solution. Smooth muscle cell membrane of the basilar artery was depolarized by 5-hydroxytryptamine (above 0.1 microM) or histamine (above 10 microM) but not by ACh (up to 100 microM) or noradrenaline (up to 10 microM). The depolarization induced by 5-hydroxytryptamine or histamine was antagonized by methysergide or mepyramine, respectively. Denervation of the vessel by storing in a cold condition (4 degrees C) decreased but did not abolish the depolarizing response. The decrease in amplitude of the depolarizing response during cold storage was attributed to associated depolarization of the smooth muscle membrane. Internal perfusion of the vessel with distilled water abolished generation of the depolarizing response, and this procedure also abolished the endothelium-dependent relaxation induced by ACh during the potassium contraction. The results suggest that the depolarizing response evoked by field stimulation is generated by substances released from non-neural components, possibly from the endothelial cells.     

A study of excitatory neuromuscular transmission in the bovine trachea

1. The excitatory innervation of bovine tracheal smooth muscle has been studied with the sucrose-gap apparatus.2. Single 2 ms electrical stimuli applied to the whole tissue excited intrinsic nerves, and produced a small transient depolarization of the smooth muscle, the excitatory junction potential (e.j.p.). The e.j.p. caused a twitch-type contraction; twitches and e.j.p.s summated during repetitive stimulation but facilitation was not observed, and action potentials were never elicited.3. The effects of electrical stimulation could be abolished by atropine (5 x 10(-7) mol/l) and augmented by neostigmine (4 x 10(-6) mol/l), and were mimicked by exogenous acetylcholine (1.0 mug/ml).4. With the electron microscope, the density of innervation was found to be low (one axon per ninety smooth muscle cells). Axons were found in small groups in the clefts between bundles of cells, but no axons penetrated within the muscle bundles. Naked axon varicosities containing agranular vesicles were seen, but no axon approached within 200 nm of a smooth muscle cell.5. It is difficult to reconcile the sparsity of innervation with the dependence of the tissue on nerve excitation to initiate activity.     

Electrical and mechanical properties and neuro-effector transmission in the smooth muscle layer of the guinea-pig ileocecal junction

Electrical and mechanical properties and neuro-effector transmission were studied in circular strips of smooth muscle taken from the ileocecal junction of guinea-pigs in relation to sphincter action, using the microelectrode, and tension recording methods. The membrane potential of the smooth muscle was low (–43 mV) compared with the membrane potential of circular muscle cells of the ileum or caecum (–58 mV or –62mV). Only small populations of the muscle cells (about 5%) generated spontaneous action potentials.Field stimulation of the tissue produced an initial slight relaxation followed by a contraction, and the mechanical responses were accompanied by membrane hyperpolarization (i. j. p.) followed by repolarization with rebound spikes. Treatment with atropine increased the amplitude of i.j. ps and decreased the amplitue of the rebound repolarization. Propranolol or phentolamine did not affect the amplitude of i. j. p., however, phentolamine slightly reduced the amplitude of the rebound repolarization.These results indicate that the ileocecal junction is predominantly controlled by non-adrenergic, non-cholinergic inhibitory nerve fibres and that the distribution of adrenergic and cholinergic excitatory nerve fibres is sparse.     

Effect of calcium on neurohumoral stimulation of feline colonic smooth muscle

The purpose of this study was to compare the effect of altering the extracellular calcium ion concentration on bethanechol or octapeptide of cholecystokinin (OP-CCK) stimulation of the isolated transverse colon of the cat. Myoelectric activity was recorded with monopolar glass-pore electrodes. Bethanechol (10(-6) M) stimulated an increase in the number of slow waves with superimposed spike potentials to 85.5 +/- 5.3% (P less than 0.001) compared with the basal spike activity (8.9 +/- 1.4%). OP-CCK (4 x 10(-9)) also increased spike activity (80.7 +/- 3.8%, P less than 0.001), which was not inhibited by atropine, phentolamine, or propranolol. Addition of 0.0 mM calcium solution to the colonic smooth muscle abolished both slow-wave and spike activity, which returned after replacing 0.25 mM calcium in the solution. Bethanechol stimulated a greater increase in spike activity as the concentration of calcium was increased. OP-CCK stimulation of colonic spike activity was more sensitive to the extracellular calcium concentration than bethanechol stimulation. Verapamil had a minimal effect on bethanechol stimulation of colonic spike activity, but it inhibited the OP-CCK stimulation. These studies suggest that 1) OP-CCK appears to stimulate colonic smooth muscle directly and 2) OP-CCK requires the presence of a greater amount of extracellular ionic calcium in order to stimulate colonic spike activity compared with bethanechol.     

Electrical responses of the smooth muscle of the guinea-pig cerebral artery to brief electrical stimulation

Electrical responses to brief electrical stimulation were investigated in the cerebral artery of a guinea-pig using a microelectrode. A single brief stimulus (0.05 ms) induced a spike potential followed by a depolarizing slow-potential, and these events were associated with muscle contraction. An outward current injected into the smooth muscle cell induced spike potential but failed to induce depolarizing slow-potential. These activities persisted in the presence of TTX (10(-6) M), guanethidine (5 X 10(-6) M), or atropin (10(-5) M). TEA (5 mM) enhanced the amplitude of the spike potential, but not that of the depolarizing slow-potential. When the external Na was reduced, the membrane transiently hyperpolarized. During this period, the depolarizing slow-potential could be evoked. In a Cl-deficient solution, the membrane depolarized and the amplitude of the depolarizing slow-potential decreased. From these observations it is believed that the contribution of K, Na, or Cl is minor. In a 20 mM-Ca solution, a brief stimulation induced neither spike potential nor depolarizing slow-potential, but did induce a hyperpolarizing slow-potential. The hyperpolarizing slow-potential was also induced in a Na-deficient solution, but only after completion of Na re-distribution across the membrane. These observations suggest that a substance released by brief stimulation produces a prolonged change in ionic conductances of the smooth muscle membrane, allowing the muscle to contract for a certain period.     

Electrical field stimulation of rat mesenteric small arteries: force and free cytosolic calcium during neurogenic contractions and mechanisms of non-neurogenic relaxations

The effect of transmural electrical field stimulation (TEFS) of rat mesenteric small arteries was studied. Stimulation parameters were selected to cause tetrodotoxin (TTX) sensitive contractions. In arteries precontracted with PGF_2α in the presence of phentolamine, TTX insensitive relaxation could be induced by TEFS. The relaxing effect of TEFS required higher stimulation amplitude and duration than the contractions. Thus, by appropriately choosing stimulation parameters, contractile responses could be elicited which were little affected by any relaxing effect, while contractions were abolished by TTX at any stimulation conditions in the present study. The contractions were abolished by cold storage and almost completely inhibited by phentolamine. Thus, contractions were neurogenic and primarily caused by noradrenaline. At low frequencies, TEFS caused phentolamine sensitive increases in free cytosolic calcium with no contractions. At higher frequencies, there was a further increase in free cytosolic calcium, associated with contraction. Only at high frequencies, noradrenaline from nerves caused sensitization of the contractile filaments to free cytosolic calcium as during stimulation with exogenous noradrenaline. The relaxations were associated with decreases in free cytosolic calcium and were probably non-neurogenic since they were resistant to TTX, cold storage, capsaicin, and repeated stimulation. Furthermore, relaxations were almost completely abolished by increasing extracellular potassium to 40 mM or by adding tetraethylammonium chloride or 4-aminopyridine. Relaxations were also reduced by ouabain and potassium free conditions.     

The effect of procaine on the mechanical and electrical activities of the smooth muscle cells of the guineal pig urinary bladder.

Procaine (1-15 mM) enhanced the spontaneous contractions of the urinary bladder smooth muscle. When a low concentration of procaine was added to normal Krebs solutions, spontaneous rhythmic contractions were enhanced. On increasing the concentration of procaine, a rise in tone (resting tension) of the preparation was observed and gradually decreased with time. The action of procaine of enhancing spontaneous contraction was observed in Na-deficient (sucrose substitutiona) and Na-free (Tris substitution) Krebs solutions. Tetrodotoxin (3 X 10(-7) G/M) DID NOT INHIBIT THE EFFECT OFPROCAINE ON MECHANICAL RESPONSE. In normal Krebs solution, procaine depolarized the membrane and increased spike frequency. The peak potential of the spike increased at 1 mM of procaine, but was suppressed at concentrations of more than 5mM. After-hyperpolization of the spike was diminished by procaine and spike duration was prolonged. The maximum rate of rise of the spike was increased immediately after application of 1 mM of procaine, but decreased wiith time. The maximum rate of fall of the spike was markedly decreased by procaine. Relative membrane resistance was increased by the application of procaine. From these results it is suggested thatprocaine mainly reduces K conductance and causes depolarization, and that enhanced spontaneous contractions are caused by depolarization and increased spike activity.     

Noradrenaline mediates slow excitatory synaptic potentials in rat dorsal raphe neurons in vitro

Repetitive focal stimulation to the slice surface within the region of the dorsal raphe (DR) nucleus of rat brain elicited a slow excitatory synaptic potential (slow EPSP), which followed a slow inhibitory synaptic potential (slow IPSP) in a majority of the DR neurons. The slow EPSPs were associated with either an increase of a decrease in membrane resistance. Noradrenaline (NA) application caused a membrane depolarization in most of the DR neurons. The NA-induced depolarization was also accompanied by either an increase or a decrease in membrane resistance. Both the slow EPSP and NA-induced depolarization were inhibited by phentolamine and prazosin but not by yohimbine and propranolol. The result suggests that slow EPSPs in rat DR neurons are mediated by NA interacting with an alpha 1-adrenoceptor.     

Direct and indirect effects of histamine on the smooth muscle cells of the guinea-pig main pulmonary artery

Electrophysiological studies of the effects of histamine on the smooth muscles in the guinea-pig main pulmonary artery revealed that this amine produced muscle contraction with an associated depolarization of the membrane. Application of cimetidine potentiated and that of mepyramine suppressed these histamine-induced responses. In the presence of mepyramine, histamine produced membrane hyperpolarization. Contractions produced by perivascular nerve stimulation were potentiated by histamine, and additional application of cimetidine further potentiated while addition of mepyramine suppressed the histamine-induced enhancement. The amplitude of excitatory junction potentials was increased by application of histamine plus cimetidine and was decreased by histamine plus mepyramine. Excitatory effects of histamine on the electrical and mechanical responses were reduced by application of tetrodotoxin, prazosin, phentolamine or guanethidine. In the presence of these drugs, histamine produced depolarization with an associated increase in membrane resistance and, in high concentrations, produced spike potentials. Electrical and mechanical responses of the smooth muscles to exogenously applied noradrenaline were potentiated by pretreatment with histamine and cimetidine, and were suppressed by histamine and mepyramine. These observations indicate that the guinesa-pig main pulmonary artery possesses two types of histamine receptor, H₁- and H₂-receptors, in the smooth muscles and in the perivascular adrenergic nerves. Stimulation of H₁ or H₂-receptor produces excitatory or inhibitory effects, respectively, on the smooth muscles and on the adrenergic nerves. Contraction of the muscle tissues produced by histamine is brought about by a direct effect on the smooth muscles and by increased release of transmitters, as a result of excitation of perivascular nerves.     

Effects of enkephalin and endorphin on the inhibitory junction potentials in the duodenal smooth muscle cells of the guinea-pig

Inhibitory junction potentials (i.j.p.s) evoked by field stimulation were recorded from the smooth muscle cells of the guinea-pig duodenum intracellularly. The membrane potential was -54.3 mV. The parameters of the i.j.p. were as follows: latency, 71 msec; time to peak, 146 msec; amplitude, 15.5 mV; rate of hyperpolarization, 107 mV/sec; and half decay time of the i.j.p., 193 msec. Met-enkephalin (10(-7)-10(-6) M) had no effect on the membrane potential and the i.j.p. The membrane potential was decreased by beta-endorphin (1.7 X 10(-7)-6.8 X 10(-7) M). Increase in the latency and the time to peak and decrease in the amplitude and the rate of hyperpolarization of the i.j.p. were observed for beta-endorphin. "Spontaneous" excitatory junction potentials (e.j.p.s) were generated by beta-endorphin. Naloxone (3.1 X 10(-6)-3.1 X 10(-4) M) hyperpolarized the membrane of the muscle cells. At high concentrations of naloxone (3.1 X 10(-4) and 3.1 X 10(-3) M), inhibition of the i.j.p. was observed. Levallorphan (2.3 X 10(-4) M) prolonged the latency and the time to peak and reduced the amplitude of the i.j.p. The membrane potential was slightly decreased by levallorphan. "Spontaneous" e.j.p.s were generated by levallorphan in a certain population of the cells. It is concluded that Met-enkephalin does not contribute to the non-adrenergic inhibitory transmission and that beta-endorphin acts as a modulator in the control mechanism of the intestinal motility. The effects of naloxone and levallorphan on the i.j.p. are discussed.