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.     

Membrane properties of the smooth muscle cells of the rat anococcygeus muscle.

1. The membrane properties of the rat anococcygeus muscle, during rest and activity, were investigated with micro-electrodes and partition stimulation. 2. Intercellular current spread occurred within the muscle and the mean length constant was 2-7 mm. The membrane showed rectification to depolarizing pulses. 3. The mean resting potential was --62-1 mV and the input resistance was 23-0 MOMEGA. Stimulation of intramural nerves produced depolarization to --21 mV and a 10% reduction in input resistance. Displacement of the membrane potential indicated that the transmembrane potential at the peak of the response was independent of the membrane potential. 4. Noradrenaline also produced depolarization and this was accompanied by a decrease in membrane resistance as indicated by a reduction in amplitude of the electrotonic potential. 5. It was concluded that the muscle possesses cable properties and that the action of the transmitter, noradrenaline, is to increase membrane permeability so that the membrane potential moves towards an equilibrium potential.     

Membrane properties of smooth muscle cells in pulmonary hypertensive rats

The membrane properties of smooth muscle cells in rat main pulmonary artery (MPA) and small pulmonary artery (SPA) were investigated during chronic normobaric hypoxia and after monocrotaline injection. As chronic pulmonary hypertension developed, pronounced differences between MPA and SPA were observed. These findings may shed light on mechanisms of smooth muscle hypertrophy. 1) The resting membrane potential of smooth muscle in MPA became less negative than the normal (depolarized), whereas the resting membrane potential of smooth muscle in SPA became more negative (hyperpolarized). 2) In MPA, both the length and time constants diminished. 3) In MPA, the maximum membrane depolarization produced by a 10-fold increase in extracellular [K+] decreased. 4) In SPA, the depolarization observed in K+-free solution was more rapid and greater in amplitude, and the transient hyperpolarization following restoration of K+-containing solution increased. 5) In SPA, initial and sustained depolarization evoked by Na+-deficient solutions were increased. 6) Depolarization in MPA was due to increased membrane permeability, perhaps to Cl-, whereas hyperpolarization in SPA could be attributed to increased activity of an electrogenic Na+-K+ pump.     

Membrane properties and innervation of smooth muscle cells in Hirschsprung's disease

Mechanical and membrane properties of smooth muscle cells and/or neuroeffector transmission in the aganglionic segment of the large intestine (Hirschsprung's disease) were compared with findings in the ganglionic segment. Tension-recording, microelectrode, and double sucrose gap methods were used. There was no difference in resting membrane potential of the longitudinal or circular muscle cell in these two segments, which were obtained at biopsy in Japanese children. In the ganglionic preparations, generations of regular prepotentials, with or without the spike, correlated well to the rhythmic contractions. However, in the aganglionic segment, irregular spike and contraction only were observed. In the circular or longitudinal muscle of the ganglionic segments, field stimulations evoked inhibitory junction potentials, excitatory junction potentials, or both and triggered initial relaxation and then a contraction of the tissue. In the aganglionic segment, however, field stimulation evoked only excitatory junction potentials followed by contraction. These results indicate that, in cases of Hirschsprung's disease, there may be a deficiency in the nonadrenergic inhibitory pathways. This is the first evidence for a lack of spasticity in muscle from the aganglionic segment of the large intestine obtained from children with Hirschsprung's disease.     

Neural control of the internal anal sphincter motility.

Control mechanism of smooth muscle movement of the internal anal sphincter (IAS) by enteric and extrinsic neuvous systems in the dog was investigated. Responses of IAS muscle strips to electrical field stimulation (EFS) and neurotransmitter agents were recorded in vitro. The contraction response to norepinephrine or to EFS was inhibited by phentolamine. The relaxation induced by EFS was not affected by phentolamine, propranolol or atropine. The mechanical activity of smooth muscle in colon and anorectum during spontaneous defecation was recorded using strain gauge force transducers. The colon and anorectum showed the characteristic motility pattern during defecation: 1) The giant migrating contraction of the colon propagated to the rectum, 2) The relaxation of the rectum prior to the contraction, and 3) The IAS muscles continued to relax while the giant contractions of the colon were migrating to the rectum. Sacral nerves were stimulated electrically and the responses of smooth muscles in the rectum and IAS were recorded. The sacral nerve stimulation induced a relaxation followed by contraction of smooth muscle in the rectum and the relaxation in IAS. The mechanical responses of smooth muscle in the IAS were modulated by alpha-adrenergic excitatory and non-adrenergic, non-cholinergic inhibitory nerves. During defecation, the relaxation of IAS smooth muscle was associated with a characteristic motility pattern of the colon and anorectum. The enteric nervous systems may be organizing the motility of these muscles by way of the motor neurones under the control the extrinsic nervous systems.     

Membrane properties of the smooth muscle of guinea-pig ureter

1. The membrane properties of the guinea-pig ureter were studied in physiological Krebs solution by intra- and extracellular stimulating methods.2. The mean membrane potential was 50 mV. Action potentials triggered by external stimulation were composed of repetitive spikes and a plateau phase.3. The effects of intracellular polarization on the membrane activity elicited by extracellular stimulation were observed. Anodal polarization enhanced the amplitude and the maximum rate of rise of the spike while cathodal polarization reduced them. The number of the spikes, the duration and amplitude of the plateau phase were not changed by polarization of any direction.4. The spikes triggered by intracellular stimulation were mostly graded, but repetitive spikes sometimes continued even after cessation of the stimulation. The effective membrane resistance was 15-23 MOmega and the time constant was 2-3 msec.5. Conduction velocity (V), chronaxie, time constant (tau) and space constant (lambda) of the tissue were measured by extracellular stimulation. These values were as follows: V, 3-6 cm/sec; chronaxie, 20-40 msec; tau, 200-300 msec; lambda, 2.5-3 mm. The conduction of excitation might be related to the cable properties of the tissue.6. The relative refractory period measured by extracellular stimulation was as long as 30 sec. During the relative refractory period dissociation of the slow depolarization and the spikes was observed by successive stimuli.7. The plateau phase was prolonged and the frequency of the spontaneous discharges was increased by treatment with Ba(2+). Tetrodotoxin had no effect on spike activity nor on the plateau phase, but Mn(2+) blocked the membrane activity.     

Properties of inhibitory junctional transmission in smooth muscle of the guinea pig lower esophageal sphincter

Inhibitory neurotransmission in guinea pig lower esophageal sphincter (LES) muscles was investigated by using electrophysiological methods. Transmural nerve stimulation (TNS) initiated an inhibitory junction potential (i.j.p.); the amplitude increased 35% by atropine (10(-6) M) and converted to a muscarinic excitatory junction potential (e.j.p.) by apamin (10(-7) M) plus Nomega-nitro-L-arginine (L-NNA, 10(-5) M). In atropinized tissue, the i.j.p. amplitude was reduced 58% by guanethidine (5 x 10(-6) M), 41% by L-NNA (10(-5) M), 57% by suramin (10(-4) M), and it was abolished by apamin (10(-7) M), suggesting that this potential was produced by ATP and nitric oxide (NO) released from adrenergic and nitrergic nerves, respectively, through the activation of Ca2+-sensitive K+ channels.Hyperpolarizations produced by ATP and NO were inhibited by apamin. The i.j.p. amplitude was reduced after desensitizing the membrane with ATP. In atropinized tissue, TNS produced a relaxation that was reduced 15% by guanethidine (5 x 10(-6) M), 50% by L-NNA (10(-5) M), and 30% by apamin (10(-7) M). Thus the LES receives cholinergic excitatory and adrenergic and nitrergic inhibitory innervations; the latter two components contribute evenly to the i.j.p. generation. The relaxation is mainly produced by NO in a membrane potential-independent way.     

Membrane potential of smooth muscle cells in K-free solution

1. The changes of the ion content, the membrane potential and of the membrane permeability of taenia coli cells have been studied during exposure to K-free solutions. The relative value of the total membrane conductance was determined by measuring the electrotonic potential during constant current pulses with an intracellular electrode. The P(K) values were calculated from (42)K-efflux in K-free solutions.2. In solutions containing penetrating anions the cells initially depolarize. Thereafter they hyperpolarize to about - 85 mV and again depolarize after 90 min to - 5 mV. These potential changes are much smaller if large anions are used as chloride substitutes. Moreover, the final depolarization is only reached after 4-5 hr. This hyperpolarization is not inhibited by 10(-5)M ouabain.3. These potential changes are accompanied by a progressive exchange of intracellular K by Na. In solutions containing chloride or nitrate the relative value of the total membrane conductance increases to a maximal value, corresponding to the peak value of the calculated P(K). Such changes of the membrane conductance and of P(K) do not occur in K-free solutions containing large anions.4. It is proposed that the initial depolarization is probably caused by an inhibition of an electrogenic Na pump. In chloride or nitrate solution the hyperpolarization is due to an increase of the [K](i)/[K](o) ratio and to an increase of the K permeability. In the presence of large anions the hyperpolarization remains small because this increase of P(K) does not occur.     

Myopathy of internal anal sphincter with polyglucosan inclusions

In two members of an affected family with a hereditary syndrome of proctalgia fugax and constipation, a hypertrophied internal anal sphincter was found with histological features suggesting a myopathy of this muscle. In these two patients, and in an unrelated patient with a similar clinical syndrome, smooth muscle fibres of the internal anal sphincter showed numerous vacuoles, many of which contained ovoid inclusion bodies. The structural features and histochemical reactions of the inclusion bodies were consistent with a polyglucosan composition. Histological examination of the internal anal sphincter may reveal smooth muscle abnormalities in functional bowel disorders.     

The sympathetic innervation of the internal anal sphincter and rectum in the cat

The distribution of the sympathetic innervation to the internal anal sphincter (IAS) and rectum and the occurrence of different types of adrenergic receptors in the two organs were investigated in anaesthetized cats. Anal pressure and rectal motility were recorded by a manometric and a volumetric method respectively. Division of both the hypogastric nerves (HGN) and the lumbar colonic nerves (LCN) reduced the anal pressure by 46 +/- 6% of the resting pressure (40.9 +/- 6.4 mmHg) and consistently increased rectal motility. Efferent electrical stimulation of the HGN as well as the LCN elicited a contraction in the anus and the rectum, which, at maximal stimulation, caused the anal pressure to reach a similar level to that recorded before division of these nerves. After injection of phentolamine the anal contraction was abolished, whereas the rectal contraction was either abolished or converted to a beta-adrenergic relaxation. Propranolol caused increased rectal contraction in response to stimulation of the HGN and the LCN, whereas the anal contraction was unaffected. The results imply that the sympathetic nerves exert a tonic excitatory effect on the IAS and a dual effect on the rectum in the cat. The results also indicate that sympathetic fibres to the IAS are conveyed in both the HGN and the LCN. Inhibitory beta-adrenergic receptors seem to be of minor importance in regulating anal pressure.     

The role of the longitudinal muscle in the anal sphincter complex

Intersphincteric resection (ISR) enables radical sphincter-preserving surgery in a subset of low rectal tumors impinging on the anal sphincter complex (ASC). Excellent anatomical knowledge is essential for optimal ISR. This study describes the role of the longitudinal muscle (LM) in the ASC and implications for ISR and other low rectal and anal pathologies. Six human adult en bloc cadaveric specimens (three males, three females) were obtained from the University of Leeds GIFT Research Tissue Programme. Paraffin-embedded mega blocks containing the ASC were produced and serially sectioned at 250 μm intervals. Whole mount microscopic sections were histologically stained and digitally scanned. The intersphincteric plane was shown to be potentially very variable. In some places adipose tissue is located between the external anal sphincter (EAS) and internal anal sphincter (IAS), whereas in others the LM interdigitates to obliterate the plane. Elsewhere the LM is (partly) absent with the intersphincteric plane lying on the IAS. The LM gave rise to the formation of the submucosae and corrugator ani muscles by penetrating the IAS and EAS. In four of six specimens, striated muscle fibers from the EAS curled around the distal IAS reaching the anal submucosa. The ASC formed a complex structure, varying between individuals with an inconstant LM affecting the potential location of the intersphincteric plane as well as a high degree of intermingling striated and smooth muscle fibers potentially further disrupting the plane. The complexity of identifying the correct pathological staging of low rectal cancer is also demonstrated. Clin. Anat. 33:567–577, 2020. © 2019 Wiley Periodicals, Inc.     

Effects of azelastine on vagal neuroeffector transmission in canine and human airway smooth muscle

Azelastine is a newly developed antiallergic drug that is reported to antagonize histamine and leukotrienes in addition to its inhibitory action on release of chemical mediators. In the present study, the effects of azelastine on neuroeffector transmission in the airway smooth muscles with double sucrose gap and isometric tension-recording methods were evaluated. Azelastine (10(-8) to 10(-6) mol/L) markedly decreased the contractile response of human bronchial and dog tracheal muscle strips to electrical field stimulation (10 pulses at 20 V, 20 Hz, 800 microseconds) in a dose-dependent manner. In parallel with actions on twitch contractions, azelastine suppressed the amplitude of excitatory junction potentials of dog trachea without changing the resting membrane potential and input resistance of smooth muscle cells. However, azelastine did not alter acetylcholine sensitivity of the smooth muscle cells. These results indicate that azelastine possesses an inhibitory action on the release of acetylcholine by vagal nerve terminals. This inhibitory effect of azelastine may contribute to the treatment of asthma in addition to its antiallergic actions.     

Junctional transmission in renin-containing and smooth muscle cells of the afferent arteriole

Intracellular recordings were done in renin-containing juxtaglomerular (JG) and vascular smooth muscle (VSM) cells of the mouse kidney afferent arteriole. Both cell types exhibited a membrane potential around –75 mV and spontaneous depolarizing transients resembling spontaneous excitatory junction potentials (SEJPs) in the arterioles of other organs. The amplitude distribution of these randomly occurring transients was skewed in both cell types with a modal value of 1.2–1.9 mV. Activation of presumably postjunctional ₁-, P₂-, ANG II- and AVP-receptors depolarized JG and VSM cells. Application of the P₁-purinoceptor agonist 2-chloroadenosine strongly increased frequency and amplitude of the SEJP-like events, whereas these transients were abolished by the P₁-purinoceptor antagonist 8-phenyltheophylline, both substances presumably acting on prejunctional receptors. The SEJP-like events were completely depressed by reserpine treatment, but not abolished by ₁-, ₂-, and P₂-antagonists. At present, it cannot be decided, whether norepinephrine is the sole transmitter in the afferent arteriole, acting on specialized junctional adrenoceptors with the P₂-purinoceptors being irrelevant for junctional transmission, or whether both substances are co-transmitters. Except norepinephrine and ATP, all other transmitter candidates tested were ruled out for various reasons.These studies were supported by the German Research Foundation within the SFB 90 Cardiovaskuläres System     

犬阴道上皮细胞和平滑肌细胞可在体外长期稳定培养

背景：体外分离培养获得足够活性良好的种子细胞是构建阴道组织工程的关键。文献报道阴道上皮细胞体外纯化培养和传代较为困难,尤其是体外长期培养犬等大动物的阴道种子细胞尚未见报道。 目的：建立体外稳定培养犬阴道上皮细胞和平滑肌细胞方法。 方法：获取犬小块阴道组织,机械分离阴道黏膜上皮,Dispase酶和胰蛋白酶分步消化收集上皮细胞,接种于无血清角化细胞培养液中培养和传代;机械分离阴道平滑肌组织后采用Ⅱ型胶原酶消化获得平滑肌细胞,在含体积分数10%胎牛血清的DMEM培养液中连续培养传代。动态观察上皮细胞和平滑肌细胞生长增殖情况,分别采用特异性抗体行细胞免疫化学染色鉴定。 结果与结论：原代培养的上皮细胞24-36 h后开始贴壁铺展,四五天后呈对数生长,七八天可达70%融合,为单一的上皮细胞,呈典型铺路石样,未见成纤维细胞混杂。每四五天可传代1次,连续传代六七次,细胞免疫化学染色角蛋白AEl/AE3抗体阳性。平滑肌细胞原代培养24 h后贴壁呈梭形,此后呈对数生长,4 d后融合呈典型的“峰和谷”样,每三四天可传代1次,连续传代七八次,细胞免疫化学染色示α-肌动蛋白染色阳性。结果证实,犬阴道上皮细胞和平滑肌细胞可在体外长期稳定培养,可为体外构建组织工程化阴道提供足够的种子细胞。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程全文链接：