TREK-1 channels do not mediate nitrergic neurotransmission in circular smooth muscle from the lower oesophageal sphincter

Background and purpose:

The ionic mechanisms underlying nitrergic inhibitory junction potentials (IJPs) in gut smooth muscle remain a matter of debate. Recently, it has been reported that opening of TWIK-related K⁺ channel 1 (TREK-1) K⁺ channels contributes to the nitrergic IJP in colonic smooth muscle. We investigated the effects of TREK-1 channel blockers on nitrergic neurotransmission in mouse and opossum lower oesophageal sphincter (LOS) circular smooth muscle (CSM).

Experimental approach:

The effects of TREK-1 channel blockers were characterized pharmacologically in murine and opossum gut smooth muscle using conventional intracellular and tension recordings.

Key results:

In LOS, L-methionine depolarized the resting membrane potential (RMP) but did not inhibit the nitrergic IJP. Cumulative application of theophylline hyperpolarized the RMP and inhibited the nitrergic IJP concentration dependently. The induced membrane hyperpolarization was prevented by pre-application of caffeine, but not by 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one. 8-Br-cAMP significantly hyperpolarized membrane potential and increased the amplitude of the nitrergic IJP. In opossum LOS muscle strips, L-methionine increased resting tone but had no effect on nerve-mediated LOS relaxation. On the other hand, theophylline markedly inhibited tone. In CSM from mouse proximal colon, L-methionine caused modest inhibition of nitrergic IJPs.

Conclusions and implications:

TREK-1 channels were not involved in the nitrergic IJP in LOS CSM. Not only does L-methionine have no effect on the nitrergic IJP or LOS relaxation, but the effect of theophylline appears to be due to interruption of Ca²⁺-releasing pathways (i.e. caffeine-like effect) rather than via blockade of TREK-1 channels.     

Regional differences in nitrergic innervation of the smooth muscle of murine lower oesophageal sphincter

BACKGROUND AND PURPOSE: Anatomical and pharmacological studies have demonstrated that the lower oesophageal sphincter (LES) is not a simple homogenous circular muscle with uniform innervation. Regional differences have been demonstrated in several species including humans. We investigated, for the first time in mice LES, regionally distinct physiological and pharmacological characteristics of the neuromusculature. EXPERIMENTAL APPROACH: Conventional intracellular recordings and pharmacological techniques were employed to evaluate electrical properties and functional innervation of smooth muscle cells. Results from CD1 (control), nNOS((-/-)) and eNOS((-/-)) genetic knockout mice were compared. KEY RESULTS: Smooth muscle of sling and clasp LES displayed unitary membrane potentials of 1- 4 mV. Transmural nerve stimulation produced a monophasic inhibitory junction potential (IJP) in the sling, whereas in the clasp a biphasic IJP, consisting of a brief IJP followed by a long-lasting slow IJP (lsIJP), was induced. Pharmacological interventions and genetically modified mice were used to demonstrate a monophasic apamin-sensitive (purinergic) component in both LES regions. However, the nitrergic IJP was monophasic in the sling and biphasic in the clasp. Unitary membrane potentials and IJPs were not different in CD1 and eNOS((-/-)) mice, suggesting no involvement of myogenic NOS. CONCLUSION AND IMPLICATIONS: These data in mouse LES indicate that there are previously unreported regional differences in the IJP and that both the apamin-resistant monophasic and biphasic IJPs are mediated primarily by nitrergic innervation.     

Mechanical and biochemical responses to endothelin-1 and endothelin-3 in bovine bronchial smooth muscle.

1. In this study, mechanical responses to endothelin-1 and endothelin-3 were examined in bovine bronchial smooth muscle. In addition, the involvement of phosphatidylinositol 4,5-bisphosphate hydrolysis (PIP2) in the responses to these peptides was assessed by measurement of inositol (1,4,5) trisphosphate (I(1,4,5)P3) production using a specific mass assay. 2. ET-1 evoked contractions of bovine bronchi which were concentration-dependent and initiated at between 10(-9) M and 10(-8) M. ET-1-evoked responses were unaffected by slight elevation of tone with potassium chloride (3 x 10(-2) M), methacholine (10(-6) M) or U46619 (10(-7) M). 3. Contractions to ET-1 were not altered by pre-incubation with atropine (10(-5) M), indomethacin (10(-5) M), nifedipine (10(-5) M), phosphoramidon (3.67 x 10(-5) M) or by removal of the epithelium. 4. ET-3 evoked small contractions which were not concentration-dependent. In the presence of phosphoramidon (3.67 x 10(-5) M) however, concentration-dependent contractions were obtained to ET-3 which were unaffected by atropine (10(-5) M) or by removal of the epithelium, but were significantly attenuated by indomethacin (10(-5) M). Nifedipine (10(-5) M) virtually abolished this response. 5. Both ET-1 and ET-3 (in the presence of phosphoramidon)-evoked contractions were significantly enhanced by the presence of the phorbol ester phorbol 12,13-dibutyrate (10(-8) M). Neither ET-1-, nor ET-3-mediated responses were antagonized by the protein kinase C (PKC) inhibitor, Ro 31-8220 (3 x 10(-9) - 3 x 10(-8) M). 6. ET-1 (3 x 10(-7) M) evoked a biphasic rise in levels of I(1,4,5)P3 which was unaltered by preincubation with atropine, whilst ET-3 (10(-10) - 3 x 10(-7) M) failed to alter levels of I(1,4,5)P3 at any time point examined, even in the presence of phosphoramidon (3.67 x 10(-5) M).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of dichloroglyoxime on isolated guinea-pig smooth muscle and atrium.

The effects of dichloroglyoxime (DCG) on isolated rings of aorta, main pulmonary artery, trachea and spontaneously-beating atrium of guines-pig were studied. DCG caused concentration- dependent relaxation of the epinephrine-contracted aortic and pulmonary artery rings and of the tone of tracheal rings. Propranolol caused a slight shift to the right in the concentration-effect curves of DCG on these preparations. Quinacrine, an inhibitor of the release of arachidonic acid and its metabolites, caused a significant shift to the right in the concentration-effect curves of DCG on the three preparations. Low concentrations of DCG increased the beating rate of the atrium, an effect which was blocked by propranolol but not by quinacrine whereas large concentrations decreased the beating rate, an effect which was not significantly affected by propranolol or by quinacrine. DCG also caused a concentration-dependent decrease in the contractility of the atrium and this effect was only slightly affected by propranolol or quinacrine. These observations suggest that the relaxant effect of dichloroglyoxime on the smooth muscle may not be mediated by the stimulation of beta adrenoceptors specifically although a nonspecific interaction with these receptors or with the contractile machinery of the cell cannot be excluded. Data with quinacrine suggest that the effects may be mediated by the release of an inhibitory metabolite of arachidonic acid. The results further suggest that in the atrium the effects of DCG may not be specific and they may be partially mediated by the release of catecholamines from the nerve endings.     

The effects of itopride on oesophageal motility and lower oesophageal sphincter function in man

Aliment Pharmacol Ther 2011; 33: 99–105

Summary

Background Itopride is a new prokinetic agent that combines antidopaminergic and cholinesterase inhibitory actions. Previous studies suggested that itopride improves heartburn in functional dyspepsia, and decreases oesophageal acid exposure in gastro‐oesophageal reflux disease. It remains unclear whether this effect is due to effects of itopride on the lower oesophageal sphincter (LES). Aims To study the effects of itopride on fasting and postprandial LES function in healthy subjects. Methods Twelve healthy volunteers (five men; 32.6 ± 2.0 years) underwent three oesophageal sleeve manometry studies after 3 days premedication with itopride 50 mg, itopride 100 mg or placebo t.d.s. Drug was administered after 30 min and a standardized meal was administered after 90 min, with measurements continuing to 120 min postprandially. Throughout the study, 10 wet swallows were administered at 30‐min intervals, and gastrointestinal symptoms were scored on 100 mm visual analogue scales at 15‐min intervals. Results Lower oesophageal sphincter resting pressures, swallow‐induced relaxations and the amplitude or duration of peristaltic contractions were not altered by both doses of itopride, at all time points. Itopride pre‐treatment inhibited the meal‐induced rise of transient LES relaxations (TLESRs). Conclusions Itopride inhibits TLESRs without significantly affecting oesophageal peristaltic function or LES pressure. These observations support further studies with itopride in gastro‐oesophageal reflux disease.     

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

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

Effects of 3,3'-di-O-methylquercetin on guinea-pig isolated smooth muscle

The effects of the flavone 3,3'-di-O-methylquercetin (DOMQ) have been examined and compared with those of quercetin, on guinea-pig isolated ileum, trachea, and main pulmonary artery (MPA). Except for transient contractions induced by low concentrations (10(-8)-3 x 10(-6) M), DOMQ and quercetin (up to 3 x 10(-4) M) caused reduction of the tone and the phasic contractions of the ileum. A23187 reversed the inhibitory effects of quercetin but not those of DOMQ. DOMQ and quercetin caused concentration-dependent relaxation of the trachea and the adrenaline-contracted MPA. DOMQ shifted to the right the concentration-effect curves induced by acetylcholine on the ileum and the trachea, and by adrenaline on MPA and those induced by CaCl2 on ileum, trachea and MPA. DOMQ also inhibited the contractions induced, in Ca2+-free EGTA-containing buffer, by histamine on ileum and by adrenaline on MPA. These observations suggest that DOMQ inhibits Ca2+ influx, Ca2+ release from intracellular stores and, more likely, Ca2+ binding to intracellular receptor proteins.     

β3-Adrenoceptors mediate smooth muscle relaxation in the rat lower oesophageal sphincter

1 The role of β₃-adrenoceptors in isoprenaline-induced relaxation of carbachol-precontracted ring segments of the rat lower oesophageal sphincter (LOS) was examined. 2 Isoprenaline (10^?8m – 10^?5m ) relaxed ring segments of the LOS in a concentration-dependent manner. Propranolol (10^?7m ) had very little antagonist effect on isoprenaline-induced relaxation. 3 Dobutamine (10^?7m – 10^?4m ), salbutamol (10^?7m – 10^?4m ) and BRL 37344 (10^?8– 10^?5m ) also all relaxed carbachol-contracted ring segments of the LOS in a concentration-dependent manner. The relaxant responses to these agonists were similarly not antagonized by propranolol (10^?7m ). 4 Cyanopindolol (10^?6m ), produced a parallel rightward displacement of isoprenaline, dobutamine, salbutamol and BRL 37344 concentration–response curves with similar potencies. The pK_B values range from 7.3 ± 0.1 to 7.7 ± 0.2. 5 The relaxant effect of isoprenaline in the rat LOS was not inhibited by N^G-nitro-l -arginine (l -NOARG; 3 × 10^?5m ), glibenclamide (10^?5m ) or tetraethylammonium (1 mm ). 6 It was concluded that β₃-adrenoceptors mediate isoprenaline-induced relaxation in rat lower oesophageal sphincter and that activation of these receptors was not linked to either ATPase-, Ca²⁺-dependent K⁺ channels or to NO release.     

Action of nicotine on guinea-pig isolated bronchial smooth muscle preparation

In the isolated bronchial preparation of the guinea-pig, nicotine induced a contraction but not a relaxation. The contractile response of the bronchial preparation to nicotine was inhibited by hexamethonium and d-tubocurarine but not influenced by atropine and tetrodotoxin. In the isolated tracheal preparation of the guinea-pig where nicotine stimulated nicotinic receptor in nervous tissues, the contractile response to nicotine as considerably accelerated by the treatment of the guinea-pig with egg-albumin, while the contractile response of the bronchial preparation to nicotine was not influenced by the same treatment. These results suggest that a possible site of action of nicotine in the isolated bronchial preparation is not on the nervous cells but on the smooth muscle cells. However, we could not rule out a contribution by chemical mediators released by nicotine in the contractile mechanisms in the bronchial preparation.     

Field stimulation-induced responses of circular smooth muscle from guinea-pig stomach

Summary Field stimulation of circular smooth muscle of guinea-pig stomach from the regions of the cardia and fundus caused contraction responses at low stimulation frequencies (0.25–1 Hz) with relaxation at higher frequencies (1–10 Hz), whilst tissues of the body and antrum responded with contraction throughout the frequency range. Atropine (10^–9–10^–8 M) antagonised the contraction responses of all tissues, with relaxation developing at higher concentrations (except for antral tissue). In contrast, metoclopramide (10^–8–10^–6 M) caused modest (cardia, fundus) or marked (body, antrum) enhancement of contractions to field stimulation, whilst domperidone (10^–8–10^–7 M), haloperidol (10^–8–10^–6 M), prazosin, propranolol and methysergide (10^–8–10^–6 M) failed to modify the contraction responses. However, whilst yohimbine and guanethidine failed to modify the contractions of the cardia, fundus and body tissues, those of the antral preparations were antagonised by nanomolar concentrations of yohimbine and by guanethidine (10^–6–5×10^–5 M). To optimise the relaxation responses for study, atropine was included in the physiological solution. Relaxation to field stimulation of preparations from the body and cardia, but not the fundus, was antagonised by reserpine pretreatment (5 mg/kg i.p., 24h), addition of guanethidine (10^–5–10^–4 M), phentolamine, prazosin or propranolol (10^–7–10^–6 M) (the effects of prazosin and propranolol being additive). Higher concentrations of haloperidol and domperidone antagonised the relaxation responses of the body preparations only. Metoclopramide, yohimbine and methysergide (10^–8–10^–6 M) were ineffective. Thus, it is concluded that the contractile effects of the 4 stomach areas to field stimulation reflects a major cholinergic involvement, with an additional ₂-adrenoceptor contractile component in antral tissue. Relaxation responses of cardia and body tissue involve ₂- and -adrenoceptors plus a further, unidentified, non-adrenergic component; the latter represents the total relaxation response of the fundic preparation.     

重组人白细胞介素-1受体拮抗药对豚鼠离体呼吸道平滑肌的影响

目的　观察人重组白细胞介素 1受体拮抗剂 (IL 1ra)对正常和卵白蛋白致敏豚鼠离体肺条、气管平滑肌的影响。方法　应用离体器官装置、张力换能器、MedLab记录系统测定肺条和气管平滑肌的张力。结果　①IL 1ra对正常豚鼠离体肺条和气管平滑肌有直接松弛作用 ,EC50 分别为1 2 9ⅹ 10 -7mol·L-1和 8 0 6× 10 -8mol·L-1;并对卵白蛋白致敏的豚鼠肺条和气管平滑肌也有直接的松弛作用 ,EC50 分别为 2 6 1× 10 -7mol·L-1和 5 88× 10 -7mol·L-1,但致敏豚鼠呼吸道平滑肌对IL 1ra的敏感性要比正常豚鼠低。②IL 1ra(10 -9～ 10 -5mol·L-1)可剂量依赖性地抑制致痉剂组胺对正常豚鼠肺条和气管平滑肌的收缩作用 (P <0 0 1)。③IL 1ra能抑制卵白蛋白攻击引起的肺条和气管平滑肌的收缩 ,IC50 分别为 7 83ⅹ 10 -7mol·L-1和 4 4 8ⅹ 10 -7mol·L-1。结论　IL 1ra对正常、痉挛及致敏状态的呼吸道平滑肌均有松弛作用     

Adenosine receptor-mediated contraction and relaxation of guinea-pig isolated tracheal smooth muscle: effects of adenosine antagonists.

1. The effects of several adenosine analogues and antagonists on guinea-pig isolated trachea have been examined. 2. 5'-N-ethylcarboxamidoadenosine (NECA), 5'-N-methylcarboxamidoadenosine (MECA) and adenosine (in the presence and absence of dipyridamole) elicited concentration-dependent tracheal relaxation. 3. The R(-)- and S(+)-enantiomers of N6-(2-phenylisopropyl)adenosine (R-PIA and S-PIA respectively), N6-cyclohexyladenosine (CHA) and 2-chloroadenosine (CADO) caused contractions at low concentrations (0.05-2.0 microM), whereas at higher concentrations, relaxation resulted. 4. For tracheal relaxation, the adenosine analogues exhibited the following rank order of potency: NECA greater than CADO greater than R-PIA = MECA greater than S-PIA greater than adenosine. The rank order of potency for inducing contractions was R-PIA greater than CHA greater than CADO greater than S-PIA. These data suggest that relaxation is mediated by adenosine A2-receptors, whereas contraction is the result of activation of A1-receptors. 5. 8-Phenyltheophylline (8-PT), aminophylline, the triazoloquinazoline CGS 15943A and NPC205 (1,3-di-n-propyl-8-(4-hydroxyphenyl)xanthine) each inhibited the R-PIA-induced contractile response, whereas enprofylline was without effect. NPC205, aminophylline and 8-PT were competitive antagonists, but CGS15943A was non-competitive. 6. That the most potent antagonist was the A1-selective agent, NPC205 (pA2 = 7.80), further suggests that the contraction is mediated by A1-receptors. Moreover, NPC205 was 13 times more potent as an antagonist of R-PIA-induced contractions (A1) than of NECA-induced relaxations (A2). 7. The antagonists were also found to relax the trachea by an unknown mechanism. That enprofylline did not antagonize the R-PIA-induced contractions, but was 3-4 times more potent a tracheal relaxant than aminophylline, further suggests that a direct effect on airway smooth muscle, rather than antagonism of endogenous adenosine, is more relevant to the bronchodilator effect of alkylxanthines in the treatment of asthma.     

Mechanical effects of some prostanoids on isolated human oesophageal submucosal veins.

The effects of prostaglandins E1, E2, F2 alpha, prostacyclin, and the thromboxane A2-mimic U46619 were investigated on isolated human oesophageal submucosal veins from the oesophageal body and the oesophagogastric junction. U46619 most potently, but also PGF2 alpha produced venocontraction without differences between preparations from the oesophageal body and the oesophagogastric junction. PGE1 and prostacyclin caused relaxation of vessels precontracted with U46619 (10(-9) M). PGE2 induced either contraction or relaxation, but biphasic effects in the same vessel segment were not seen. Indomethacin 10(-6) M inhibited the contractile responses to both noradrenaline and K+ (124 mM), suggesting that the agonists induced synthesis or release of vasoconstrictor prostanoids. Prostanoids exert potent mechanical effects in submucosal oesophageal veins and may be of physiological importance.     

Relaxant effects of lithium on guinea-pig tracheal smooth muscle in vitro.

The effect of lithium was studied on resting tension of guinea-pig spiral tracheal strips and their responses to carbachol and histamine in vitro. Lithium reversibly relaxed tracheal smooth muscle in a dose-dependent manner. In addition, lithium increased the ED50 for carbachol 10 fold and that for histamine by over 100 fold; maximum responses for each agonist were also reduced. Lithium-induced relaxation of tracheal smooth muscle was unaffected by changes in extracellular calcium concentration over the range 0-11 mM, verapamil, ouabain, procaine, propranolol or reduction in extracellular sodium concentration. The ability of lithium to reduce carbachol- and histamine-induced contraction of tracheal smooth muscle was also not altered by propranolol, procaine, ouabain, verapamil or lowered extracellular sodium. We conclude that lithium acts directly on tracheal smooth muscle to relax it by an as yet unknown mechanism. This may or may not be related to the ability of this agent to alter agonist-induced contraction of tracheal smooth muscle.     

The effects of metoclopramide on acetylcholine release and on smooth muscle response in the isolated guinea-pig ileum

Summary The effects of metoclopramide on smooth muscle contraction and on release of acetylcholine were studied in the guinea-pig myenteric plexus longitudinal muscle preparation. Acetylcholine was determined either as endogenous acetylcholine, or as labelled transmitter from strips preloaded with ³H-choline.Metoclopramide caused an increase in resting tension of longitudinal muscle as well as an increase in resting output of either endogenous or labelled acetylcholine. Tetrodotoxin abolished the metoclopramide-evoked increase in transmitter release. The increase in smooth muscle tension was clearly related to the increase in resting output. The effects of metoclopramide on both longitudinal muscle contraction and resting release of labelled acetylcholine were prevented in the presence of a concentration of 5-hydroxytryptamine (5-HT) that desensitized 5-HT receptors. This suggests that metoclopramide stimulates neuronal 5-HT receptors and, thereby, facilitates acetylcholine release.Metoclopramide augmented the twitch-like contractions induced by field stimulation at 0.1 Hz. Contractions elicited at 1 Hz were only slightly enhanced. Similarly, metoclopramide facilitated only the release of labelled acetylcholine evoked by electrical stimulation at 0.1 Hz, but not that at 1 Hz. The facilitatory effetts of metoclopramide on twitch height and evoked release could not be attributed to a blockade of presynaptic inhibitory -adrenoceptors, dopamine or muscarine receptors.     

Lack of direct antiarrhythmic electrophysiological effects of salicylate on isolated guinea-pig myocardium

Conventional microelectrode techniques were used to study the influence of Na-salicylate, Na-benzoate, Na-2,6-dihydroxybenzoate and 2,4-dinitrophenol (2,4-DNP) on the action potential (AP) of guinea-pig papillary muscles and atria. In papillary muscles, Na-salicylate (0.19-1.87 mmol/l) concentration-dependently shortened the AP duration and the functional refractory period. The AP amplitude decreased slightly with the largest concentration, while the resting potential and the maximum depolarisation velocity (Vmax) were not affected. A concentration-dependent negative inotropic effect was also seen. All drug effects were reversible after washout. In atria, 6.24 mmol/l Na-salicylate induced a slight shortening of the AP duration, a decrease of the AP amplitude and Vmax, but no decrease of the contractile force. The effects of the uncoupling agent, 2,4-DNP (10 mumol/l), were similar to those of the largest concentration of Na-salicylate in papillary muscles and in atria. Na-benzoate and Na-2,6-dihydroxybenzoate had no significant influence on AP duration, AP amplitude, resting potential, Vmax, refractory period or force of contraction of either papillary muscles or atria. These results suggest that Na-salicylate exerts its effects on isolated guinea-pig myocardium by uncoupling the oxidative phosphorylation, whereas two other possible mechanisms of action, namely an increase of membrane surface charge and an inhibition of prostaglandin synthesis, seem to be of minor importance.     

The isolated circular muscle layer of the vas deferens of the guinea-pig.

1 A method is described for the removal of the outer longitudinal layer of muscle from the guinea-pig isolated vas deferens. The remaining tube of circular muscle was perfused at constant flow for recording changes in pressure in response to transmural electrical stimulation or to drugs. 2 The response to transmural stimulation was a rise in perfusion pressure at frequencies of stimulation of 5 to 50 Hz for trains of 16 to 256 pulses. At some frequencies and train lengths a second rise in pressure (the after-response) occurred after the cessation of the stimulus train. 3 Perfused vasa stripped of their longitudinal muscle did not develop longitudinal tension on electrical stimulation nor on intraluminal or extraluminal exposure to agonists. 4 Stripped perfused vasa gave concentration-dependent pressure rises to noradrenaline but not to acetylcholine. 5 Responses to transmural stimulation and to noradrenaline were antagonized by thymoxamine. Cocaine or desmethylimipramine increased the duration of the after-response to transmural stimulation. Reserpine pretreatment almost abolished the response and after-response. The after-response, but not the response, was increased by physostigmine and antagonized by atropine. 6 The results are discussed in relation to the known histochemical and electronmicroscopical demonstrations of dense noradrenergic and cholinergic populations of nerve terminals in the circular layer. It is suggested that mechanisms may exist for the separate control of the longitudinal and circular layers as a basis for propulsive activity.