Secondary nicotinic synapses on sympathetic B neurons and their putative role in ganglionic amplification of activity.

The strength and number of nicotinic synapses that converge on secretomotor B neurons were assessed in the bullfrog by recording intracellularly from isolated preparations of paravertebral sympathetic ganglia 9 and 10. One input to every B neuron invariably produced a suprathreshold EPSP and was defined as the primary nicotinic synapse. In addition, 93% of the cells received one to four subthreshold inputs that were defined as secondary nicotinic synapses. This contradicts the prevailing view, which has long held that amphibian B neurons are singly innervated. More important, the results revealed that B cells provide the simplest possible experimental system for examining the role of secondary nicotinic synapses on sympathetic neurons. Combining the convergence data with previous estimates of divergence indicates that the average preganglionic B neuron forms connections with 50 ganglionic B neurons and that the majority of these nicotinic synapses are secondary in strength. Secondary EPSPs evoked by low-frequency stimulation ranged from 0.5 to 10 mV in amplitude and had an average quantal content of 1. Nonetheless, secondary synapses could trigger action potentials via four mechanisms: spontaneous fluctuations of EPSP amplitude, two-pulse facilitation, coactivation with other secondary synapses, and coactivation with a slow peptidergic EPSP. The data were used to formulate a stochastic theory of integration, which predicts that ganglia function as amplifiers of the sympathetic outflow. In this two-component scheme, primary nicotinic synapses mediate invariant synaptic gain, and secondary nicotinic synapses mediate activity-dependent synaptic gain. The model also provides a common framework for considering how facilitation, metabotropic mechanisms, and preganglionic oscillators regulate synaptic amplification in sympathetic ganglia.

Primary structure, distribution, and effects on motility of CGRP in the intestine of the cod Gadus morhua.

Calcitonin gene-related peptide (CGRP) was isolated from an extract of the intestine of the cod Gadus morhua. The primary structure of this 37-amino acid peptide was established as follows: ACNTA TCVTH RLADF LSRSG GIGNS NFVPT NVGSK AF-NH2. The peptide shows close structural similarities to other nonmammalian (3-4 amino acid substitutions) and mammalian (5-8 amino acid substitutions) CGRPs, and it contains the two residues Asp14 and Phe15 that seem to be characteristic for CGRP in nonmammalian vertebrates. Cod CGRP (10(-9)-10(-7) M) inhibited the motility of spontaneously active ring preparations from the cod intestine and was significantly (P < 0.05) more potent than rat alpha-CGRP. Neither prostaglandins nor nitric oxide is involved in the inhibitory response produced by cod CGRP, and the lack of effect of tetrodotoxin suggests an action of CGRP on receptors on the intestinal smooth muscle cells. The competitive CGRP antagonist human alpha-CGRP-(8-37) significantly (P < 0.05) reduced the response to cod CGRP. Immunohistochemistry demonstrated CGRP-immunoreactive neurons intrinsic to the intestine, and a dense innervation with immunoreactive nerve fibers was observed in the myenteric plexus and the circular muscle layer. Myotomy studies show that CGRP-containing nerves project orally and anally in the myenteric plexus, whereas nerve fibers in the circular muscle layer project mainly anally, indicating a role for CGRP in descending inhibitory pathways of the cod intestine.

Projections and actions of tachykininergic, cholinergic, and serotonergic neurones in the intestine of the atlantic cod.

The native tachykinins cod neurokinin A and cod substance P, serotonin and acetylcholine have excitatory effects on the circular smooth muscle of the cod intestine. Furthermore, immunoreactivities to the cod tachykinins, serotonin and two markers for cholinergic neurones, viz. choline acetyltransferase and vesicular acetylcholine transporter, have been demonstrated in myenteric neurones of the cod intestine. In order to elucidate whether the neurones containing these substances project orally and thus might be involved in the ascending excitatory reflex of peristalsis, myotomy operations have been performed on the cod intestine. The immunoreactive areas of the myenteric plexus immediately oral and anal to the myotomy operations have been measured by using confocal laser scanning microscopy. Large accumulations of immunoreactivity to the tachykinins are found on the anal side of the myotomies, indicating oral projections of tachykininergic neurones. The areas immunoreactive to serotonin and choline acetyltransferase are of equal size on the oral and anal sides. Since the tachykinin containing neurones of the intestine project orally, and since cod neurokinin A and cod substance P have excitatory effects on circular smooth muscle, we conclude that tachykininergic neurones are involved in the ascending excitatory reflex of peristalsis in the cod intestine.

Nitric oxide synthase- and neuropeptide Y-containing subpopulations of sympathetic neurons in the coeliac ganglion of the Atlantic cod, Gadus morhua, revealed by immunohistochemistry and retrograde tracing from the stomach.

In this study retrograde tracing was used to locate sympathetic ganglion cells innervating the stomach of a teleost fish, Gadus morhua. A subpopulation of small neurons in the coeliac ganglion was retrogradely labelled after Fast Blue injection in the stomach wall. Neurons projecting to the myenteric plexus and muscle layers contained tyrosine hydroxylase immunoreactivity, and neurons projecting to submucosal layers and blood vessels contained neuropeptide Y immunoreactivity in addition to being tyrosine hydroxylase immunoreactive. A population of nitric oxide synthase containing tyrosine hydroxylase immunoreactive neurons was also found in the coeliac ganglion. These neurons were not frequently labelled after injection in any layer of the stomach. The presence of entero-enteric pathways was also surveyed, but too few enteric neurons were labelled with Fast Blue after injection in the coeliac ganglion to indicate a presence of an entero-enteric reflex. We conclude that in teleost fish, as previously reported in a variety of mammals, a pattern of target specific chemical coding of sympathetic neurons exists, but that all reflex systems of mammalian vertebrates are perhaps not present in fish.

Anally projecting neurons exhibiting immunoreactivity to galanin, nitric oxide synthase and vasoactive intestinal peptide, detected by confocal laser scanning microscopy, in the intestine of the Atlantic cod, Gadus morhua.

The projections of enteric neurons showing immunoreactivity for vasoactive intestinal peptide (VIP), nitric oxide synthase (NOS) and galanin were investigated in the myenteric plexus of the intestine of the Atlantic cod, Gadus morhua. Quantification of immunoreactive material on the proximal and distal side of a myotomy was performed by means of confocal laser scanning microscopy. NOS immunoreactivity was reduced anal to the myotomy, whereas there was an accumulation of immunoreactivity for VIP and for galanin oral to the cut. These results suggest the presence of VIP, NOS and galanin in neurons with oral-to-anal projections along the intestine of the cod. Since descending neurons in the myenteric plexus of many other vertebrates also contain these substances, we conclude that the oral-to-anal projections of neurons containing VIP, NOS and galanin are highly conserved features and important for the descending phase of intestinal peristalsis on an evolutionary basis.

Dendritic morphology of neurons in sympathetic ganglia of the goldfish, Carassius auratus.

We used intracellular dye injection to examine the dendritic morphology of postganglionic neurons in the coeliac ganglion of goldfish. About 80% of the neurons had at least one dendrite, with the mean number of dendrites per cell being 7.8 +/- 5.5 (+/- SD, n = 37 cells). Dendrites varied in length from a few microns to more than 400 microns. Around 37% of the neurons possessed axon collateral in addition to dendrites. These results show that postganglionic sympathetic neurons of goldfish can have a complex morphology, more like the sympathetic neurons of small mammals than those of amphibians. This raises the possibility that at least some sympathetic ganglion cells of teleost fish receive multiple convergent preganglionic inputs, suggesting a hitherto unsuspected level of complexity in these pathways.

Neuropeptide immunoreactivity and co-existence in cardiovascular nerves and autonomic ganglia of the estuarine crocodile, Crocodylus porosus, and cardiovascular effects of neuropeptides.

The two aortas of the crocodile are in open connection at two sites, the foramen of Panizzae immediately outside the ventricles, and the arterial anastomosis at the level of the gut. The present study was performed to elucidate the innervation of the cardiovascular structures of the crocodile, in part to provide a further basis for the assumption that the apertures of the foramen and the anastomosis may be altered, possibly leading to changes in the flow profiles of the central vessels. The presence of smooth muscle arranged at the circumference of the foramen and in the walls of the anastomosis was demonstrated. The cardiovascular structures were innervated by nerves containing co-existing tyrosine hydroxylase, NPY and somatostatin immunoreactivities, which also occurred in neurons of the sympathetic ganglia. CGRP and substance P immunoreactive material co-existed in cardiovascular nerves, and in the nodose ganglion. In addition, bombesin, VIP and galanin immunoreactive nerves were found. Effects of neuropeptides on blood flows and blood pressures were studied in vivo. Substance P increased all blood flows measured, NPY increased the flow through the arterial anastomosis while neurotensin caused an initial decrease in the flow through the arterial anastomosis. In conclusion, there is a rich innervation of the heart and major vessels of the estuarine crocodile, including the foramen of Panizza and the arterial anastomosis. These nerves possibly regulate the distribution of blood in the cardiovascular system, which is further suggested by the results of the injection of neuropeptides.

Enteric reflexes and nitric oxide in the fish intestine

The aim of the present study was to elucidate the possible regulation of peristalsis in the intestine of the Atlantic cod Gadus morhua. For this purpose, the mid intestine was dissected out and placed in a partitioned bath. Balloon distension (0.1­0.4 ml) and intramural field stimulation (8 Hz, 10 V) were carried out and the responses of the circular muscle were recorded 1.5 cm orally and anally to the stimulus using force transducers. The preparations developed spontaneous contractions propagating in the anal direction with a frequency of about one contraction per 2 min. Distension of the muscle wall with a balloon did not evoke any recordable peristaltic reflexes. Intramural stimulation caused a contraction oral to the stimulation and a relaxation anal to the stimulation in most cases. Tetrodotoxin abolished the responses to electrical stimulation in both directions. Atropine reduced and methysergide abolished the oral contractions caused by electrical stimulation. Administration of the nitric oxide synthesis inhibitor l-NG-nitro-arginine methyl ester (l-NAME) abolished the anal relaxation caused by electrical stimulation and augmented the oral contractions. The results indicate the presence in teleost fish intestine of an ascending excitatory peristaltic reflex which involves a cholinergic­serotonergic pathway and a descending inhibitory reflex involving a nitrergic pathway. These observations suggest a high degree of conservation of peristaltic mechanisms during vertebrate evolution.

Neuropeptides in the Australian lungfish Neoceratodus forsteri: effects in vivo and presence in autonomic nerves.

The Australian lungfish Neoceratodus forsteri is one of the few extant species of a phylogenetically ancient group. Immunohistochemistry showed the presence of galanin-, vasoactive intestinal polypeptide (VIP)-, neurotensin-, substance P-, and calcitonin gene-related peptide (CGRP)-like immunoreactivities in nerve fibers in the heart, lung, and gut, with a coexistence of VIP-, galanin-, and somatostatin-like immunoreactivity in the lung and galanin- and somatostatin-like immunoreactivity in the gut. About 20% of the substance P-immunoreactive fibers in gut and lung contained CGRP-like material. Major vessels showed a sparse innervation. In free-swimming unanesthetized fish, neurotensin (1 nmol/kg), galanin (1 nmol/kg), and bombesin (10 nmol/kg) reduced the heart rate. In two specimens tested, the effect of neurotensin was partially antagonized by atropine. Galanin and bombesin reduced and cholecystokinin 8 (CCK-8-S) increased blood flow to the lung. Neurotensin decreased, CCK-8-S increased, and substance P had no effect on dorsal aortic pressure, and all three decreased flow to the gut. It can be concluded from the present study that the general vertebrate pattern of cardiovascular and visceral nervous control by several neuropeptides is present also in Neoceratodus.

Galanin-like immunoreactivity in extrinsic and intrinsic nerves to the gut of the Atlantic cod, Gadus morhua, and the effect of galanin on the smooth muscle of the gut.

The presence of galanin-like immunoreactivity in nerves to the stomach of the Atlantic cod has been investigated by immunohistochemistry. The distribution of ganglion cells showing galanin-like immunoreactivity was compared with the total distribution in nerves and ganglia. Projection studies were made to determine the origin of the galanin neurons. The effect of galanin was studied in smooth muscle strip preparations of the gut wall and arteries. Galanin-like immunoreactive ganglion cells frequently occurred along the vagal branches to the stomach. Most of them projected cranially. Immunoreactive nerve fibers were present in all layers of the gut and around arterial branches on the surface of the stomach. Ligations of the vagus and splanchnic nerves produced accumulations of immunoreactive material on both sides of the ligature. Galanin produced weak contractile effects unaffected by tetrodotoxin on the gut wall and on gut arteries. It is concluded that a population of the ganglion cells along the vagus nerve in the Atlantic cod contains a galanin-like peptide. Some of these cells may be parts of autonomic parasympathetic pathways innervating the gut of the Atlantic cod, having direct excitatory effects on the smooth muscles of the gut wall and gut arteries.

Effects of substance P and distribution of substance P-like immunoreactivity in nerves supplying the stomach of the cod, Gadus morhua.

The innervation of the cod stomach by neurons showing substance P-like immunoreactivity (SPLI), and the effect and mechanism of action of substance P (SP) on the vascularly perfused cod stomach and on isolated muscle strip preparations from the pyloric sphincter have been investigated.Infusion of SP produced a contraction of the stomach wall, which could not be blocked by tetrodotoxin, atropine or methysergide, indicating a direct effect on the stomach smooth muscle. Similarly, the contraction produced by SP on preparations from the pyloric sphincter was unaffected by tetrodotoxin.Nerves showing SPLI were frequent in the myenteric plexus of the whole stomach, and in the submucosa and mucosa of the pyloric part of the stomach. SPLI was also observed in fibres in the intestinal branch of the vagus and occasionally in the splanchnic nerves. Ligation of the nerves showed an accumulation of SPLI above as well as below the ligature, being more prominent proximal to the ligature in the vagus and distal to the ligature in the splanchnic nerve. In the vagus nerve, descending and ascending SPLI-fibres were seen surrounding non-reactive cell bodies. No reduction in intensity of the immunoreaction of the neurons in the stomach wall was observed after ligation or sectioning of the vagosympathetic trunk or the splanchnic nerves, nor were SP-levels measured by radioimmunoassay reduced. After denervation of vagal branches close to the stomach wall an insignificant decrease of immunoreactivity was observed in the myenteric plexus. Capsaicin treatment had no conclusive effect on the distribution of SPLI.It is concluded that the innervation showing SPLI may be of intrinsic as well as extrinsic origin, with pathways in both vagal and splanchnic branches. Only a direct effect of SP on the smooth muscle could be demonstrated.

A randomized hemodynamic comparison of intravenous amiodarone with and without Tween 80.

In 20 patients undergoing coronary arteriography, the hemodynamic effects of an experimental preparation of i.v. amiodarone 5 mg kg-1 without Tween 80 (N) (10 patients) were compared with those of the commercial form with Tween 80 (A) (10 patients). Analysis of variance demonstrated differences during the 3 min of injection and for 3 min afterwards: left ventricular systolic pressure decreased from 110 + 11 to 86 +/- 11 mmHg (P = 0.001) after A and from 114 +/- 22 to 106 +/- 19 (P = 0.05) after N (comparison P = 0.01) while related tachycardia was also more pronounced after A (comparison P = 0.001). Left ventricular end diastolic pressure transiently decreased after A while continuously increasing after N (P = 0.05). During the following 30 min both A and N caused similar bradycardia, increase in ventricular filling pressure, vascular resistance and decrease in cardiac and contractility indexes. Amiodarone blood levels were similar after A or N. These data document a significant initial short duration vasoplegia, mainly related to Tween 80, after A, when amiodarone itself after producing a similar very slight effect causes bradycardia, and a moderate and progressive negative inotropic effect. It was concluded that while the experimental form would be of interest, the risk of severe hypotension after i.v. Cordarone can be largely avoided by using a slower rate of infusion, especially in patients with hypovolemic status.

[Simultaneous appearance of precordial pain and effort-induced left bundle branch block. A study of 6 patients with normal coronary angiography]. / Apparition simultanée de précordialgie et d'un bloc de branche gauche à l'effort. Etude chez 6 patients avec coronarographie normale.

6 patients, 4 men and 2 women, aged 38 to 60 years, were hospitalised for chest pain on effort, labelled as angina. The resting electrocardiogram was normal, except in 2 cases with incomplete left bundle branch block. Exercise stress testing induced simultaneous chest pain and complete left bundle branch block (LBBB) (at a rate of 15 to 160 beats/min, mean 115). The pain lasted as long as the LBBB was present and eased as it disappeared. All patients had normal coronary angiography and negative provocative tests of coronary spasm with atrial stimulation (4 cases), isoprenaline test (2 cases), atropine injection (2 cases) and nitroglycerin (1 case). A 2D echocardiogram performed in all the patients, showed no signs of myocardial disease. Myocardial perfusion on effort was studied by Thallium 201 scintigraphy in all 6 patients. There were no perfusion defects. An exercise stress test was performed after beta-blocker treatment in 3 patients. LBBB appeared each time and the pain was always present. The induction of bradycardia with propranolol (5 cases) or verapamil (3 cases) did not reduce the frequency of chest pain. No serious complications were observed during follow-up (myocardial infarction, death). The syndrome of painful LBBB is not related exclusively to tachycardia. Myocardial ischaemia does not seem to be the underlying mechanism judged by the negative thallium studies.