Excitatory Effects Of Acetylcholine Research Articles

EFFECTS OF ACETYLCHOLINE, VAGAL STIMULATION AND TYRAMINE ON THE ISOLATED ATRIA OF THE TORTOISE.

Abstract In the vagus-atrial preparation of the tortoise (Amyda japonica), a cold-blooded animal, acetylcholine and vagal stimulation, in the presence of atropine, produced positive inotropic effects which were inhibited by dichloroisoprenaline or pronethalol. Hexamethonium blocked the excitatory effect of acetylcholine but only slightly inhibited that of vagal stimulation. Tyramine exerted a positive inotropic action in this preparation, but had little effect on atria prepared from reserpine-treated animals. Tyramine was shown to release a substance from the tortoise atria which caused contraction of aortic strips taken from reserpine-treated rabbits, and which appeared to be a catecholamine.

Dose-dependence of the excitatory effects of acetylcholine on common snail neurons after orthodromic tetanization.

Studies of the cellular mechanisms of learning and memory often use neuronal analogs of these processes. Post-tetanic changes in the reactivity of nerve cells provide one such model. Orthodromic tetanic stimulation of nerves evokes increases in the cholinosensitivity of the bodies of neurons LPa3 and RPa3 in the common snail [3]. The likely mechanism for post-tetanic increases in neuron cholinosensitivity may consist of changes in the number of binding centers for the mediator in the cholinoreceptor molecule, which is characterized by the Hill coefficient [4]. The aim of the present work was to compare the Hill coefficient for somatic extrasynaptic cholinoreceptors on neurons LPa3 and RPa3 in the common snail before and during post-tetanic increases in the cholinosensitivity of these cells. The Hill coefficient was calculated by constructing dose-response curves for the effects of acetylcholine (ACh) on neurons. Experiments were conducted using semi-intact “CNS-visceral sac” preparations from the common snail Helix lucorum. Methods for obtaining these preparations, applying acetylcholine, and recording transmembrane currents have been described in detail elsewhere [3]. Intracellular electrodes were made from Pyrex glass and filled with 2 M KCl or 2 M potassium acetate (electrode resistance was 20.61 ± 0.04 MΩ). The mean amplitude of the excitatory postsynaptic current was 18.0 ± 3.0 nA. Local iontophoretic application of ACh was to the dorsal surface of the cells from glass micropipettes filled with 1 M ACh chloride (Sigma, USA) and having a resistance of 37.29 ± 0.04 MΩ. Cationic phoretic currents were 0.1‐1.5 μA with injections lasting 0.2‐2 sec. The mean amplitude of the ACh response was 14.9 ± 1.6 nA, with a latency of 1.5 ± 0.2 sec. Electrical tetanic stimulation of the intestinal nerve was applied via Nichrome electrodes (resistance 200 MΩ) using square-wave current impulses (10.5 mA, 100 msec); tetanization lasted 1‐2 min (1‐2 per sec). The Hill coefficient was determined as the tangent of the angle of the slope on plots of the relationship between the magnitude of the response and concentration of agonist in double logarithmic coordinates at the lowest concentrations [4]. The measure of the quantity of ACh applied to membranes used in the present work was the strength of the phoretic current through the ACh-containing injection pipette. The size of the phoretic current was directly proportional to the quantity of acetylcholine released in strictly uniform conditions (phoresis time, micropipette resistance) in each individual experiment. Results were obtained from 37 neurons (17 LPa3 and 20 RPa3) in 37 preparations. Cell membrane potentials was ‐50.31 ± 0.05 mV. Recording and measurement of the amplitudes of ACh-evoked currents (ACh currents) were performed using a IBM-486 PC running the program CONAN 3.0. Results were analyzed statistically used in Excel, Stadia, and Statistica 4.3. The significance of effects was assessed using Student’s and Wilcoxon’s tests. The strength of the iontophoretic current was selected at the beginning of the experiment such that the amplitude of the ACh response was maximal. The interval between mediator injections was 10 min. The mean of the three maximal ACh responses was taken as 100%. The dose-response relationship was then determined for the effects of ACh: ACh was injected by phoretic currents of increasing magnitude (0.1‐1.5 μA). After control applications of ACh, experimental series were performed in which tetanic stimulation was applied to the intestinal nerve (1‐2 per sec, 1‐2 min).

Acetylcholine attenuates synaptic GABA release to supraoptic neurons through presynaptic nicotinic receptors

Both inhibitory GABAergic and excitatory glutamatergic inputs to supraoptic nucleus (SON) neurons can influence the release of vasopressin and oxytocin. Acetylcholine is known to excite SON neurons and to increase vasopressin release. The functional significance of cholinergic receptors, located at the presynaptic nerve terminals, in the regulation of the excitability of SON neurons is not fully known. In this study, we determined the role of presynaptic cholinergic receptors in regulation of the inhibitory GABAergic inputs to the SON neurons. The magnocellular neurons in the rat hypothalamic slice were identified microscopically, and the spontaneous miniature inhibitory postsynaptic currents (mIPSCs) were recorded using the whole-cell voltage-clamp technique. The mIPSCs were abolished by the GABA A receptor antagonist, bicuculline (10 μM). Acetylcholine (100 μM) significantly reduced the frequency of mIPSCs of SON neurons from 3.59±0.36 to 1.62±0.20 Hz ( n=37), but did not alter the amplitude and the decay time constant of mIPSCs. Furthermore, the nicotinic receptor antagonist, mecamylamine (10 μM, n=13), eliminated the inhibitory effect of acetylcholine on mIPSCs of SON neurons. The muscarinic receptor antagonist, atropine (100 μM), did not alter significantly the effect of acetylcholine on mIPSCs in most of the 17 SON neurons studied. These results suggest that the excitatory effect of acetylcholine on the SON neurons is mediated, at least in part, by inhibition of presynaptic GABA release. Activation of presynaptic nicotinic receptors located in the GABAergic terminals plays a major role in the cholinergic regulation of the inhibitory GABAergic input to SON neurons.

Modulation of ganglion cell activity in the pineal gland of the rainbow trout: effects of cholinergic, catecholaminergic, and GABAergic receptor agonists.

Second order neurons within intact isolated pineal glands of the rainbow trout were explored by extracellular recordings to investigate modulatory effects of putative intrapineal neurotransmitters. Acetylcholine, dopamine, and norepinephrine were found to increase ganglion cell activity in a majority of cells tested. The excitatory effects of acetylcholine, dopamine, and norepinephrine were mimicked by muscarinic, dopamine D2, and beta-adrenergic receptor agonists and significantly increased with the applied light intensity, resulting in an attenuation of the ganglion cell response to light. GABA decreased discharge activity in most cells tested. This effect, which could be mimicked with the GABAA receptor agonist piperidine, was independent from the adaptive status. Acetylcholine and GABA were still active if applied during synaptic blockade with low Ca++ high Mg(++)-perfusion medium, whereas dopamine and norepinephrine exhibited no effects if applied during synaptic blockade, suggesting a differential cellular distribution of neurotransmitter receptors in the trout pineal gland. These data demonstrate that ganglion cell activity in the trout pineal gland is under the influence of several neurotransmitters, including acetylcholine, dopamine, norepinephrine, and GABA, which is in contrast to the originally proposed simple bineuronal transduction pathway from photoreceptors onto ganglion cells. Since the above-mentioned neurotransmitters are believed to be released from pineal interneurons, we may conclude that ganglion cell activity in the teleost pineal gland is, similarly to the retina, the product of photoreceptor signals and a modulatory active interneuronal system.

Catecholamine and acetylcholine sensitivity of rat lateral hypothalamic neurons related to learning.

1. Unit activity in the rat lateral hypothalamus (LHA) was recorded during discrimination learning of cue tone (CTS) or cue light (CL) stimulation that predicted reward by glucose or intracranial self stimulation (ICSS), or aversion by weak electric shock or tail pinch. Roles of the catecholaminergic and cholinergic systems in the LHA were investigated by electrophoretic application of dopamine (DA), norepinephrine (NE), acetylcholine (ACh), and their antagonists [spiperone (SPP), phenoxybenzamine (PBZ), phentolamine, propranolol, and atropine (Atr)]. 2. Activity of 264 LHA neurons was recorded. Of these, 234 (89%) responded during CTS learning in one or more phases. Of 121 neurons tested by both rewarding and aversive stimuli, 86 (71%) discriminated reward and aversion and their respective CTSs. 3. Effects of DA on 138, NE on 134, and ACh on 73 neurons were tested. Among these, 67 were tested with all three. DA inhibited 40 and excited 14. NE inhibited 74 and excited 10. ACh excited 35 and inhibited 3. DA-sensitive neurons responded to both NE (P less than 0.001) and ACh (P less than 0.05) more often than DA-insensitive neurons. In most cases, the effect of DA was similar to the effect of NE, and opposite to the effect of ACh. The inhibitory effect of DA was blocked by SPP, a D2 antagonist, and the excitatory effect of ACh was blocked by Atr. The inhibitory effect of NE was blocked by the beta-antagonist, propranolol, and enhanced by the alpha-antagonist, phentolamine. 4. DA-sensitive neurons responded to both rewarding and aversive stimuli and respective CTS+ and CTS- more often than DA-insensitive neurons (P less than 0.01). The effect of DA was usually similar to the effect of rewarding stimuli and their predicting CTS+ and was opposite to the effect of aversive stimuli and their predicting CTS-. 5. The proportion of NE-sensitive neurons that responded to rewarding and aversive stimuli was the same as the proportion of NE-insensitive neurons that responded to the same stimuli. NE-sensitive neurons responded to CTS+ and CTS- more often than NE-insensitive neurons (P less than 0.01). The effect of NE was usually similar to the effect of rewarding stimuli and predicting CTS+, and opposite to the effect of aversive stimuli and predicting CTS-. 6. ACh-sensitive neurons responded to aversive stimuli and predicting CTS- more often than ACh-insensitive neurons (P less than 0.01), but the response ratio of ACh-sensitive neurons to rewarding stimuli was similar to that of ACh-sensitive neurons.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of acetylcholine and substance P on electrical activity of intact toad gastric muscles

By using isolated gastric muscle cells of Bufo marinus, others have studied the mechanism of action of muscarinic agonists and substance P (SP). To compare responses of isolated cells with those of intact muscles, we have studied the effects of acetylcholine (ACh) and SP on membrane potentials of circular muscle cells in strips of intact muscle from the toad gastric corpus region. These cells had average resting potentials of -69 +/- 0.7 mV. Membrane potential rhythmically depolarized, producing slow waves at an average frequency of 1/min and average amplitude of 25 +/- 2.2 mV. The major effect of ACh (10(-7) to 10(-4) M) was chronotropic; the frequency of slow waves was increased by 88 +/- 11% by 10(-6) M ACh. The amplitudes and rates of rise of slow waves were decreased by ACh. SP had effects similar to ACh; its major effect was chronotropic. The data suggest that ACh and SP primarily affect the pacemaker mechanism in gastric muscles. Since rhythmicity is apparently not expressed in isolated gastric myocytes, it is possible that this effect of these agonists may have been missed in studies of dispersed cells. Our data suggest that the excitatory effects of ACh and SP on contractions may be due to summation of Ca2+ signals, a partial tetanus-like effect.

Role of acetylcholine in induction of repetitive activity in human atrial fibers.

The actions of acetylcholine and its interactions with epinephrine were studied in human atrial tissues by recording transmembrane potentials and contractile force. Acetylcholine (0.55-5.5 microM) reduced force, shortened the duration and shifted to more negative values the plateau of action potentials, abolished phase 4 depolarization, and suppressed the activity of spontaneous fibers. During the recovery, often there was a rebound increase in some parameters of the action potential and in force. Epinephrine (0.3-2.8 microM) induced oscillatory potentials and aftercontractions and acetylcholine abolished them. However, during the washout of acetylcholine in the presence of epinephrine, the oscillatory potentials and aftercontractions were larger than before acetylcholine, and repetitive activity was often induced. The inhibitory and excitatory effects of acetylcholine were mimicked by methacholine (5.1 microM) and abolished by atropine (1.5 microM). The postacetylcholine rebound was also potentiated by theophylline (0.6-2 mM) but was not blocked by propranolol (1-3.4 microM), prazosin (1 microM), and diltiazem (0.1 microM). It is concluded that in human atrial fibers acetylcholine has inhibitory as well as excitatory effects that are exaggerated in the presence of epinephrine and are mediated by the activation of the muscarinic receptor. The interaction between acetylcholine and epinephrine involves an antagonism at an intracellular level.

Xylazine-evoked depression of rat cerebral cortical neurons: a pharmacological study

1. 1. Iontophoretic application of the veterinary tranquilizer xylazine suppressed the spontaneous firing of 134 137 neurons tested, an effect not antagonized by adenosine receptor and α-adrenergic receptor blocking agents. 2. 2. Depressions in neuronal activity evoked by norepinephrine, γ-aminobutyric acid or adenosine failed to be potentiated by xylazine. 3. 3. However, the excitatory effects of acetylcholine and glutamate were both inhibited by xylazine. 4. 4. Thus, the depressant effect of locally applied xylazine may involve a decrease in the excitability of the neuronal membrane.

Pertussis toxin blocks the inhibitory effect of adenosine on rat cerebral cortical neurons

Rat cerebral cortex was injected with 2 μg of purified pertussis toxin, which inactivates various guanine nucleotide-dependent regulatory proteins. The spontaneous firing of cortical neurons in toxin-treated animals was unresponsive to adenosine or adenosine 5′- N-ethylcarboxamide applied iontophoretically in coparison with the pronounced inhibitory effects of these purines observe on neurons in the contralateral, saline-injected, hemisphere. Excitatory effects of acetylcholine were also reduced in the toxin-treated cortex. These findings implicate guanine nucleotide-dependent regulatory proteins (G proteins) in the inhibitory actions of adenosine on transmitter release.

Modulatory effect of serotonin on the acetylcholine sensitivity of identified neurons in the brain of Helix pomatia L.

1. 1. Excitatory effect of acetylcholine (ACh) was modified by serotonin (5-HT) on identified neurons of intact brain with partially preserved functional connections at Helix pomatia L. 2. 2. Depolarization caused by ACh on neuron LPa3 and on several spontaneously active cells of the visceral ganglion was enhanced by brief application of 5-HT to the soma of these neurons. Guanosine triphosphate (GTP) and theophylline caused similar effects to the serotonin changes in ACh sensitivity of these cells. 3. 3. ACh sensitivity of neurons RPa3 and RPa2 was reduced following application of 5-HT, GTP and theophylline. 4. 4. Similarly, the effect of 5-HT tactile stimulation of the heart caused decrease in ACh sensitivity of neuron RPa2.

Competitive inhibition by dimethylsulfoxide of molluscan and vertebrate acetylcholinesterase

Anticholinesterase-like effects of dimethylsulfoxide (DMSO) were demonstrated on a variety of invertebrate muscles. The excitatory effects of acetylcholine (ACh) on the isolated preparations of the Geukensia demissa heart and anterior byssus retractor muscle (ABRM), and of the Busycon contrarium radula protractor muscle, were potentiated by DMSO (1–5 μl/ml; 1 μl/ml; = 14 mM). The negative chronotropic effects of ACh, but not of 4-ketoamyltrimethylammonium, were potentiated by DMSO (1–5 μl/ml) on the isolated heart of the oyster Crassostrea virginica. These four muscles have acetylcholinesterase enzymes of high activity. In contrast, Mercenaria mercenaria hearts have weak cholinesterase activity, and the effects of ACh on this isolated myocardium were not potentiated by DMSO (2–20 μl/ml). DMSO (0.1–15 μl/ml) was a competitive inhibitor of both a crude preparation of oyster heart acetylcholinesterase (AChE) (the K m increased 24-fold with DMSO at 15 μl/ml; the I 50 was 1.3 μl/ml DMSO when [ACh] = K m ) and a purified Electrophorus AChE (the Km increased 4.5-fold when DMSO was 10 μl/ml; the I 50 was 10 μl/ml DMSO near [ACh] = K m ). The same doses of DMSO were needed to potentiate the pharmacological effects of ACh on the oyster heart, as to inhibit the AChE of this tissue.

Heterogeneous behavior of the canine arterial and venous wall. Importance of the endothelium.

Experiments were designed to determine the contribution of endothelial cells to the heterogeneous behavior of the arterial and venous wall. Rings of canine femoral, pulmonary, saphenous, and splenic arteries and veins, with and without endothelium, were mounted for isometric tension recording in Krebs-Ringer bicarbonate solution. Endothelium-dependent inhibitory responses to acetylcholine, adenosine triphosphate, bovine thrombin, and arachidonic acid were prominent in the arteries. In the veins, only transient endothelium-dependent relaxations to these substances were observed. Removal of the endothelium decreased the augmentation of the response to norepinephrine caused by anoxia in both arteries and veins. In the veins, arachidonic acid and thrombin caused endothelium-dependent increases in tension during contractions evoked by norepinephrine. The endothelium-independent inhibitory effects of isoproterenol and adenosine and the excitatory effects of acetylcholine and ATP were more pronounced in the veins than in the arteries. These experiments demonstrate that in the arterial and venous wall the endothelial cells can contribute to both inhibitory and excitatory responses of the smooth muscle cells of the media. Inhibitory endothelial responses prevail in the arteries, and excitatory ones in the veins.

Excitatory effect of acetylcholine on different types of neurons in the first somatosensory neocortex of the rat: Laminar distribution and pharmacological characteristics

In rats anaesthetized with either urethane, pentobarbital or fluothane the effects of acetylcholine, cholinergic agonists and antagonists (applied by iontophoresis) were studied on single cortical neurons of first somatosensory region. The laminar distribution of the neurons excited by acetylcholine was determined by the reconstruction of each electrode track based on a dye deposit made at the last recording site. Neurons were identified using antidromic stimulation of the pyramidal tract, the ventrobasal thalamus and the corpus callosum. Neurons excited by acetylcholine could be segregated into two groups: one encompassing layer Vb and the upper part of layer VI, the other more deeply located at the limit between the cerebral cortex and the subjacent white matter. Neuronal responses to glutamate and nicotine, unlike those to acetylcholine were evenly distributed in the cortex. Pyramidal tract neurons and corticothalamic neurons were frequently excited by acetylcholine and were shown to be located with the first group of acetylcholine sensitive neurons. Commissural neurons were rarely excited by acetylcholine and were not restricted to either group. The analysis of neuronal responses to acetylcholine and various agonists (carbachol, nicotine, acetyl-β-methylcholine, carbamyl-β-methylcholine, butyrylcholine) and antagonists (atropine, mecamylamine) revealed a prominent but not exclusive muscarinic character. It is concluded (i) that cortical neurons of first somatosensory cortex which are excited by acetylcholine belong to two populations, one consisting, at least in part, of projection neurons (upper group) and the other of interneurons (lower group); (ii) that cortical acetylcholine receptors are of a ‘mixed’ type strongly weighted toward the muscarinic side.

Modulation of cholinergic transmission by substance P.

Substance P administered iontophoretically to Renshaw cells in the cat had a dual effect, sometimes causing excitation and at other times inhibiting the excitatory effect of acetylcholine (ACh). The inhibitory effect was selective for the nicotinic receptors on Renshaw cells and the excitatory effect seemed to be due to the release of ACh from cholinergic terminals. It has not been possible to demonstrate a similar inhibitory effect on nicotinic receptors at the neuromuscular junction in frogs or in the chick, although a small agonist effect was occasionally observed. In the atropinized cat, intra-arterial injections of ACh to the superior cervical ganglion cause both a rise in blood pressure and contractions of the nictitating membrane which are abolished by hexamethonium. Intra-arterial injections of substance P partially blocked these nicotinic actions of ACh, but no excitatory effect of substance P was observed. These observations are discussed in relation to other studies and indicate that the polypeptide could function as an inhibitory or facilitatory regulator of cholinergically mediated responses at some but not all cholinergic synapses.

Effect of prostaglandins F2? and E2 on sensitivity of rabbit cortical neurons to acetylcholine and noradrenalin

The effect of prostaglandins F2α and E2 on unit activity and sensitivity of somatosensory cortical neurons of rabbits to acetylcholine and noradrenalin was investigated by microiontophoresis. Prostaglandin F2α predominantly inhibited, whereas E2 increased the spike activity of the neurons. F2α usually modified the sensitivity of the neurons to acetylcholine, E2 to noradrenalin. The influence of F2α on the excitatory effects of acetylcholine, and of E2 on the inhibitory effects of noradrenalin as a rule was characterized by weakening of the action of the mediators or a change in the sign of the initial responses. It is suggested that prostaglandins of the F group participate in cholinergic, and those of the E group in adrenergic transmission. The prostaglandins studied evidently exert their action through selective inhibition of the activity of adenylate or guanylate cyclases, leading to a decrease in the synthesis of secondary transmitters, namely cyclic AMP and cyclic GMP.

Noise and relaxation studies of acetylcholine induced currents in the presence of procaine.

1. The effects of procaine on the excitatory effects of acetylcholine (ACh) on Aplysia neurones were studied by analysing the current relaxations induced by voltage steps. Procaine was applied by perfusion and ACh was applied iontophoretically under two sets of experimental conditions designed to produce very different ACh concentrations at the membrane. 2. When ACh was applied at high concentration, the current relaxation following a hyperpolarizing step showed two components at procaine concentrations of about 2 X 10(-5) M; a fast increase was followed by a slow decrease. At high procaine concentrations (greater than 10(-4) M), the relaxation has only one component, corresponding to an exponential current decrease. 3. When ACh was applied at low concentrations, the current relaxation usually showed two components. At low procaine concentrations a fast increase was followed by a slower one; at higher procaine concentrations, a fast decrease preceded the slow increase. 4. All the results can be accounted for by assuming that procaine binds preferentially to the activated receptor-channel complex and converts it into a non-conducting complex. The association and dissociation constants were calculated to be (at 12 degrees C, -80 mV) 1.3 X 10(6) M-1 sec-1 and 10 sec-1.

Dual excitatory of acetylcholine in the mouse vas deferens.

SummaryRelatively low concentrations of acetylcholine ( 10-4M) may be due to release of norepinephrine following interaction of acetylcholine with nicotinic receptors on adrenergic nerve terminals.

Excitatory effect of acetylcholine on the reflexogenic zone of the epicardium and pericardium in vagotomized animals

Excitatory effect of acetylcholine on the reflexogenic zone of the epicardium and pericardium in vagotomized animals