Synaptic transmission at nicotinic acetylcholine receptors in rat hippocampal organotypic cultures and slices.
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1. Whole-cell clamp recordings of the compound synaptic current elicited by afferent stimulation of Schaffer collaterals showed that blockade of the NMDA, AMPA and GABAA receptor-mediated components by 6-nitro-7-sulphamoyl- benzo(f)quinoxaline-2,3-dione (NBQX), 3-((R)-2-carboxypiperazine-4-yl)propyl-1-phosphonate (R-CPP) and picrotoxin, respectively, left a small residual current in 39 out of 41 CA1 pyramidal neurones in organotypic cultures and 9 out of 16 CA1 cells in acutely prepared slices. 2. This current represented 2. 9 +/- 0.4 % of the compound evoked synaptic response in organoypic cultures and 1.4 +/- 0.5 % in slices. It was characterized by a slightly rectifying I-V curve and a reversal potential of 3.4 +/- 5. 1 mV. 3. This residual current was insensitive to blockers of GABAB, purinergic, muscarinic and 5-HT3 receptors, but it was essentially blocked by the nicotinic receptor antagonist d-tubocurarine (91 +/- 4 % blockade; 20 microM), and partly blocked by alpha-bungarotoxin (200 nM) and methyllycaconitine (10 nM), two antagonists with a higher selectivity for alpha7 subunit-containing nicotinic receptors (48 +/- 3 % and 55 +/- 11 % blockade, respectively). 4. The residual current was of synaptic origin, since it occurred after a small delay; its amplitude depended upon the stimulation intensity and it was calcium dependent and blocked by the sodium channel antagonist tetrodotoxin. 5. We conclude that afferent stimulation applied in the stratum radiatum evokes in some hippocampal neurones a small synaptic current mediated by activation of neuronal nicotinic receptors. (+info)
Nicotinic acetylcholine receptors assembled from the alpha7 and beta3 subunits.
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Intracellular recordings were performed in voltage-clamped Xenopus oocytes upon injection with a mixture of cDNAs encoding the beta3 and mutant alpha7 (L247Talpha7) neuronal nicotinic acetylcholine receptor (nAChR) subunits. The expressed receptors maintained sensitivity to methyllycaconitine and to alpha-bungarotoxin but exhibited a functional profile strikingly different from that of the homomeric L247Talpha7 receptor. The heteromeric L247Talpha7beta3 nAChR had a lower apparent affinity and a faster rate of desensitization than L247Talpha7 nAChR, exhibited nonlinearity in the I-V relationship, and was inhibited by 5-hydroxytryptamine, much like wild type alpha7 (WTalpha7) nAChR. Single channel recordings in cell-attached mode revealed unitary events with a slope conductance of 19 picosiemens and a lifetime of 5 ms, both values being much smaller than those of the homomeric receptor channel. Upon injection with a mixture of WTalpha7 and beta3 cDNAs, clear evidence was obtained for the plasma membrane assembly of heteromeric nAChRs, although ACh could not activate these receptors. It is concluded that beta3, long believed to be an orphan subunit, readily co-assembles with other subunits to form heteromeric receptors, some of which may be negative regulators of cholinergic function. (+info)
Effects of Delphinium alkaloids on neuromuscular transmission.
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The Delphinium alkaloids methyllycaconitine (MLA), nudicauline, 14-deacetylnudicauline (14-DN), barbinine, and deltaline were investigated for their effects on neuromuscular transmission in lizards. The substituent at C14 provides the only structural difference among the alkaloids MLA, nudicauline, 14-DN, and barbinine. Deltaline lacks the N-(methylsuccinyl)anthranilic acid at C18 common to the other four alkaloids. Each alkaloid reversibly reduced extracellularly recorded compound muscle action potential (CMAP) amplitudes in a concentration-dependent manner. The IC(50) values for CMAP blockade were between 0.32 and 13.2 microM for the N-(methylsuccinimido)anthranoyllycacotonine-type alkaloids and varied with the C14 moiety; the IC(50) value for deltaline was 156 microM. The slopes of the concentration-response curves for CMAP blockade were similar for each alkaloid except barbinine, whose shallower curve suggested alternative or additional mechanisms of action. Each alkaloid reversibly reduced intracellularly recorded spontaneous, miniature end-plate potential (MEPP) amplitudes. Alkaloid concentrations producing similar reductions in MEPP amplitude were 0.05 microM for 14-DN, 0.10 microM for MLA, 0.50 microM for barbinine, and 20 microM for deltaline. Only barbinine altered the time constant for MEPP decay, further suggesting additional or alternative effects for this alkaloid. MLA and 14-DN blocked muscle contractions induced by exogenously added acetylcholine. All five alkaloids are likely nicotinic receptor antagonists that reduce synaptic efficacy and block neuromuscular transmission. The substituent at C14 determines the potency and possibly the mechanism of nicotinic acetylcholine receptor blockade for MLA, nudicauline, 14-DN, and barbinine at neuromuscular synapses. The lower potency of deltaline indicates that the N-(methylsuccinyl)anthranilic acid at C18 affects alkaloid interactions with nicotinic acetylcholine receptors at neuromuscular junctions. (+info)
Cooperative activation of action potential Na+ ionophore by neurotoxins.
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Four neurotoxins that activate the action potential Na+ ionophore of electrically excitable neuroblastoma cells interact with two distinct classes of sites, one specific for the alkaloids veratridine, batrachotoxin, and aconitine, and the second specific for scorpion toxin. Positive heterotropic cooperativity is observed between toxins bound at these two classes of sites. Tetrodotoxin is a noncompetitive inhibitor of activation by each of these toxins (KI = 4-8 nM). These results suggest the existence of three functionally separable components of the action potential Na+ ionophore: two regulatroy components, which bind activating neurotoxins and interact allosterically in controlling the activity of a third ion-transport component, which binds tetrodotoxin. (+info)
Nicotinic receptor activation in human cerebral cortical interneurons: a mechanism for inhibition and disinhibition of neuronal networks.
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Cholinergic control of the activity of human cerebral cortical circuits has long been thought to be accounted for by the interaction of acetylcholine (ACh) with muscarinic receptors. Here we report the discovery of functional nicotinic receptors (nAChRs) in interneurons of the human cerebral cortex and discuss the physiological and clinical implications of these findings. The whole-cell mode of the patch-clamp technique was used to record responses triggered by U-tube application of the nonselective agonist ACh and of the alpha7-nAChR-selective agonist choline to interneurons visualized by means of infrared-assisted videomicroscopy in slices of the human cerebral cortex. Choline induced rapidly desensitizing whole-cell currents that, being sensitive to blockade by methyllycaconitine (MLA; 50 nM), were most likely subserved by an alpha7-like nAChR. In contrast, ACh evoked slowly decaying whole-cell currents that, being sensitive to blockade by dihydro-beta-erythroidine (DHbetaE; 10 microM), were most likely subserved by an alpha4beta2-like nAChR. Application of ACh (but not choline) to the slices also triggered GABAergic postsynaptic currents (PSCs). Evidence is provided that ACh-evoked PSCs are the result of activation of alpha4beta2-like nAChRs present in preterminal axon segments and/or in presynaptic terminals of interneurons. Thus, nAChRs can relay inhibitory and/or disinhibitory signals to pyramidal neurons and thereby modulate the activity of neuronal circuits in the human cerebral cortex. These mechanisms, which appear to be retained across species, can account for the involvement of nAChRs in cognitive functions and in certain neuropathological conditions. (+info)
Anti-arrhythmic effects of sophoridine and oxysophoridine.
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AIM: To compare the effects of oxysophoridine (Oxy) and sophoridine (Sop) on experimental arrhythmias and myocardial physiologic properties. METHODS: Arrhythmias were induced by drugs and myocardial ischemia. Physiologic properties were determined on isolated heart atria. RESULTS: Oxy 500 mg.kg-1 (1/6 LD50) decreased the incidence of ventricular arrhythmias induced by aconitine (P < 0.01), increased the threshold dose of ouabain-induced ventricular premature (VP, P < 0.05), ventricular tachycardia (VT, P < 0.05), ventricular fibrillation (VF, P < 0.01), and cardiac arrest, (P < 0.01). After i.v. Oxy 500 mg.kg-1 into the rats with ligation of left anterior descending coronary artery, the total numbers of ectopic beats were decreased (P < 0.05), the incidence of VF was lowered, and the duration of VT was shortened (P < 0.01). Oxy 250 mg.kg-1 (1/13 LD50) i.v. shortened the duration of arrhythmias induced by BaCl2 (P < 0.01) and delayed the onset of arrhythmias induced by chloroform-epinephrine (P < 0.05). Oxy produced dose-dependent positive inotropic effects in the isolated left atrial of guinea pigs, increased the concentration of epinephrine to elicit automaticity in left atria, decreased slightly the excitability, and prolonged the functional refractory period. Sop produced the similar effects on arrhythmias as Oxy. CONCLUSION: Oxy produced the similar anti-arrhythmic effects as Sop did at the equivalent effective dose. (+info)
Inhibition and disinhibition of pyramidal neurons by activation of nicotinic receptors on hippocampal interneurons.
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Nicotinic acetylcholine receptors (nAChRs) are expressed in the hippocampus, and their functional roles are beginning to be delineated. The effect of nAChR activation on the activity of both interneurons and pyramidal neurons in the CA1 region was studied in rat hippocampal slices. In CA1 stratum radiatum with muscarinic receptors inhibited, local pressure application of acetylcholine (ACh) elicited a nicotinic current in 82% of the neurons. The majority of the ACh-induced currents were sensitive to methyllycaconitine, which is a specific inhibitor of alpha7-containing nAChRs. Methyllycaconitine-insensitive nicotinic currents also were present as detected by a nonspecific nAChR inhibitor. The ACh-sensitive neurons in the s. radiatum were identified as GABAergic interneurons by their electrophysiological properties. Pressure application of ACh induced firing of action potentials in approximately 70% of the interneurons. The ACh-induced excitation of interneurons could induce either inhibition or disinhibition of pyramidal neurons. The inhibition was recorded from the pyramidal neuron as a burst of GABAergic synaptic activity. That synaptic activity was sensitive to bicuculline, indicating that GABA(A) receptors mediated the ACh-induced synaptic currents. The disinhibition was recorded from the pyramidal neuron as a reduction of spontaneous GABAergic synaptic activity when ACh was delivered onto an interneuron. Both the inhibition and disinhibition were sensitive to either methyllycaconitine or mecamylamine, indicating that activation of nicotinic receptors on interneurons was necessary for the effects. These results show that nAChRs are capable of regulating hippocampal circuits by exciting interneurons and, subsequently, inhibiting or disinhibiting pyramidal neurons. (+info)
Two distinct classes of functional 7-containing nicotinic receptor on rat superior cervical ganglion neurons.
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Nicotinic acetylcholine receptors (nAChRs) that bind alpha-bungarotoxin (alpha Bgt) were studied on isolated rat superior cervical ganglion (SCG) neurons using whole-cell patch clamp recording techniques. Rapid application of ACh onto the soma of voltage clamped neurons evoked a slowly desensitizing current that was reversibly blocked by alpha Bgt (50 nM). The toxin-sensitive current constituted on average about half of the peak whole-cell response evoked by ACh. Nanomolar concentrations of methyllycaconitine blocked the alpha Bgt-sensitive component of the ACh-evoked current as did intracellular dialysis with an anti-alpha 7 monoclonal antibody. The results indicate that the slowly reversible toxin-sensitive response elicited by ACh arises from activation of an unusual class of alpha 7-containing receptor (alpha 7-nAChR) similar to that reported previously for rat intracardiac ganglion neurons. A second class of functional alpha 7-nAChR was identified on some SCG neurons by using rapid application of choline to elicit responses. In these cases a biphasic response was obtained, which included a rapidly desensitizing component that was blocked by alpha Bgt in a pseudo-irreversible manner. The pharmacology and kinetics of the responses resembled those previously attributed to alpha 7-nAChRs in a number of other neuronal cell types. Experiments measuring the dissociation rate of 125I-labelled alpha Bgt from SCG neurons revealed two classes of toxin-binding site. The times for toxin dissociation were consistent with those required to reverse blockade of the two kinds of alpha Bgt-sensitive response. These results indicate that rat SCG neurons express two types of functional alpha 7-nAChR, differing in pharmacology, desensitization and reversibility of alpha Bgt blockade. (+info)