Cholinergic effect on the dopamine turnover in the rat corpus striatum]

The peripheral administration of oxotremorine caused a significant increase in dihydroxyphenylacetic acid (DOPAC) in the striatum of rats, dopamine (DA) level was unaffected. Injection of oxotremorine into the substantia nigra failed to change the content of dopamine and its acid metabolites homovanillic acid (HVA) and DOPAC in striatum. Injection of oxotremorine or carbachol into the substantia nigra or into the caudate nucleus did not significantly influence the DA-turnover. The partly inconsistent results are discussed in connection with literature data in regard to the existence of excitatory as well as inhibitory cholinergic systems, which are located differently and are involved in the regulation of DA-turnover.     

Stereotyped behaviour after cholinergic, but not dopaminergic, stimulation of the substantia nigra in rats

Stereotyped behaviour of the rat was measured after intracerebral drug application in an objective and quantitative way by means of a new method developed in this laboratory. Bilateral intranigral injection of apomorphine /APO/, a specific dopaminergic agonist, did not evoke any signs of stereotyped behaviour. Also ineffective was the application of APO in the amygdaloid nucleus. Dopaminergic blockade of the substantia nigra by topical application of triperidol, a potent dopaminergic antagonist, failed to influence the stereotypy elicited by systematic APO administrationDirect cholinergic stimulation of the substantia nigra with carbachol resulted in a dose-related stereotyped behaviour not distinguishable by sight from that evoked by systematic APO administration. The effect of intranigral carbachol was antagonized by a previous intraperitoneal injection of 10 mg/kg of atropine. Stereotypy could easily be produced also by intracaudate application of APO. Topical triperidol blockade of the caudate nucleaus prevented the stereotypy caused by intraperitoneal application of APO.It is concluded that at least a part of nigral neurons cannot be directly excited by apomorphine. However, they can be excited by carbacol and seem thus, to contain muscarinic receptors. The stimulation of these receptors results in an excitation of the neurons involved and produces marked stereotyped behaviour.     

A noncholinergic action of acetylcholinesterase (AChE) in the brain: From neuronal secretion to the generation of movement

1. In various brain regions, there is a puzzling disparity between large amounts of acetylcholinesterase and low levels of acetylcholine. One such area is the substantia nigra. Furthermore, within the substantia nigra, a soluble form of acetylcholinesterase is released from the dendrites of dopamine-containing nigrostriatal neurons, independent of cholinergic transmission. These two issues have prompted the hypothesis that acetylcholinesterase released in the substantia nigra has an unexpected noncholinergic function. 2. Electrophysiological studies demonstrate that this dendritic release is a function, not of the excitability of the cell from which the acetylcholinesterase is released, but of the inputs to it. In order to explore this phenomenon at the behavioral level, a novel system has been developed for detecting release of acetylcholinesterase "on-line." It can be seen that release of this protein within the substantia nigra can reflect, but is not causal to, movement. 3. Once released, the possible actions of acetylcholinesterase can be studied at both the cellular and the behavioral level. Independent of its catalytic site, acetylcholinesterase has a "modulatory" action on nigrostriatal neurons. The functional consequences of this modulation would be to enhance the sensitivity of the cells to synaptic inputs. 4. Many basic questions remain regarding the release and action of acetylcholinesterase within the substantia nigra and, indeed, within other areas of the brain. Nonetheless, tentative conclusions can be formulated that begin, in a new way, to provide a link between cellular mechanisms and the control of movement.     

Cholinergic receptors in the Aplysia gill

Acetylcholine has been suggested as a neurotransmitter released in the Aplysia gill by peripheral afferents of central neurons and by peripheral neurons within the gill. The perfused gill, isolated from the abdominal ganglion, was examined. At concentrations greater than 1 microM, acetylcholine elicited a slowly developing tonic contraction of the afferent vein that reversed upon washout. This effect was observed on both quiescent and active preparations. At concentrations less than 1 microM, acetylcholine perfusion resulted in a reduction of gill tone. The excitatory effect of acetylcholine was reduced 80 and 60% by the cholinergic antagonists atropine and hexamethonium, respectively. The acetylcholine-evoked contraction was potentiated 2.5-fold when curare was coinfused. Carbachol did not mimic the excitatory effects of acetylcholine. At all concentrations examined (1-100 microM), carbachol infusion reduced baseline tension, the amplitude of spontaneous contractions and contractions evoked by FMRFamide and dopamine. Contractions evoked by perfusion of p-chlorophenylthiocyclic AMP were greatly reduced when carbachol was added to the perfusate. Further addition of curare reversibly blocked carbachol inhibition of the cyclic AMP-evoked contractions. These findings suggest that excitatory and inhibitory cholinergic receptors are involved in the regulation of gill contractile behavior by acetylcholine.     

Modulation of cerebral acetylcholine metabolism by the dorsal diencephalic conduction system in the rat

We investigated the effects of interruption of the impulse flow in the habenulopeduncular pathways by local infusion of tetrodotoxin on the acetylcholine and choline content in selected dopamine rich regions in the forebrain and midbrain in rats. The tetrodotoxin infusion caused a marked increase in acetylcholine content in the medial frontal cortex, striatum and ventral tegmental area+interpeduncular nucleus, but not in the limbic area or the substantia nigra, whereas choline content was reduced only in both the striatum and ventral tegmental area+interpeduncular nucleus. There was an increase in 3,4-dihydroxyphenylacetic acid content in the striatum after the manipulation. These findings suggest that the dorsal diencephalic conduction system may be involved in the integration of the activity of cholinergic neurons in the forebrain and midbrain regions and striatal dopanine neurons may play a role in the modulation of cholinergic neurons.     

Tremor following intracerebral carbachol injection. 1.Carbachol sensitivity of various brain structures]

Tremor produced by intracerebral injection of carbachol. I. Susceptibility of different brain areas to carbachol Microinjections of carbachol into the lateral ventricle of rats caused tremor depending on dose. Intensity and duration of motor effects after injection of carbachol (30 mug/3 mul bilateral) into different brain areas were found to depend on localization: strongest tremor was elicited by injections into the nucleus caudatoputamen and cortex cerebri, moderate tremor by administration into the substantia nigra reticularis, globus pallidus and thalamic brain regions. Target areas of mean sensitivity were demonstrated in more rostral and caudal parts of the formatio reticularis. The injection of carbachol into the nucleus ruber, nucleus linearis and substantia nigra compacta brought about lowest tremor values. Ablation of the site of injection from the remaining brain abolished tremor induced by carbachol contrary to the tremor induced by oxotremorine.     

Dual muscarinic and nicotinic action on a motor program in Drosophila.

The effect of cholinergic agonists and antagonists on the central pattern generator of the pharyngeal muscles has been studied in third instar larvae of Drosophila. The pharyngeal muscles are a group of rhythmically active fibers involved in feeding. Bath application of the cholinergic agonists carbachol, muscarine, pilocarpine, and acetylcholine (ACh) to a semiintact preparation including the pharyngeal muscles and the central nervous system (CNS), initiated long-lasting endogenous-like bursting activity in the muscles. The muscarinic antagonists, atropine and scopolamine, blocked these responses as well as endogenous activity. Perfusion with nicotine elicited a short, tonic response that was marginally blocked by mecamylamine but not by curare, alpha-bungarotoxin, hexamethonium, or the muscarinic antagonists. This is the first time that a response to cholinergic drugs has been examined in Drosophila. The pharyngeal muscle preparation may prove to be a valuable system for studying mutations of cholinergic metabolism, receptors, and second messengers.     

Dual muscarinic and nicotinic action on a motor program in Drosophila

The effect of cholinergic agonists and antagonists on the central pattern generator of the pharyngeal muscles has been studied in third instar larvae of Drosophila. The pharyngeal muscles are a group of rhythmically active fibers involved in feeding. Bath application of the cholinergic agonists carbachol, musarine, pilocarpine, and acetylcholine (ACh) to a semiintact preparation including the pharyngeal muscles and the central nervous system (CNS), initiated long-lasting endogenous-like bursting activity in the muscles. The muscarinic antagonists, atropine and scopolamine, blocked these responses as well as endogenous activity. Perfusion with nicotine elicited a short, tonic response that was marginally blocked by mecamylamine but not by curare, α-bungarotoxin, hexamethonium, or the muscarinic antagonists. This is the first time that a response to cholinergic drugs has been examined in Drosophila. The pharyngeal muscle preparation may prove to be a valuable system for studying mutations of cholinergic metabolism, receptors, and second messengers.     

Perikaryal RNA changes within neurons of the caudate-putamen and substantia nigra in soman intoxicated rats

Quantitative azure B cytophotometry was employed to monitor neuronal ribonucleic acid (RNA) metabolism within cholinergic and noncholinergic brain compartments following single sc injections of either 0.5, 0.9 or 1.5 LD₅₀ soman (pinacolyl methylphosphonofluoridate). Dose-dependent losses in neuronal RNA were observed within the cholinergic caudate-putamen (CP) and dopaminergic substantia nigra pars compacta (SNPC), whereas levels of RNA were generally maintained or elevated within the gabaergic substantia nigra pars reticulata (SNPR). CP acetylcholinesterase was inhibited in a dose-dependent fashion. These neuronal RNA changes are perhaps related to seizure activity. The overall data lend support to the hypothesis that alterations in noncholinergic activity contribute to certain manifestations of soman neurotoxicity, such as seizures, which probably stem from an impairment in the functional integrity of both excitatory and inhibitory neuronal elements.     

Pineal muscarinic phosphoinositide response: pertussis toxin resistant signaling with very low receptor number

Autoradiography revealed very low density of muscarinic receptors in the rat pineal gland. Yet, the magnitude of phosphoinositide hydrolysis elicited with 0.1 mM carbachol was similar to that seen with 1 mM norepinephrine. The cholinergic and adrenergic phosphoinositide responses were fully additive. The cholinergic signal was insensitive to pertussis toxin both in vivo and in vitro and persisted in pineals cultured for 5 days. The data expand our previous finding on functional muscarinic acetylcholine receptors in the rat pineal gland.     

Role of the substantia nigra in the behavioural-cardiovascular integration in the cat

Electrical stimulation of the substantia nigra elicited complex patterns of response composed of somatomotor and circulatory changes. Increase in blood pressure associated with acceleration in heart rate was consistently produced by substantia nigra stimulation both in conscious and in anaesthetized cats. The respiration was either accelerated or arrested by the stimulation. Also self-stimulation of the substantia nigra elicited pressor responses. Electrical stimulation of the nucleus of the tractus solitarius paired with self-stimulation of the substantia nigra, produced regular changes in the lever-pressing rate for self-stimulation. It is suggested that the substantia nigra is probably involved in the neural mechanisms coupling the circulatory changes with the somatomotor responses.     

Acetylcholine activates cerebral interneurons and feeding motor program in Limax maximus

The cellular and network effects of acetylcholine (ACh) on the control system for feeding in Limax maximus were measured by intracellular recordings from feeding command-like interneurons and whole nerve recordings from buccal ganglion motor nerve roots that normally innervate the ingestive feeding muscles. The buccal ganglion motor nerve root discharge pattern that causes rhythmic feeding movements, termed the feeding motor program (FMP), was elicited either by attractive taste solutions applied to the lip chemoreceptors or by ACh applied to the cerebral ganglia. The ability of exogenous ACh applied to the cerebral ganglia to trigger FMP was blocked by the cholinergic antagonists curare and atropine. If the strength of the lip-applied taste stimulus was in the range of 1-2 times threshold, cerebral application of the cholinergic antagonists blocked or greatly decreased the ability of lip-applied taste solutions to trigger FMP (5 of 8 trials). The cerebral feeding interneurons, some of which activate FMP when stimulated intracellularly, are excited by small pulses of ACh applied directly to the cell body from an ACh-filled micropipette. A pulse of ACh that activates several of the feeding interneurons simultaneously triggers FMP. The data suggest that under certain stimulus conditions an obligatory set of cholinergic synapses onto the feedininterneurons must be activated for taste inputs to trigger ingestion. The determination of ACh's action within the feeding control system is necessary for understanding how enhanced cholinergic transmission leads to prolonged associative memory retention (Sahley, et al., 1986).     

Interaction between the Cholinergic and Purinergic Control of Transmitter Release in the Neuromuscular Junction

The interaction between the cholinergic and purinergic receptors in the frog neuromuscular junction was studied using a standard microelectrode technique. The inhibitory action of an acetylcholine analog, carbachol, on transmitter release virtually disappeared when the releasing machinery was initially blocked by adenosine, indicating the existence of a functional cross-talk between the purinergic and cholinergic receptors.     

Cholinergic regulation of the corpora allata in adult male loreyi leafworm Mythimna loreyi

The direct effect of acetylcholine on the activation of the corpora allata (CA) was investigated in the adult male loreyi leafworm, Mythimna loreyi. Acetylcholine, in the presence of the choline esterase inhibitor physostigmine (50 microM), elicited a stimulatory effect on juvenile hormone acids (JHAs) release from the CA. Maximum effect was obtained at concentrations of 10 and 50 microM. Repeated administration of 10 microM acetylcholine on the same CA did not elicit similar stimulatory effect. Since JHA release can be significantly activated by carbachol and not by nicotine, this cholinergic effect is likely to belong to the muscarinic type. The effect of acetylcholine was significantly antagonized by gallamine triethiodide (M(2) antagonist) and 4-DAMP (M(3) antagonist), pirenzepine (M(1) antagonist), and tropicamide (M(4) antagonist) were ineffective. It is concluded that in the adult male M. loreyi, the cholinergic regulation of CA is most likely via M(2) and M(3) muscarinic receptors.     

Detection of basal and potassium-evoked acetylcholine release from embryonic DRG explants

Spontaneous and potassium-induced acetylcholine release from embryonic (E12 and E18) chick dorsal root ganglia explants at 3 and 7 days in culture was investigated using a chemiluminescent procedure. A basal release ranging from 2.4 to 13.8 pm/ganglion/5 min was detected. Potassium application always induced a significant increase over the basal release. The acetylcholine levels measured in E12 explants were 6.3 and 38.4 pm/ganglion/5 min at 3 and 7 days in culture, respectively, while in E18 explant cultures they were 10.7 and 15.5 pm/ganglion/5 min. In experiments performed in the absence of extracellular Ca2+ ions, acetylcholine release, both basal and potassium-induced, was abolished and it was reduced by cholinergic antagonists. A morphometric analysis of explant fibre length suggested that acetylcholine release was directly correlated to neurite extension. Moreover, treatment of E12 dorsal root ganglion-dissociated cell cultures with carbachol as cholinergic receptor agonist was shown to induce a higher neurite outgrowth compared with untreated cultures. The concomitant treatment with carbachol and the antagonists at muscarinic receptors atropine and at nicotinic receptors mecamylamine counteracted the increase in fibre outgrowth. Although the present data have not established whether acetylcholine is released by neurones or glial cells, these observations provide the first evidence of a regulated release of acetylcholine in dorsal root ganglia.     

Differential regulation of somatodendritic and nerve terminal dopamine release by serotonergic innervation of substantia nigra

Nigrostriatal dopaminergic neurons release dopamine from dendrites in substantia nigra and axon terminals in striatum. The cellular mechanisms for somatodendritic and axonal dopamine release are similar, but somatodendritic and nerve terminal dopamine release may not always occur in parallel. The current studies used in vivo microdialysis to simultaneously measure changes in dendritic and nerve terminal dopamine efflux in substantia nigra and ipsilateral striatum respectively, following intranigral application of various drugs by reverse dialysis through the nigral probe. The serotonin releasers (+/-)-fenfluramine (100 micro m) and (+)-fenfluramine (100 micro m) significantly increased dendritic dopamine efflux without affecting extracellular dopamine in striatum. The non-selective serotonin receptor agonist 1-(m-chlorophenyl)-piperazine (100 micro m) elicited a similar pattern of dopamine release in substantia nigra and striatum. NMDA (33 micro m) produced an increase in nigral dopamine of a similar magnitude to mCPP or either fenfluramine drug. However, NMDA also induced a concurrent increase in striatal dopamine. The D2 agonist quinpirole (100 micro m) had a parallel inhibitory effect on dopamine release from dendritic and terminal sites as well. Taken together, these data suggest that serotonergic afferents to substantia nigra may evoke dendritic dopamine release through a mechanism that is uncoupled from the impulse-dependent control of nerve terminal dopamine release.     

Neurochemical evidence for denervation supersensitivity: the effect of unilateral substantia nigra lesions on apomorphine-induced increases in neostriatal acetylcholine levels

Unilateral lesions of the nigro-neostriatal dopaminergic projection were induced by injections of 6-hydroxydopamine into the zona compacta of the substantia nigra. This resulted in a reduction of neostriatal dopamine to less than 6 percent of the control side. Two months later intraperitoneal injections of apomorphine (1 mg/kg) produced contralateral turning and a significant increase in neostriatal acetylcholine levels. The increase was significantly greater in the neostriatum ipsilateral to the lesion than in the intact side. Haloperidol (1 mg/kg) produced a significant decrease in neostriatal acetylcholine but this decrease did not differ between the “denervated” and intact neostriata. The nigral 6-hydroxydopamine lesions did not by themselves affect neostriatal acetylcholine levels. The fact that apomorphine produces a greater increase in neostriatal acetylcholine after lesions of the dopaminergic nigro-neostriatal projection supports earlier behavioral data suggestive of denervation supersensitivity of neostriatal dopaminergic receptors after these lesions.     

Direct interaction of LSD with central "beta"-adrenergic receptors.

There is already some evidence for a cholinergic system in the substantia nigra. In this paper we have studied the uptake of ³HCh and the subsequent release of radio label. The uptake is sodium dependent and the potassium evoked release is predominantly ³HACH. This release is strongly calcium dependent. These data support the existence of a cholinergic neuronal system in the substantia nigra.     

Changes in Levels of Dopamine and Tyramine in the Rat Caudate Nucleus Following Alterations of Impulse Flow in the Nigrostriatal Pathway

Following electric stimulation of the substantia nigra for 1 h there was a substantial increase in dopamine (DA) turnover in the rat caudate nucleus evidenced by an increase in its acid metabolite homovanillic acid (HVA). Concurrently there was an increase in striatal m-tyramine (mTA) and a substantial decrease in p-tyramine (pTA). Lesioning the substantia nigra to decrease impulse flow resulted in a buildup of striatal DA and mTA, but again a decrease in pTA. Following pretreatment with a tyrosine hydroxylase inhibitor, the effects of stimulation of the nigra on mTA were reversed, there being a significant decrease in this amine. The decrease of pTA in response was partially prevented by tyrosine hydroxylase inhibition. The effects of stimulation or substantia nigra lesions on pTA levels were reversed, however, by tyrosine hydroxylase inhibition, a significant increase in this amine being recorded. mTA and DA levels were largely unaffected by a combination of lesion and tyrosine hydroxylase inhibition. The results provide insight into the possible biosynthetic interrelationships between DA and the tyramine isomers in the rat caudate nucleus.     

Neurotensin Regulation of Endogenous Acetylcholine Release from Rat Striatal Slices Is Independent of Dopaminergic Tone

The effects of neurotensin (NT) alone or in combination with the dopamine antagonist sulpiride were tested on the release of endogenous acetylcholine (ACh) from striatal slices. NT enhanced potassium (25 mM)-evoked ACh release from striatal slices in a dose-dependent manner. This effect was tetrodotoxin-insensitive, suggesting an action directly on cholinergic elements. The dopamine antagonist sulpiride (5 x 10(-5) M) significantly increased (63%) potassium-evoked ACh release from striatal slices; potassium-evoked ACh release was further increased (90%) in the presence of NT (10(-5) M) and sulpiride (5 x 10(-5) M). The second set of experiments tested the effects of 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra on NT-induced increases of potassium-evoked ACh release. These lesions did not alter the NT regulation of potassium-evoked ACh release from striatal slices, but did significantly increase spontaneous (33%) and potassium-evoked (40%) ACh release from striatal slices. Striatal choline acetyltransferase activity was not affected by 6-OHDA lesions. In addition, following 6-OHDA lesions, sulpiride was ineffective in altering ACh release from striatal slices. Furthermore, evoked ACh release in the presence of the combination of NT and sulpiride was not different from that in the presence of NT alone. These results suggest that in the rat striatum, NT regulates cholinergic interneuron activity by interacting with NT receptors associated with cholinergic elements. Moreover, the NT modulation of cholinergic activity is independent of either an interaction of NT with D2 dopamine receptors or the sustained release of dopamine.