Acetylcholinesterase in central vocal control nuclei of the zebra finch (<Emphasis Type="Italic">Taeniopygia guttata</Emphasis>)

The distribution of acetylcholinesterase (AChE) in the central vocal control nuclei of the zebra finch was studied using enzyme histochemistry. AChE fibres and cells are intensely labelled in the forebrain nucleus area X, strongly labelled in high vocal centre (HVC) perikarya, and moderately to lightly labelled in the somata and neuropil of vocal control nuclei robust nucleus of arcopallium (RA), medial magnocellular nucleus of the anterior nidopallium (MMAN) and lateral magnocellular nucleus of the anterior nidopallium (LMAN). The identified sites of cholinergic and/or cholinoceptive neurons are similar to the cholinergic presence in vocal control regions of other songbirds such as the song sparrow, starling and another genus of the zebra finch (Poephila guttata), and to a certain extent in parallel vocal control regions in vocalizing birds such as the budgerigar. AChE presence in the vocal control system suggests innervation by either afferent projecting cholinergic systems and/or local circuit cholinergic neurons. Co-occurrence with choline acetyltransferase (ChAT) indicates efferent cholinergic projections. The cholinergic presence in parts of the zebra finch vocal control system, such as the area X, that is also intricately wired with parts of the basal ganglia, the descending fibre tracts and brain stem nuclei could underlie this circuitry’s involvement in sensory processing and motor control of song.     

Development of neurons containing acetylcholinesterase and cholinacetyltransferase in dispersed cell culture of rat cerebellum

Summary Cells from one-day-old cerebellum were grown for up to 30 days in dispersed cell culture. The characteristic neurons (deep cerebellar, Golgi and Purkinje cells) maintained their properties. It was found histochemically that some of the large cells display strong AChE activities in the perikaryon and in some processes, while biochemically the specific activities of the marker enzymes of the acetylcholine system, AChE (EC 3.1.1.7) and ChAc (EC 2.3.1.6), were increased and unchanged, respectively. During cultivation, the number of AChE-positive neurons increased. It can be inferred from these studies that, besides the AChE-positive (cholinoceptive) cells, ChAc-active (cholinergic) neurons (possibly Golgi II. type cells and some neurons in the deep cerebellar nuclei) are present in the cerebellum of the rat.     

Development of neurons containing acetylcholinesterase and cholinacetyltransferase in dispersed cell culture of rat cerebellum.

Cells from one-day-old cerebellum were grown for up to 30 days in dispersed cell culture. The characteristic neurons (deep cerebellar, Golgi and Purkinje cells) maintained their properties. It was found histochemically that some of the large cells display strong AChE activities in the perikaryon and in some processes, while biochemically the specific activities of the marker enzymes of the acetylcholine system, AChE (EC 3.1.1.7) and ChAc (EC 2.3.1.6), were increased and unchanged, respectively. During cultivation, the number of AChE-positive neurons increased. It can be inferred from these studies that, besides the AChE-positive (cholinoceptive) cells, ChAc-active (cholinergic) neurons (possibly Golgi II. type cells and some neurons in the deep cerebellar nuclei) are present in the cerebellum of the rat.     

Cholinergic neurons in an association cortex slab chronically isolated from the cat

Small numbers of short- and long-axon cholinergic interneurons were revealed on a slab of association cortex three weeks after (neuronal) isolation from the cat by means of a histochemical acetylcholinesterase reaction. Short-axon neurons are located at layers II–VI and take the form of mainly spindle-shaped medium sized cells with their axons forming synaptic terminals on pyramidal and stellate neurons of the isolated section. Typical positioning of cholinergic terminals on the perikaryon and proximal portions of cholinoceptive neuron dendrites was noted. Pyramidal cholinoceptive cells may be classed as noncholinergic cells, whereas stellate cells may be either cholinergic or noncholinergic. Long-axon cholinergic interneurons of different shapes and sizes are situated at layers I and VI. Neuronal axons located in these layers run within fibers of the first and subcortical layers, establishing intracortical connections beyond the confines of the isolated section.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 21, No. 1, pp. 60–66, January–February, 1989.     

Influence of cholinergic and adrenergic agents of the electric activity of the brain and the heart under hypoxia

Injection of adrenergic and cholinergic agents to animals in the normal athmospheric conditions did not tigger drastic changes on the electric activity of the brain and heart. Acutehipoxia demands high adaptability from the body. In such conditions stimulation of reticular formation and hypothalamus produces different changes in the EEG and ECG activity whith injecting adrenergic and cholinergic agents. It was determined that cholinergic influence are effective in the regulation of electrical brain activity while adrenergics are more important for the realization of descending influences of the truncus cerebri vegetative centers and are less active in the modulation of the cerebral cortex activity.     

Early changes in GABA and glutamate levels and aminotransferase activity in parts of the rat brain following whole-body gamma irradiation at an absolutely lethal dose

The contents of gamma-aminobutyric acid (GABA) and glutamate (GL) as well as GABA-aspartate- and alanine aminotransferase activities were measured in rat cerebellum, cerebral cortex and truncus cerebri 1, 3, 6, 24 and 48 hr following total-body gamma-irradiation (60Co) with a dose of 30 Gy. All the indices under study changed in a similar way in the cortex and truncus cerebri while in the cerebellum, GABA level increased and GABA-alpha-ketoglutarate aminotransfearse activity decreased 60 min after irradiation. The levels of GABA and GL in the cortex and truncus cerebri decreased immediately and increased 24 hr after irradiation. Activity of aminotransferases changed in a phase manner: changes in aspartate- and alanine aminotransferase activity were more pronounced than those of GABA-alpha-ketoglutarate aminotransferase activity and correlated with the glutamate level changes.     

Pontine reticular origin of cholinergic excitatory afferents to the locus coeruleus controlling the gain of vestibulospinal and cervicospinal reflexes in decerebrate cats

1. Previous experiments had shown that the medullary inhibitory reticulospinal (mRS) neurons act 180 degrees out-of-phase with respect to the excitatory vestibulospinal (VS) neurons during the vestibular and the neck reflexes involving the limb extensor motoneurons. This finding suggested that the higher the firing rate of the medullary inhibitory RS neurons in the animal at rest, the greater the disinhibition which affects the limb extensor motoneurons during side-down roll tilt of the animal or side-up neck rotation, thus leading to an increased gain of response of limb extensors to sinusoidal stimulation of labyrinth and neck receptors. The gain of these postural reflexes would then represent a sensitive test to evaluate the background discharge of the inhibitory reticulospinal system of the medulla. 2. The discharge of the inhibitory mRS neurons is under the tonic excitatory control of cholinergic pontine reticular formation (pRF) neurons which are also self-excitatory, while these cholinergic pontine neurons are in turn inhibited by the norepinephrine (NE)-containing locus coeruleus (LC) neurons, which are also self-inhibitory due to mechanisms of recurrent and/or lateral inhibition. The present experiments were performed to find out whether cholinergic and cholinoceptive pontine reticular neurons, which are under the inhibitory control of the LC neurons, also send axons to the LC on which they may exert an excitatory influence. This excitatory effect would then counteract the self-inhibitory influence mediated by the NE, which acts on the alpha 2-adrenoceptors distributed on the somatodendritic membrane of the LC neurons. 3. In precollicular decerebrate cats, local injection into the dorsal aspect of the pontine tegmentum of 0.25 microliter of a solution of the muscarinic blocker atropine sulphate at the concentration of 6 micrograms/microliter of sterile saline did neither modify the postural activity in the ipsilateral limbs nor the response gain of the ipsilateral forelimb extensor triceps brachii to sinusoidal stimulation of labyrinth receptors (roll tilt of the animal at 0.15 Hz, +/- 10 degrees). These negative results were attributed to the fact that in these preparations the activity of the cholinergic and cholinoceptive pRF neurons and the related inhibitory mRS neurons is very low, due to the tonic discharge of the NE-containing LC neurons, which exert a prominent inhibitory influence on the underlying reticular structures.(ABSTRACT TRUNCATED AT 400 WORDS)     

METABOLISM OF PUTATIVE TRANSMITTERS IN INDIVIDUAL NEURONS OF APLYSIA CALIFORNICA

The activities of acetylcholinesterase (acetylcholine acetyl-hydrolase; EC 3.1.1.7) and catechol-O-methyl transferase (S-adenosylmethionine: catechol-O-methyl transferase; EC 2.1.1 .a) were measured in the various ganglia of the nervous system of Aplysia californica and in some of the individually identifiable neurons in these ganglia. All of the neurons studied had measurable levels of activity for both enzymes. Since different individual neurons exhibited approximately the same level of activity we concluded that neither of these enzymes could be used to classify neurons as ‘cholinergic’ vs. ‘aminergic’ or ‘cholinoceptive’ vs. ‘aminoceptive'. The ubiquitous distribution of either or both of these enzymes in different single neurons may be related to glial contamination.     

Topography of cholinergic neurons in the brain stem of the human fetus

Cholinacetyltransferase (ChAT) activity has been studied in 56 nuclei of the cerebral trunk in human fetuses at the age of 6-8 lunar months. Cytoplasmic and synaptic ChAT activity has been revealed and three types of neurons for cholinergic synaptic transmission has been distinguished. There are only cholinergic-noncholinoceptive neurons in five macrocellular nuclei of the cranial nerves. In 25 nuclei (paravicellular, reticular, pigmented, sensitive nuclei of the cranial nerves, nuclei of the funiculi posterior and some other switching centres) there are only noncholinergic-cholinoceptive neural cells. In 16 nuclei there are three, and in 8 nuclei--two types of cells. Either noncholinergic-cholinoceptive or cholinergic-noncholinoceptive cells predominate; there is no predominance of cholinergic-cholinoceptive neurons in any of the nuclei. Mapping on the position of the cholinergic synaptic transmission neurons in the cerebral trunk is composed.     

Neurohistological and histochemical characteristics of Loeschcke's zone of the cat medulla

Neuronal organization and transmitter profile of the chemosensitive area on the ventral medulla surface (zone L) were studied in the cat using neurohistological and histochemical techniques. Neurons of different shapes and sizes distributed through this area are concentrated mainly at the level of the medial hypoglossal nerve root, with their numbers gradually decreasing rostrally and caudally from this nerve. Only an insignificant proportion of zone L neurons are cholinergic and monoaminergic, while the remainder probably employ other biologically active substances as transmitters. Cholinergic and noncholinergic cholinoceptive neurons were found in zone L, as well as neurons sensitive to catecholamines and serotonin. It is postulated that the catecholaminergic, serotoninergic, and cholinergic fibers observed in zone L may be of local origin or may originate from other structures.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 18, No. 2, pp. 219–226, March–April, 1986.     

Cholinergic Control of Nerve Growth Factor in Adult Rats: Evidence from Cortical Cholinergic Deafferentation and Chronic Drug Treatment

Abstract: It is well documented that nerve growth factor (NGF) plays an important role in maintaining functions of cholinergic basal forebrain neurons. In the present study, we tested the hypothesis that cholinergic activity controls NGF levels in cholinoceptive neurons of the cerebral cortex and hippocampus. To address that question, we used both cholinergic deafferentation of cerebral cortex and hippocampus by cholinergic immunolesion with 192IgG-saporin and chronic pharmacological treatment of sham-treated and immunolesioned rats with the cholinergic agonist pilocarpine and the cholinergic antagonist scopolamine. We observed an increase in NGF protein levels in the cortex and hippocampus after cholinergic immunolesions and also after muscarinic receptor blockade by chronic intracerebroventricular scopolamine infusion in sham-treated rats after 2 weeks. There was no further increase in the accumulation of NGF after scopolamine treatment of immunolesioned rats. Chronic infusion of pilocarpine had no effect on cortical and hippocampal NGF protein levels in sham-treated rats. In rats with cholinergic immunolesions, however, pilocarpine did prevent the lesion-induced accumulation of NGF. There was no effect of cholinergic lesion and drug treatment on cortical or hippocampal NGF mRNA levels, consistent with the importance of NGF retrograde transport as opposed to its de novo synthesis. This study provides strong evidence for the hypothesis that there is cholinergic control of cortical and hippocampal NGF protein but not mRNA levels in adult rats.     

Cholinergic microstimulation of the peribrachial nucleus in the cat. I. Immediate and prolonged increases in ponto-geniculo-occipital waves.

The cholinergic agonist carbachol was injected into the pontine Pb area where PGO bursting cells have been recorded. When microinjections were localized to the ventrolateral aspect of the caudal Pb nucleus near aggregates of ChAT immunolabeled cholinergic neurons, carbachol produced an immediate onset of state-independent PGO waves in the ipsilateral LGB. These state-independent PGO waves persisted for 3-4 days. After the first 24 hrs PGO wave activity increasingly became associated with REM sleep and with REM transitional SP sleep as both of these PGO-related states increased in amount to 3-4 times baseline levels. The increase in amount of PGO-related states peaked on days 2-4 following one carbachol injection and persisted for 10-12 days. These results suggest a two stage process: stage one, PGO enhancement, is the direct consequence of the membrane activation of cholinoceptive PGO burst neurons by carbachol; stage two, REM enhancement, is the consequence of metabolic activation of endogenous cholinergic neurons. This experimental preparation is a useful model for the study of the electrophysiology and functional significance of PGO wave and REM sleep generation.     

THE DISTRIBUTION OF MUSCARINIC RECEPTOR SITES IN THE NERVOUS SYSTEM OF THE DOG

Abstract— The concentration of muscarinic receptors has been measured in 22 areas of the dog nervous system by measuring the atropine-sensitive uptake of tritium-labelled propylbenzilylcholine mustard. The highest concentration of receptor was found in the caudate nucleus, intermediate concentrations were found in five areas of cerebral cortex, the other basal ganglia and the superior colliculus. Significant concentrations were found in the corpus callosum and subcortical white matter, and are believed to be on axons derived from cholinoceptive neurons. The results are discussed in relation to other evidence concerning cholinergic transmission in the nervous system.     

Fusion of diphtheria toxin and urotensin II produces a neurotoxin selective for cholinergic neurons in the rat mesopontine tegmentum

Urotensin II is a neuropeptide first isolated from fish and later found in mammals: where it has potent cardiovascular, endocrine and behavioral effects. In rat brain the urotensin II receptor (UII-R) is predominately expressed in the cholinergic neurons of the pedunculopontine (PPTg) and laterodorsal tegmental nuclei. Typically, the function of the PPTg has been examined using excitotoxins, destroying both cholinergic and non-cholinergic neurons, which confounds interpretation. We took advantage of UII-R's unique expression profile, by combining UII with diphtheria toxin, to engineer a toxin specific for cholinergic neurons of the PPTg. In vitro, two different toxin constructs were shown to selectively activate UII-R (average EC50 approximately 30 nmol/L; calcium mobility assay) and to be 10,000-fold more toxic to UII-R expressing CHO cells, than wildtype cells (average LD50 approximately 2 nmol/L; cell viability). In vivo, pressure injection into the PPTg of rats, resulted in specific loss of choline transporter and NADPH diaphorase positive neurons known to express the UII-R. The lesions developed over time, resulting in the loss of over 80% of cholinergic neurons at 21 days, with little damage to surrounding neurons. This is the first highly selective molecular tool for the depletion of mesopontine cholinergic neurons. The toxin will help to functionally dissect the pedunculopontine and laterodorsal tegmental nuclei, and advance the understanding of the functions of these structures.     

Demonstration of muscarinic acetylcholine receptor-like immunoreactivity in the rat forebrain and upper brainstem

Summary The distribution of muscarinic acetylcholine receptor protein (mAChR) in the rat forebrain and upper brainstem was described by using a monoclonal antibody (M35) raised against mAChR purified from bovine forebrain homogenates. A method is investigated for light microscopic (LM) and electronmicroscopic (EM) immunocytochemical visualization of reactivity to mAChR-proteins. Putative cholinoceptive neurons including their dendrites were found immunoreactive in the cortical mantle, hippocampus, basal ganglia, amygdala, thalamus and several midbrain regions. In the neocortex, immunoprecipitate with M35 was mainly present in layer 5 pyramidal cells, some layer 3 pyramidal neurons and layer 2 stellate cells, all including their characteristic dendritic profiles of both basal and apical dendrites. In the hippocampus, a variety of pyramidal, granular and non-pyramidal celltypes were stained in various hippocampal cell layers, in the dentate hilus and in stratum oriens of cornu ammonis. Moreover, positively reacting cells occurred in central and lateral amygdala, all parts of the basal ganglia and ventral pallidum. The thalamus was very richly provided with labeled neurons in several nuclei but notably numerous in the ventrolateral, anteroventral and geniculate nuclei. In cortex and hippocampus also some staining of astrocytes occurred. Electron microscopic study of the intracellular distribution of M35 immunoreactivity in all cases showed dense precipitates in the soma cytoplasm in close association with the golgi apparatus, but conspicuous absence near the endoplasmic reticulum. Immunoprecipitate can be followed within the dendritic tree along the microtubular transport system, up to proximal and distal postsynaptic membrane positions, apposing non labeled presynaptic endings. Muscarinic receptor subtype recognition by M35 will be discussed by comparing M35 distribution with cholinergic innervation patterns, muscarinic receptor ligand binding studies and localization of muscarinic receptor subtype mRNAs.     

Demonstration of muscarinic acetylcholine receptor-like immunoreactivity in the rat forebrain and upper brainstem

The distribution of muscarinic acetylcholine receptor protein (mAChR) in the rat forebrain and upper brainstem was described by using a monoclonal antibody (M35) raised against mAChR purified from bovine forebrain homogenates. A method is investigated for light microscopic (LM) and electronmicroscopic (EM) immunocytochemical visualization of reactivity to mAChR-proteins. Putative cholinoceptive neurons including their dendrites were found immunoreactive in the cortical mantle, hippocampus, basal ganglia, amygdala, thalamus and several midbrain regions. In the neocortex, immunoprecipitate with M35 was mainly present in layer 5 pyramidal cells, some layer 3 pyramidal neurons and layer 2 stellate cells, all including their characteristic dendritic profiles of both basal and apical dendrites. In the hippocampus, a variety of pyramidal, granular and non-pyramidal celltypes were stained in various hippocampal cell layers, in the dentate hilus and in stratum oriens of cornu ammonis. Moreover, positively reacting cells occurred in central and lateral amygdala, all parts of the basal ganglia and ventral pallidum. The thalamus was very richly provided with labeled neurons in several nuclei but notably numerous in the ventrolateral, anteroventral and geniculate nuclei. In cortex and hippocampus also some staining of astrocytes occurred. Electron microscopic study of the intracellular distribution of M35 immunoreactivity in all cases showed dense precipitates in the soma cytoplasm in close association with the golgi apparatus, but conspicuous absence near the endoplasmic reticulum. Immunoprecipitate can be followed within the dendritic tree along the microtubular transport system, up to proximal and distal postsynaptic membrane positions, apposing non labeled presynaptic endings. Muscarinic receptor subtype recognition by M35 will be discussed by comparing M35 distribution with cholinergic innervation patterns, muscarinic receptor ligand binding studies and localization of muscarinic receptor subtype mRNAs.     

Distribution of cholineacetyltransferase histochemistry in isthmus and medulla of Onchorynchus masu. Tract-tracing observation on the ascending meso-pontine cholinergic system

The distribution of cholinergic neurons was studies in the brain steam, medulla and rostral spinal cord of the salmon Onchorynchus masu using histochemical choline acetyltransferase (ChAT) detection. Cholinergic neurons were observed in the isthmus, cranial nerve motor nuclei and spinal cord. In order to characterize several cholinergic nuclei observed in the isthmus of O. masu, their projections were studied by application of 1,1'-dioctadecyl-3,3,3',3,'-tetramethylindocarbocyanine perchlorate (DiI) to selected structures of the brain. The secondary gustatory nucleus projected mainly to the lateral hypothalamic lobes, whereas the nucleus isthmi projected to the optic tectum and parvocellular superficial pretectal nucleus, as it was earlier described for the other teleost group. In addition, the other isthmic cholinergic nuclei in O. masu may be homologous to the meso-pontine system of mammals. We conclude that the cholinergic systems of teleosts show many primitive features that have been presented during evolution, together with exclusive to the group characteristics.     

Anatomical funneling, sparse connectivity and redundancy reduction in the neural networks of the basal ganglia

The major anatomical characteristics of the main axis of the basal ganglia are: (1) Numerical reduction in the number of neurons across layers of the feed-forward network, (2) lateral inhibitory connections within the layers, and (3) neuro-modulatory effects of dopamine and acetylcholine, both on the basal ganglia neurons and on the efficacy of information transmission along the basal ganglia axis. We recorded the simultaneous activity of neurons in the output stages of the basal ganglia as well as the activity of dopaminergic and cholinergic neurons during the performance of a probability decision-making task. We found that the functional messages of the cholinergic and dopaminergic neurons differ, and that the cholinergic message is less specific than that of the dopaminergic neurons. The output stage of the basal ganglia showed uncorrelated neuronal activity. We conclude that despite the huge numerical reduction from the cortex to the output nuclei of the basal ganglia, the activity of these nuclei represents an optimally compressed (uncorrelated) version of distinctive features of cortical information.     

Alpha-adrenergic receptor (α2A ) is colocalized in basal forebrain cholinergic neurons: A light and electron microscopic double immunolabeling study

A variety of data suggest that noradrenaline and acetylcholine may interact in the basal forebrain, however no morphological studies have addressed whether indeed cholinergic neurons express adrenergic receptors. We have investigated the presence of alpha-adrenergic receptor subtype α_2A -AR in cholinergic neurons of the basal forebrain. Cholinergic neurons were identified with an antibody against choline acetyltransferase and the receptor with a polyclonal antibody raised against a 47 amino acid fragment of the third intracellular loop of the α_2A -AR. For double labeling at the light microscopic level the Ni-DAB/DAB technique was used, and for electron microscopy an immunoperoxidase/immunogold method was applied. We detected the α_2A -AR protein in cholinergic as well as in non-cholinergic neurons. Almost half of all cholinergic neurons contained this adrenergic receptor. Double-labeled neurons were distributed throughout the rostro-caudal extent of the basal forebrain cholinergic continuum, including the medial septum, vertical and horizontal diagonal band nuclei, pallidal regions, substantia innominata and the internal capsule. Non-cholinergic neurons that expressed the α_2A -AR outnumbered cholinergic/α_2A -AR neurons by several factors. Electron microscopy confirmed the presence of α_2A -AR in cholinergic neurons in the medial septum, vertical and horizontal diagonal band nuclei. Gold particles (10 nm) indicative of α_2A -AR were diffusely distributed in the cytoplasm and accumulated in cytoplasmic areas near the Golgi complex and cysterns of the endoplasmic reticulum and were associated with the cellular membranes at synaptic and non-synaptic locations. Since many of the α_2A -AR+/non-cholinergic neurons we detected are likely to be GABAergic cells, our data support the hypothesis that noradrenaline may act via basal forebrain cholinergic and non-cholinergic neurons to influence cortical activity.