Transplantation of embryonic ventral forebrain neurons to the neocortex of rats with lesions of nucleus basalis magnocellularis--I. Biochemical and anatomical observations

Unilateral ibotenic acid lesions of the rat nucleus basalis magnocellularis produce approximately 60% depletion of choline acetyltransferase activity in ipsilateral frontal and frontoparietal neocortex. This depletion, which represents the loss of most of the extrinsic neocortical cholinergic input, is stable for at least 6 months. Embryonic ventral forebrain neurons survive transplantation to such cholinergically denervated neocortex. Cholinergic cells abound within these transplants and appear able to reinnervate the cholinergically depleted host cortex, as assessed histochemically and by measurement of choline acetyltransferase activity. Outgrowing fibres may extend beyond 2 mm from the grafts and often appear to be organized in an appropriate laminar pattern within the host cortex. Peptidergic neurons are sparse within the grafts and their fibres frequently appear unable to grow into the host tissue. Control grafts of non-cholinergic embryonic hippocampal cells survive well but have no effect on cortical depletions of acetylcholinesterase or choline acetyltransferase activity. Reconstruction of the extrinsic cholinergic input to the cortex by transplantation provides a useful tool for understanding the functions of this pathway.     

Regional decreases of cortical choline acetyltransferase after lesions of the septal area and in the area of nucleus basalis magnocellularis

Choline acetyltransferase and [³H]choline uptake have been measured in neocortical regions and hippocampus one week after lesions which destroyed the septum bilaterally, and after unilateral lesions in the area of nucleus basalis magnocellularis. Lesions of the septal area, which severely decreased choline acetyltransferase in hippocampus, only moderately decreased choline acetyltransferase in a posterior cortical region and had no effect in frontal and parietal regions. In contrast, lesions which included nucleus basalis magnocellularis decreased choline acetyltransferase markedly in frontal and parietal regions and had less of an effect in the posterior cortical regions. Lesion-induced decreases of [³H]choline uptake paralleled those of choline acetyltransferase. Lesions which included nucleus basalis magnocellularis had no effect on choline acetyltransferase in hippocampus, nucleus accumbens, olfactory tubercle, midbrain or pons-medulla.These results suggest the existence of topographically distinct cholinergic inputs to neocortex. In agreement with previous studies, cholinergic projections from the peripallidal region of nucleus basalis magnocellularis are predominantly to frontal and parietal neocortex. In contrast to previous suggestions, cholinergic projections to neocortex from the septal area are limited to the posterior regions of neocortex.     

Basal forebrain neurons provide major cholinergic innervation of primate neocortex

In 3 monkeys, lesions were made in the basal forebrain by microinjections of ibotenic acid into the nucleus basalis. Bilateral samples of multiple neocortical gyri were assayed for the activity of choline acetyltransferase. Compared to control hemispheres, enzyme activity was reduced up to 69% in the neocortex ipsilateral to the lesion; in addition, acetylcholinesterase staining was decreased at the lesioned site and in the ipsilateral cortex. These results support the concept that the principal cholinergic innervation of the primate neocortex is derived from axons and nerve terminals of neurons whose perikarya are located in the basal forebrain, particularly the nucleus basalis.     

Neocortical cholinergic innervation: A description of extrinsic and intrinsic components in the rat

Electrothermic lesion of the peri-pallidal region of the rat caused a marked reduction in the activity of choline acetyltransferase in the ipsilateral fronto-parietal cortex without affecting the activity of glutamate decarboxylase. Only lesions that involved the ventral globus pallidus significantly reduced cortical choline acetyltransferase activity; and lesions limited to the thalamus, internal capsule, pyriform cortex or zone incerta were ineffective. Excito-toxin lesions of the ventral globus pallidus caused 45-5% reductions in all presynaptic markers for cholinergic neurons but did not significantly decrease presynaptic markers for noradrenergic, serotonergic or histaminergic neurons in the cortex. The maximal reductions in cortical choline acetyltransferase activity achieved with the pallidal lesion was 70%; and enzyme activity reached its nadir by four days after placement of the lesion. The pallidal lesion, which ablated the large isodendritic acetylcholinesterase positive neuronal perikarya, resulted in a profound loss in histochemically stained acetylcholinesterase-reactive fibers in the fronto-parietal cortex but not in the cingulate, pyriform and occipital cortex or hippocampal formation; analysis of the subregions in choline acetyltransferase activity. The kainate lesion of the parietal cortex to ablate intrinsic neurons did not reduce the activity of tyrosine hydroxylase, a marker for noradrenergic terminals, but depressed glutamate decarboxylase by 68%; in contrast choline acetyltransferase activity fell only 29%. The results indicate that approximately 70% of the cholinergic innervation in the fronto-parietal cortex is derived from acetylcholinesterase positive neurons in the peripallidal nucleus basalis, whereas the remainder appears to be localized in cortical intrinsic neurons.     

Atlas of cholinergic neurons in the forebrain and upper brainstem of the macaque based on monoclonal choline acetyltransferase immunohistochemistry and acetylcholinesterase histochemistry

Choline acetyltransferase immunohistochemistry was used to map the cholinergic cell bodies in the forebrain and upper brainstem of the macaque brain. Neurons with choline acetyltransferase-like immunoreactivity were seen in the striatal complex, in the septal area, in the diagonal band region, in the substantia innominata, in the medial habenula, in the pontomecencephalic tegmentum and in the oculomotor and trochlear nuclei. The ventral striatum contained a higher density of cholinergic cell bodies than the dorsal striatum. All of the structures that contained the choline acetyltransferase positive neurons also had acetylcholinesterase-rich neurons. Choline acetyltransferase positive neurons were not encountered in the cortex. Some perikarya in the midline, intralaminar, reticular and limbic thalamic nuclei as well as in the hypothalamus were rich in acetylcholinesterase but did not give a positive choline acetyltransferase reaction. A similar dissociation was observed in the substantia nigra, the raphe nuclei and the nucleus locus coeruleus where acetylcholinesterase-rich neurons appeared to lack perikaryal choline acetyltransferase activity.     

Serotonergic input to cholinergic neurons in the substantia innominata and nucleus basalis magnocellularis in the rat.

The aim of the present study was to determine, at the light microscopic level, whether the serotonergic fibers originating from the dorsal raphe nucleus (B7), median raphe nucleus (B8) and ventral tegmentum (B9) make putative synaptic contacts with cholinergic neurons of the nucleus basalis magnocellularis and substantia innominata. For this purpose, we utilized: (i) the anterograde transport of Phaseolus vulgaris leucoagglutinin combined with choline acetyltransferase immunohistochemistry; (ii) choline acetyltransferase/tryptophan hydroxylase double immunohistochemistry; and (iii) the FluoroGold retrograde tracer technique combined with tryptophan hydroxylase immunohistochemistry. Following iontophoretic injections of Phaseolus vulgaris leucoagglutinin in the dorsal raphe nucleus, labeling was observed primarily in the ventral aspects of the nucleus basalis magnocellularis and in the intermediate region of the substantia innominata. When Phaseolus vulgaris leucoagglutinin was combined with choline acetyltransferase immunohistochemistry, a close association between the Phaseolus vulgaris leucoagglutinin-positive fibers and cholinergic neurons was observed, even though the majority of the Phaseolus vulgaris leucoagglutinin-immunoreactive terminals seemed to establish contact with non-cholinergic elements. Following Phaseolus vulgaris leucoagglutinin injection in the median raphe nucleus, very few labeled fibers with no evident close contact with nucleus basalis magnocellularis and substantia innominata cholinergic neurons were observed. After tryptophan hydroxylase/choline acetyltransferase double immunohistochemistry, a plexus of serotonergic (tryptophan hydroxylase-positive) fibers in the vicinity of choline acetyltransferase-immunoreactive neurons of the substantia innominata and nucleus basalis magnocellularis was observed, and some serotonergic terminals have been shown to come into very close contact with the cholinergic cells. Most of the tryptophan hydroxylase-immunoreactive terminals seem to establish contacts with non-cholinergic cells. Following FluoroGold injection in the nucleus basalis magnocellularis and substantia innominata, the majority of retrogradely labeled neurons was observed mainly in the ventromedial cell group of the dorsal raphe nucleus. In this area, a minority of the FluoroGold-positive neurons was tryptophan hydroxylase immunoreactive. These findings show that serotonergic terminals, identified in very close association with the cholinergic neurons in the substantia innominata and nucleus basalis magnocellularis, derive primarily from the B7 serotonergic cell group of the dorsal raphe nucleus, and provide the neuroanatomical evidence for a direct functional interaction between these two neurotransmitter systems in the basal forebrain.     

Distribution of catecholaminergic afferent fibres in the rat globus pallidus and their relations with cholinergic neurons

The topographical distribution of catecholaminergic nerve fibres and their anatomical relationship to cholinergic elements in the rat globus pallidus were studied. Peroxidase–antiperoxidase and two-colour immunoperoxidase staining procedures were used to demonstrate tyrosine hydroxylase (TH), dopamine β-hydroxylase (DBH), phenylethanolamine N-methyltransferase (PNMT) and choline acetyltransferase (ChAT) immunoreactivities, combined with acetylcholinesterase (AChE) pharmacohistochemistry. TH immunoreactive nerve fibres were seen to enter the globus pallidus from the medial forebrain bundle. The greatest density of such fibres was found in the ventral region of the globus pallidus, which was also characterized by the greatest density of ChAT immunoreactive neurons. TH immunoreactive nerve fibres showed varicose arborizations and sparse boutons, which were occasionally seen in close opposition to cholinergic structures. In all regions of the globus pallidus, there were also larger, smooth TH immunoreactive nerve fibres of passage to the caudate putamen. A smaller number of DBH immunoreactive nerve fibres and terminal arborizations were found in the substantia innominata, internal capsule and in the globus pallidus bordering these structures. A few PNMT immunoreactive nerve fibres in the substantia innominata and internal capsule did not enter the globus pallidus. Electron microscopy revealed TH immunoreactive synaptic profiles in the ventromedial area of the globus pallidus corresponding to the nucleus basalis magnocellularis of Meynert (nBM). These made mainly symmetrical and only a few asymmetrical synaptic contacts with dendrites containing AChE reaction product. The results indicate that cholinergic structures in the nBM are innervated by dopaminergic fibres and terminals, with only a very small input from noradrenergic fibres.     

Separate neuronal populations of the rat globus pallidus projecting to the subthalamic nucleus, auditory cortex and pedunculopontine tegmental area.

The topographic arrangement of globus pallidus neurons sending axons to the subthalamic nucleus, auditory cortex and pedunculopontine tegmental nucleus was studied in the rat using retrograde fluorescent tracers. Neurons projecting to the subthalamic nucleus were localized in the rostral part of the globus pallidus, while neurons projecting to the auditory cortex and to the pedunculopontine tegmental nucleus were located in the caudal part. The two populations of pallidocortical and pallidotegmental neurons were also distributed in a separate manner within the caudal globus pallidus. The former neurons were large and located more ventromedially, whereas the latter were medium-sized and located more dorsolaterally. Using a retrograde fluorescent tracing technique combined with choline acetyltransferase immunofluorescence histochemistry, it was found that a vast majority of pallidocortical neurons expressed choline acetyltransferase immunoreactivity, and that pallidotegmental neurons rarely exhibited choline acetyltransferase immunoreactivity. A method of retrograde tracing with wheatgerm agglutinin conjugated with horseradish peroxidase associated to immunohistochemistry for glutamate decarboxylase confirmed the GABAergic nature of the pallidotegmental pathway. The present study revealed the independent nature of the globus pallidus neurons projecting to the subthalamic nucleus, auditory cortex and pedunculopontine tegmental nucleus. Within this cellular arrangement, the presence of functionally distinct neuronal populations at the caudal pallidal level was also identified, with large cholinergic cells innervating the neocortex and medium-sized GABAergic cells "feeding" the mesencephalic tegmentum.     

Immunocytochemical localization of cholinergic terminals in the region of the nucleus basalis magnocellularis of the rat: a correlated light and electron microscopic study

The cholinergic circuitry in the nucleus basalis magnocellularis of the rat was investigated in a correlated light and electron microscopic study by using monoclonal antibodies against the acetylcholine-synthesizing enzyme, choline acetyltransferase, following the unlabelled antibody peroxidase-antiperoxidase immunocytochemical procedure. After the immunocytochemical approach, large cholinergic cells and a few immunoreactive fibres exhibiting a varicose appearance, were detected by light microscopy in portions of the nucleus basalis magnocellularis located within the anatomical limits of the globus pallidus, mostly in its ventromedial part. Cholinergic neurons and fibre-like structures were also found within the substantia innominata on the edge of globus pallidus. The same material studied by light microscopy was analysed with the electron microscope. At the ultrastructural level, the immunopositive neurons showed the same cytological characteristics and pattern of synaptic input as cholinergic basal forebrain cells. Additionally, scarce immunoreactive preterminal axons and terminal boutons were detected in the region. The immunoreactive terminals were scattered or formed occasional clusters and appeared as heavily immunostained vesicle-filled boutons making exclusively axodendritic synaptic contacts principally with immunonegative distal dendrites. Both symmetric and asymmetric synaptic contacts established between these structures were detected, although the symmetric contacts were the more numerous. The surface of postsynaptic immunonegative dendrites in asymmetric synaptic contact with immunoreactive terminals was generally covered by terminals that lacked detectable immunoreactivity. In contrast, those in symmetric synaptic contact with labelled terminals showed much sparser input from immunonegative terminals, suggesting that they may belong to interneurons. Very rarely, cholinergic terminals were detected in asymmetric synaptic contact with dendrites which also contained positive immunoreaction product. Asymmetric contacts were frequently characterized by the presence of subjunctional dense bodies. The detection of cholinergic terminals in the region of the nucleus basalis magnocellularis of the rat indicates that this region not only contains cholinergic projecting neurons, but receives a cholinergic input itself. Results of this study provide evidence of the existence of a cholinergic transmission in the basal forebrain of the rat, and also that acetylcholine might play a role in the regulation of the extrinsic cortical cholinergic innervation. The possible sources of this innervation are discussed.     

Choline acetyltransferase-immunoreactive neurons in the rat entopeduncular nucleus.

Using a monoclonal antibody against choline acetyltransferase, neurons of the rat entopenduncular nucleus were found to express choline acetyltransferase immunoreactivity. These cholinergic cells were located mostly in the rostral portion of the entopeduncular nucleus with a marked decrease towards its caudal portion. To identify their target sites, a retrograde fiber tracing technique was combined with immunohistochemistry for choline acetyltransferase. After injection of wheatgerm agglutinin conjugated with horseradish peroxidase into the habenula, some of the entopedunculo-habenular cells were found to be immunoreactive for choline acetyltransferase. The cells in the peripallidal region (the substantia innominata, nucleus basalis magnocellularis and ansa lenticularis) with choline acetyltransferase immunoreactivity did not contain horseradish peroxidase. Following injection of fluorescent tracer into the frontal cerebral cortex, retrogradely labeled cells were observed in the rostral part of the entopedunucular nucleus. A majority of these entopedunculo-cortical cells exhibited choline acetyltransferase immunoreactivity, similar to the cells of the peripallidal region projecting to the neocortex. Employing two different fluorescent tracers, entopedunculo-cortical cells were shown to constitute a distinct cell population from the numerous entopedunculo-habenular cells. The present study demonstrated, in the rat entopeduncular nucleus, the presence of cholinergic neurons that projected to the neocortex and habenula.     

Effects of treatment with microencapsulated monosialoganglioside GM1 on cortical and striatal acetylcholine release in rats with cortical devascularizing lesions

The present study shows a novel administration form of the monoganglioside GM1, which following microencapsulation in human serum albumin was topically applied on cortical regions damaged by devascularization in rats. The effects of microencapsulated GM1 on extracellular levels of acetylcholine, choline and dopamine in the cortex and in the striatum were analyzed using in vivo microdialysis. Cholinergic neurons in the nucleus basalis magnocellularis were studied immunohistochemically using monoclonal antibodies raised against choline acetyltransferase (ChAT). It was found that cortical devascularizing lesions produced a decrease in extracellular levels of cortical acetylcholine and choline, and retrograde morphological changes in cholinergic neurons in the nucleus basalis magnocellularis. GM1 promoted (1) recovery of the retrograde morphological changes produced by the decortication in the nucleus basalis magnocellularis and (2) a parallel increase in cortical acetylcholine release. No changes were observed in the striatum, nor on cortical or striatal dopamine levels simultaneously measured in the same perfusates.     

Learning and memory deficits after lesions of the nucleus basalis magnocellularis: reversal by physostigmine

The role of the cholinergic nucleus basalis magnocellularis in spatial learning and memory was studied in the rat. Animals received bilateral injections of ibotenic acid (5 micrograms/microliters) into the region of the nucleus basalis magnocellularis. Six weeks postoperatively they were deprived of food and trained for 5 weeks in a 16-arm radial maze in which 9 of the arms were baited with food. The nucleus basalis magnocellularis-lesioned animals showed significant deficits in the acquisition of the task. Further analysis of the data indicated that this was due primarily to a deficit in reference (long-term) as opposed to working (short-term) memory. After the 5-week training period the nucleus basalis magnocellularis-lesioned animals received intraperitoneal injections of physostigmine sulphate (0.5 mg/kg) 30 min before each daily trial for 1 week. This treatment resulted in a significant improvement in the performance of the spatial memory task on all three measures. The ibotenate lesions reduced the activity of choline acetyltransferase by about 40% in the anterior cortex and by 15% in the posterior cortex. Hippocampal choline acetyltransferase activity was not affected, indicating that the septohippocampal cholinergic projection was spared by the lesions. The activity of glutamate decarboxylase was not affected in any of these regions. These results suggest that the cholinergic projections of the nucleus basalis magnocellularis play an important role in the acquisition of a spatial memory task.     

Delayed treatment with nerve growth factor improves acquisition of a spatial task in rats with lesions of the nucleus basalis magnocellularis: evaluation of the involvement of different neurotransmitter systems.

Rats received bilateral lesions of the nucleus basalis magnocellularis by infusion of ibotenic acid. Fourteen days later, osmotic minipumps releasing human recombinant nerve growth factor (0.3 micrograms/day) were implanted subcutaneously. Starting one month after the lesion, spatial learning of the animals was tested using the Morris water maze. Acquisition of the task was impaired by the lesion, but treatment with nerve growth factor reduced the average latency to find the platform by approximately 9 s, which represents 28% of the lesion-induced behavioral deficit. Retention of this task and spatial acuity, tested in a trial in which the platform was not present, did not show a statistically significant improvement. Lesions of the nucleus basalis magnocellularis reduced the choline acetyltransferase activity in the neocortex, but not in the hippocampus. Treatment with nerve growth factor increased the choline acetyltransferase activity in the neocortex but not in the hippocampus. There was no significant difference in the levels of norepinephrine, dopamine, serotonin or their metabolites in the cortex or hippocampus between nerve growth factor-treated animals and lesioned control animals. There was no significant correlation between any of these neurochemical changes and behavioral performance (acquisition and spatial acuity). Treatment with nerve growth factor did not increase the number or the size of nerve growth factor receptor-immunoreactive neurons in the nucleus basalis magnocellularis. These data suggest that delayed treatment with nerve growth factor results in an improvement of spatial learning in rats with lesions of the nucleus basalis magnocellularis. A possible role for cholinergic mechanisms in this effect is discussed.     

Nucleus basalis lesions in neonate rats induce a selective cortical cholinergic hypofunction and cognitive deficits during adulthood

Summary Ibotenic acid was infused into the nucleus basalis magnocellularis (nBM) of 2-day old rats to eliminate immature cholinergic neurons before they develop functional synaptic connections in the neocortex. For bilaterally lesioned neonates, cognitive testing was initiated 2 months after lesioning and animals were sacrificed at 8 or 12 months of age. Lesioned animals exhibited a marked deficit in the retention of passive avoidance behavior, as well as in the acquisition of 2-way active avoidance behavior. Lesioned animals also made significantly more alternation errors than control animals in the Lashley III spatial maze and showed severe impairments in general learning, reference memory and working memory during 17-arm radial maze testing. For all 4 tasks, neonatally lesioned animals did not show any recovery to the performance level of control animals. Histological analysis of the subcortex from lesioned animals during adulthood revealed: (1) a substantial reduction in acetylcholinesterase-positive cells (presumably cholinergic) within the nucleus basalis, (2) decreased acetylcholinesterase staining in neocortex and (3) a gliosis essentially restricted to the globus pallidus. Surrounding brain regions were apparently not damaged as a direct result of excitotoxin infusion. Neurochemically, neonate nBM lesioning produced a long term cholinergic hypofunction as evidenced by significant reductions of 25% and 18% in frontal cortex chorine acetyltransferase (CAT) activity at 12 and 8 months of age, respectively. By contrast, prefrontal cortical concentrations of biogenic amines and their metabolites were unaffected, thus indicating a degree of neurochemical specificity for these neonatal nBM lesions. The persistant cortical cholinergic hypofunction in lesioned animals may be related to the long term deficits in learning/memory abilities that were also observed. It is suggested that neonatal nBM lesioning could provide a useful animal model for elucidating the plasticity of the developing brain after cortical anervation.     

Rivastigmine antagonizes deficits in prepulse inhibition induced by selective immunolesioning of cholinergic neurons in nucleus basalis magnocellularis

Impairments of cortical cholinergic inputs from the nucleus basalis magnocellularis fundamentally alter information processing and attentional function, thereby advancing the severity of psychopathology in major neuropsychiatric disorders. It was previously shown in adult rats that bilateral 192 IgG saporin-induced selective immunolesioning of the cholinergic neurons in the nucleus basalis produces pronounced and long-lasting deficits in sensorimotor gating measured by prepulse inhibition of the startle reflex. This behavioral paradigm is considered a valid model of sensorimotor gating deficits in the psychotic spectrum and efforts to analyze the significance of the cholinergic basal forebrain in this context are of great interest. In the present study the predictive value of the selective cholinergic immunolesioning model was tested by examining the ability of the cholinesterase inhibitor rivastigmine to restore prepulse inhibition in immunolesioned rats. We report here a pronounced restoring effect of acute (0.75 or 1.5 mg/kg s.c.) as well as repeated (0.75 mg/kg s.c. b.i.d., for 10 days) treatment with rivastigmine in this model of disrupted prepulse inhibition. Intra-nucleus basalis magnocellularis infusions of 192 IgG saporin resulted in extensive loss of basal-cortical cholinergic neurons as shown by the marked decrease in basal telencephalic choline acetyltransferase immunopositive neurons and cortical choline acetyltransferase activity. In this condition, rivastigmine was found to significantly increase cortical acetylcholine extracellular levels in lesioned animals measured by in vivo microdialysis. Taken together, our results strengthen the proposal that the nucleus basalis represents a critical station of the startle gating circuitry. In addition, our findings strongly indicate that even after dramatic decrease of cholinergic neurons, inhibition of acetylcholinesterase restores the cholinergic synaptic function to a point approaching normalization of experimentally induced psychopathology.     

Distribution of enkephalin-immunoreactive nerve fibres and terminals in the region of the nucleus basalis magnocellularis of the rat: a light and electron microscopic study

Summary This investigation was carried out on the distribution of enkephalin-containing nerve fibres and terminals in the region of the nucleus basalis magnocellularis (NBM) of the rat. At the light microscope (LM) level, enkephalin-immunoreactive sites and endogenous choline acetyltransferase (ChAT) were demonstrated by employing the two-colour immunoperoxidase staining technique, using highly specific monoclonal antibodies against enkephalin and ChAT. A pharmacohistochemical procedure to reveal acetylcholinesterase (AChE)-synthesizing neurons combined with the peroxidase-antiperoxidase (PAP) immunocytochemical technique to detect endogenous enkephalins, provided ultrastructural data on the relationships of neuronal elements containing AChE and enkephalins in the region of the NBM.At the LM level, cholinergic neurons of the NBM were surrounded by a dense network of enkephalin-immunoreactive nerve fibres. Electron microscopic (EM) observations of histochemically characterized structures, that were first identified in the LM, revealed that intensely AChE-stained structures in the region of the NBM received sparse synaptic inputs from enkephalin immunoreactive terminals. Synaptic inputs of immunoreactive terminals onto intensely AChE-stained neuron cell bodies were not detected. Synaptic contacts onto proximal AChE-positive dendrites were sparse, but the density increased on more distal regions of the dendrites. All immunoreactive boutons studied established symmetrical synaptic contacts with AChE-positive post-synaptic structures. The pattern of the synaptic input to these cells differs strikingly from that onto typical globus pallidus neurons. The perikarya and dendrites of the latter neurons were characteristically ensheathed in immunoreactive synaptic boutons.Results are consistent with the view that enkephalin-like substances in the rat might be synaptic transmitters or neuromodulators in the area of the NBM and that cholinergic neurons of the NBM (Ch4) are integrated into the circuitry of the basal ganglia. Enkephalins may play an important role regulating the extrinsic cholinergic innervation of the neocortex.     

Cholinergic neurons expressing neuromedin K receptor (NK3) in the basal forebrain of the rat: a double immunofluorescence study

By using a double immunofluorescence method we have examined the distribution of cholinergic neurons expressing neuromedin K receptor (NK3) in the rat brain and spinal cord. The distribution of neuromedin K receptor-like immunoreactive neurons completely overlapped with that of choline acetyltransferase-positive neurons in certain regions of the basal forebrain, e.g. the medial septal nucleus, nucleus of the diagonal band of Broca, magnocellular preoptic nucleus and substantia innominata. Partially overlapping distributions of neuromedin K receptor-like immunoreactive and choline acetyltransferase-positive neurons were found in the basal nucleus of Meynert, globus pallidus, ventral pallidum of the forebrain, tegmental nuclei of the pons and dorsal motor nucleus of the vagus. Neurons showing both neuromedin K receptor-like and choline acetyltransferase immunoreactivities, however, were found predominantly in the medial septal nucleus, nucleus of the diagonal band of Broca and magnocellular preoptic nucleus of the basal forebrain: 66-80% of these choline acetyltransferase-positive neurons displayed neuromedin K receptor-like immunoreactivity. Neurons showing both neuromedin K receptor-like and choline acetyltransferase immunoreactivities were hardly detected in other aforementioned regions of the forebrain, brainstem and spinal cord. The present study has provided morphological evidence for direct physiological modulation or regulation of cholinergic neurons by tachykinins through the neuromedin K receptor in the basal forebrain of rats.     

Comparative effects of quisqualic and ibotenic acid-induced lesions of the substantia innominata and globus pallidus on the acquisition of a conditional visual discrimination: differential effects on cholinergic mechanisms

Two experiments tested the hypothesis that the deficits in conditional discrimination learning produced by ibotenic acid-induced lesions of the ventral pallidum and substantia innominata are produced by loss of the magnocellular cholinergic cells in the nucleus basalis and adjacent regions. Experiment 1 replicated the previously reported deficit in conditional learning produced by ibotenate-induced lesions of the ventral pallidum/substantia innominata, but failed to demonstrate any restoration of learning by a subchronic regimen of the acetylcholinesterase inhibitor physostigmine sufficient to produce significant (30%), but equivalent, degrees of inhibition in the frontal cortex of ventral pallidum/substantia innominata-lesioned or sham-operated rats. Experiment 2 examined the effects of quisqualic acid-induced lesions of the ventral pallidum/substantia innominata. According to most of the measures of learning employed, the quisqualic acid-induced lesion of the ventral pallidum/substantia innominata failed to impair conditional learning, even though the quisqualate-induced lesion produced greater degrees of cholinergic neuron destruction than the ibotenate-induced lesion, as measured in terms of reductions in cortical choline acetyltransferase activity (44% vs 27%). Although consideration of individual data suggested that very high (60%) levels of choline acetyltransferase reduction in Experiment 2 might have detrimental effects of conditional learning, the overall failure of the quisqualate-induced lesions of the ventral pallidum/substantia innominata to impair learning is to be contrasted with the significant behavioural effects of ibotenate-induced lesions. Histological and immunocytochemical analysis showed that the quisqualate-induced lesion, unlike that produced by ibotenate, tended to produce less damage to the overlying dorsal globus pallidus and to parvocellular neurons of the ventral pallidum/substantia innominata, thus implicating these nonspecific effects of ibotenate-induced lesions in their behavioural effects. The present results question previous interpretations of the behavioural effects of ibotenate-induced lesions of the ventral pallidum/substantia innominata in terms of damage inflicted on the cortically-projecting cholinergic cells of the nucleus basalis, and suggest that quisqualic acid, although also nonspecific in its excitotoxic effects, is nevertheless more selective for producing damage to cholinergic neurons in the ventral pallidum/substantia innominata than ibotenic acid.     

Limited neocortical devascularizing lesions causing deficits in memory retention and choline acetyltransferase activity--effects of the monosialoganglioside GM1

In mature rats with unilateral cortical lesions, choline acetyltransferase activity was found to decrease by 40% in the ipsilateral nucleus basalis magnocellularis, compared with control animals. Intraventricular administration of the monosialoganglioside GM1 (5 mg/kg per day), via minipumps, over a period of 14 days prevented this fall in choline acetyltransferase activity. The activity of this enzyme in the sham operated subjects was not significantly different from that in the unoperated group. This biochemical data is in full agreement with regards to the protective action of monosialoganglioside GM1 on forebrain cholinergic neurons. In this study the behavioral effects of these cortical lesions were investigated for the first time. Results show that these lesions, with their associated retrograde neuronal degenerative effects, altered a variety of sensorimotor and memory-based behaviors. In particular, the limited unilateral devascularization of the neocortex increased baseline locomotor activity above control; affected motor coordination; impaired passive avoidance retention and reacquisition; and decreased the retention of learnt Morris water-maze information. Infusion of the monosialoganglioside into the ventricles, whilst having no effect on the increased motor activity or motor coordination deficit, did enhance the reacquisition of information in both memory-related tasks.     

Spatial relationship between neurotensinergic axons and cholinergic neurons in the rat basal forebrain: a light microscopic study with three-dimensional reconstruction

Cholinergic neurons of the basal forebrain are known to project to the hippocampus and cerebral cortex wherein they play an important role in cortical activation, attention and memory. These neurons have been shown to possess neurotensin binding sites and to respond electrophysiologically to local application of neurotensin, indicating the presence of functional receptors on their membrane. In the present light microscopic study, the spatial relationship between neurotensinergic axons and cholinergic nerve cell bodies and proximal dendrites was investigated in the basal forebrain of the rat by dual immunostaining for neurotensin and choline acetyltransferase. Rostrally, neurotensinergic fibres were concentrated in the lateral septum and anterior substantia innominata, whereas cholinergic neurons were located in the medial septum, diagonal band of Broca and magnocellular preoptic nucleus. At high magnification, a few neurotensinergic axonal varicosities were observed in the region of cholinergic neurons, and fewer still in close proximity to cholinergic perikarya and proximal dendrites. Caudally, neurotensinergic fibres formed a dense plexus of varicose axons in the same region where cholinergic neurons were located in the posterior substantia innominata and in the ventral and caudal aspects of the globus pallidus. At high magnification, many of these neurotensinergic varicosities were seen in close proximity to the cholinergic perikarya. These results suggest that cholinergic cells receive a much denser neurotensinergic innervation in the caudal than in the rostral aspect of the basal forebrain. This differential distribution is not reflected in the uniform density of neurotensin receptors and potent responses to neurotensin through the cholinergic cell population, suggesting the possibility that neurotensin's effects are mediated in part by a paracrine mechanism.