Mouse nerve growth factor prevents degeneration of axotomized basal forebrain cholinergic neurons in the monkey

NGF, a trophic polypeptide, is necessary for the normal development and survival of certain populations of neurons in the CNS and PNS. In the CNS, cholinergic neurons of the basal forebrain magnocellular complex (BFMC) are prominent targets of NGF. During rat development, NGF increases the activity of ChAT in these neurons. In adult rats with experimental injury of axons in the fimbria-fornix, NGF prevents degenerative changes in axotomized cholinergic BFMC neurons in the medial septal nucleus (MSN). Because the amino acid sequences of NGF and its receptor (NGF-R) are highly conserved across species, we hypothesized that mouse NGF would also prevent degeneration of cholinergic BFMC neurons in nonhuman primates. Therefore, the present study was designed to test whether fimbria-fornix lesions result in retrograde degenerative changes in basal forebrain cholinergic neurons in macaques, whether these changes are prevented by mouse NGF, and whether the protective effect of NGF is selective for cholinergic neurons of the basal forebrain. Following unilateral complete transection of the fornix, animals were allowed to survive for 2 weeks, during which time half of the subjects received intraventricular NGF in vehicle and the other half received vehicle alone. In animals receiving vehicle alone, there was a 55% reduction in the number of ChAT-immunoreactive cell bodies within the MSN ipsilateral to the lesion; loss of immunoreactive somata was more severe in caudal planes of the MSN. Remaining immunoreactive neurons appeared smaller than those in control, unoperated animals. In Nissl stains, there was no apparent loss of basophilic profiles in the MSN, but cells showed reduced size and intensity of basophilia. Treatment with NGF almost completely prevented reductions in the number and size of cholinergic neurons and had a significant general effect in preventing atrophy in basophilic magnocellular neurons of the MSN, though some basophilic neurons in the MSN did not appear to respond to NGF. Adjacent 7-microns-thick sections stained with ChAT and NGF-R immunocytochemistry revealed that these markers are strictly colocalized in individual neurons in the MSN in controls and in both groups of experimental animals. Thus, mouse NGF profoundly influences the process of axotomy-induced retrograde degeneration in cholinergic BFMC neurons in primates. The in vivo effectiveness of mouse NGF on primate BFMC neurons suggests that mouse or human recombinant NGF may be useful in ameliorating the ACh-dependent, age-associated memory impairments that occur in nonhuman primates.(ABSTRACT TRUNCATED AT 400 WORDS)     

Recombinant human nerve growth factor prevents retrograde degeneration of axotomized basal forebrain cholinergic neurons in the rat

Cholinergic neurons in the basal forebrain magnocellular complex (BFMC) respond to nerve growth factor (NGF) during development and in adult life, and it has been suggested that the administration of NGF might ameliorate some of the abnormalities that occur in neurological disorders associated with degeneration of this population of neurons. A prerequisite for the introduction of NGF in clinical trials is the availability of active recombinant human NGF (rhNGF). The present investigation was designed to test, in vivo, the efficacy of a preparation of rhNGF. Axons of cholinergic neurons of the BFMC in the rat were transected in the fimbria-fornix; this manipulation alters the phenotype and, eventually, causes retrograde degeneration of these neurons. Our investigation utilized two lesion paradigms (resection and partial transection of fibers in the fimbria-fornix), two different strains of rats, and two delivery systems. Following lesions, animals were allowed to survive for 2 weeks, during which time one group received intraventricular mouse NGF (mNGF), a second group received rhNGF, and a third group received vehicle alone. In animals receiving vehicle, there was a significant reduction in the number (resection: 70%; transection: 50%) and some reduction in size of choline acetyltransferase- or NGF receptor-immunoreactive cell bodies within the medial septal nucleus ipsilateral to the lesion. Treatment with either mNGF or rhNGF completely prevented these alterations in the number and size of cholinergic neurons. The rhNGF was shown to be equivalent in efficacy with mNGF. Thus, rhNGF is effective in preventing axotomy-induced degenerative changes in cholinergic neurons of the BFMC. Our results, taken together with the in vitro effects of rhNGF (42), indicate that an active rhNGF is now available for further in vivo studies in rodents and primates with experimentally induced or age-associated lesions of basal forebrain cholinergic neurons. These investigations provide essential information for the consideration of future utilization of rhNGF for treatment of human neurological disorders, including Alzheimer's disease.     

GABAergic and cholinergic neurons exhibit high-affinity nerve growth factor binding in rat basal forebrain

We have used dissociated, rat basal forebrain cultures to identify specific cell types that are potentially responsive to nerve growth factor (NGF). Expression of high-affinity NGF binding sites was examined. A subpopulation of cells containing choline acetyltransferase (CAT), the acetylcholine-synthesizing enzyme, exhibited high-affinity binding, employing combined immunocytochemistry and 125I-NGF radioautography. Unexpectedly, a gamma-aminobutyric acid (GABA)-containing cell group also expressed high-affinity binding. These cells that exhibit high-affinity binding appear to be neurons since they stain positively with the neuron marker, neuron-specific enolase, and negatively with the nonneuron marker, glial fibrillary acidic protein. Our observations suggest that NGF may regulate multiple brain systems and functions that have yet to be explored. Conversely, only subsets of cholinergic or GABA neurons expressed high-affinity binding, suggesting that these transmitter populations are composed of differentially responsive subpopulations.     

The role of nerve growth factor receptors in cholinergic basal forebrain degeneration in prodromal Alzheimer disease

Dysfunction of nerve growth factor (NGF) and its high (TrkA) and low (p75NTR) affinity receptors has been suggested to underlie the selective degeneration of the nucleus basalis (NB) cholinergic cortical projection neurons in end stage Alzheimer disease (AD). Whether the NGF system is dysfunctional during the prodromal stages of AD has only recently been evaluated. Surprisingly, the number of choline acetyltransferase-containing neurons remains stable despite a significant reduction in NGF receptor-positive cells in people with mild cognitive impairment (MCI), suggesting a phenotypic NGF receptor downregulation but not a frank loss of NB neurons during prodromal AD. Moreover, there is a loss of cortical TrkA in the face of stable p75NTR and increased proNGF levels, the precursor molecule of mature NGF, in early AD. Depending upon the cellular context these changes may result in increased pro-apoptotic signaling, cell survival, or a defect in retrograde transport mechanisms. Alterations in NGF and its receptors within the cholinotrophic NB system in early AD suggest that NGF-mediated cell signaling is required for the longterm survival of these neurons. Therapeutic neurotrophic intervention might delay or prevent NB neuron degeneration and preserve cholinergic cortical function during prodromal AD.     

Hypothesis linking plasticity,vulnerability and nerve growth factor to basal forebrain cholinergic neurons

The hypothesis is advanced that two striking features of basal forebrain cholinergic neurons (BFCN)—their role in memory and their vulnerability in disease—are related to their dependence on nerve growth factor (NGF). Numerous studies, involving retrograde transport, receptor localization and responses to exogenous growth factor, have demonstrated a relatively selective association of NGF with these neurons in adult brain. The role of BFCN innervating cortical areas such as the hippocampus (and other cortical areas) in memory is widely supported by the results of surgical or pharmacological manipulation in experimental animals. Neuronal degeneration, indicated by decreased cortical cholinergic activity and/or subcortical cell loss, is also evident in a wide variety of cerebral disorders involving memory loss and dementia (Alzheimer's and Parkinson's disease, subacute sclerosing panencephalitis, Lewy body dementia and olivopontocerebellar atrophy are some examples) and to a lesser extent in normal ageing itself, in which memory is particularly vulnerable. Evidence is provided to support the idea that activity-dependent uptake of NGF underlies both the memory function of BFCN and their susceptibility to disease. It is suggested that therapeutic strategies in dementia may need to be directed towards stimulation (pharmacological or physiological) of surviving BFCN.     

Age-related loss of nerve growth factor sensitivity in rat basal forebrain neurons

Nerve growth factor (NGF) has recently been implicated as a trophic agent in the survival and maintenance of basal forebrain cholinergic neurons. To test the hypothesis that NGF may play a role in the age-related degeneration of basal forebrain neurons and decline of cerebral cholinergic function, we have used a monoclonal antibody to the NGF receptor, 192 IgG, to immunocytochemically visualize and compare rat basal forebrain neurons responsive to NGF in aged (30 months) and young adult (10 months) rats. In a subpopulation of aged rats, NGF receptor-immunoreactive cells in the basal forebrain appear vacoulated and shrunken, and the neuropil staining is markedly reduced. While no substantial decline in cell density is apparent in Nissl-stained sections, the number of NGF receptor-positive cell profiles within the vertical limb of diagonal band nuclei is reduced by an average of 32% in aged rats. Marked reduction in the expression of NGF receptors in aged rats may signify loss of capacity of the basal forebrain neurons to bind and transport NGF from their terminals in the hippocampus and cortex, subsequent decrease in NGF delivered to the cell bodies, and eventual cellular dysfunction and death of neurons in aging.     

Acidic fibroblast growth factor mRNA is expressed by basal forebrain and striatal cholinergic neurons

Evidence for the importance of the basal forebrain cholinergic system in the maintenance of cognitive function has stimulated efforts to identify trophic mechanisms that protect this cell population from atrophy and dysfunction associated with aging and disease. Acidic fibroblast growth factor (aFGF) has been reported to support cholinergic neuronal survival and has been localized in basal forebrain with the use of immunohistochemical techniques. Although these data indicate that aFGF is present in regions containing cholinergic cell bodies, the actual site of synthesis of this factor has yet to be determined. In the present study, in situ hybridization techniques were used to evaluate the distribution and possible colocalization of mRNAs for aFGF and the cholinergic neuron marker choline acetyltransferase (ChAT) in basal forebrain and striatum. In single-labeling preparations, aFGF mRNA-containing neurons were found to be codistributed with ChAT mRNA⁺ cells throughout all fields of basal forebrain, including the medial septum/diagonal band complex and striatum. By using a double-labeling (colormetric and isotopic) technique, high levels of colocalization (over 85%) of aFGF and ChAT mRNAs were observed in the medial septum, the diagonal bands of Broca, the magnocellular preoptic area, and the nucleus basalis of Meynert. The degree of colocalization was lower in the striatum, with 64% of the cholinergic cells in the caudate and 33% in the ventral striatum and olfactory tubercle labeled by the aFGF cRNA. These data demonstrate substantial regionally specific patterns of colocalization and support the hypothesis that, via an autocrine mechanism, aFGF provides local trophic support for cholinergic neurons in the basal forebrain and the striatum. © 1996 Wiley-Liss, Inc.     

Neurofibrillary degeneration of cholinergic and noncholinergic neurons of the basal forebrain in Alzheimer's disease

Two principal features of Alzheimer's disease (AD) are (1) the occurrence of neurofibrillary tangles (NFTs) and senile plaques, and (2) the loss of cortical cholinergic activity because of dysfunction of neurons in the basal forebrain cholinergic system. The relationship of these two abnormalities is an unresolved issue in the pathology of AD. We used polyclonal antibodies specific for paired helical filaments (PHFs), combined with acetylcholinesterase (AChE) histochemistry, to assess the cytoskeletal changes of cholinergic and noncholinergic neurons in the basal forebrain in AD. In both sporadic and familial AD, the nucleus basalis of Meynert (nbM) showed a marked decrease in AChE-positive (AChE+) perikarya and abundant immunoreactive NFTs. In double-labeling studies of the nbM, PHF reactivity was found both in surviving AChE+ neurons and in many AChE- NFTs that were not associated with microscopically recognizable cell structures. Some surviving AChE+ perikarya did not contain NFTs. Numerous NFTs and senile plaques were identified by PHF immunoreactivity in other basal forebrain areas, including subnuclei of the amygdala that showed low or absent AChE activity. We conclude that the dysfunction and death of cholinergic neurons in the nbM is associated with extensive NFT formation, including apparently residual NFTs in loci where nbM neurons once existed; and many noncholinergic neurons and neurites in the basal forebrain show NFT and senile plaque formation. The cytopathology of AD involves neurons of varying transmitter specificities, including cholinergic neurons in the basal forebrain.     

Neurogenesis of basal forebrain cholinergic neurons in rat

The basal forebrain cholinergic system embodies a heterogeneous group of neurons distributed in the basal telencephalon that project topographically to the cortical mantle. We sought to examine the generation of these neurons to determine whether basal forebrain neurons have unique patterns of neurogenesis or, if, in contrast, they are born along general neurogenic gradients. The techniques of tritiated thymidine autoradiography and choline acetyltransferase (ChAT) immunocytochemistry were combined to determine the birthdays and neurogenic gradients of cholinergic cells in this region of rat brain. Cholinergic neurogenesis throughout the basal forebrain ranged from embryonic days 12 to 17 (E12-17). Neurogenesis in the nucleus basalis magnocellularis occurred over E12-16, with a peak day of generation on E13. The horizontal limb nucleus of the diagonal band which is located rostral to the nucleus basalis was generated over E12-17, with the majority of cells arising on E14-15. The rostral-most nuclei of the basal forebrain cholinergic system, the vertical limb of the diagonal band and the medial septum, were generated between E13-17, with peak days of neurogenesis on E15 and E15-16, respectively. These results were evaluated quantitatively and demonstrated that the basal forebrain cholinergic neurons were generated along the general caudal-to-rostral gradient previously described for all neurons in this brain region. The results of this study, in combination with those of similar investigations, emphasize that position-dependent epigenetic factors appear to be more potent determinants of the time of neuronal origin than factors which influence a cell's transmitter phenotype.     

Nerve growth factor induces Fos-like immunoreactivity within identified cholinergic neurons in the adult rat basal forebrain

Immunocytochemical techniques were used to examine and compare the effects of intracerebroventricular administration of nerve growth factor (NGF) on Fos expression within identified cholinergic and non-cholinergic neurons located in different regions of the adult rat basal forebrain. Animals were killed 1, 3, 6, and 12 h after receiving NGF (0.5 or 5.0 μg) or vehicle into the left lateral ventricle and sections through the medial septum, diagonal band of Broca, nucleus basalis magnocellularis, and striatum were processed for the combined immunocytochemical detection of Fos and choline acetyltransferase (a marker for cholinergic neurons), or Fos and parvalbumin (a marker for gamma aminobutyric acid (GABA)-containing neurons). NGF produced a significant increase in the percentage of cholinergic neurons containing Fos-like immunoreactivity within all four regions examined. The largest increases were detected in the medial septum (47.8%) and the horizontal limb of the diagonal band of Broca (67.7%). In these areas, NGF-mediated induction of Fos-like immunoreactivity was detected as early as 3 h, peaked at 6 h, and was reduced by 12 h, postinfusion. Small but significant increases in the percentage of cholinergic neurons containing Fos-like immunoreactivity were also detected in the striatum (4.2%) and in the nucleus basalis magnocellularis (19.2%) 3–12 h following administration of the higher dose of NGF. No evidence for an NGF-mediated induction of Fos within parvalbumin-containing neurons was detected in any of the four regions at any of the time-points examined; however, evidence for an NGF-mediated induction of Fos within epithelial cells lining the lateral ventricle was observed. These data demonstrate that NGF induces Fos expression within cholinergic, and not parvalbumin-containing (GABAergic), neurons in the basal forebrain, and furthermore that intracerebroventricular administration of NGF influences the different subgroups of basal forebrain cholinergic neurons to different degrees. ©1977 Elsevier Science B.V. All rights reserved.     

Nerve growth factor mRNA-containing cells are distributed within regions of cholinergic neurons in the rat basal forebrain

It has been proposed that nerve growth factor (NGF) provides critical trophic support for the cholinergic neurons of the basal forebrain and that it becomes available to these neurons by retrograde transport from distant forebrain targets. However, neurochemical studies have detected low levels of NGF mRNA within basal forebrain areas of normal and experimental animals, thus suggesting that some NGF synthesis may actually occur within the region of the responsive cholinergic cells. In the present study with in situ hybridization and immunohistochemical techniques, the distribution of cells containing NGF mRNA within basal forebrain was compared with the distribution of cholinergic perikarya. The localization of NGF mRNA was examined by using a ³⁵S-labeled RNA probe complementary to rat preproNGF mRNA and emulsion autoradiography. Hybridization of the NGF cRNA labeled a large number of cells within the anterior olfactory nucleus and the piriform cortex as well as neurons in a continuous zone spanning the lateral aspects of both the horizontal limb of the diagonal band of Broca and the magnocellular preoptic nucleus. In the latter regions, large autoradiographic grain clusters labeled relatively large Nissl-pale nuclei; it did not appear that glial cells were autoradiographically labeled. Comparison of adjacent tissue sections processed for in situ hybridization to NGF mRNA and immunohistochemical localization of choline acetyltransferase (ChAT) demonstrated overlapping fields of cRNA-labeled neurons and ChAT-immunoreactive perikarya in both the horizontal limb of the diagonal band and magnocellular preoptic regions. However, no hybridization of the cRNA probe was observed in other principal cholinergic regions including the medial septum, the vertical limb of the diagonal band, or the nucleus basalis of Meynert. These results provide evidence for the synthesis of NGF mRNA by neurons within select fields of NGF-responsive cholinergic cells and suggest that the generally accepted view of “distant” target-derived neurotrophic support should be reconsidered and broadened.     

Fiber pathways of basal forebrain cholinergic neurons in monkeys

In rhesus monkeys, autoradiographic tracing methods, complemented by immunocytochemical and histochemical techniques, were used to delineate pathways by which cholinergic neurons of the nucleus basalis of Meynert (nbM) and nucleus of the diagonal band of Broca (ndbB) project to forebrain targets. Following injections of [3H]amino acids into these nuclei, 5 major fiber pathways were identified: axons of the nbM and ndbB project medially, principally within the cingulum bundle, to dorsomedial portions of the hemispheres; nbM and ndbB fibers exit laterally beneath the pallidum and striatum, enter the external and extreme capsules, and pass within the corona radiata to terminate in lateral and caudal regions of neocortex; axons coursing ventrally from the nbM project to portions of the temporal lobe, including the amygdala; some fibers pass through the fibrae pass orbitofrontales to the orbitofrontal cortex; and, finally axons of the nbM/ndbB project via the fimbria/rornix and a ventral pathway to the hippocampus. The presence of these 5 radiolabeled pathways arising from basal forebrain cholinergic neurons was confirmed by acetylcholinesterase histochemistry and choline acetyltransferase immunocytochemistry.     

GABAergic control of basal forebrain cholinergic neurons and memory

The involvement of the GABAergic innervation of basal forebrain neurons in the rats' conditional visual discrimination performance was examined. Performance in such a task is based on the subjects's ability to retrieve information about response rules, and previous experiments have demonstrated that basal forebrain lesions interfere with this ability. Following the acquisition of the task, chronic guide cannulae were stereotaxically implanted into the substantia innominata of both hemispheres, and the animals were retrained. Administration of the GABAA-agonist muscimol into the substantia innominata (0, 25, 50 ng/0.5 microliters/hemisphere) dose-dependently decreased the number of correct responses, increased the number of errors of omission, increased response latency, but did not affect side bias. Systemic co-administration of the cholinesterase inhibitor physostigmine (0, 0.1, 0.2 mg/kg; i.p.) exclusively interacted with the effects of muscimol on correct responding. Specifically, physostigmine dose-dependently intensified and attenuated the muscimol-induced reduction in correct responding. Although it cannot be excluded that alternative neuronal mechanisms were involved in the mediation of the effects of muscimol and their interaction with physostigmine, these findings support previous evidence indicating that the activity of basal forebrain cholinergic neurons is controlled by a GABAergic input, and that this neuronal link is involved in mnemonic processing.     

Nerve growth factor receptor-containing cholinergic neurons of the basal forebrain project to the thalamic reticular nucleus in the rat

The origin of nerve growth factor receptor-immunoreactive (NGFr-ir) fibers innervating the thalamic reticular nucleus (Rt) was here investigated in the rat using retrograde tracers in combination with immunocytochemistry. Neurons retrogradely labeled from Rt were scattered ipsilaterally throughout the medial septal nucleus and the other cell groups of the basal forebrain, which contained NGFr-ir cells; 10–20% of these retrogradely labeled neurons were also NGFr-ir. Furthermore, a few retrogradely labeled NGFr-ir cells were detected in the basal forebrain on the contralateral side. Retrograde tracing combined with a double immunocytochemical procedure revealed that all the NGFr-ir neurons labeled from Rt also displayed immunoreactivity for choline acetyltransferase. The present results demontrate that the NGFr-ir neurons of the basal forebrain which project to Rt are cholinergic. The possible functional implications of these findings are discussed.     

Pituitary adenylate cyclase-activating polypeptide promotes the survival of basal forebrain cholinergic neurons in vitro and in vivo: comparison with effects of nerve growth factor

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a member of the vasointestinal polypeptide gene family for which neurotrophic activity has been postulated. PACAP mRNA is expressed in the developing and adult hippocampus, which is the principal target region of septal cholinergic neurons. We therefore studied the effects of PACAP on septal cholinergic neurons. In primary cultures from septum of embryonic and postnatal rats, PACAP increased the number of neurons immunohistochemically stained for the low-affinity nerve growth factor (NGF) receptor p75 and for the enzyme choline acetyltransferase (ChAT). PACAP also caused a corresponding increase in ChAT activity. In comparison, NGF had a greater effect than PACAP on the number of p75- and ChAT-positive neurons in these cultures. In vivo, following fimbria fornix transection, the number of immunohistochemically stained septal cholinergic neurons fell significantly to 18% in rats given continuous intracerebroventricular infusion of vehicle, whereas in rats given NGF the number of these neurons did not differ significantly from unoperated controls. In PACAP-treated rats the number was 48% of unoperated values, which represented a significant increase compared with vehicle-treated rats and a significant decrease compared with NGF-treated rats or unoperated controls. Double-staining experiments revealed that most ChAT-positive neurons in rat medial septum also express PACAP receptor 1. Together the results show that PACAP promotes the survival of septal cholinergic neurons in vitro, and after injury in vivo, suggesting that PACAP acts as a neurotrophic factor influencing the development and maintenance of these neurons.     

Cultured basal forebrain cholinergic neurons in contact with cortical cells display synapses,enhanced morphological features,and decreased dependence on nerve growth factor

Prior studies examining the dependence of basal forebrain cholinergic neurons (BFCNs) on nerve growth factor (NGF) for survival have reached differing conclusions depending on the experimental paradigm employed, suggesting the importance of environmental and developmental variables. The present study examined the NGF dependence of BFCNs and modulatory effects of target (cortical) neurons under the controlled conditions of dissociated cell cultures. Initial experiments found BFCNs (identified by using choline acetyltransferase immunocytochemistry) in pure basal forebrain (BF) cultures to be dependent on NGF between the 2nd and 4th week in vitro. During that developmental period, NGF deprivation for 3 days, induced by application of anti-NGF antibody, resulted in degeneration of over 80% of BFCNs, whereas at earlier or later times, BFCNs were largely resistant to NGF deprivation. When BF neurons were plated together with cortical neurons (as dissociated co-cultures), the BFCNs grew neuritic processes (labeled with acetylcholinesterase histochemistry) that appeared to specifically target cortical neurons; electron microscopy revealed that synapses formed between these cells. BFCNs in co-cultures were more resistant to NGF deprivation, were larger, and had much more extensive neuritic growth than BFCNs in pure BF cultures. The resistance of BFCNs to NGF deprivation provided by cortical neurons could be largely reproduced by addition of other trophic factors (brain-derived neurotrophic factor, BDNF; neurotrophin 3, NT3; neurotrophin 4/5, NT4/5; or glial-derived neurotrophic factor, GDNF) during NGF deprivation in pure BF cultures. These results suggest that developing BFCNs undergo a critical period requiring trophic influences that may be provided by NGF or other trophic factors, as well as unknown factors derived from cortical neurons. © 1996 Wiley-Liss, Inc.     

Nerve growth factor receptor immunoreactive profiles in the normal, aged human basal forebrain: colocalization with cholinergic neurons

A monoclonal antibody raised against the receptor for nerve growth factor (NGF) has been used to map the distribution of NGF receptor-containing profiles within the human basal forebrain of four male and three female elderly patients without neurologic or psychiatric illness. Immunohistochemically processed tissue reveals a continuum of NGF receptor-positive neurons located within the medial septum, vertical and horizontal limb nuclei of the diagonal band, and nucleus basalis. NGF receptor-containing neurons are also found within the bed nucleus of the stria terminalis, the anterior commissure, the internal capsule, and the internal and external medullary laminae of the globus pallidus. Virtually all (greater than 95%) NGF receptor-containing neurons colocalize with the specific cholinergic marker choline acetyltransferase (ChAT) or the nonspecific marker acetylcholinesterase (AChE). Conversely, a few cholinergic perikarya are found which are not NGF receptor positive (and vice versa). These findings demonstrate that human basal forebrain neurons on which NGF receptor immunoreactivity is detected are primarily cholinergic and analogous to the nonhuman primate Ch1-Ch4 subgroups of Mesulam et al. (J. Comp. Neurol., 214:170-197, '83). NGF receptor-containing fiber tracts are observed emanating from the medial septum and vertical limb nucleus of the diagonal band coursing medially within the fornix. Another fascicle originating mainly from the nucleus basalis and travelling within the external capsule enroute to the cortex is observed innervating all cortical layers. Comparison of NGF receptor- and ChAT-containing neurons reveals cholinergic perikarya within the striatal complex, whereas virtually no NGF receptor-containing neurons are found in these structures. An occasional displaced NGF receptor-containing neurons is seen in the ventrolateral portion of the putamen and the white matter underlying the nucleus accumbens. These data are discussed in terms of the relationship of NGF receptor- and ChAT-containing neurons within the basal forebrain and in terms of the possible functional significance of NGF in normal and diseased brain.     

Time of origin of cholinergic neurons in the rat basal forebrain

The timing of the final mitotic division of basal forebrain cholinergic neurons was studied by injecting [3H]thymidine into timed pregnant rats and processing the brains of their progeny as young adults for immunohistochemistry with a monoclonal antibody to choline acetyltransferase (ChAT) followed by autoradiography. ChAT-positive neurons located caudally in the basal forebrain were found to become postmitotic mostly on embryonic (E) days 12 and 13, whereas the peak final mitosis of more rostrally located ChAT-positive neurons occurred increasingly later, with the most rostral ChAT-immunoreactive neurons leaving their final mitotic cycles on E15 and E16. In all basal forebrain regions, cholinergic neurogenesis was complete by E17. These results indicate that the cholinergic neurons in the basal forebrain become postmitotic in a caudal-to-rostral gradient over about 5 days. The continuity of the gradient suggests that these cholinergic neurons may derive from the same germinal source.     

Nerve growth factor increases choline acetyltransferase activity in developing basal forebrain neurons

Nerve growth factor (NGF) is a neuronotrophic protein. Its effects on developing peripheral sensory and sympathetic neurons have been extensively characterized, but it is not clear whether NGF plays a role during the development of central nervous system neurons. To address this point, we examined the effect of NGF on the activity of neurotransmitter enzymes in several brain regions. Intracerebroventricular injections of highly purified mouse NGF had a marked effect on the activity of choline acetyltransferase (ChAT), a selective marker of cholinergic neurons. NGF elicited prominent increases in ChAT activity in the basal forebrain of neonatal rats, including the septum and a region which contains neurons of the nucleus basalis and substantia innominata. NGF also increased ChAT activity in the hippocampus and neocortex, terminal regions for the fibers of basal forebrain cholinergic neurons. In analogy with the response of developing peripheral neurons, the NGF effect was shown to be selective for basal forebrain cholinergic cells and to be dose-dependent. Furthermore, septal neurons closely resembled sympathetic neurons in the time course of their response to NGF. These observations suggest that endogenous NGF does play a role in the development of basal forebrain cholinergic neurons.     

Heterogeneity and selectivity of the degeneration of cholinergic neurons in the basal forebrain of patients with Alzheimer's disease

Cholinergic neurons were studied by immunohistochemistry, with an antiserum against choline acetyltransferase (ChAT), in the basal forebrain (Ch1 to Ch4) of four patients with Alzheimer's disease (AD) and four control subjects. ChAT-positive cell bodies were mapped and counted in Ch1 (medial septal nucleus), Ch2 (vertical nucleus of the diagonal band), Ch3 (horizontal nucleus of the diagonal band) and Ch4 (nucleus basalis of Meynert). Compared to controls, the number of cholinergic neurons in AD patients was reduced by 50% on average. The interindividual variations in cholinergic cell loss were high, neuronal loss ranging from moderate (27%) to severe (63%). Despite the small number of brains studied, a significant correlation was found between the cholinergic cell loss and the degree of intellectual impairment. To determine the selectivity of cholinergic neuronal loss in the basal forebrain of AD patients, NPY-immunoreactive neurons were also investigated. The number of NPY-positive cell bodies was the same in controls and AD patients. The results (1) confirm cholinergic neuron degeneration in the basal forebrain in AD and the relative sparing of these neurons in some patients, (2) indicate that degneration of cholinergic neurons in the basal forebrain contributes to intellectual decline, and (3) show that, in AD, such cholinergic cell loss is selective, since NPY-positive neurons are preserved in the basal forebrain.