Combined 192 IgG-saporin and 5,7-dihydroxytryptamine lesions in the male rat brain: a neurochemical and behavioral study

In a previous experiment [Eur J Neurosci 12 (2000) 79], combined intracerebroventricular injections of 5,7-dihydroxytryptamine (5,7-DHT; 150 microg) and 192 IgG-saporin (2 microg) in female rats produced working memory impairments, which neither single lesion induced. In the present experiment, we report on an identical approach in male rats. Behavioral variables were locomotor activity, T-maze alternation, beam-walking, Morris water-maze (working and reference memory) and radial-maze performances. 192 IgG-saporin reduced cholinergic markers in the frontoparietal cortex and the hippocampus. 5,7-DHT lesions reduced serotonergic markers in the cortex, hippocampus and striatum. Cholinergic lesions induced motor deficits, hyperactivity and reduced T-maze alternation, but had no other effect. Serotonergic lesions only produced hyperactivity and reduced T-maze alternation. Beside the deficits due to cholinergic lesions, rats with combined lesions also showed impaired radial-maze performances. We confirm that 192 IgG-saporin and 5,7-DHT injections can be combined to produce concomitant damage to cholinergic and serotonergic neurons in the brain. In female rats, this technique enabled to show that interactions between serotonergic and basal forebrain cholinergic mechanisms play an important role in cognitive functions. The results of the present experiment in male rats are not as clear-cut, although they are not in obvious contradiction with our previous results in females.     

Brain cholinergic vulnerability: relevance to behavior and disease

The major populations of cholinergic neurons in the brain include two "projection" systems, located in the pontine reticular formation and in the basal forebrain. These two complexes comprise, in part, the anatomical substrates for the "ascending reticular activating system" (ARAS). The pontine cholinergic system relays its rostral influences mainly through thalamic intralaminar nuclei, but it also connects to the basal forebrain and provides a minor innervation of cortex. The basal forebrain cholinergic complex (BFCC) projects directly to cortex and hippocampus, and has a minor connection with the thalamus. Recent data reveal that a parallel system of basal forebrain GABAergic projection neurons innervates cortex/hippocampus in a way that seems to complement the BFCC. Generally, the picture developed from more than 50 years of research is consistent with a "global" influence of these two ascending cholinergic projections on cortical and hippocampal regions. Seemingly, the BFCC acts in tandem or in parallel with the pontine cholinergic projection to activate the electro-encephalogram, increase cerebral blood flow, regulate sleep-wake cycling, and modulate cognitive function. There are quite a number and variety of human brain conditions, notably including Alzheimer's disease, in which degeneration of basal forebrain cholinergic neurons has been documented. Whether the corticopetal GABA system is affected by disease has not been established. Studies of degeneration of the pontine projection are limited, but the available data suggest that it is relatively preserved in Alzheimer's disease. Hypotheses of BFCC degeneration include growth factor deprivation, intracellular calcium dysfunction, amyloid excess, inflammation, and mitochondrial abnormalities/oxidative stress. But, despite considerable research conducted over several decades, the exact mechanisms underlying brain cholinergic vulnerability in human disease remain unclear.     

Galanin: neurobiologic mechanisms and therapeutic potential for Alzheimer's disease

The neuropeptide galanin (GAL) is widely distributed in the mammalian CNS. Several lines of evidence suggest that GAL may play a critical role in cognitive processes such as memory and attention through an inhibitory modulation of cholinergic basal forebrain activity. Furthermore, GAL fibers hyperinnervate remaining cholinergic basal forebrain neurons in Alzheimer's disease (AD). This suggests that GAL activity impacts cholinergic dysfunction in advanced AD. Pharmacological and in vitro autoradiographic studies indicate the presence of heterogeneous populations of GAL receptor (GALR) sites in the basal forebrain which bind GAL with both high and low affinity. Interestingly, we have recently observed that GALR binding sites increase in the anterior basal forebrain in late-stage AD. Three G protein-coupled GALRs have been identified to date that signal through a diverse array of effector pathways in vitro, including adenylyl cyclase inhibition and phospholipase C activation. The repertoire and distribution of GALR expression in the basal forebrain remains unknown, as does the nature of GAL and GALR plasticity in the AD basal forebrain. Recently, GAL knockout and overexpressing transgenic mice have been generated to facilitate our understanding of GAL activity in basal forebrain function. GAL knockout mice result in fewer cholinergic basal forebrain neurons and memory deficits. On the other hand, mice overexpressing GAL display hyperinnervation of basal forebrain and memory deficits. These data highlight the need to explore further the putative mechanisms by which GAL signaling might be beneficial or deleterious for cholinergic cell survival and activity within basal forebrain. This information will be critical to understanding whether pharmacological manipulation of GALRs would be effective for the amelioration of cognitive deficits in AD.     

Nerve growth factor: structure, function and therapeutic implications for Alzheimer's disease

Over the past decade, neurotrophic factors have generated much excitement for their potential as therapy for neurological disorders. In this regard, nerve growth factor (NGF), the founding member of the neurotrophin family, has generated great interest as a potential target for the treatment of Alzheimer's disease (AD). This interest is based on the observation that cholinergic basal forebrain (CBF) neurons which provide the major source of cholinergic innervation to the cerebral cortex and hippocampus undergo selective and severe degeneration in advanced AD and that these neurons are dependent upon NGF and its receptors for their survival. In fact, NGF transduces its effects by binding two classes of cell surface receptors, TrkA and p75(NTR), both of which are produced by CBF neurons. This review focuses on NGF/receptor binding, signal transduction, regulation of specific cellular endpoints, and the potential use of NGF in AD. Alterations in NGF ligand and receptor expression at different stages of AD are summarized. Recent results suggest that cognitive deficits in early AD and mild cognitive impairment (MCI) are not associated with a cholinergic deficit. Thus, the earliest cognitive deficits in AD may involve brain changes other than simply cholinergic system dysfunction. Recent findings indicate an early defect in NGF receptor expression in CBF neurons; therefore treatments aimed at facilitating NGF actions may prove highly beneficial in counteracting the cholinergic dysfunction found in end-stage AD and attenuating the rate of degeneration of these cholinergic neurons.     

Cholinergic but not monoaminergic denervation increases nerve growth factor content in the adult rat hippocampus and cerebral cortex

Summary We have investigated whether degeneration of basal forebrain cholinergic neurons is a potential trigger for increased NGF production in the adult rat brain. Electrolytic lesions of cholinergic neurons in the septum-diagonal band and in the nucleus basalis of Meynert induced a transient increase in NGF in the ventral hippocampus (+70%) and cerebral cortex (+125%), respectively. In contrast, selective aminergic denervation of the forebrain by electrolytic lesion of the medial forebrain bundle, did not increase NGF levels in hippocampus and cerebral cortex. Thus, a cholinergic mechanism appears to regulate NGF production in adult rat basal forebrain.     

Muscarinic cholinergic contribution to memory consolidation: with attention to involvement of the basolateral amygdala

The central cholinergic system and muscarinic cholinergic receptor (mR) activation have long been associated with cognitive function. And degeneration of the cholinergic basal forebrain (CBF) neurons is a pronounced hallmark of Alzheimer's Disease (AD). However, CBF immunolesions as animal models of AD cholinergic degeneration have not replicated the robust memory deficits of nonselective excitotoxic lesions. The less studied cholinergic projections to the amygdala, which are affected in AD but unaffected by immunolesions, may be more important in memory storage than previously suspected. The sparing of these amygdalopetal projections may help explain the dissociation between excitotoxic and immunotoxic CBF lesions. The CBF projections to cortex have since been shown to be important for attentional processes, which may contribute indirectly to memory. Nonetheless, there are conditions under which their selective ablation produces clear memory deficits. For example, memory enhancement induced by posttraining basolateral amygdalar activation is ineffective when corticopetal cholinergic projections are lesioned. Moreover, posttraining cholinergic agonism enhances long-term memory. Such findings suggest that cholinergic innervation of the cortex may be particularly important during modulation of memory storage for stressful and/or arousing events. In concordance, mR agonism facilitates neuronal plasticity and can induce expression of memory-associated immediate early genes. The present article reviews the behavior, physiology and inducible genetic expression literatures which together suggest that the early CBF lesion data were not a red herring but rather that CBF projections not only to cortex but also to the amygdala may in fact have important neuromodulatory functions in memory consolidation processes.     

Pharmaceutical research strategies in Alzheimer's disease

The hypothesis that the symptomatology of Alzheimer's disease is attributable to loss of cortical cholinergic innervation is supported by post-mortem studies demonstrating both reduced choline acetyltransferase (CAT) activity and reduced high-affinity choline uptake (HAChT) in the cortex, and neuron cell loss in the basal forebrain nucleus basalis of Meynert. Cholinergic denervation occurs early in the course of the disease and correlates with the severity of cognitive impairment. Pharmaceutical research strategies based on the cholinergic hypothesis and focusing on the development of acetylcholine esterase inhibitors and muscarinic agonist will be discussed. An alternative hypothesis to explain the dementia of Alzheimer's disease is the glutamatergic hypothesis. This is based on postmortem evidence indicating loss of glutamate, reduced binding and uptake of the neuro-transmitter and loss of pyramidal cells in the cortex. The possibility of developing glutamatergic drugs for treating the symptomatology of Alzheimer's disease will be considered. Studies have also indicated that the cellular processes involved in Alzheimer's disease occur in non-neuronal cells as well as in the brain. The possibility that Alzheimer's disease is a systemic disease and that drugs can be developed to treat the disease process will be considered.     

Systemic and intrabasalis administration of the orexin-1 receptor antagonist,SB-334867, disrupts attentional performance in rats

Rationale  

Orexin neurons project to a number of brain regions, including onto basal forebrain cholinergic neurons. Basal forebrain corticopetal cholinergic neurons are known to be necessary for normal attentional performance. Thus, the orexin system may contribute to attentional processing.     

Activating the damaged basal forebrain cholinergic system: tonic stimulation versus signal amplification

The hypothesis that the cognitive decline in senile dementia is related to the loss of cortical cholinergic afferent projections predicts that pharmacological manipulations of the remaining cholinergic neurons will have therapeutic effects. However, treatment with cholinesterase inhibitors or muscarinic agonists has been, for the most part, largely unproductive. These drugs seem to disrupt the normal patterning of cholinergic transmission and thus may block proper signal processing. An alternative pharmacological strategy which focuses on the amplification of presynaptic activity without disrupting the normal patterning of cholinergic transmission appears to be more promising. Such a strategy may make use of the normal GABAergic innervation of basal forebrain cholinergic neurons in general, and in particular of the inhibitory hyperinnervation of remaining cholinergic neurons which may develop under pathological conditions. Disinhibition of the GABAergic control of cholinergic activity is assumed to intensify presynaptic cortical cholinergic activity and to enhance cognitive processing. Although the extent to which compounds such as the benzodiazepine receptor antagonist-carboline ZK 93 426 act via the basal forebrain GABA-cholinergic link is not yet clear, the available data suggest that the beneficial behavioral effects of this compound established in animals and humans are based on indirect cholinomimetic mechanisms. It is proposed that an activation of residual basal forebrain cholinergic neurons can be achieved most physiologically via inhibitory modulation of afferent GABAergic transmission. This modulation may have a therapeutic value in treating behavioral syndromes associated with cortical cholinergic denervation.     

Galactose counteracts hypoglycemia-induced decline of cholinergic neurons at low pH in organotypic rat brain slices of the basal nucleus of Meynert

A growing body of evidence indicates that hypoglycemia and acidosis may contribute to the development of Alzheimer's disease (AD). The cell death of basal forebrain cholinergic neurons constitutes a hallmark of AD and directly correlates with cognitive impairment. The aim of the present study was to investigate, in an organotypic rat brain slice model of the basal nucleus of Meynert, the effects of glucose deprivation on cholinergic neurons under normal and acidic conditions. Furthermore, we were interested to explore whether different saccharides (galactose, fructose, saccharose, lactose) can replace glucose under low pH conditions. Our data show a pH-dependent survival of cholinergic neurons at a high (37.1 mmol/l) glucose level, which was markedly decreased at a low (5.6 mmol/l) glucose level. Galactose (+31.5 mmol/l) significantly counteracted the loss of choline acetyltransferase-positive neurons in low-glucose-treated slices, while fructose, lactose and saccharose only partly protected cholinergic neurons. In conclusion, our results indicate that replacement of glucose with different saccharides, but most potently with galactose, protects cholinergic neurons against hypoglycemia at a low pH.     

Cholecystokinin protects cholinergic neurons against basal forebrain lesion.

Alzheimer's Disease (AD) patients have a severe degeneration of cholinergic neurons in their cerebral cortices. Basal forebrain (BF)-lesioned rat is used as a model animal of a cholinergic deficit in the cerebral cortex. Cholinergic markers were decreased in the cerebral cortex of BF-lesioned rats. Intracerebroventricular continuous infusion of cholecystokinin octapeptide (CCK8) following BF lesion obviously preserved these cholinergic markers. These results suggest that CCK8 prevents the degeneration of cholinergic neurons in the cerebral cortex following BF lesion.     

Combined damage to entorhinal cortex and cholinergic basal forebrain neurons, two early neurodegenerative features accompanying Alzheimer's disease: effects on locomotor activity and memory functions in rats.

In Alzheimer's disease (AD), cognitive decline is linked to cholinergic dysfunctions in the basal forebrain (BF), although the earliest neuronal damage is described in the entorhinal cortex (EC). In rats, selective cholinergic BF lesions or fiber-sparing EC lesions may induce memory deficits, but most often of weak magnitude. This study investigated, in adult rats, the effects on activity and memory of both lesions, alone or in combination, using 192 IgG-saporin (OX7-saporin as a control) and L-N-methyl-D-aspartate to destroy BF and EC neurons, respectively. Rats were tested for locomotor activity in their home cage and for working- and/or reference-memory in various tasks (water maze, Hebb-Williams maze, radial maze). Only rats with combined lesions showed diurnal and nocturnal hyperactivity. EC lesions impaired working memory and induced anterograde memory deficits in almost all tasks. Lesions of BF cholinergic neurons induced more limited deficits: reference memory was impaired in the probe trial of the water-maze task and in the radial maze. When both lesions were combined, performance never improved in the water maze and the number of errors in the Hebb-Williams and the radial mazes was always larger than in any other group. These results (i) indicate synergistic implications of BF and EC in memory function, (ii) suggest that combined BF cholinergic and fiber-sparing EC lesions may model aspects of anterograde memory deficits and restlessness as seen in AD, (iii) challenge the cholinergic hypothesis of cognitive dysfunctions in AD, and (iv) contribute to open theoretical views on AD-related memory dysfunctions going beyond the latter hypothesis.     

Orexin/hypocretin modulation of the basal forebrain cholinergic system: insights from in vivo microdialysis studies

Since its discovery less than a decade ago, interest in the hypothalamic orexin/hypocretin system has blossomed due to the diversity and importance of the roles played by these neuropeptides. Orexin neurons have widespread projections throughout the central nervous system and intense research has focused on elucidating the pathways and mechanisms by which orexins exert their diverse array of functions. Our group has recently focused on orexin inputs to the basal forebrain cholinergic system, which plays a crucial role in cognitive--particularly attentional--function. Orexin cells provide a robust input to cholinergic neurons in the basal forebrain and act here to modulate cortical acetylcholine release. Orexin A also increases local glutamate release within the basal forebrain, suggesting an additional, indirect effect of orexins on basal forebrain cholinergic activity. Orexin activation of the basal forebrain cholinergic system appears to be especially relevant in the context of homeostatic challenges, such as food deprivation. Thus, orexins can stimulate cortical cholinergic transmission which, in turn, may promote the detection and selection of stimuli related to physiological needs. In this manner, orexin interactions with the basal forebrain cholinergic system are likely to form a link between arousal and attention in support of the cognitive components of motivated behavior.     

α7 nicotinic acetylcholine receptors in Alzheimer's disease: neuroprotective, neurotrophic or both?

One of the early signs of Alzheimer's disease is the impairment in hippocampus-based episodic memory function, which is improved through the enhancement of cholinergic transmission. Several studies suggest that α7 nicotinic receptor (nAChR) activation represents a useful therapeutic strategy for the cognitive impairments associated with early Alzheimer's disease as the α7 subtype of nicotinic acetylcholine receptors are expressed by basal forebrain cholinergic projection neurons as well as by their targets in the hippocampus. The current model for the cholinergic deficit in Alzheimer's disease posits that inappropriate accumulation of misfolded oligomeric aggregates of β-amyloid peptide leads to the dysfunction of the signaling mechanisms that support the cholinergic phenotype; this is manifested as an altered function of nicotinic acetylcholine receptors and the nerve-growth factor trophic support system that results in the loss of cholinergic markers and eventually cholinergic neurons from the basal forebrain cholinergic system. A view was confounded by the fact that α7 nAChRs and β-amyloid peptides have been shown to interact in vitro and in vivo, including human post-mortem AD brain. This review will begin with a brief overview of the basal forebrain cholinergic system, followed by a discussion of the current knowledge of the cholinergic deficit in Alzheimer's disease, then a summary of the cholinergic phenotype observed in transgenic Alzheimer's disease mouse models. We will also present our recent findings that support our hypothesis that the α7 nicotinic acetylcholine receptor performs both the neurotrophic and neuroprotective roles in the maintenance of the cholinergic phenotype and discusses potential mechanisms and implications for Alzheimer's disease therapy.     

Oral administration of idebenone induces nerve growth factor in the brain and improves learning and memory in basal forebrain-lesioned rats

Nerve growth factor plays an important role in the survival and maintenance of cholinergic neurons in the central neuronal system. In senile dementia of the Alzheimer type, learning and memory are impaired by the loss of neurons in the magnocellular cholinergic neuronal system. It is, therefore, of interest to investigate the role of nerve growth factor in this degenerative disorder. Since nerve growth factor does not cross the blood-brain barrier and is easily metabolized by peptidases when administered peripherally, it can be used for medical treatment only when directly injected into the brain.We demonstrate here that the oral administration of idebenone, a potent in vitro nerve growth factors synthesis stimulator, induced an increase in nerve growth factor protein and mRNA, and in choline acetyltransferase activity, in basal forebrain lesioned rats, but not in intact rats. Idebenone also ameliorated the behaviroral deficits in habituation, water maze, and passive avoidance tasks in these animals.These results suggest that idebenone stimulated nerve growth factor synthesis in vivo and ameliorates the behavioral defitics which were accompanied with the recovery of the reduced choline acetyltransferase activity in the basal forebrain-lesioned rats. Correspondence to. T. Nabeshima at the above address     

神经生长因子治疗阿尔采末病的胆碱能神经机制

神经生长因子 (nervegrowthfactor,NGF)是中枢胆碱能神经元存活和功能维持最重要的神经营养因子之一 ,对阿尔采末病 (Alzheimersdisease,AD)的治疗潜力已引起人们极大兴趣。投射于大脑皮质和海马的基底前脑胆碱能神经元退变是AD早期病变 ,也是导致患者认知功能降低的主要原因。NGF可通过兴奋残存神经元上高亲和性TrkA受体 ,促进中枢胆碱能神经元的存活和正常功能的发挥 ,同时神经元激活也使其自身免受AD的有害作用 ,即所谓“useitorloseit”现象。然而 ,NGF不能透过血脑屏障 ,如何使外源性NGF到达脑内靶区是亟待解决的难题 ,一旦获得理论和技术上的突破 ,NGF防治AD的临床应用才更具价值     

Reduced [125I]-Bolton Hunter substance P binding (NK1 receptors) in the basal forebrain nuclei of aged rats

1. Quantitative autoradiography and homogenate radioligand binding of [125I]-Bolton Hunter substance P ([125I]-BHSP) were used to compare brain NK1 receptors in young (2 months) and aged (18-20 months) rats. 2. The autoradiographic distribution and density of [125I]-BHSP binding sites was similar in all cortical regions of young and aged rats. In contrast, the density of [125I]-BHSP binding sites was significantly (P < 0.05) lower in the basal forebrain nuclei (intermediate part of the lateral septal nuclei, medial septal nucleus and horizontal and vertical nuclei of the diagonal band) of aged rats. In all other brain regions examined, binding densities were almost identical in young and aged rats. 3. Because a population of NK1 receptors ([125I]-BHSP binding sites) in the basal forebrain nuclei is associated with cholinergic neurons, the decrease in NK1 receptors in aged rats may reflect degeneration of cholinergic neurons and contribute to the motor and cognitive deficiencies that occur with ageing.     

慢性低灌注对大鼠基底前脑胆碱能系统及认知功能的影响

目的探讨慢性低灌注状态对大鼠基底前脑胆碱能神经系统及认知功能的影响。方法永久结扎SD系大鼠双侧颈总动脉建立大鼠前脑慢性低灌注状态模型,于1、2及4个月时观察基底前脑组织病理学变化、应用免疫组织化学方法检测基底前脑胆碱乙酰转移酶(Cholineacetyltransferase,ChAT)的表达水平、应用改良的MG-2型“Y”型迷宫检测大鼠学习和记忆能力。结果与对照组相比,术后第1、2和4个月时,基底前脑神经元和胶质细胞结构紊乱、呈片状和灶性坏死,ChAT阳性神经元及纤维显著减少(P<0.01),并随时间延长逐渐加重;大鼠全天总反应时间(Totalreactiontime,TRT)明显延长(P<0.01);并且以上两者呈显著负相关(r=-0.83、P<0.01)。结论慢性低灌注状态时,大鼠基底前脑发生缺血性病理改变,导致基底前脑胆碱能神经系统损害,从而在整体水平出现学习记忆障碍。低灌注状态时,基底前脑胆碱能神经系统损伤是认知功能障碍形成的重要机制。     

The cholinergic deficit in Alzheimer's disease: impact on cognition, behaviour and function

Although the neurodegeneration occurring in Alzheimer's disease (AD) affects multiple neurotransmitters, the cholinergic system has received the greatest attention. Acetylcholine (ACh) is fundamental to mnemonic function, assisting in the septal hippocampal pathway and facilitating cortical activation. One of the earliest pathological events in AD is the degeneration of ACh-synthesizing neurons in the subcortical nuclei of the human basal forebrain. Indeed, the loss of cholinergic function in AD is correlated with the density of histopathological markers of AD, the severity of cognitive dysfunction and disease duration. However, the precise mechanism by which the cholinergic system influences cognition, and behaviour, is unknown. Recent preliminary data from functional imaging and ligand-binding studies implicate a dynamic interaction between the nicotinic-muscarinic cholinergic receptor systems. The relatively preserved thalamic nicotinic system, compared with the dysfunctional cortical muscarinic system, may facilitate thalamocortical metabolic excitation in the failing AD brain. Thus, it is hypothesized that thalamic influence within frontal-subcortical circuits is augmented in AD patients who demonstrate a marked improvement to cholinesterase inhibitor therapy. Understanding the cholinergic basis of the cognitive, functional and behavioural deficits in AD, and the differential treatment response to various agents, will ultimately improve patient care and neuropharmacological insights. This paper reviews the current understanding of the cholinergic influence in cognition, behaviour and, as a result, function in AD patients.     

Neuronal nicotinic acetylcholine receptors serve as sensitive targets that mediate β-amyloid neurotoxicity

Alzheimer's disease (AD) is the most common form of brain dementia characterized by the accumulation of beta-amyloid peptides (Abeta) and loss of forebrain cholinergic neurons. Abeta accumulation and aggregation are thought to contribute to cholinergic neuronal degeneration, in turn causing learning and memory deficits, but the specific targets that mediate Abeta neurotoxicity remain elusive. Recently, accumulating lines of evidence have demonstrated that Abeta directly modulates the function of neuronal nicotinic acetylcholine receptors (nAChRs), which leads to the new hypothesis that neuronal nAChRs may serve as important targets that mediate Abeta neurotoxicity. In this review, we summarize current studies performed in our laboratory and in others to address the question of how Abeta modulates neuronal nAChRs, especially nAChR subunit function.