Lesions of the nucleus basalis magnocellularis (Meynert) induce enhanced somatosensory responses and tactile hypersensitivity in rats.

We used the immunotoxin 192 immunoglobulin G-saporin to produce a selective cholinergic lesion in the nucleus basalis of Meynert (NBM) of rats and investigated whether the NBM lesion led to tactile hypersensitivity in the forepaw. The paw mechanical threshold test showed that the lesioned rats had a decreased threshold compared to the control. Surprisingly, there was a significant positive correlation between mechanical threshold and survival rate of NBM cholinergic neurons. Furthermore, using local field potential (LFP) recordings and voltage-sensitive dye (VSD) imaging, we found that the forepaw-evoked response in the primary somatosensory cortex (S1) was significantly enhanced in both amplitude and spatial extent in the NBM-lesioned rats. The neurophysiological measures of S1 response, such as LFP amplitude and maximal activated cortical area depicted by VSD, were also correlated with withdrawal behavior. Additional pharmacological experiments demonstrated that forepaw-evoked responses were increased in naive rats by blocking S1 cholinergic receptors with mecamylamine and scopolamine, while the response decreased in NBM-lesioned rats with the cholinergic agonist carbachol. In addition, NBM burst stimulation, which facilitates acetylcholine release in the S1, suppressed subsequent sensory responses to forepaw stimulation. Taken together, these results suggest that neuronal loss in the NBM diminishes acetylcholine actions in the S1, thereby enhancing the cortical representation of sensory stimuli, which may in turn lead to behavioral hypersensitivity.

The effects of NBM- lesion on synaptic plasticity in rats.

The nucleus basalis magnocellularis (NBM) is a major source of cholinergic projections to the neocortex that is vulnerable to degeneration in Alzheimer's disease. Despite numerous anatomical, pharmacological, behavioral, and physiological investigations of NBM, there is no in vivo study of its effects on short- or long-term synaptic plasticity. Hence, this study was devoted to the assessment of the effects of bilateral lesion of the NBM on synaptic plasticity in the dentate gyrus of the hippocampus using electrophysiological techniques. For this purpose, twenty-five male Wistar rats were randomly allocated into the three Control, Sham, and NBM-lesioned groups. Lesion was made via bilateral injections of 5µg/µl ibotenic acid. After twenty-one days, the input-output functions, paired-pulse facilitation/inhibition, and long-term potentiation (LTP) were evaluated in the dentate gyrus while the perforant pathway was stimulated. NBM lesion was found to attenuate the basal synaptic responsiveness, paired-pulse responses, and LTP in the rats' dentate gyrus, indicating that lesions of this cholinergic nucleus affects both short- and long-term neural plasticity in the dentate gyrus although NBM does not send direct cholinergic projections to the hippocampus.

Event-related oscillations (ERO) during an active discrimination task: Effects of lesions of the nucleus basalis magnocellularis.

The cholinergic system in the brain is involved in attentional processes that are engaged for the identification and selection of relevant information in the environment and the formation of new stimulus associations. In the present study we determined the effects of cholinergic lesions of nucleus basalis magnocellularis (NBM) on amplitude and phase characteristics of event related oscillations (EROs) generated in an auditory active discrimination task in rats. Rats were trained to press a lever to begin a series of 1kHz tones and to release the lever upon hearing a 2kHz tone. A time-frequency based representation was used to determine ERO energy and phase synchronization (phase lock index, PLI) across trials, recorded within frontal cortical structures. Lesions in NBM produced by an infusion of a-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) resulted in (1) a reduction of the number of correct behavioral responses in the active discrimination task, (2) an increase in ERO energy in the delta frequency bands, (3) an increase in theta, alpha and beta ERO energy in the N1, P3a and P3b regions of interest (ROI), and (4) an increase in PLI in the theta frequency band in the N1 ROIs. These studies suggest that the NBM cholinergic system is involved in maintaining the synchronization/phase resetting of oscillations in different frequencies in response to the presentation of the target stimuli in an active discrimination task.

Imidafenacin has no influence on learning in nucleus basalis of Meynert-lesioned rats.

The prevalence of overactive bladder (OAB) and Alzheimer's disease (AD) increases with age, and much attention has been paid to the risk of cognitive impairment which may be induced by antimuscarinics used for OAB in patients with AD. Imidafenacin, an antimuscarinic agent for OAB treatment, has been reported not to affect learning in normal animals. However, under the condition in which sensitivity to learning impairment by antimuscarinics is increased, it remains unclear whether imidafenacin still does not impair the learning. Therefore, the influences of imidafenacin on passive avoidance response were investigated in sham-operated and nucleus basalis of Meynert (nbM)-lesioned rats and compared with oxybutynin hydrochloride and tolterodine tartrate. The learning-inhibitory doses of intravenous oxybutynin hydrochloride and tolterodine tartrate were 0.3 and 3 mg/kg in sham-operated rats and 0.1 and 1 mg/kg in nbM-lesioned rats, respectively. Thus, the learning impairments by those antimuscarinics were more sensitive in nbM-lesioned rats than in sham-operated rats. On the other hand, intravenous administration of imidafenacin had no influence on learning in either case of the rats. In normal rats, however, intracerebroventricular administration of imidafenacin impaired learning to the same degree as that of oxybutynin hydrochloride. Thus, the present study suggests that imidafenacin, unlike the other antimuscarinics used, has no significant risk of enhancing learning impairment even in models whose sensitivity to learning impairment by antimuscarinics is commonly increased, probably because of its low brain penetration.

High volume microinfusion suppresses local astrocyte response within nucleus basalis of rat.

Our study investigates the impact of different volume sham control and excitotoxin microinfusions in vivo on local reactive astroglial response within rat nucleus basalis (NB). We followed the effects of unilateral 200, 100, and 50 nL of sham-control (phosphate buffer PBS) versus ibotenic acid (IBO) microinfusions, mechanical NB lesion (10 µL Hamylton syringe needle positioned into NB for 5 min), or physiological control (intact brain), on the local reactive astroglial response within the NB site, by immunoreactivity against glial fibrillary acidic protein (GFAP). NB lesions were identified by NADPHdiaphorase histochemistry. Local astrocytes responses within NB were suppressed by both high volume microinfusions, PBS and IBO (200 and 100 nL) versus mechanical lesion. Our study has proved, for the first time, the volume of microinfusion as critical for any selective pharmacological stimulation or lesion in vivo, and suggest the microinfusion volume less than 50nL as protective for physiological astroglial reactivity.

Cholinergic deafferentation of the neocortex using 192 IgG-saporin impairs feature binding in rats.

The binding problem refers to the fundamental challenge of the CNS to integrate sensory information registered by distinct brain regions to form a unified neural representation of a stimulus. Although the human cognitive literature has established that attentional processes in frontoparietal cortices support feature binding, the neurochemical and specific downstream neuroanatomical contributions to feature binding remain unknown. Using systemic pharmacology in rats, it has been shown that the neuromodulator acetylcholine is essential for feature binding at encoding, but the neural source of such critical cholinergic neurotransmission has yet to be identified. Cholinergic efferents from the nucleus basalis magnocellularis (NBM) of the basal forebrain provide the majority of the cholinergic input to the neocortex. Accordingly, it was hypothesized that the NBM is the neural source that provides the critical neuromodulatory support for feature binding. To test this hypothesis, rats received bilateral 192 IgG-saporin lesions of the NBM, and their feature binding performance was tested using a forced-choice digging paradigm. Relative to sham-lesioned rats, NBM-lesioned rats were significantly impaired at acquiring a crossmodal feature conjunction (FC) stimulus set that required feature binding, whereas their ability to retrieve an FC stimulus set and to acquire two crossmodal feature singleton stimulus sets, one of greater difficulty than the other but neither requiring feature binding, remained intact. These behavioral findings, along with histological analyses demonstrating positive relationships between feature-binding acquisition and markers of cholinergic activity in frontoparietal regions, reveal the importance of neocortical cholinergic input from the NBM to feature binding at encoding.

The nucleus basalis magnocellularis contributes to feature binding in the rat.

The binding problem refers to the fundamental challenge of the central nervous system to integrate sensory information registered by multiple brain regions to form a unified neural representation of a stimulus. Although the human cognitive literature has yielded substantial insights into the attention-dependent nature and general cortical networks involved in feature binding, the specific downstream neuroanatomical modulatory contributions to feature binding remain unknown. We hypothesized that the nucleus basalis magnocellularis (NBM) of the basal forebrain would be critical for feature binding given the NBM's widespread neuromodulatory projections to regions of the neocortex important for attentional processing, such as the frontal and parietal cortices. Accordingly, we tested the ability of rats with bilateral excitotoxic (quisqualic acid) lesions of the NBM to acquire a crossmodal Feature-Conjunction (FC) task that required feature binding and a Feature-Singleton (FS) task that did not require feature binding. Additionally, rats retrieved a FC stimulus set they had acquired prior to surgery. Relative to sham-lesioned controls, NBM-lesioned rats were significantly impaired at acquiring and retrieving the FC task, while their ability to acquire the FS task remained intact. These findings provide insight into the functional role of the NBM and establish the importance of this basal forebrain structure to the fundamental cognitive process of feature binding.

Nucleus basalis magnocellularis and substantia innominata corticopetal cholinergic lesions attenuate freezing induced by predator odor.

Previous studies have demonstrated that corticopetal cholinergic lesions applied to the nucleus basalis magnocellularis and substantia innominata (NBM/SI) attenuate operant suppression induced by aversive events. However, these lesions have no effect on open-arm behavior in the elevated plus-maze or changes in startle reactivity induced by bright light. This raises the possibility that NBM/SI corticopetal cholinergic lesions alter operant behavior and/or appetitive state, as opposed to the aversive state operant suppression is supposed to index. To address this concern, the authors documented the effect of NBM/SI corticopetal cholinergic lesions on freezing induced by a component of fox feces (2,4,5-trimethylthiazoline [TMT]), a paradigm that does not involve food deprivation or operant performance. TMT presentation induced freezing behavior, and this effect was attenuated by NBM/SI corticopetal cholinergic lesions. Because predator odor presentation, but not presentation of a predator, induces defense behaviors that are sensitive to anxiolytic drugs, the results of the study suggest that NBM/SI corticopetal cholinergic lesions attenuate anxiety-like states.

Attenuation of the blood flow response to physostigmine in the rat cortex deafferented from the basal forebrain.

Previous functional investigations in rats failed to demonstrate that the classical cholinesterase inhibitor, physostigmine, can compensate for cortical cholinergic deficit induced by deafferentation from the nucleus basalis magnocellularis (NBM). As these studies were carried out shortly after NBM lesion (1-2 weeks), we sought to determine whether compensatory effects of physostigmine would appear at a longer postlesion time (3-5 weeks). Cerebral blood flow was used as a quantitative measure of brain function. At 3-5 weeks after unilateral NBM lesion, interhemispheric comparisons in resting conditions showed that the cortical cholinergic deficit was still present and that blood flow was lower in cortical areas on the lesion side, similarly to what was observed after 1-2 weeks, while basal blood flow in intact hemispheres remained unchanged. In contrast, under physostigmine, blood flow became significantly lower in deafferented cortical areas at 3-5 weeks postlesion time, whereas there were no significant interhemispheric differences in the short term. Comparisons with saline-infused rats showed reduced blood flow responses to physostigmine in forebrain regions, e.g. in the parietal cortex from 83% to 25% at 1-2 and 3-5 weeks postlesion, respectively. These changes cannot be ascribed to a global loss of reactivity, since responses in brainstem regions (medulla, cerebellum) remained unchanged statistically. The results demonstrate a reduced responsiveness to physostigmine at the longer postlesion time, and support the existence of a cholinosensitive mechanism antagonizing NBM influence. This mechanism may limit the activating effects of cholinergic agonists in the forebrain after NBM deafferentation.

Lesions to the nucleus basalis magnocellularis lower performance but do not block the retention of a previously acquired learning set.

Rats were first trained to acquire an olfactory discrimination learning set (ODLS) on 40 olfactory-unique discrimination problems. Following acquisition of ODLS, animals were lesioned bilaterally in the nucleus basalis magnocellularis (nBM) using either quisqualic acid (QUIS) or 192 IgG-saporin (SAP). QUIS animals performed significantly worse than control animals following surgery and SAP animals performed transiently worse than control animals. Despite lowered performances, both QUIS and SAP animals performed significantly better than expected by chance on trial 2 indicating retention of the ODLS previously acquired. Implications for the role of the nBM in aspects of cognitive flexibility and its role in acquisition versus retention are discussed.

Effect of nucleus basalis magnocellularis cholinergic lesions on fear-like and anxiety-like behavior.

Previous research has suggested that cholinergic neurons in the nucleus basalis magnocellularis and substantia innominata (NBM/SI) may be important in mediating aversive states. The authors investigated the effect of NBM/SI cholinergic lesions, induced with 192 IgG saporin, on behavioral measures of aversive states in rats. Behavior in the elevated plus maze and behavioral suppression induced by 2 fear-conditioned stimuli, a tone and a light, were evaluated. Lesions had no effect on any measures in the elevated plus maze but attenuated operant suppression induced by the light and attenuated freezing induced by the tone, though this last effect was not statistically significant. The results of the study suggest that NBM/SI cholinergic neurons may be important in mediating selective aspects of aversive states.

Selective lesions of the nucleus basalis magnocellularis impair cognitive flexibility.

The authors tested the hypothesis that the cholinergic nucleus basalis magnocellularis (NBM) is involved in solving problems requiring cognitive flexibility. Rats with 192 IgG-saporin lesions of the NBM were assessed for perseveration (i.e., cognitive inflexibility) in the serial reversal of an operant discrimination and during subsequent extinction testing. It was hypothesized that the NBM lesion and control groups would not differ in the acquisition of the initial, simple discrimination, because this task does not demand cognitive flexibility. In contrast, it was hypothesized that the NBM lesion group would show perseveration during serial reversal and extinction testing. Results generally supported these hypotheses, suggesting that the NBM plays an important role in mediating cognitive flexibility.

Verapamil prevents, in a dose-dependent way, the loss of ChAT-immunoreactive neurons in the cerebral cortex following lesions of the rat nucleus basalis magnocellularis.

In the present study we analysed the neuroprotective effect of the L-type voltage-dependent calcium channel antagonist verapamil on cholineacetyltransferase (ChAT)-immunoreactive neurons in the cerebral cortex of rats with bilateral electrolytic lesions of the nucleus basalis magnocellularis (NBM). Treatment with verapamil (1.0, 2.5, 5.0 and 10.0 mg/kg/12 h i.p.) started 24 h after NBM lesions and lasted 8 days. Animals were sacrificed on day 21 after NBM-lesions. The bilateral NBM-lesions produced significant loss of ChAT-immunoreactive neurons in frontal, parietal and temporal cortex. Although the number of ChAT-positive neurons was significantly higher in NBM-lesioned animals treated with verapamil at a dose of 2.5, 5.0 and 10.0 mg/kg than in saline treated ones, the most significant effect was obtained at a dose of 5 mg/kg. This is, to our knowledge, the first report showing an inverted U-shape mode of neuroprotective action of the calcium antagonist verapamil, at morphological level in this particular model of brain damage. The demonstrated beneficial effect of verapamil treatment suggests that the regulation of calcium homeostasis during the early period after NBM lesions might be a possible treatment to prevent neurodegenerative processes in the rat cerebral cortex.

Ciliary neurotrophic factor is expressed in the magnocellular neurosecretory system of the rat in vivo: evidence for injury- and activity-induced upregulation.

Although ciliary neurotrophic factor (CNTF) has been shown to promote the survival of magnocellular neurons when applied exogenously to explants of the paraventricular and supraoptic nuclei (SON) in vitro, little is known regarding its expression or regulation in the adult magnocellular neurosecretory system (MNS) following injury in vivo. Therefore, we utilized in situ hybridization and immunocytochemical analysis in conjunction with quantitative optical densitometric analysis to identify the cellular source of CNTF and examine the temporal pattern of its expression, following unilateral transection of the hypothalamo-neurohypophysial tract in the adult rat. In intact rats, CNTF immunoreactivity (CNTF-ir) was predominantly localized within identified astrocytes within the ventral glial limitans subjacent to the SON. Quantitative optical densitometric analysis of CNTF-ir levels in the axotomized SON demonstrated that the proportional area of CNTF-ir was significantly elevated between 3 and 30 days following injury. A significant but more limited increase was also observed in the non-injured contralateral SON. In situ hybridization confirmed the expression and upregulation of CNTF in the axotomized SON. These results demonstrate the expression of CNTF in the adult rodent MNS in vivo and provide evidence that levels of CNTF are upregulated in response to both direct injury, and heightened metabolic activity, within the lesioned and sprouting SON, respectively.

Behavioral effects of basal forebrain cholinergic lesions in young adult and aging rats.

The interactive effects of age and cholinergic damage were assessed behaviorally in young and middle-aged rats. Rats were lesioned at either 3 or 17 months of age by injection of 192 IgG-saporin immunotoxin into the medial septum and the nucleus basalis magnocellularis, and they were then tested on a range of behavioral tasks: a nonmatching-to-position task in a T-maze, an object-recognition task, an object-location task, and an open-field activity test. Depending on the task used, only an age or a lesion effect was observed, but there was no Age X Lesion interaction. Middle-aged and young rats responded to the cholinergic lesions in the same manner. These results show that in the middle-aged rats in which cholinergic transmission was affected, additional injury to the system was not always accompanied by major cognitive dysfunctions.

Further analysis of the effects of immunotoxic lesions of the basal nucleus of Meynert reveals substantial impairment on visual discrimination learning in monkeys.

In this paper we undertake a combined analysis of several studies in which marmoset monkeys received immunotoxic lesions of the cortical cholinergic projections from the basal nucleus of Meynert (NBM) bilaterally and/or in combination with immunotoxic lesions of other parts of the cholinergic system or ablations of the target inferotemporal neocortical area. Analysis of the mean learning scores across all visual discriminations learning tasks for each lesion combination revealed highly significant impairments where the NBM was lesioned bilaterally or where an NBM lesion in one hemisphere was crossed with an inferotemporal cortical ablation in the other hemisphere. This demonstrates that the cholinergic projection from the NBM to the major target area of neocortex involved in visual discrimination learning, i.e. the inferotemporal cortex, makes an important contribution to the perceptuo-mnemonic processes necessary for this type of learning. A new study demonstrates a significant effect of a subtotal bilateral cholinergic lesion confined to the NBM on a concurrent object-reward association task using black objects which is perceptually and mnemonically demanding. These results do not preclude the possibility that cholinergic projections from the NBM to other parts of the neocortex make a contribution to other cortical functions which are not mnemonic. It is well established that lesions of the cholinergic projection from the diagonal band of Broca disrupts the mnemonic functions of the hippocampus. The results described here suggest that degeneration of the cholinergic projections in Alzheimer's disease and other dementias will contribute to the loss of those mnemonic functions which are dependent on the neocortex.

Decreased neurogenesis after cholinergic forebrain lesion in the adult rat.

Adult neurogenesis has been shown to be regulated by a multitude of extracellular cues, including hormones, growth factors, and neurotransmitters. The cholinergic system of the basal forebrain is one of the key transmitter systems for learning and memory. Because adult neurogenesis has been implicated in cognitive performance, the present work aims at defining the role of cholinergic input for adult neurogenesis by using an immunotoxic lesion approach. The immunotoxin 192IgG-saporin was infused into the lateral ventricle of adult rats to selectively lesion cholinergic neurons of the cholinergic basal forebrain (CBF), which project to the two main regions of adult neurogenesis: the dentate gyrus and the olfactory bulb. Five weeks after lesioning, neurogenesis, defined by the number of cells colocalized for bromodeoxyuridine (BrdU) and the neuronal nuclei marker NeuN, declined significantly in the granule cell layers of the dentate gyrus and olfactory bulb. Furthermore, immunotoxic lesions to the CBF led to increased numbers of apoptotic cells specifically in the subgranular zone, the progenitor region of the dentate gyrus, and within the periglomerular layer of the olfactory bulb. We propose that the cholinergic system plays a survival-promoting role for neuronal progenitors and immature neurons within regions of adult neurogenesis, similar to effects observed previously during brain development. As a working hypothesis, neuronal loss within the CBF system leads not only to cognitive deficits but may also alter on a cellular level the functionality of the dentate gyrus, which in turn may aggravate cognitive deficits.

Choline pivaloyl esters improve in rats cognitive and memory performances impaired by scopolamine treatment or lesions of the nucleus basalis of Meynert.

The effects of two choline pivaloyl esters, [2-(2,2-dimethylpropionyloxy)ethyl]trimethylammonium iodide (1) and [2-(2,2-dimethylpropionyloxy)ethyl]trimethylammonium 2,2-dimethylpropionate (2), on learning and memory impairments induced in rats by scopolamine or lesions of nucleus basalis magnocellularis (NBM) have been evaluated by object recognition and Morris water maze tests in comparison with Tacrine (THA). Both 1 and 2 restored discrimination in object recognition test for assessing working-episodic memory and improved spatial memory in scopolamine or NBM-lesioned rats as well. The positive effects produced by 1 and 2 on cognitive and memory deficits were well comparable with those evoked by THA, used as reference compound.

Parafascicular electrical stimulation attenuates nucleus basalis magnocellularis lesion-induced active avoidance retention deficit.

Previous experiments from our laboratory showed that retention of two-way active avoidance learning is improved by post-training intracranial electrical stimulation (ICS) of the parafascicular nucleus (PF) and impaired by pre-training electrolytic lesions of the nucleus basalis magnocellularis (NBM). The question investigated here was whether post-training PF ICS is able to attenuate the active avoidance retention deficit observed in rats lesioned pre-training in the NBM. To this goal, the following experimental design was used: rats bilaterally lesioned in the NBM and stimulated in the PF, rats lesioned in the NBM, rats stimulated in the PF, control rats implanted in the PF, and sham-operated rats were first trained in a shuttle-box for a single 30-trial session and tested again following two successive retention intervals (24 h and 11 days). The results showed that: (1) NBM lesions impaired the 11-day performance without affecting either the acquisition or the 24-h retention of the avoidance learning; (2) PF ICS treatment in unlesioned rats improved performance in both retention sessions only when the stimulation was applied in the posterior region of the nucleus; and (3) stimulation of the posterior PF compensated the 11-day retention impairment induced by NBM lesions. These results are discussed in relation to the interaction of arousal systems in the modulation of cognitive processes.