References

Lee R, Gallegos RA, Crawford EF, Wills DN, Zhukov VI, Huitron-Resendiz S, Criado JR, Henriksen SJ (2003) Ventral tegmental area lesions alter EEG power spectrum across the sleep/wake cycle. Neuroscience 2003 Abstracts 616.5. Society for Neuroscience, New Orleans, LA.

Summary: The ventral tegmental area (VTA) has long been implicated in reward and drug abuse. We previously demonstrated (Lee et al, J. Neurosci. 2001) a role for VTA GABAergic neurotransmission in REM sleep. In continuing studies the potential role of the VTA in modulating electroencephalogram (EEG) activation was explored by selectively lesioning mu-opioid receptor expressing cells, or NMDA-lesioning cells, in the VTA. Under sodium pentobarbital anesthesia rats received either (1) a sham operation (2) a saporin injection (3) an injection of a dermorphin-saporin (DERM-SAP) conjugate (Advanced Targeting Systems, San Diego, CA) (4) or a bilateral VTA injection of NMDA. All injections were delivered in a volume of 0.5 to 1.0 µL over 4 to 8 minutes. Animals were also implanted with electrodes for recording the EEG & EMG. The filtered EEG & EMG were recorded continuously for 24 hours beginning 21 days after surgery. Frequency analysis of the EEG in 15-sec epochs revealed differences in the distribution of relative power in the DERM-SAP or NMDA-lesioned animals, compared to controls. Higher frequency components (12-25 Hz) were reduced in DERM-SAP lesioned animals during waking and slow wave sleep. Histology demonstrated gliosis of GAD-stained neurons in the VTA 3 to 4 weeks after injection of DERM-SAP. These data suggest that long-projecting GABA neurons of the VTA have a desynchronizing influence on cortical EEG arousal mechanisms. This is supported by anatomical evidence of both direct and indirect non-thalamic GABAergic projections to widespread areas of cortex in the rodent. Supported by: DA08301 to SJH.

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Reynolds JN, Sutherland JM, Sutherland RJ (2003) A novel nitrate ester enhances performance in a spatial memory task in rats with forebrain cholinergic depletion. Neuroscience 2003 Abstracts 626.11. Society for Neuroscience, New Orleans, LA.

Summary: Forebrain acetylcholine (ACh) depletion is associated with a variety of cognitive problems, including memory deficits. Here we evaluate the efficacy of a novel nitrate ester, GT 1061, a potential cognition enhancer, in reversing the memory deficit produced by ACh depletion. Long-Evans hooded rats were stereotaxically injected with 192-IgG-saporin intraventricularly or into basal forebrain cell regions to effect loss of cholinergic cells in the basal forebrain and depletion of cholinergic input to neocortex and hippocampus. The rats were postoperatively tested in a version of the Morris water task in which the location of the hidden platform was changed every second day. This version allows for repeated testing of new spatial learning and 24-hr memory retention. ACh depletion causes statistically reliable deficits in new learning and retention components of the task. We examined the effects of oral (0.5, 1, 5, and 10 mg/kg) and intraperitoneally injected (1, 10, 25 and 50 mg/kg) GT 1061 and donepezil oral (0.05, 0.5, 0.1, 1 mg/kg) and intraperitoneal (0.5 mg/kg) on performance of ACh depleted rats. Both oral and injected GT1061 (10 mg/kg oral, and 1 mg/kg i.p.) and donepezil improved new learning and retention performance. The improvement was especially evident during the 24-hr retention tests when GT1061 treated rats performed as well as normal rats. On this measure 10 mg/kg GT 1061 and 1 mg/kg donepezil administered orally were equipotent.

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Dudkin KN, Chueva IV, Makarov FN, Beach TG, Roher AE (2003) Impairments of working memory processes on a model of Alzheimer’s disease in monkeys. Neuroscience 2003 Abstracts 626.8. Society for Neuroscience, New Orleans, LA.

Summary: We have investigated the characteristics of visual working memory in a delayed-discrimination task in a model of Alzheimer’s disease (AD) in rhesus monkeys. Three animals received unilateral stereotaxic intracerebroventricular injection of the nucleus basalis of Meynert and three monkeys received sterile saline injections and thus served as controls. The lesioning agent consisted of a ribosomal toxin, saporin, conjugated to monoclonal antibodies against (the nbM lesion) the p75 neurotrophin receptor (p75NTR), which is expressed almost exclusively on cholinergic neurons of the nbM. The rationale for the model is the same as for a rabbit model of AD (Roher et al, Ann. NY Acad Sci. 2000). The monkeys were trained to discriminate stimuli with different types of visual information (spatial frequency gratings, color, spatial choice, spatial relationships between components of objects). The data obtained demonstrate that the nbM lesioning agent had a weak effect on visual differentiation without delay (long-term memory), but significantly decreased the duration of information storage (by a factor of 2 – 3) in working memory later two months after injection. These changes depended on temporal stage after injection and stimulus properties, and were accompanied by increase of motor reaction time and of refusal of task decision. In monkeys that were sham injected, there were no alterations in working memory characteristics. The results suggest that considerable worsening of the working memory characteristics for monkeys after lesion of the nbM reflects the formation of an AD model in these monkeys. The principles of functional organization of working memory and role of pathology of the cortical mechanisms in an impairment of memory characteristics are discussed.

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Kim I, Wilson RE, Wellman CL (2003) Cholinergic deafferentation increases expression of the glur-1 subunit of the AMPA receptor in frontal cortex of young adult but not aging rats. Neuroscience 2003 Abstracts 633.9. Society for Neuroscience, New Orleans, LA.

Summary: Previously, we demonstrated that plasticity of frontal cortex is altered in aging rats: cholinergic lesions of the nucleus basalis magnocellularis (NBM) produce larger declines in dendritic morphology in frontal cortex of aged rats compared to young adults. In addition, these lesions result in upregulation of dendritic spines in frontal cortex of young adult but not aging rats. To begin to identify possible mechanisms underlying age-related differences in plasticity after NBM lesion, we assessed immunohistochemical labeling of the AMPA receptor subunit GluR1 in young adult and aging rats after either sham or 192 IgG saporin lesions of the NBM. Young adult (N=17), middle-aged (N=16), and aged rats (N=13) received unilateral sham or 192 IgG saporin lesions of the NBM. Two weeks later, brains were processed for immunohistochemical labeling of the GluR1 subunit of the AMPA receptor. An unbiased stereological technique was used to estimate density of labeled neurons in layer II-III of frontal cortex. Cells were identified as neurons based on standard morphological criteria and counted. Using a computerized image analysis system interfaced with a microscope, the average optical density of the white matter below frontal cortex was determined; neurons with optical densities at least one standard deviation above this mean were identified as intensely labeled. In young adult rats, lesions produced a 55% increase in the density of intensely GluR1-immunopositive neurons in frontal cortex. On the other hand, lesions had no effect on counts of GluR1-immunoreactive neurons in middle-aged and aged rats. This age-related difference in lesion-induced expression of AMPA receptor subunit protein could underlie the age-related differences in dendritic plasticity after NBM lesions.

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Fletcher BR, Baxter MG, Rapp PR, Shapiro ML (2003) 192-IgG saporin lesions of the medial septum and vertical diagonal band impair cognitive flexibility. Neuroscience 2003 Abstracts 425.8. Society for Neuroscience, New Orleans, LA.

Summary: Learning and memory remain largely intact following selective basal forebrain cholinergic lesions. By comparison, single unit recording studies have documented reliable effects of such lesions, including abnormally rigid hippocampal place fields when animals are confronted with changes in the configuration of the testing environment. The present experiment tested the prediction that cholinergic lesions of the basal forebrain would impair performance of tasks requiring cognitive flexibility. Rats received 192-IgG saporin or control vehicle injections into the medial septal nucleus and vertical diagonal band, and were tested on cued and spatial delayed match-to-place tasks in a radial arm water maze. Test sessions consisted of four sample trials in which animals searched for a cued or hidden escape platform located in a fixed position at the end of one arm (60 sec cutoff, inter-trial interval = 15 sec). A memory delay was imposed by returning rats to the home cage for a variable delay (15 sec. – 6 hrs), followed by two test trials. The lesion and control groups learned at similar rates in both versions of the task, and performed comparably on the critical test trials, independent of the length of the retention interval. However, lesioned rats were impaired during the transition from the cued to spatial variants of testing. Specifically, the lesion group made significantly more errors on an early sample trial in the spatial task, returning to the location that was previously correct during cued training. Pending histological confirmation of the extent and selectivity of the experimental lesions, this pattern of results suggests that damage to the basal forebrain cholinergic system spares spatial learning but impairs cognitive flexibility when task contingencies are changed.

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Nelson LE, Franks NP, Maze M (2003) Discrete lesioning of orexin (hypocretin)-containing neurons potentiates dexmedetomidine- but not pentobarbital-induced hypnosis. Neuroscience 2003 Abstracts 426.13. Society for Neuroscience, New Orleans, LA.

Summary: Introduction: Recent work suggests that anesthetics putatively modulated by the α2-adrenoceptor (e.g. dexmedetomidine (DEX)) or the GABAA receptor (e.g. pentobarbital (PTB)) elevate and depress c-Fos expression, respectively, in the orexinergic perifornical area (PeF)1. Here the hypnotic effects of DEX and PTB are assessed after selective lesion of the PeF by orexin-B conjugated to saporin (OX-SAP). Methodology: Anesthetized Fischer rats were administered stereotaxic PeF injections of saline (0.5µl/side) or OX-SAP (490ng/0.5μl/side; as described2). Loss of righting reflex (LORR) induced by DEX (150μg/kg, SC), PTB (50mg/kg, SC), and saline was tested 1 day pre- and 1, 4, 8, and 12 days post-surgery, then lesions were assessed histologically. All data are presented as means±SEMs (n=6; comparisons by unpaired t-tests and ANOVA, Newman-Keuls). Results: Bilateral PeF lesions enhanced DEX-induced hypnosis at days 8 (281.2±15.8 min; p<0.05) and 12 (322.7±15.93 min; p<0.001) as compared to naïve (234.5±9.0 min) and saline sham animals (day 8, 236.7±9.8 min; day 12, 242.8±10.80 min). In contrast, PTB-induced LORR remained unaffected at day 12 (124.8±6.8 min) relative to naïve (119.4±4.6 min) and sham (120.8±5.3 min). These results agree with previous reports that by day 12, PeF-microinjected OX-SAP induces roughly 80% cell loss2. Conclusion: The absence of a functional PeF potentiates hypnosis induced by DEX but not PTB, as perturbation of the PeF by microinjected GABAA receptor antagonist gabazine is known to 1. References: 1 Nelson et al. (2002) SfN abstract 776.14/M20; 2 Geraschenko et al. (2001) J Neurosci 21:7273-83.

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Sherren N, Pappas BA (2003) Dendritic dysgenesis in midline cortical regions following selective acetylcholine and dopamine lesions in neonatal rats. Neuroscience 2003 Abstracts 457.11. Society for Neuroscience, New Orleans, LA.

Summary: Both acetylcholine (ACh) and dopamine (DA) afferents reach their cortical targets during periods of synaptogenesis, and are perfectly positioned to influence the cytoarchitectural development of cortical neurons. Thus the behavioural outcomes of these lesions may be related to the development of appropriate dendritic morphology in neurons from cortical regions involved in cognition. Previous studies have either used non-specific lesion techniques or have not examined long-term effects. We lesioned rat pups at P7 with either 600 ng of the selective immunotoxin 192 IgG-saporin, or 150 ug of 6-hydroxydopamine preceded by desmethylimipramine, or both, and aged them to four months. One squad of rats was sacrificed for neurochemistry and another was prepared for morphological analysis using Golgi-Cox stain. The ACh lesion caused a 32% decrease in choline acetyltransferase activity in the frontal/cingulate cortex and a 72% reduction in retrosplenial cortex (RSC). This was associated with reductions in total dendritic length of the apical tree of layer V pyramidal cells in the medial prefrontal cortex (mPFC), the apical tree of layer III pyramidal cells in the anterior cingulate cortex (ACC), and the basal tree of layer III pyramidal cells in RSC. The DA lesion caused a 76% reduction in DA levels in frontal/cingulate cortex and no change in RSC levels. This was associated with reductions in total dendritic length of the basal and apical trees of layer V pyramidal cells in mPFC, and the basal tree of layer III pyramidal cells in ACC. No changes in layer III pyramidal cells were noted in RSC following the DA lesion. These data demonstrate that ascending ACh and DA afferents play a vital role in the cytoarchitectural development of the cortex. This is particularly important considering that hypofunction in these systems is a characteristic feature of neurodevelopmental disorders involving mental retardation, such as Rett and Down syndrome.

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Neale JJ, Goodchild AK, Dampney RAL, Pilowsky PM (2003) Destruction of brainstem catecholamine neurons attenuates somatosympathetic reflex and responses to cholecystokinin. Neuroscience 2003 Abstracts 501.11. Society for Neuroscience, New Orleans, LA.

Summary: The integrity of the rostral ventrolateral medulla (RVLM) is essential for the expression of many sympathetic reflexes and the maintenance of vasomotor tone. The RVLM contains bulbospinal neurons, of which about half are catecholaminergic (C1). Destruction of bulbospinal C1 neurons leads to attenuation or abolition of the sympathetic baroreflex and chemoreflex, respectively. This study examines the effects of such destruction on blood pressure (BP), the somatosympathetic reflex and responses to intravenous (i.v) cholecystokinin (CCK) in urethane-anaesthetised, paralysed and ventilated Sprague-Dawley rats. Eighty percent of the spinally projecting C1 neurons in the RVLM were destroyed by bilateral microinjections of the immunotoxin, anti-DBH-saporin (12ng/100nl), into the intermediolateral cell column of the thoracic spinal cord (T1-2). Following treatment with the neurotoxin, systolic BP was measured for 3-5 weeks before testing the reflexes. No significant changes in systolic BP were observed. In the present study destruction of bulbospinal C1 neurons attenuated the baroreflex, replicating the findings of Schreihofer and Guyenet (2000, Am J Physiol 279:R729-R742). Activation of the somatosympathetic reflex by electrical stimulation of the tibial nerve normally elicits two peaks in averaged splanchnic sympathetic nerve activity. Following destruction of C1 neurons, the threshold voltage was reduced and the second peak was either markedly attenuated or abolished at two times threshold voltage. Intravenous injection of CCK (1, 10 and 100mg/kg) elicited depressor and sympathoinhibitory responses that were significantly reduced following destruction of bulbospinal C1 neurons. These results demonstrate a key role of bulbospinal C1 neurons in the somatosympathetic reflex and the sympathetic responses to i.v CCK but not in the tonic control of blood pressure.

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Recabarren MP, Valdes JL, Seron-Ferre M, Torrealba F (2003) Tuberomammillary nucleus lesion decreases the anticipatory events induced by restricted feeding in rats. Neuroscience 2003 Abstracts 510.19. Society for Neuroscience, New Orleans, LA.

Summary: Our previous studies indicate that the histamine-containing neurons of the tuberomammillary nucleus (TMN) become active in anticipation to feeding time in rats under a restricted feeding schedule. To assess the role of the TMN in this anticipatory activity in rats, we lesioned the TMN bilaterally with stereotaxic injections of 50ng ORX-SAP (Advanced Targeting System, CA). We analyzed the locomotor activity, core temperature and the feeding frequency exhibited by these animals during a restricted feeding protocol, where food was available between 10:00 h and 12:00 h for at least 2 weeks. Rats were implanted in the abdominal cavity with telemetric sensors (Minimitter, OR) to measure locomotor activity and core temperature. During the whole experiment rats were maintained in individual cages and under controlled photoperiod of 12 hours light and 12 hours dark, light were on at 07:00 h. We analyzed the 3 hours preceding food arrival. We checked the extent of TMN destruction by immunostaining the brain sections with antibody against adenosine diaminase (ADA), which colocalize with histaminergic neurons in the TMN. Control rats were subjected to the same procedures except for the injection of the ORX-SAP toxin. Results: Lesion rats showed a significant decrease in the number of ADA-ir neurons in the TMN, as well as a decreased anticipatory activity under restricted feeding in comparison with control rats. Lesion rats although awake before food arrival, were less eager to feed compared to controls, as assessed by food bin approaches. Control rats were slightly more active than lesion rats during restriction. In conclusion, the functional integrity of the TMN is required for the full expression of the anticipatory events that are stimulated by a restricted feeding schedule.

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Hunter CL, Quintero EM, Gilstrap L, Bhat NR, Granholm AE (2003) Neuroinflammatory response to mu p75-saporin immunotoxin-induced degeneration of basal forebrain cholinergic neurons. Neuroscience 2003 Abstracts 527.15. Society for Neuroscience, New Orleans, LA.

Summary: Basal forebrain cholinergic neurons, which provide the major cholinergic innervation to the cortical regions and play a key role in the processing of information involved in cognitive processes, degenerate during both normal aging and Alzheimer’s disease. Neuroinflammation, specifically the activation of microglia, is known to affect the progression of neuronal degeneration. Activated microglia produce inflammatory mediators that have neuropathic as well as neuroprotective actions, and it has been suggested that inflammatory mediators produced by activated microglia may play a role in the decline of specific neuronal sub-types in neurodegenerative diseases. The immunotoxin mu p75-SAP has been shown to selectively destroy cholinergic neurons in the basal forebrain of mice, resulting in reduced choline acetyl-transferase activity and cognitive impairments. To characterize the inflammatory response to mu p75-SAP lesions, 3 month-old mice received icv injections of mu p75-SAP (3.6 mg) followed by treatment with an anti-inflammatory agent, minocycline (45 mg/kg i.p.), or saline. Seven days after lesioning, immunohistochemistry was used to analyze markers for cholinergic and non-cholinergic neurons and inflammation. Cholinergic lesioning resulted in a dramatic increase in CD45, a microglial marker, but no change in GFAP, an astroglial marker, in the basal forebrain region. Lesioned animals had elevated levels of phosphorylated p38, a MAP kinase protein involved in inflammatory pathways. Minocycline treatment reduced this inflammatory response. Furthermore, preliminary results suggest that animals treated with minocycline after mu p75-SAP lesioning are partially protected from cholinergic degeneration.

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