Leptin receptor-mediated regulation of cholinergic neurotransmitter phenotype in cells of central nervous system origin.
(49/969)
Leptin is an adipocyte-secreted hormone that regulates body weight and exerts effects on hematopoiesis, reproduction, and immunity. The leptin receptor (OBR) shares sequence similarity and signaling capabilities with receptors for cytokines of the ciliary neurotrophic factor (CNTF) family. Our previous finding that CNTF and leptin exert similar anti-obesity effects and activate common neuronal signaling pathways, prompted us to investigate whether leptin may share with CNTF the ability to regulate the expression of specific neuronal genes. To this end, we established a cell line, derived from the murine septal cholinergic neuronal cell line SN-56, which stably expresses OBR. In this cell line, termed SN-56/OBR, leptin induces STAT transcription factor activation and STAT-dependent reporter gene expression in a manner similar to that of CNTF. Furthermore, in SN-56/OBR cells both CNTF and leptin produce changes in neurotransmitter and neuropeptide phenotype characteristic of cholinergic neurons, such as an increase in choline acetyltransferase and vasoactive intestinal polypeptide, and a decrease in neuropeptide Y expression. SN-56/OBR cells thus constitute an interesting new model system to investigate leptin action in cells of central nervous system origin. Possible physiological implications of OBR's intrinsic ability to regulate cholinergic phenotypic markers are discussed. (+info)
Alzheimer-like neurodegeneration in aged antinerve growth factor transgenic mice.
(50/969)
Neurotrophin nerve growth factor (NGF) has been suggested to be involved in age-related neurodegenerative diseases, but no transgenic model is currently available to study this concept. We have obtained transgenic mice expressing a neutralizing anti-NGF recombinant antibody, in which the levels of antibodies are three orders of magnitude higher in adult than in newborn mice [F.R., S. C. , A.C., E. Di Daniel, J. Franzot, S. Gonfloni, G. Rossi, N. B. & A. C. (2000) J. Neurosci., 20, 2589-2601]. In this paper, we analyze the phenotype of aged anti-NGF transgenic mice and demonstrate that these mice acquire an age-dependent neurodegenerative pathology including amyloid plaques, insoluble and hyperphosphorylated tau, and neurofibrillary tangles in cortical and hippocampal neurons. Aged anti-NGF mice also display extensive neuronal loss throughout the cortex, cholinergic deficit in the basal forebrain, and behavioral deficits. The overall picture is strikingly reminiscent of human Alzheimer's disease. Aged anti-NGF mice represent, to our knowledge, the most comprehensive animal model for this severe neurodegenerative disease. Also, these results demonstrate that, in mice, a deficit in the signaling and/or transport of NGF leads to neurodegeneration. (+info)
Spinal cholinergic neurons activated during locomotion: localization and electrophysiological characterization.
(51/969)
The objective of the present study was to determine the location of the cholinergic neurons activated in the spinal cord of decerebrate cats during fictive locomotion. Locomotion was induced by stimulation of the mesencephalic locomotor region (MLR). After bouts of locomotion during a 7-9 h period, the animals were perfused and the L(3)-S(1) spinal cord segments removed. Cats in the control group were subjected to the same surgical procedures but no locomotor task. The tissues were sectioned and then stained by immunohistochemical methods for detection of the c-fos protein and choline acetyltransferase (ChAT) enzyme. The resultant c-fos labeling in the lumbar spinal cord was similar to that induced by fictive locomotion in the cat. ChAT-positive cells also clearly exhibited fictive locomotion induced c-fos labeling. Double labeling with c-fos and ChAT was observed in cells within ventral lamina VII, VIII, and possibly IX. Most of them were concentrated in the medial portion of lamina VII close to lamina X, similar in location to the partition and central canal cells found by Barber and collaborators. The number of ChAT and c-fos-labeled neurons was increased following fictive locomotion and was greatest in the intermediate gray, compared with dorsal and ventral regions. The results are consistent with the suggestion that cholinergic interneurons in the lumbar spinal cord are involved in the production of fictive locomotion. Cells in the regions positive for double-labeled cells were targeted for electrophysiological study during locomotion, intracellular filling, and subsequent processing for ChAT immunohistochemistry. Three cells identified in this way were vigorously active during locomotion in phase with ipsilateral extension, and they projected to the contralateral side of the spinal cord. Thus a new population of spinal cord cells can be defined: cholinergic partition cells with commissural projections that are active during the extension phase of locomotion. (+info)
Expression, purification and characterization of recombinant human choline acetyltransferase: phosphorylation of the enzyme regulates catalytic activity.
(52/969)
Choline acetyltransferase synthesizes acetylcholine in cholinergic neurons and, in humans, may be produced in 82- and 69-kDa forms. In this study, recombinant choline acetyltransferase from baculovirus and bacterial expression systems was used to identify protein isoforms by two-dimensional SDS/PAGE and as substrate for protein kinases. Whereas hexa-histidine-tagged 82- and 69-kDa enzymes did not resolve as individual isoforms on two-dimensional gels, separation of wild-type choline acetyltransferase expressed in insect cells revealed at least nine isoforms for the 69-kDa enzyme and at least six isoforms for the 82-kDa enzyme. Non-phosphorylated wild-type choline acetyltransferase expressed in Escherichia coli yielded six (69 kDa) and four isoforms (82 kDa) respectively. Immunofluorescent labelling of insect cells expressing enzyme showed differential subcellular localization with the 69-kDa enzyme localized adjacent to plasma membrane and the 82-kDa enzyme being cytoplasmic at 24 h. By 64 h, the 69-kDa form was in cytoplasm and the 82-kDa form was only present in nucleus. Studies in vitro showed that recombinant 69-kDa enzyme was a substrate for protein kinase C (PKC), casein kinase II (CK2) and alpha-calcium/calmodulin-dependent protein kinase II (alpha-CaM kinase), but not for cAMP-dependent protein kinase (PKA); phosphorylation by PKC and CK2 enhanced enzyme activity. The 82-kDa enzyme was a substrate for PKC and CK2 but not for PKA or alpha-CaM kinase, with only PKC yielding increased enzyme activity. Dephosphorylation of both forms of enzyme by alkaline phosphatase decreased enzymic activity. These studies are of functional significance as they report for the first time that phosphorylation enhances choline acetyltransferase catalytic activity. (+info)
Activity-dependent neurotrophic factor: intranasal administration of femtomolar-acting peptides improve performance in a water maze.
(53/969)
Activity-dependent neurotrophic factor (ADNF) is a glia-derived protein that is neuroprotective at femtomolar concentrations. A nine-amino acid peptide derived from ADNF (Ser-Ala-Leu-Leu-Arg-Ser-Ile-Pro-Ala; ADNF-9) captured the activity of the parent protein and has been reported to protect cultured neurons from multiple neurotoxins. Antibodies recognizing ADNF-9 produced neuronal apoptosis, and identified an additional, structurally related, glia-derived peptide, Asn-Ala-Pro-Val-Ser-Ile-Pro-Gln (NAP). Previous comparative studies have characterized s.c.-injected NAP as most efficacious in protecting against developmental retardation and learning impairments in apolipoprotein E-deficient mice. This study was designed to assess 1) neuroprotection after intranasal administration of ADNF-9 and NAP to rats treated with the cholinotoxin ethylcholine aziridium; and 2) bioavailability and pharmacokinetics after intranasal administration. Results showed significant improvements in short-term spatial memory, as assessed in a water maze, after daily intranasal administration of 1 microg of peptide (ADNF-9 or NAP) per animal. However, a 5-day pretreatment with ADNF-9 did not improve performance measured after cessation of treatment. Compared with rats treated with ADNF-9, NAP-pretreated animals exhibited a significantly better performance. Furthermore, NAP (and not ADNF-9) protected against loss of choline acetyl transferase activity. Significant amounts of (3)H-labeled NAP reached the brain, remained intact 30 min after administration, and dissipated 60 min after administration. This study revealed efficacy for ADNF-related peptides in rodent models for neurodegeneration. The small size of the molecules, the low dosage required, the noninvasive administration route, and the demonstrated activity in a relevant paradigm suggest NAP as a lead compound for future drug design. (+info)
Induction and maintenance of the neuronal cholinergic phenotype in the central nervous system by BMP-9.
(54/969)
Bone morphogenetic proteins (BMPs) have multiple functions in the developing nervous system. A member of this family, BMP-9, was found to be highly expressed in the embryonic mouse septum and spinal cord, indicating a possible role in regulating the cholinergic phenotype. In cultured neurons, BMP-9 directly induced the expression of the cholinergic gene locus encoding choline acetyltransferase and the vesicular acetylcholine transporter and up-regulated acetylcholine synthesis. The effect was reversed upon withdrawal of BMP-9. Intracerebroventricular injection of BMP-9 increased acetylcholine levels in vivo. Although certain other BMPs also up-regulated the cholinergic phenotype in vitro, they were less effective than BMP-9. These data indicate that BMP-9 is a differentiating factor for cholinergic central nervous system neurons. (+info)
Complete and long-term rescue of lesioned adult motoneurons by lentiviral-mediated expression of glial cell line-derived neurotrophic factor in the facial nucleus.
(55/969)
To date, delivery of neurotrophic factors has only allowed to transiently protect axotomized facial motoneurons against cell death. In the present report, long-term protection of these neurons was evaluated by continuously expressing the neurotrophic factor glial cell line-derived neurotrophic factor (GDNF) within the facial nucleus using a lentiviral vector system. The viral vector was injected unilaterally into the facial nucleus of 4-month-old Balb/C mice. In contrast to axotomy in other adult rodents, facial nerve lesion in these animals leads to a progressive and sustained loss and/or atrophy of >50% of the motoneurons. This model thus represents an attractive model to evaluate potential protective effects of neurotrophic factors for adult-onset motoneuron diseases, such as amyotrophic lateral sclerosis. One month after unilateral lentiviral vector injection, the facial nerve was sectioned, and the animals were killed 3 months later. Viral delivery of the GDNF gene led to long-term expression and extensive diffusion of GDNF within the brainstem. In addition, axotomized motoneurons were completely protected against cell death, because 95% of the motoneurons were present as demonstrated by both Nissl staining and choline acetyltransferase immunoreactivity. Furthermore, GDNF prevented lesion-induced neuronal atrophy and maintained proximal motoneuron axons, despite the absence of target cell reinnervation. This is the first evidence that viral-mediated delivery of GDNF close to the motoneuron cell bodies of the facial nucleus of adult mice can lead to complete and long-term protection against lesion-induced cell death. (+info)
Synaptic integration mediated by striatal cholinergic interneurons in basal ganglia function.
(56/969)
The physiological role of striatal cholinergic interneurons was investigated with immunotoxin-mediated cell targeting (IMCT). Unilateral cholinergic cell ablation caused an acute abnormal turning behavior. These mice showed gradual recovery but displayed abnormal turning by both excess stimulation and inhibition of dopamine actions. In the acute phase, basal ganglia function was shifted to a hyperactive state by stimulation and suppression of striatonigral and striatopallidal neurons, respectively. D1 and D2 dopamine receptors were then down-regulated, relieving dopamine-predominant synaptic perturbation but leaving a defect in controlling dopamine responses. The acetylcholine-dopamine interaction is concertedly and adaptively regulated for basal ganglia synaptic integration. (+info)