Ovarian steroid and neurotoxin models of brain aging in rodents.
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Aging↗
Biomedical subjects
G M Pasinetti
Publications and source records attributed to G M Pasinetti.
Complement C1qB and C4 mRNAs responses to lesioning in rat brain.
These data show the presence of mRNAs for two complement components (C) in the adult rat brain and describe their responses to experimental lesions. Cortical deafferentation caused elevations in striatal C1qB and C4 mRNAs that coincided temporally and overlapped anatomically with the course of degeneration of corticostriatal afferent fibers. By in situ hybridization, C1qB mRNA in the lesioned striatum was colocalized to cells immunoreactive for CR3, a complement receptor found on microglia-macrophages. The mRNA for SGP-2, a putative C inhibitor in rat, showed parallel changes. Similarly, in hippocampus and other brain regions, kainic acid lesions increased C1qB mRNA. The data suggest that microglia-macrophages and possibly other cells in rat brain rapidly up-regulate C-mRNAs in response to deafferentation and local neuron injury. These experimental responses provide models to analyze changes in C components during Alzheimer's disease and other chronic neurodegenerative conditions.
Animals↗
Amyloid precursor protein mRNA encoding the Kunitz protease inhibitor domain is increased by kainic acid-induced seizures in rat hippocampus.
A 168-nucleotide exon, found in alternatively spliced amyloid precursor protein (APP) mRNAs, encodes a Kunitz protease inhibitor (KPI) domain. Kainic acid (ip) caused a selective increase of KPI mRNA in rat hippocampus. By in situ hybridization, KPI mRNA was induced in the neuronal layers of the hippocampus 11-12 h after the onset of kainate-induced seizures. The kainate-induced elevation of the KPI-containing APP-770 mRNA was blocked by pretreatment with the anticonvulsant pentobarbital. These data suggest that kainate-induced seizures cause alterations in APP RNA stability and/or processing in rat hippocampal neurons.
Amino Acid Sequence↗
Slow changes of tyrosine hydroxylase gene expression in dopaminergic brain neurons after neurotoxin lesioning: a model for neuron aging.
Slow neuron regression develops during the adult phase of life in select brain systems of mammals. We describe a model in adult rats that resolves several phases in a slow atrophic process that differentially influences levels of mRNA and protein for tyrosine hydroxylase (TH). Responses of striatal dopaminergic markers to 6-hydroxydopamine (6-OHDA) lesions in rats indicated that the striatal terminals maintained TH protein, despite greater than 3-fold loss of TH mRNA in the substantia nigra pars compacta (SNC) cell bodies whose axons project to the striatum. The loss of TH mRNA/cell was progressive up to 9 months, whereas SNC cell body shrinkage stabilized by 3 months post-lesioning. Consideration of possible mechanisms in protein turnover motivated a search for PEST motifs in the TH of rats and other vertebrates that could be a point of regulation by altering the rate of TH protein turnover.
3,4-Dihydroxyphenylacetic Acid↗
Regionally specific and rapid increases in brain-derived neurotrophic factor messenger RNA in the adult rat brain following seizures induced by systemic administration of kainic acid.
In situ hybridization techniques were used to analyse the spatiotemporal pattern of brain-derived neurotrophic factor messenger RNA elevation associated with kainic acid-induced seizure activity in the rat. Pronounced increases in hippocampal brain-derived neurotrophic factor messenger RNA levels were observed as early as 30 min following the onset of behavioral seizures. The greatest increase (10-fold) occurred in the dentate granule cell layer, while pyramidal layers CA1, CA3, and CA4 exhibited increases of two- to six-fold. Peak elevation of brain-derived neurotrophic factor messenger RNA in CA1 hippocampal region was evident at 4 h in CA3, and in the dentate granule layer at 30 min postseizure. Elevations persisted in the dentate and hilar regions to four days, while the increases in CA1 and CA3 returned to control levels by 16 h following seizure. Significant increases in brain-derived neurotrophic factor messenger RNA were also observed in the superficial layers of cortex (II and III) and in the piriform cortex which reached peak elevations by 8 h. No detectable changes were observed in the dorsomedial thalamus. Although histologically defined pyramidal and granule cell layers displayed relatively uniform increases in brain-derived neurotrophic factor messenger RNA in response to kainate, a closer examination of the labeling patterns using emulsion autoradiography revealed discrete areas of high grain densities overlapping uniform, moderate hybridization densities in the dentate granule cell layer and CA3, suggesting that the capacity to upregulate brain-derived neurotrophic factor messenger RNA in these regions may differ among individual neurons. In conclusion, our studies revealed that brain-derived neurotrophic factor messenger RNA induction in response to systemic kainate administration differs in hippocampal and cortical areas, in magnitude, time of onset and duration. The observed temperospatial pattern does not correspond in a simple way to increases in metabolic or electrical activity associated with seizures or neuronal vulnerability coincident with the seizures.
Animals↗
Decortication and striatal mRNA: increases of mRNA for fibronectin, but not of NCAM or alpha-1 tubulin.
Ipsilateral frontal cortex lesions damaged the corticostriatal input. Northern hybridization analysis showed increases in fibronectin (FN) mRNA, but not changes in mRNA for neural cell adhesion molecule (NCAM), neuro-filament-68 (NF-68) or alpha-tubulin (alpha-1T) 72 h post-lesion. In situ hybridization resolved a different spatial-temporal distribution. The superficial cell layer beneath the wound cavity showed transient elevations of FN mRNA that peaked at 72 h post-lesion. However, in the ipsilateral striatum, FN mRNA was maximal at later times than in the wound cavity, at 240 h post-lesion. Changes in NCAM and alpha-tubulin mRNAs in response to decortication occur only around the wound cavity but not in the deafferentated striatum. The different time courses of mRNA revealed by Northern blot analysis and in situ hybridization are most probably due to contamination of the ipsilateral striatum at dissection with superficial tissue adjacent to the wound cavity. These results suggest that cellular responses to ipsilateral decortication consist of two phases: (i) a wound healing process; and (ii) striatal responses to deafferentation.
Animals↗
Striatal responses to decortication. I. Dopaminergic and astrocytic activities.
Unilateral ablation of the frontal cortex induced 30%-50% decrease of dopamine (DA) concentration in the ipsilateral striatum at 10 and 27 days after lesioning. There were increased ratios of dihydroxyphenylacetic acid (DOPAC): DA and homovanillic acid (HVA): DA by 20%-60% at 10 days post-lesioning, which suggest compensatory increases of DA metabolism. While no change in total striatal tyrosine hydroxylase (TH) polypeptide concentration was found at any post-lesion time, TH catalytic activity was decreased slightly (-25%) at 10 days. Among individual rats, at 3, 10 and 27 days post-lesioning, striatal DA concentration was inversely related to striatal glial fibrillary acidic protein (GFAP) concentration, a marker of astrocytic activity. The loss of DA was observed whether or not DA was normalized to striatal protein, which suggests that DA loss cannot be simply attributed to increased astrocytic proteins. These data suggest reciprocal relationships between the extent of astrocytic reactions after cortical deafferentation and striatal DA loss, which could involve local remodelling without primary damage to the nigro-striatal terminals.
3,4-Dihydroxyphenylacetic Acid↗
Sulfated glycoprotein-2 (SGP-2) mRNA is expressed in rat striatal astrocytes following ibotenic acid lesions.
A combination of in situ hybridization and immunocytochemistry (ICC) was used to identify the cells that contained mRNA for sulfated glycoprotein-2 (SGP-2) in adult male rats after striatal ibotenic acid (IA) lesioning. Astrocytic responses were monitored by ICC for glial fibrillary acidic protein (GFAP). IA lesioning that caused death of striatal neurons also stimulated astrocytic responses as monitored by GFAP and SGP-2. The SGP-2 immunoreactivity showed punctate deposits in the lesioned striatum without any apparent cellular localization. By in situ hybridization combined with ICC, SGP-2 mRNA was localized in astrocytes that were GFAP-immunopositive. These data suggest that reactive astrocytes may express SGP-2 which may be eventually secreted.
Animals↗
Molecular and morphological correlates following neuronal deafferentation: a cortico-striatal model.
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Afferent Pathways↗
Disappearance of GAD-mRNA and tyrosine hydroxylase in substantia nigra following striatal ibotenic acid lesions: evidence for transneuronal regression.
Transneuronal regression in substantia nigra reticulata (SNR) and substantia nigra compacta (SNC) neurons was studied in Fischer 344 male rats by immunocytochemistry and by in situ hybridization. Three months after striatal lesioning by ibotenic acid, there was a shrinkage (30%) of the SNR region cross-sectional area and a 50% disappearance of neurons that contain glutamic acid decarboxylase (GAD)-mRNA, but only in the ventromedial portion of this nucleus. Loss of dopaminergic neurons, as recognized by tyrosine hydroxylase immunoreactivity, occurred only in caudal portions of the SNC and SNR. These findings suggest that lesions in reciprocally connected pathways, like the nigrostriatal and striatonigral systems, may produce a vicious cycle (feedforward cascade) of neurodegeneration due to interference with retrograde ana anterograde influences.
Animals↗
Tyrosine hydroxylase mRNA concentration in midbrain dopaminergic neurons is differentially regulated by reserpine.
Tyrosine hydroxylase (TH)-mRNA, assayed by in situ hybridization combined with TH immunocytochemistry, showed a selective increase in the ventral tegmental area (A-10) but not in the substantia nigra (A-9) midbrain dopaminergic (DAergic) neurons 3 days after reserpine treatment. TH-mRNA in locus ceruleus noradrenergic (A-4) neurons was increased by reserpine, as confirmed by RNA blot hybridization. These findings show that TH-mRNA is differentially regulated in midbrain DAergic neurons in response to reserpine.
Animals↗
Chronic lesions differentially decrease tyrosine hydroxylase messenger RNA in dopaminergic neurons of the substantia nigra.
Long-term effects of lesions were analyzed in terms of gene expression. Nine months after unilateral 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra pars compacta (s. nigra), the remaining dopaminergic (DAergic) neurons (tyrosine hydroxylase (TH) cells determined by immunocytochemistry (ICC] on the lesioned side were atrophic with smaller nucleoli. By in situ hybridization, the DAergic neurons on the lesioned side had a 50% smaller TH-mRNA concentration than on the contralateral non-lesioned side. However, beta-tubulin mRNA concentration in DAergic neurons was unaffected by the lesion. The lesions did not alter TH-mRNA concentration in the contralateral non-lesioned side by comparison with unoperated controls. We propose that chronic lesions have long-term effects on gene expression because of damage sustained during compensatory hyperactivity after the lesion, or because of decreased trophic support from other neurons.
Animals↗
Selective reduction of mRNA for the beta-amyloid precursor protein that lacks a Kunitz-type protease inhibitor motif in cortex from Alzheimer brains.
In poly(A) RNA from cerebral cortex obtained postmortem from victims of Alzheimer's disease (AD), an alternatively spliced mRNA for the amyloid precursor protein (APP-695 mRNA) was shown to be decreased by 65%. Another form (APP-751 mRNA) with an additional exon encoding a Kunitz-type (serine) protease inhibitor motif did not change appreciably (less than 30% decrease) in AD cortex. If this twofold increase in the APP-751 mRNA/APP-695 mRNA ratio results in a corresponding increase in the APP-751/APP-695 protein ratio, this would support the hypothesis that impaired proteolysis promotes the accumulation of abnormal proteins in the brain during AD. In the two previously known, major alternatively spliced forms of ca. 3.3 and 3.5 kb, we resolved doublet RNAs for each form that are consistent with sequence data showing multiple polyadenylation sites (J. Kang et al., 1987, Nature (London) 325, 733-736.). Smaller APP-related transcripts were also found (1.1, 1.0, 0.8, and 0.3 kb), some of which are selectively altered in AD.
Alzheimer Disease↗
Complement mRNA in the mammalian brain: responses to Alzheimer's disease and experimental brain lesioning.
This study describes evidence in the adult human and rat brain for mRNAs that encode two complement (C) proteins, C1qB and C4. C proteins are important effectors of humoral immunity and inflammation in peripheral tissues but have not been considered as normally present in brain. Previous immunocytochemical studies showed that C proteins are associated with plaques, tangles, and dystrophic neurites in Alzheimer's disease (AD), but their source is unknown. Combined immunocytochemistry and in situ hybridization techniques show C4 mRNA in pyramidal neurons and C1qB mRNA in microglia. Primary rat neuron cultures also show C1qB mRNA. In the cortex from AD brains, there were two- to threefold increases of C1qB mRNA and C4 mRNA, and increased C1qB mRNA prevalence was in part associated with microglia. As a model for AD, we examined entorhinal cortex perforant path transection in the rat brain, which caused rapid increases of C1qB mRNA in the ipsilateral, but not contralateral, hippocampus and entorhinal cortex. The role of brain-derived acute and chronic C induction during AD and experimental lesions can now be considered in relation to functions of C proteins that pertain to cell degeneration and/or cell preservation and synaptic plasticity.
Aged↗
Combined in situ hybridization and immunocytochemistry in the assay of pharmacological effects on tyrosine hydroxylase mRNA concentration.
An assay for tyrosine hydroxylase (TH) mRNA by in situ hybridization in combination with immunocytochemistry (ICC) for TH on the same section is described. The in situ hybridization protocol was optimized for [35S]cRNA (complementary RNA, i.e. anti-sense strand) probe concentration and time of hybridization. The specificity of hybridization was measured by several critera. The advantage of measuring grain density versus grains per cell is discussed for quantitation of in situ autoradiography. Finally, the reserpine-induced increase in adrenal TH mRNA was used to validate quantitative aspects of the in situ hybridization technique by comparison with blot hybridization. In contrast to the adrenal, reserpine did not increase TH mRNA in substantia nigra (s. nigra) neurons as measured by either technique.
Animals↗