RESUMO
Basal forebrain cholinergic neurons, which innervate the hippocampus and cortex, have been implicated in many forms of cognitive function. Immunolesion-based methods in animal models have been widely used to study the role of acetylcholine (ACh) neurotransmission in these processes, with variable results. Cholinergic neurons have been shown to release both glutamate and ACh, making it difficult to deduce the specific contribution of each neurotransmitter on cognition when neurons are eliminated. Understanding the precise roles of ACh in learning and memory is critical because drugs that preserve ACh are used as treatment for cognitive deficits. It is therefore important to define which cholinergic-dependent behaviors could be improved pharmacologically. Here we investigate the contributions of forebrain ACh on hippocampal synaptic plasticity and cognitive behavior by selective elimination of the vesicular ACh transporter, which interferes with synaptic storage and release of ACh. We show that elimination of vesicular ACh transporter in the hippocampus results in deficits in long-term potentiation and causes selective deficits in spatial memory. Moreover, decreased cholinergic tone in the forebrain is linked to hyperactivity, without changes in anxiety or depression-related behavior. These data uncover the specific contribution of forebrain cholinergic tone for synaptic plasticity and behavior. Moreover, these experiments define specific cognitive functions that could be targeted by cholinergic replacement therapy.
Assuntos
Comportamento Animal , Potenciação de Longa Duração , Memória , Prosencéfalo/metabolismo , Proteínas Vesiculares de Transporte de Acetilcolina/metabolismo , Animais , Western Blotting , Imunofluorescência , Aprendizagem , Camundongos , Camundongos Knockout , Plasticidade Neuronal , Reação em Cadeia da PolimeraseRESUMO
Stress-inducible phosphoprotein 1 (STI1) is part of the chaperone machinery, but it also functions as an extracellular ligand for the prion protein. However, the physiological relevance of these STI1 activities in vivo is unknown. Here, we show that in the absence of embryonic STI1, several Hsp90 client proteins are decreased by 50%, although Hsp90 levels are unaffected. Mutant STI1 mice showed increased caspase-3 activation and 50% impairment in cellular proliferation. Moreover, placental disruption and lack of cellular viability were linked to embryonic death by E10.5 in STI1-mutant mice. Rescue of embryonic lethality in these mutants, by transgenic expression of the STI1 gene, supported a unique role for STI1 during embryonic development. The response of STI1 haploinsufficient mice to cellular stress seemed compromised, and mutant mice showed increased vulnerability to ischemic insult. At the cellular level, ischemia increased the secretion of STI1 from wild-type astrocytes by 3-fold, whereas STI1 haploinsufficient mice secreted half as much STI1. Interesting, extracellular STI1 prevented ischemia-mediated neuronal death in a prion protein-dependent way. Our study reveals essential roles for intracellular and extracellular STI1 in cellular resilience.
Assuntos
Embrião de Mamíferos/metabolismo , Proteínas de Choque Térmico/metabolismo , Isquemia/metabolismo , Chaperonas Moleculares/metabolismo , Príons/metabolismo , Animais , Blastocisto/metabolismo , Western Blotting , Fator de Transcrição CDX2 , Células Cultivadas , Feminino , Proteínas de Choque Térmico/genética , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Técnicas In Vitro , Isquemia/genética , Camundongos , Camundongos Mutantes , Chaperonas Moleculares/genética , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Gravidez , Príons/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Acetylcholine (ACh) has been suggested to facilitate plasticity and improve functional recovery after different types of brain lesions. Interestingly, numerous studies have shown that striatal cholinergic interneurons are relatively resistant to acute ischemic insults, but whether ACh released by these neurons enhances functional recovery after stroke is unknown. We investigated the role of endogenous striatal ACh in stroke lesion volume and functional outcomes following middle cerebral artery occlusion to induce focal ischemia in striatum-selective vesicular acetylcholine transporter-deficient mice (stVAChT-KO). As transporter expression is almost completely eliminated in the striatum of stVAChT-KO mice, ACh release is nearly abolished in this area. Conversely, in other brain areas, VAChT expression and ACh release are preserved. Our results demonstrate a larger infarct size after ischemic insult in stVAChT-KO mice, with more pronounced functional impairments and increased mortality than in littermate controls. These changes are associated with increased activation of GSK-3, decreased levels of ß-catenin, and a higher permeability of the blood-brain barrier in mice with loss of VAChT in striatum neurons. These results support a framework in which endogenous ACh secretion originating from cholinergic interneurons in the striatum helps to protect brain tissue against ischemia-induced damage and facilitates brain recovery by supporting blood-brain barrier function.
Assuntos
Acetilcolina/metabolismo , Corpo Estriado/metabolismo , Infarto da Artéria Cerebral Média/metabolismo , Acidente Vascular Cerebral/metabolismo , Acetilcolina/genética , Animais , Corpo Estriado/patologia , Modelos Animais de Doenças , Infarto da Artéria Cerebral Média/genética , Infarto da Artéria Cerebral Média/patologia , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Acidente Vascular Cerebral/genética , Acidente Vascular Cerebral/patologia , Proteínas Vesiculares de Transporte de Acetilcolina/deficiência , Proteínas Vesiculares de Transporte de Acetilcolina/genéticaRESUMO
Stress-inducible phosphoprotein I (STIP1, STI1 or HOP) is a co-chaperone intermediating Hsp70/Hsp90 exchange of client proteins, but it can also be secreted to trigger prion protein-mediated neuronal signaling. Some mothers of children with autism spectrum disorders (ASD) present antibodies against certain brain proteins, including antibodies against STIP1. Maternal antibodies can cross the fetus blood-brain barrier during pregnancy, suggesting the possibility that they can interfere with STIP1 levels and, presumably, functions. However, it is currently unknown whether abnormal levels of STIP1 have any impact in ASD-related behavior. Here, we used mice with reduced (50%) or increased STIP1 levels (fivefold) to test for potential ASD-like phenotypes. We found that increased STIP1 regulates the abundance of Hsp70 and Hsp90, whereas reduced STIP1 does not affect Hsp70, Hsp90 or the prion protein. Interestingly, BAC transgenic mice presenting fivefold more STIP1 show no major phenotype when examined in a series of behavioral tasks, including locomotor activity, elevated plus maze, Morris water maze and five-choice serial reaction time task (5-CSRTT). In contrast, mice with reduced STIP1 levels are hyperactive and have attentional deficits on the 5-CSRTT, but exhibit normal performance for the other tasks. We conclude that reduced STIP1 levels can contribute to phenotypes related to ASD. However, future experiments are needed to define whether it is decreased chaperone capacity or impaired prion protein signaling that contributes to these phenotypes.
Assuntos
Transtorno do Deficit de Atenção com Hiperatividade/metabolismo , Comportamento Animal , Proteínas de Choque Térmico/deficiência , Animais , Transtorno do Deficit de Atenção com Hiperatividade/genética , Transtorno do Deficit de Atenção com Hiperatividade/psicologia , Modelos Animais de Doenças , Predisposição Genética para Doença , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas de Choque Térmico/genética , Masculino , Aprendizagem em Labirinto , Camundongos Endogâmicos C57BL , Camundongos Knockout , Atividade Motora , Fenótipo , Proteínas PrPC/metabolismo , Tempo de Reação , Natação , Fatores de TempoRESUMO
Tissue pH is an indicator of altered cellular metabolism in diseases including stroke and cancer. Ischemic tissue often becomes acidic due to increased anaerobic respiration leading to irreversible cellular damage. Chemical exchange saturation transfer (CEST) effects can be used to generate pH-weighted magnetic resonance imaging (MRI) contrast, which has been used to delineate the ischemic penumbra after ischemic stroke. In the current study, a novel MRI ratiometric technique is presented to measure absolute pH using the ratio of CEST-mediated contrast from amine and amide protons: amine/amide concentration-independent detection (AACID). Effects of CEST were observed at 2.75 parts per million (p.p.m.) for amine protons and at 3.50 p.p.m. for amide protons downfield (i.e., higher frequency) from bulk water. Using numerical simulations and in vitro MRI experiments, we showed that pH measured using AACID was independent of tissue relaxation time constants, macromolecular magnetization transfer effects, protein concentration, and temperature within the physiologic range. After in vivo pH calibration using phosphorus ((31)P) magnetic resonance spectroscopy ((31)P-MRS), local acidosis is detected in mouse brain after focal permanent middle cerebral artery occlusion. In summary, our results suggest that AACID represents a noninvasive method to directly measure the spatial distribution of absolute pH in vivo using CEST MRI.