RESUMO
Apotransferrin injected intracranially into young rats has been shown in our laboratories to induce an early differentiation of oligodendroglial cells and an increased deposition of myelin. The expression of some myelin-specific proteins such as myelin basic protein (MBP) and 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) and of their mRNAs were significantly increased in these animals. Also, in the cytoskeleton obtained from isolated myelin, it was found that several microtubule associated proteins (MAPs), particularly the stable tubule only peptide (STOP) and MAP 1B, as well as actin and tubulin were markedly increased. In the present paper, we compare the changes in expression of brain and myelin cytoskeletal proteins in a newly generated transferrin transgenic mouse (Tg), overexpressing the human transferrin gene, with the results obtained in aTf-injected rats. In the myelin cytoskeletal fraction of Tg mice there was a significant increase in the expression of MBP, tubulin, tau and STOP, similarly to what was previously found in the aTf-injected rats. Immunohistochemical studies showed that a variance with what occurs in the aTf-injected model, in which the above mentioned changes were limited to the corpus callosum, in the Tg mice the changes in expression of cytoskeletal proteins were observed in the various anatomical areas studied such as cerebral cortex, brain stem and cerebellum. There was also an increased expression of neurofilaments in the Tg animals, in contrast with results obtained in aTf-injected rats, suggesting that in the Tg mice, the continuous overexpression of Tf might also induce some neuronal changes. Changes in tau, total and acetylated tubulin and MAP 1B were observed in both neurons and OLGc. The increase in STOP was more significant in OLGc while the changes in MAP2 were exclusively found in neurons.
Assuntos
Encéfalo/metabolismo , Proteínas do Citoesqueleto/biossíntese , Bainha de Mielina/metabolismo , Transferrina/genética , Animais , Encéfalo/efeitos dos fármacos , Proteínas do Citoesqueleto/efeitos dos fármacos , Proteínas do Citoesqueleto/genética , Feminino , Humanos , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Transgênicos , Proteínas Associadas aos Microtúbulos/biossíntese , Proteínas Associadas aos Microtúbulos/efeitos dos fármacos , Proteínas Associadas aos Microtúbulos/genética , Proteína Básica da Mielina/biossíntese , Proteína Básica da Mielina/efeitos dos fármacos , Proteína Básica da Mielina/genética , Bainha de Mielina/efeitos dos fármacos , Bainha de Mielina/genética , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/metabolismo , Ratos , Ratos Wistar , Transferrina/farmacologiaRESUMO
To determine whether neonatal intracranial injection of apotransferrin (aTf), which increases myelin deposition, has behavioral effects in rats, 3-day-old rats were intracranially injected with 350 ng of aTf and tested at 25 and 60 days of age. An anxiolytic-like behavior was observed in aTf-treated rats, evidenced by an increase in the exploration of open arms in the plus maze test without changes in the locomotor activity. This behavioral profile persists until adulthood. Intraperitoneal injection of 0.75 mg/kg of picrotoxin, a GABA(A) receptor channel antagonist, abolished this anxiolytic-like behavior, indicating that neonatal aTf induces a long-lasting increase in GABA(A) receptor functionality.
Assuntos
Apoproteínas/metabolismo , Apoproteínas/farmacologia , Comportamento Animal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Bainha de Mielina/efeitos dos fármacos , Oligodendroglia/efeitos dos fármacos , Transferrina/metabolismo , Transferrina/farmacologia , Animais , Animais Recém-Nascidos , Ansiedade/tratamento farmacológico , Ansiedade/fisiopatologia , Comportamento Animal/fisiologia , Encéfalo/citologia , Encéfalo/crescimento & desenvolvimento , Antagonistas GABAérgicos/farmacologia , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Aprendizagem em Labirinto/fisiologia , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Bainha de Mielina/metabolismo , Oligodendroglia/citologia , Oligodendroglia/metabolismo , Picrotoxina/farmacologia , Ratos , Ratos Wistar , Receptores de GABA-A/efeitos dos fármacos , Receptores de GABA-A/metabolismoRESUMO
We have recently shown that sustained neonatal hyperthyroidism in the rat activates apoptosis of oligodendroglial cells (OLGc) and that inhibition of the proteasome-ubiquitin (Ub) pathway by lactacystin produces increased apoptosis in cerebellar granule cells (CGC). In the present study we have analyzed the relationship between the activation of the Ub-dependent pathway, the expression of the Ub genes and programmed cell death in neurons of the rat cerebellum and cerebral cortex and in OLGc. This study was carried out in normal animals, in rats submitted to sustained neonatal hyperthyroidism and in cell cultures treated with an excess of thyroid hormones. In neurons of the cerebral cortex, thyroid hormone produces an increase of Ub-protein conjugates, an enhancement in the expression of the Ub genes and an increase in apoptosis, while the opposite results are obtained in CGC. These results indicate that in neurons, the changes in the cell death program produced by thyroid hormone run in parallel with those occurring in the Ub-dependent pathway. In OLGc, thyroid hormone increases apoptosis but does not produce changes in the Ub pathway. Preliminary studies indicate that in coincidence with what occurs in optic nerves, the sciatic nerves both in controls and in hyperthyroid animals are unable to form Ub-protein conjugates. These results indicate that in cells of the CNS such as neurons, in which the Ub-dependent pathway is actively expressed, it appears to be closely correlated with apoptosis.
Assuntos
Apoptose/fisiologia , Encéfalo/fisiologia , Oligodendroglia/fisiologia , Nervo Isquiático/fisiologia , Tri-Iodotironina/farmacologia , Ubiquitinas/metabolismo , Envelhecimento , Animais , Animais Recém-Nascidos , Apoptose/efeitos dos fármacos , Encéfalo/citologia , Encéfalo/efeitos dos fármacos , Tronco Encefálico/citologia , Tronco Encefálico/efeitos dos fármacos , Tronco Encefálico/fisiologia , Ciclo Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/ultraestrutura , Células Cultivadas , Córtex Cerebral/citologia , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/fisiologia , Feminino , Masculino , Oligodendroglia/citologia , Oligodendroglia/efeitos dos fármacos , Ratos , Ratos Wistar , Nervo Isquiático/citologia , Nervo Isquiático/efeitos dos fármacos , Transcrição Gênica , Ubiquitinas/genéticaRESUMO
In the present paper we first studied the brain distribution and the time and dose dependent effects of apotransferrin, after its intracranial injection into young rats and at different post-natal ages. Its action upon the transferrin receptor (TfR) and upon the expression of brain transferrin, as well as its effect on the proliferation and differentiation of oligodendroglial cells (OLGc) was one of the main objectives of our investigation. Total DNA and BrdU labeling, as an index of cellularity and proliferation, respectively, were the same in the control and experimental groups of rats. A significant increase in the MBP+ and CA II+ OLGc, and a decrease in the more immature (A2B5+) OLGc were found in the aTf injected rats. At 10 and 17 days of age, Tf-mRNA decreased to around 20% of the amount present in control animals. The TfR-mRNA in the animals receiving a single dose of aTf at 3 days of age showed an increase in its expression at 10 and 17 days of age, coincident with a higher immunoreactivity of the TfR itself of neurons, choroid plexus and brain capillaries in different brain areas. Although TfR+ OLGc were present up to 7 days of age in controls and in the Tf injected rats, no positive cells were observed at 17 days of age, even in the aTf injected rats. Our results give support to the hypothesis that aTf is an important factor necessary for the maturation of the OLGc, and that the effects that it produces in the OLGc-myelin unit after its intracranial injection in young rats are not due to an increase in proliferation, but to an accelerated differentiation of Tf-sensitive OLGc.
Assuntos
Apoproteínas/farmacologia , Encéfalo/metabolismo , Oligodendroglia/citologia , Transferrina/farmacologia , Animais , Apoproteínas/administração & dosagem , Apoproteínas/genética , Encéfalo/citologia , Diferenciação Celular , Relação Dose-Resposta a Droga , Feminino , Expressão Gênica , Humanos , Injeções , Radioisótopos do Iodo/metabolismo , Masculino , Oligodendroglia/metabolismo , RNA Mensageiro , Ratos , Ratos Wistar , Receptores da Transferrina/genética , Crânio , Fatores de Tempo , Transferrina/administração & dosagem , Transferrina/genéticaRESUMO
We have carried out a study of the effects of sustained neonatal hyperthyroidism on myelin and on the oligodendroglial cells, in an effort to obtain further insight into the molecular mechanisms underlying the action of thyroid hormones on the central nervous system (CNS). Expression of the mRNAs of myelin basic protein (MBP) myelin proteolipid protein (PLP), 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase), transferrin, and c-Jun was investigated in 10- and 17-day-old normal and hyperthyroid rats, using Northern blot analysis. At 10 days of age, the levels of all the explored mRNAs were markedly higher in the experimental animals. The mRNA of transferrin showed a ninefold increase over control values, suggesting the possibility that this putative trophic factor might act as one of the mediators in the action of thyroid hormones. At 17 days of age on the other hand, the levels of all the mRNAs decreased markedly, reaching values below control, except for c-Jun, which remained higher than in normals. At 70 days of age, hyperthyroid rats showed clear evidence of myelin deficit, in agreement with previous results of our laboratories (Pasquini et al.: J Neurochem 57: Suppl S124, 1991). Immunocytochemistry of 70-day-old rat brain tissue sections showed a substantial reduction in the amount of MBP-reacting structures and a marked decrease in the number of oligodendroglial cells. Although the above-mentioned results could be the consequence, as proposed by Barres et al. (Development 120:1097-1108, 1994) and Baas et al. (Glia 19:324-332, 1997) of a premature arrest in oligodendroglial cell proliferation followed by early differentiation, the persistent high levels of expression of c-Jun, together with the dramatic decrease in the number of oligodendrocytes, suggested the possibility that prolonged hyperthyroidism could activate apoptotic mechanisms in the myelin forming cells. Using propidium iodide-labeled isolated oligodendroglial cells, we found, by flow cytometry, a significant increase in the number of apoptotic/hypo-diploid propidium iodide-positive cells. These results indicate that one of the actions of sustained levels of thyroid hormones in the neonate rat is to increase oligodendroglial cell death by apoptosis.