RESUMO
Antisense oligodeoxynucleotides have been highly successful agents at modulating gene expression in the adult brain and widely exploited in the field of neuroendocrinology. We have also used this technique in the developing brain to explore the role of select proteins during sensitive periods of development, particularly those influenced by steroid hormones. Presented here are the technical details of using antisense oligodeoxynucleotides in the neonatal brain, as well as a review of some of our successes and failures. Our goal is to illustrate the relative ease of use of this technique in neonates and demonstrate the power such an approach offers so that other investigators will also begin to take advantage of this useful tool.
Assuntos
Neuroendocrinologia/métodos , Oligodesoxirribonucleotídeos Antissenso/farmacologia , Animais , Animais Recém-Nascidos , Comportamento Animal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/crescimento & desenvolvimento , Encéfalo/fisiologia , Feminino , Glutamato Descarboxilase/antagonistas & inibidores , Glutamato Descarboxilase/genética , Glutamato Descarboxilase/metabolismo , Infusões Parenterais , Masculino , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Oligodesoxirribonucleotídeos Antissenso/administração & dosagem , Oligodesoxirribonucleotídeos Antissenso/genética , RatosRESUMO
Calbindin-D28k (calbindin) is a 28 kilodalton calcium binding protein which potentially plays a role in neuroprotection. We report here the normal development and gonadal steroid modulation of a sexually dimorphic group of calbindin immunoreactive cells within the sexually dimorphic nucleus of the preoptic area (SDN) which we call the calbindin-immunoreactive SDN or CALB-SDN. Beginning on PN2, a faintly immunoreactive CALB-SDN is present, however, the volume is not sexually dimorphic. On PN4, the staining of the CALB-SDN appears more robust but the volume is still not sexually dimorphic. By PN8 and extending through PN12 and PN26, the latest age analysed, the volume of the CALB-SDN is larger in males by two- to fourfold. Cresyl violet counterstain reveals a similar developmental profile of the SDN as well as clusters of darkly staining calbindin immunonegative cells which lie around the CALB-SDN. Castration of males on PN0 decreases the volume of the CALB-SDN by PN12 and administration on the day of birth and PN1 of either testosterone propionate or oestradiol benzoate, but not dihydrotestosterone propionate to females increases the volume of the CALB-SDN by PN12. By demonstrating the sexual dimorphism and gonadal steroid modulation of the CALB-SDN, we hereby establish that calbindin is a specific marker of a subdivision of the SDN and can be used as such in future studies.
Assuntos
Biomarcadores/análise , Hormônios Esteroides Gonadais/farmacologia , Área Pré-Óptica/efeitos dos fármacos , Área Pré-Óptica/crescimento & desenvolvimento , Proteína G de Ligação ao Cálcio S100/análise , Caracteres Sexuais , Envelhecimento , Animais , Animais Recém-Nascidos , Calbindina 1 , Calbindinas , Estradiol/análogos & derivados , Estradiol/farmacologia , Feminino , Hormônios Esteroides Gonadais/fisiologia , Masculino , Proteínas do Tecido Nervoso/análise , Orquiectomia , Gravidez , Área Pré-Óptica/química , Ratos , Ratos Sprague-Dawley , Testosterona/farmacologiaRESUMO
The proteins calbindin-D(28K) and calretinin buffer intracellular calcium and are speculated to be involved in the integration of neuronal signaling. Using Western blot analysis, we compared the levels of calbindin-D(28K) and calretinin in the developing male and female rat hypothalamus on postnatal days (PN) 0, PN2, PN4, PN6, PN8, and PN10. Analysis of variance (ANOVA) of mean calbindin levels indicated a significant effect of sex (p
Assuntos
Envelhecimento/metabolismo , Hipotálamo/fisiologia , Proteína G de Ligação ao Cálcio S100/metabolismo , Análise de Variância , Animais , Animais Recém-Nascidos , Calbindina 2 , Calbindinas , Feminino , Ácido Glutâmico/metabolismo , Hipotálamo/citologia , Hipotálamo/crescimento & desenvolvimento , Imuno-Histoquímica , Masculino , Proteínas do Tecido Nervoso/análise , Proteínas do Tecido Nervoso/metabolismo , Área Pré-Óptica/fisiologia , Ratos , Ratos Sprague-Dawley , Proteína G de Ligação ao Cálcio S100/análise , Caracteres Sexuais , Núcleo Hipotalâmico Ventromedial/fisiologia , Ácido gama-Aminobutírico/metabolismoRESUMO
The skeleton is a major repository for divalent cations, including toxic heavy metals such as lead. Unfortunately the effects of such agents on bone (and cartilage) have been minimally investigated in the past. With the current level of understanding of the mechanisms of bone formation and cartilage development it is now appropriate to begin to research the effects of lead on cellular processes. The following discussion describes some of the points of regulation in bone and cartilage formation where interference in metabolic processes could compromise the development of normal tissues as well as affect the homeostatic mechanisms of the skeleton.
