RESUMO
PURPOSE: To verify the capability of rabbit and rat ciliary body to synthesize and secrete ceruloplasmin. METHODS: Isolated ciliary body (CB) was cultured in the presence of [35S]-methionine, and the incubation medium was processed for immunoprecipitation. Total RNA from CB was processed for RT-PCR, and the amplification products were sequenced. Also, sections of CB were immunostained for the localization of ceruloplasmin. RESULTS: A labeled peptide, having a molecular weight of about 135 kDa, the expected size of ceruloplasmin, was immunopurified in the incubation media from both animal species. The RT-PCR and sequencing experiments detected the presence of ceruloplasmin mRNA in rat samples. Both layers of rabbit and rat ciliary epithelium (CE) exhibited ceruloplasmin reactivity after immunohistochemical processing. CONCLUSIONS: Taken altogether, these results indicate the CB, particularly its epithelium, as one of the possible sources of the ocular intrinsic ceruloplasmin.
Assuntos
Ceruloplasmina/genética , Ceruloplasmina/metabolismo , Corpo Ciliar/enzimologia , Animais , Técnicas Imunoenzimáticas , Imunoprecipitação , Masculino , Técnicas de Cultura de Órgãos , RNA Mensageiro/metabolismo , Coelhos , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transferrina/genéticaRESUMO
The concept of the blood-aqueous barrier is largely based on the use of horseradish peroxidase (HRP). The present investigation was designed to check its reliability as a macromolecular tracer, especially with regard to the transport of plasma proteins. Rabbits were killed 5 min to 24 h after being intravenously injected with HRP. The tracer diffused rapidly, reaching the aqueous humor of the eye in 3 min or less and was detected at high concentration in the narrow space between the outer epithelial layer of the ciliary epithelium and the wall of the pervious capillaries in the stroma of the processes. HRP appeared to migrate from the blood to the posterior chamber, permeating the tight junctions, viz., the anatomical basis of the blood-aqueous barrier. It was detected at higher concentration at the anterior surface of the iris, at short time intervals; this was interpreted as penetration of the tracer from the aqueous humor of the anterior chamber. The choroid was also labeled in continuation with the reaction in the stroma of the pars plana of the ciliary body which, in turn, sometimes reached the iris root. Therefore, the pervious blood vessels of the choroid could be a source of macromolecules for the iris root. HRP also induced the formation of lysosomes in the ciliary epithelium. This can hardly be accepted as the way in which plasma proteins are physiologically transported to the aqueous humor. However, the pathway of HRP migration over short time intervals seems to be in agreement with previous research indicating that the entrance of serum albumin into the posterior chamber is the first step of its incorporation into the aqueous humor.
Assuntos
Humor Aquoso/metabolismo , Peroxidase do Rábano Silvestre , Hidrolases Anidrido Ácido/metabolismo , Animais , Proteínas Sanguíneas/metabolismo , Corpo Ciliar/enzimologia , Epitélio/enzimologia , Secções Congeladas , Peroxidase do Rábano Silvestre/farmacocinética , Masculino , Microtomia , CoelhosRESUMO
The effects of pertussis toxin on the intraocular pressure (IOP) lowering effect of clonidine and isoproterenol as well as on the inhibitory effects of clonidine and neuropeptide Y on adenylate cyclase activity of ciliary processes were studied in albino rabbits. I.v. administered pertussis toxin elicited transient changes in IOP which, however, returned to control values during 2-3 days. In the following days the IOP lowering effect of the alpha 2-adrenergic agonist clonidine was abolished and that of the nonselective beta-adrenergic agonist isoproterenol was attenuated. At the same time, the inhibitory effects of clonidine and neuropeptide Y on basal as well as stimulated adenylate cyclase activities in homogenates of ciliary processes were grossly diminished. The effects of pertussis toxin on the IOP lowering action of adrenergic agonists and on the inhibitory action of clonidine and neuropeptide Y on adenylate cyclase activity were ascribed to an impairment of the function of a G protein in ciliary processes, probably G(i) protein. It is suggested that the decrease of IOP induced by clonidine is due to inhibition of adenylate cyclase.