Abstract
Abstract: :
Purpose: NT–4 is a neurotrophic factor which is expressed in the CNS together with its receptor TrkB. TrkB binds both NT–4 and BDNF, another neurotrophin expressed in several areas of the brain, including the retina. Previous reports showed that BDNF plays an important role in the maturation of the retina. To understand if endogenous NT–4 is expressed and has a function in mammalian retina we analysed the retina of mice with a deleted gene for nt–4 (nt–4–/– , nt–4 ko). Methods: Using RT–PCR and ELISA we investigated the gene expression and NT–4 protein in the retina of control wild type mice. We performed immunocytochemistry (ICCH) on retinal sections in adult and P15 nt–4 ko and wild type mice using retinal cell–type specific antibodies. Quantitative analysis of tyrosine hydroxylase (TH) immunoreactivity in the OPL and IPL was carried out using confocal microscopy and Metamorph® software. TH immunopositive cell bodies and varicosities were quantified in retinal whole mount. Data were analysed using the Wilcoxon–Mann–Whitney statistic test (U–test). Results: With RT–PCR and ELISA we showed that nt–4 mRNA and protein are expressed in the retina of adult and P15 wild type mice. TH ICCH was altered in adult and P15 nt–4 ko mice while other markers such as PKC, calbindin, Dab–1 and parvalbumin resulted normal. Quantitative analysis of TH immunoreactivity performed with Metamorph® software in the IPL of adult mice did not reveal significant difference between nt–4+/+ and nt–4–/– mice whereas an increase of 154% in the area occupied by dopaminergic fibers was present in the OPL of nt–4 ko adult mice (U–test, p=0.003). In P15 nt–4–/– mice, quantitative analysis of TH immunoreactivity revealed a 74% increase in the area occupied by dopaminergic fibers in the IPL (U–test, p=0.05) and a 281% increase in the area occupied by dopaminergic fibers in the OPL (U–test, p=0.004) respect to nt–4+/+ mice. The number of TH–immunopositive cells was not significantly different between nt–4–/– (611±76) and nt–4+/+ mice (588±115). Conclusions: These results demonstrate that NT–4 is expressed in the retina and plays a role in the development and maintenance of retinal dopaminergic cells. Notably, NT–4 inhibits and BDNF stimulates retinal dopaminergic network, respectively, thus exerting opposite actions in mammalian retina.
Keywords: retinal connections, networks, circuitry • retina: proximal (bipolar, amacrine, and ganglion cells)