Suppression of trk(B) expression by antisense oligonucleotides alters a neuronal phenotype in the rod pathway of the developing rat retina

trk(B) is the high-affinity receptor for brain-derived neurotrophic factor (BDNF), a trophic molecule with demonstrated effects on the survival and differentiation of a wide variety of neuronal populations. In the mammalian retina, trk(B) is localized to both ganglion cells and numerous cells in the inner nuclear layer, Much information on the role of BDNF in neuronal development has been derived from the study of trk(B)- and BDNF-deficient mutant mice, This includes an attenuation of the numbers of cortical neurons immunopositive for the calcium-binding proteins, parvalbumin, and calbindin. Unfortunately, these mutant animals typically fail to survive for >24-48 hr after birth. Since most retinal neuronal differentiation occurs postnatally, we have devised an alternative scheme to suppress the expression of trk(B) in the retina to examine the role of BDNF on the postnatal development of neurons of the inner retina, Neonatal rats were treated with intraocular injection of an antisense oligonucleotide (1-2 mu l of 10-100 mu M solution) targeted to the trk(B) mRNA, Immunohistochemistry with a polyclonal antibody to trk(B) showed that the expression of trk(B) in retinal neurons was suppressed 48-72 hr following a single injection, Northern blot analysis demonstrated that antisense treatment had no effect on the level of trk(B) mRNA, even after multiple injections. This suggests an effect of trk(B) antisense treatment on protein translation, but not on RNA transcription, No alterations were observed in the thickness of retinal cellular or plexiform layers, suggesting that BDNF is not the sole survival factor for these neurons, There were, however, alterations in the patterns of immunostaining for parvalbumin, a marker for the narrow-field, bistratified AII amacrine cell-a central element of the rod (scotopic) pathway, This was evidenced by a decrease in both the number of immunostained somata (>50%) and in the intensity of immunolabeling, However, the immunostaining pattern of calbindin was not affected. These studies suggest that the ligands for trk(B) have specific effects on the neurochemical phenotypic expression of inner retinal neurons and in the development of a well-defined retinal circuit.