The purpose of this study was to investigate whether signaling through insulin, Fibroblast Growth Factor (FGF) receptors, and Mitogen-Activated Protein Kinase (MAPK) pathways protect retinal neurons against excitotoxicity and regulate the proliferation of Müller glia. We found that intraocular injections of insulin or FGF2 had variable effects upon the phosphorylation of ERK1/2, p38 MAPK and CREB, and the expression of immediate early genes, cFos and Egr1. Accumulations of pERK1/2, p38 MAPK, pCREB, cFos and Egr1 in response to insulin or FGF2 were confined to Müller glia, whereas retinal neurons did not appear to respond to growth factors. Unlike FGF2, insulin stimulated microglia-like cells to up-regulate the intermediate filament transitin and lysosomal membrane glycoprotein (LMG). The affects of insulin and FGF2 upon microglia-like cells and/or MAPKsignaling in Müller glia had profound effects upon numbers of dying neurons in response to excitotoxic damage. Although FGF2 significantly reduced numbers of dying neurons, insulin significantly increased numbers of dying neurons. In addition to neuroprotective affects, FGF2 also "primed" the Müller glia to proliferate following retinal damage, whereas insulin had no effect upon glial proliferation. Further, we found that FGF receptor isoform 1 (FGFR1) and FGFR3 are prominently expressed in the retina, whereas the insulin receptor and FGFR2 are not expressed, or are expressed at very low levels. We conclude that MAPK-signaling through FGF receptors stimulates Müller glia to become more neuroprotective and progenitor-like, whereas insulin acting on Müller and microglia-like cells through unidentified receptors had the opposite effect.

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Bullwhip and mini-bullwhip cells are unconventional types of retinal neurons that utilize the neuropeptides glucagon, glucagon-like peptide 1 (GLP1) and substance P. These cells have been implicated in regulating the proliferation of neural progenitors in the circumferential marginal zone (CMZ) of the chicken retina. The purpose of this study was to investigate the roles of the bullwhip cells in regulating ocular size and shape. We found that intravitreal delivery of colchicine at postnatal day 7 destroys the vast majority (approximately 98%) of the bullwhip and mini-bullwhip cells and their peptidergic terminals that are concentrated in the CMZ near the equator of the eye. Interestingly, colchicine-treatment resulted in excessive ocular growth that involved the expansion of equatorial diameter, but not axial length. Intraocular injections of glucagon completely prevented the equatorial expansion that occurs with colchicine-treatment. In eyes with undamaged retinas, exogenous glucagon suppressed equatorial eye growth, whereas glucagon receptor antagonists caused excessive equatorial growth. Furthermore, visual stimuli that increase or decrease rates of ocular growth caused a down- or up-regulation, respectively, of the immediate early gene Egr1 in the bullwhip cells; indicating that the activity of the bullwhip cells is regulated by growth-guiding visual cues. We found that the glucagon receptor was expressed by cells in the fibrous and cartilaginous sclera in equatorial regions of the eye. Taken together, these findings suggest that glucagon peptide released from the terminals of the bullwhip and mini-bullwhip cells regulates the growth of the equatorial sclera in a vision-dependent manner. Although the bullwhip and mini-bullwhip cells are not abundant, less than 1000 cells per retina, their influence on the development of the eye is substantial and includes vision-guided ocular growth.

Different growth factors have been shown to influence the development of form-deprivation myopia and lens-induced ametropias. However, growth factors have relatively little effect on the growth of eyes with unrestricted vision. We investigate whether the combination of insulin-like growth factor 1 (IGF1) and fibroblast growth factor 2 (FGF2) influence ocular growth in eyes with unrestricted vision. Different doses of IGF1 and FGF2 were injected into the vitreous chamber of postnatal chicks. Measurements of ocular dimensions and intraocular pressure (IOP) were made during and at the completion of different treatment paradigms. Histological and immunocytochemical analyses were performed to assess cell death, cellular proliferation and integrity of ocular tissues. Treated eyes had significant increases in equatorial diameter and vitreous chamber depth. With significant variability between individuals, IGF1/FGF2-treatment caused hypertrophy of lens and ciliary epithelia, lens thickness was increased, and anterior chamber depth was decreased. Treated eyes developed myopia, in excess of 15 diopters of refractive error. Shortly after treatment, eyes had increased intraocular pressure (IOP) which was increased in a dose-dependent manner. Seven days after treatment with IGF1 and FGF2 changes to anterior chamber depth, lens thickness and elevated IOP were reduced, whereas increases in the vitreous chamber were persistent. Some damage to ganglion cells was detected in peripheral regions of the retina at 7 days after treatment. We conclude that the extreme myopia in IGF1/FGF2-treated eyes results from increased vitreous chamber depth, decreased anterior chamber depth, and changes in the lens. We propose that factor-induced ocular enlargement and myopia result from changes to the sclera, lens and anterior chamber depth.