Ocular diseases such as glaucoma, age-related macular degeneration (AMD), diabetic retinopathy, and retinitis pigmentosa require drug management in order to prevent blindness and affecting millions of adults in US and worldwide. There is an increasing need to develop devices for drug delivery to address ocular diseases. This research focused on an implantable ocular drug delivery device design, simulation and experiments with design requirements including constant diffusion rate, extended period of time operation, the smallest possible volume of device and reservoir. The drug delivery device concept uses micro-/nano-channels module embedded between top and bottom covers with a drug reservoir. Several microchannel design configurations were developed and simulated using commercial finite element software (ANSYS and COMSOL), with a goal to investigate how the microchannel dimensions affect the diffusion characteristics. In addition to design simulations, various microchannel configurations were fabricated on silicon wafer using photolithography techniques as well as 3D printing. Also, the top and bottom covers of the device were fabricated from PDMS through replica molding techniques. These fabricated microchannel design configurations along with top and bottom covers were all integrated into the device. Both single straight microchannels (nine different sizes of width and depth) as well as four micro-channel configurations were tested using citric acid (pH changes) and Brimonidine drug (concentration changes using the Ultra-Violet Visible Spectrophotometer) for their diffusion characteristics. Experiments were conducted to obtain the diffusion rates through various single micro-channels as well as micro-channel configurations using the change in pH neutral solution to verify the functionality and normalized diffusion rate of microchannels and configurations. The results of experimental data of diffusion rate were compared with those obtained from simulations, and a good agreement was found. The results showed the diffusion rate and the optimum size of microchannel in conjunction with the required drug release time. The results obtained also indicate that even though specific diffusion rates can be obtained but delivering the drug with constant amount needs a mechanism at the device outlet with some control mechanism. For future studies, this result may be used as a baseline for developing a microfabricated device that allows for accurate drug diffusion in many drug delivery applications.

Retinal ganglion cells play a crucial role in the relay of visual signals from the eye to the brain. This cell type is affected and eventually lost in the eye disease glaucoma, resulting in progressive and irreversible loss of vision. Studies of the molecular mechanisms leading to retinal ganglion cell death are important for the understanding of the disease and for designing future treatments. This thesis addresses and studies these molecular mechanisms, including alterations in gene expression after experimental retinal injuries. The effects of a neuroprotective drug, brimonidine, after transient retinal ischemia were also studied in order to help explain the mechanisms behind the protective properties of this drug. Several methods, including quantitative reverse transcriptase PCR, micro-arrays, western blot and immunohistochemistry, were used. The results showed that transient retinal ischemia triggers cell division in Müller cells and alters the gene expression of growth factors, their receptors, and intermediate filaments in the retina. Several genes related to the apoptosis process were less affected. Pre-treatment with brimonidine increased the levels of certain growth factors (BDNF, NT3, CNTF, FGF9) compared with vehicle. Brimonidine also had marked effects on genes related to progenitor cells, among them the recognized neural stem cell marker nestin. The increase in levels of nestin after ischemia was countered by brimonidine treatment. Moreover, retinal ganglion cell death following either optic nerve transection or optic nerve crush appears to involve the extrinsic apoptotic pathway although the gene expression response appears to differ between these injuries. The results obtained in this work contribute to an increased understanding of retinal injuries and highlight the importance of Müller cells in the endogenous defense against retinal injuries.

Müller cells are major glial cells in the retina and have a broad range of functions that are vital for the retinal neurons. During retinal injury gliotic response either leads to Müller cell dedifferentiation and formation of a retinal progenitor or to maintenance of mature Müller cell functions. The overall aim of this thesis was to investigate the intra- and extracellular signaling of Müller cells, to understand how Müller cells communicate during an injury and how their properties can be regulated after injury. Focus has been on the α2-adrenergic receptor (α2-ADR) and endothelin receptor (EDNR)-induced modulation of Müller cell-properties after injury. The results show that α2-ADR stimulation by brimonidine (BMD) triggers Src-kinase mediated ligand-dependent and ligand-independent transactivation of epidermal growth factor receptor (EGFR) in both chicken and human Müller cells. The effects of this transactivation in injured retina attenuate injury-induced activation and dedifferentiation of Müller cells by attenuating injury-induced ERK signaling. The attenuation was concomitant with a synergistic up-regulation of negative ERK- and RTK-feedback regulators during injury. The data suggest that adrenergic stress-signals modulate glial responses during retinal injury and that α2-ADR pharmacology can be used to modulate glial injury-response. We studied the effects of this attenuation of Müller cell dedifferentiation on injured retina from the perspective of neuroprotection. We analyzed retinal ganglion cell (RGC) survival after α2-ADR stimulation of excitotoxically injured chicken retina and our results show that α2-ADR stimulation protects RGCs against the excitotoxic injury. We propose that α2-ADR-induced protection of RGCs in injured retina is due to enhancing the attenuation of the glial injury response and to sustaining mature glial functions. Moreover, we studied endothelin-induced intracellular signaling in Müller cells and our results show that stimulation of EDNRB transactivates EGFR in Müller cells in a similar way as seen after α2-ADR stimulation. These results outline a mechanism of how injury-induced endothelins may modulate the gliotic responses of Müller cells. The results obtained in this thesis are pivotal and provide new insights into glial functions, thereby uncovering possibilities to target Müller cells by designing neuroprotective treatments of retinal degenerative diseases or acute retinal injury.