Reactive
gliosis is a hallmark of many
retinal neurodegenerative conditions, including
glaucoma. Although a majority of studies to date have concentrated on reactive
gliosis in the optic nerve head, very few studies have been initiated to investigate the role of reactive
gliosis in the retina. We have previously shown that reactive glial cells synthesize elevated levels of
proteases, and these
proteases, in turn, promote the death of retinal ganglion cells (RGCs). In this investigation, we have used two glial toxins to inhibit reactive
gliosis and have evaluated their effect on
protease-mediated death of RGCs.
Kainic acid was injected into the vitreous humor of C57BL/6 mice to induce reactive
gliosis and death of RGCs. C57BL/6 mice were also treated with glial toxins,
alpha-aminoadipic acid (AAA) or
Neurostatin, along with KA. Reactive
gliosis was assessed by immunostaining of
retinal cross sections and
retinal flat-mounts with
glial fibrillary acidic protein (GFAP) and
vimentin antibodies. Apoptotic cell death was assessed by TUNEL assays. Loss of RGCs was determined by immunostaining of flat-mounted retinas with Brn3a
antibodies. Proteolytic activities of
matrix metalloproteinase-9 (MMP-9),
tissue plasminogen activator (tPA), and
urokinase plasminogen activator (uPA) were assessed by zymography assays. GFAP-immunoreactivity indicated that KA induced reactive
gliosis in both
retinal astrocytes and in Muller cells. AAA alone or in combination with KA decreased GFAP and
vimentin-immunoreactivity in Mϋller cells, but not in astrocytes. In addition AAA failed to decrease KA-mediated
protease levels and apoptotic death of RGCs. In contrast,
Neurostatin either alone or in combination with KA, decreased reactive
gliosis in both astrocytes and Mϋller cells. Furthermore,
Neurostatin decreased
protease levels and prevented apoptotic death of RGCs. Our findings, for the first time, indicate that inhibition of reactive
gliosis decreases
protease levels in the retina, prevents apoptotic death of retinal neurons, and provides substantial neuroprotection.