Neville N. Osborne

Neuroprotection in relation to retinal ischemia and relevance to glaucoma.

Management of glaucoma is directed at the control of intraocular pressure (IOP), yet it is recognized now that increased IOP isjust an important risk factor in glaucoma. Therapy that prevents the death of ganglion cells is the main goal of treatment, but an understanding of the causes of ganglion cell death and precisely how it occurs remains speculative. Present information supports the working hypothesis that ganglion cell death may result from a particular form of ischemia. Support for this view comes from the fact that not all types of retinal ischemia lead to the pathologic findings seen in glaucomatous retinas or to cupping in the optic disk area. Moreover, in animal experiments in which ischemia is caused by elevated IOP, a retinal abnormality similar to that seen in true glaucoma is produced, whereas after occlusion of the carotid arteries a different pattern of damage is found. In ischemia, glutamate is released, and this initiates the death of neurons that contain ionotropic glutamate (NMDA) receptors. Elevated glutamate levels exist in the vitreous humor of patients with glaucoma, and NMDA receptors exist on ganglion cells and a subset of amacrine cells. Experimental studies have shown that a variety of agents can be used to prevent the death of retinal neurons (particularly ganglion cells) induced by ischemia. These agents are generally those that block NMDA receptors to prevent the action of the released glutamate or substances that interfere with the subsequent cycle of events that lead to cell death. The major causes of cell death after activation of NMDA receptors are the influx of calcium into cells and the generation of free radicals. Substances that prevent this cascade of events are, therefore, often found to act as neuroprotective agents. For a substance to have a role as a neuroprotective agent in glaucoma, it would ideally be delivered topically to the eye and used repeatedly. It is, therefore, of interest that betaxolol, a beta-blocker presently used to reduce IOP in humans, also has calcium channel-blocking functions. Moreover, experimental studies show that betaxolol is an efficient neuro protective agent against retinal ischemia in animals, when injected directly into the eye or intraperitoneally.

Cellular signaling and factors involved in Müller cell gliosis: Neuroprotective and detrimental effects

Müller cells are active players in normal retinal function and in virtually all forms of retinal injury and disease. Reactive Müller cells protect the tissue from further damage and preserve tissue function by the release of antioxidants and neurotrophic factors, and may contribute to retinal regeneration by the generation of neural progenitor/stem cells. However, Müller cell gliosis can also contribute to neurodegeneration and impedes regenerative processes in the retinal tissue by the formation of glial scars. This article provides an overview of the neuroprotective and detrimental effects of Müller cell gliosis, with accounts on the cellular signal transduction mechanisms and factors which are implicated in Müller cell-mediated neuroprotection, immunomodulation, regulation of Müller cell proliferation, upregulation of intermediate filaments, glial scar formation, and the generation of neural progenitor/stem cells. A proper understanding of the signaling mechanisms implicated in gliotic alterations of Müller cells is essential for the development of efficient therapeutic strategies that increase the supportive/protective and decrease the destructive roles of gliosis.

A hypothesis to explain ganglion cell death caused by vascular insults at the optic nerve head: possible implication for the treatment of glaucoma

Some apparent characteristics of ganglion cell death in glaucoma Glaucoma is a progressive optic neuropathy with characteristic optic disc changes and associated visual field defects.1 2 The pattern and progression of visual field loss due to ganglion cell death varies between glaucoma patients suggesting that there is some variability in the magnitude of the insult responsible for the cell loss. Ganglion cell death with a spatial and temporal distribution typical of “glaucoma” can be experimentally induced in animals.3-5 One way to try and mimic glaucoma in experimental animals is to raise the intraocular pressure (IOP).3-6 It is clear from such studies that ganglion cells do not all die at the same time.4-8Furthermore, the rate of deterioration of ganglion cells is proportional to the magnitude of the insult.9 The reason for the initiation of ganglion cell death in glaucoma is unknown, but a number of explanatory theories have been proposed with the vasogenic theory perhaps the most widely accepted hypothesis.2 10-12 Raised IOP is not the sole factor responsible for glaucomatous retinal damage but an important one of a number that have been implicated (Fig 1).12-17 Only 10% of patients with increased IOP (⩾22 mm Hg) have glaucoma and between one third and one half of patients with glaucoma initially do not have elevated IOP.1 18 19 Furthermore, as many as one sixth of patients with glaucomatous damage do not appear to have elevated IOP.1 It is therefore clear that raised IOP is not synonymous with having glaucoma.1 20 21 Nevertheless, high IOP is arguably the most important risk factor (see Fig 1), and it is clearly associated with ganglion cell death in glaucoma patients. Figure 1 Possible causes of ganglion cell death in glaucoma. It is suggested that activation of …

Ganglion cell death in glaucoma: what do we really know?

Glaucomatous optic neuropathy is a chronic process which progresses over many years. Data derived from clinical observations and from animal experiments suggest that the axons of the optic nerve and the retinal ganglion cell somata do not die at the same time but that death can vary between months and many years.1 2 Glaucoma patients have characteristic fields of visual loss which enlarge as the disease progresses. Thus, glaucomatous optic neuropathy may not be a chronic degeneration of the whole of the optic nerve and ganglion cell somata but rather a series of acute losses of individual, or groups of, ganglion cells. It seems therefore reasonable to assume that when a patient is diagnosed initially as having glaucoma, only some ganglion cells are dead, whereas others may range from being “unhealthy” to being “slightly sick” while others are “perfectly normal”. It seems also reasonable to argue that if a neuroprotectant (a substance which reaches the retina to elicit an effect: Table 1) can be applied at some stage before blindness occurs it could be of benefit to the glaucoma patient by either “slowing down” the death process of neurons that has already been initiated to die or perhaps by preventing the initiation of death signals to perfectly healthy ganglion cells. This argument can be made despite the lack of success in the use of neuroprotectants for a variety of other diseases (stroke, epilepsy, Parkinson’s disease, AIDS dementia) where neuronal death is a characteristic.1 3 It is the apparent nature of ganglion cell death in glaucoma, very slow and variable, that makes it more likely that the use of neuroprotectants could be successful. View this table: Table 1 The term neuroprotection in glaucoma implies that the substance used to “protect” the ganglion cells reaches the retina (“direct neuroprotection”) to have an effect. In contrast, …

GABA neurones in retinas of different species and their postnatal development in situ and in culture in the rabbit retina

SummaryThe localisation of GABA immunoreactive neurones in retinas of a variety of animals was examined. Immunoreactivity was associated with specific populations of amacrine neurones in all species examined, viz. rat, rabbit, goldfish, frog, pigeon and guinea-pig. All species, with the exception of the frog, possessed immunoreactive perikarya in their retinal ganglion cell layers. These perikarya are probably displaced amacrine cells because GABA immunoreactivity was absent from the optic nerves and destruction of the rat optic nerve did not result in degeneration of these cells. GABA immunoreactivity was also associated with the outer plexiform layers of all the retinas studied; these processes are derived from GABA-positive horizontal cells in rat, rabbit, frog, pigeon and goldfish retinas, from bipolar-like cells in the frog, and probably from interplexiform cells in the guinea-pig retina.The development of GABA-positive neurones in the rabbit retina was also analysed. Immunoreactivity was clearly associated with subpopulations of amacrine and horizontal cells on the second postnatal day. The immunoreactivity at this stage is strong, and fairly well developed processes are apparent. The intensity of the immunoreactivity increases with development in the case of the amacrine cells. The immunoreactive neurones appear fully developed at about the 8th postnatal day, although the immunoreactivity in the inner plexiform layer becomes more dispersed as development proceeds. The immunoreactive horizontal cells become less apparent as development proceeds, but they can still be seen in the adult retina.The GABA immunoreactive cells in rabbit retinas can be maintained in culture. Cultures of retinal cells derived from 2-day-old animals can be maintained for up to 20 days and show the presence of GABA-positive cells at all stages. In one-day-old cultures the GABA immunoreactive cells lacked processes but within three days had clearly defined processes. After maintenance for 10 days a meshwork of GABA-positive fibres could also be seen in the cultures.

Serotonin-Containing Neurones in Vertebrate Retinas

Abstract: It has been established by a combination of HPLC and electrochemical detection that frog, lizard, goldfish, rabbit, and bovine retinas contain both dopamine and serotonin. Immunohistological and immunoradiographical methods show that serotonin is localised in amacrine perikarya and processes situated in the inner plexiform layers of frog, lizard, and goldfish retinas. The amount of serotonin in the mammalian retina appears to be too low for detection in neurones. The serotonin in the bovine retina is located mainly in the inner nuclear and plexiform layers, suggesting that the amine is present in the same types of cells as found for frog, lizard, and goldfish retinas. Retinas incubated in [3H]serotonin showed that radioactivity is associated with processes in the inner plexiform layer and amacrine perikarya. These results suggest that the neuronal elements that contain endogenous serotonin also have the capacity to accumulate exogenous amine and are consistent with the opinion that serotonin has a neuronal function in retinas of a variety of vertebrates.

In vivo and in vitro experiments show that betaxolol is a retinal neuroprotective agent

The aim of the study was to determine whether betaxolol is a neuroprotective agent and can therefore slow down the changes seen in the retina following ischaemia/reperfusion. Ischaemia was induced in one rat eye by raising the intraocular pressure for 45 min. Three days later electroretinograms were recorded from both eyes and the retinas were examined immunohistochemically for the localisation of calretinin and choline acetyltransferase (ChAT) immunoreactivities. The effect of glutamate agonists, hypoxia or experimental ischaemia was examined on the GABA immunoreactivity, lactate dehydrogenase (LDH) and internal calcium levels ([Ca2+]i) of the isolated rabbit retina, rat cortical cultures and chick retinal cell cultures respectively. Betaxolol was tested to see whether it can attenuate the influence of the glutamate agonists, hypoxia or experimental ischaemia. Ischaemia for 45 min causes a change in the nature of the normal calretinin immunoreactivity, an obliteration of the ChAT immunoreactivity and a drastic reduction in the b-wave of the electroretinogram after 3 days of reperfusion. When betaxolol was injected i.p. into the rats before ischaemia and on the days of reperfusion the changes to the calretinin and ChAT immunoreactivities were reduced and the reduction of the b-wave was prevented. Rabbit retinas incubated in vitro in physiological solution lacking oxygen/glucose or containing the glutamate agonists kainate or NMDA caused a change in the nature of the GABA immunoreactivity. Inclusion of betaxolol partially prevented the changes caused by NMDA and lack of oxygen/glucose. Rat cortical cultures exposed to glutamate or hypoxia/reoxygenation resulted in a release of LDH. The release of the enzyme was almost completely attenuated when betaxolol was included in the culture medium. Kainate increased the [Ca2+]i in chick retinal cultures, as measured with Indo-1. In a medium with sodium, this kainate-induced elevation of [Ca2+]i was significantly reduced by betaxolol. The combined data show that betaxolol is a neuroprotective agent and attenuates the effects on the retina induced by raising the intraocular pressure to simulate an ischaemic insult as may occur in glaucoma.

Nadph Diaphorase Localization and Nitric-Oxide Synthetase-Activity in the Retina and Anterior Uvea of the Rabbit Eye

The distribution of the enzyme nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase was examined histochemically in the retina, iris, ciliary processes, cornea and conjunctiva of the rabbit eye. The epithelial cells of the ciliary process, iris, conjunctiva and, to a lesser extent, the cornea all showed intense staining. In the retina, staining for NADPH diaphorase was intense in the inner segments of the photoreceptors and a sparsely distributed population of amacrine cells. In addition, another population of amacrine cells, some presumed ganglion cells as well as a number of horizontal cells, stained less intensely for the enzyme. The retina, ciliary processes and, as a comparison, the cerebellum of the rabbit all contain nitric oxide synthetase (NOS) activity, as each tissue can metabolize citrulline from arginine. This process is Ca2+ dependent and is reduced by the NOS inhibitor, NG-monomethyl-L-arginine. The presence of NOS activity in the ciliary processes and the localization of NADPH diaphorase in the ciliary epithelial cells are of significance as they suggest that the ciliary epithelial cells may contain NOS which would imply a role for nitric oxide in aqueous humour production.

Mitochondria: Their role in ganglion cell death and survival in primary open angle glaucoma

Retinal ganglion cell axons within the globe are functionally specialised being richly provided with many mitochondria. Mitochondria produce the high energy that is required for nerve conduction in the unmylenated part of the ganglion cell axons and for the maintenance of optimum neuronal function. We proposed that in the initiation of glaucoma (POAG) an alteration in the quality of blood flow dynamics in the optic nerve head results in sustained or intermittent ischemia of a defined nature. This results in normal mitochondrial function being negatively affected and as a consequence retinal ganglion cell function is compromised. Ganglion cells in this state are now susceptible to secondary insults which they would normally tolerate. One secondary insult to ganglion cell mitochondria in such a state might be light entering the eye. Other insults to the ganglion cells might come from substances such as glutamate, prostaglandins and nitric oxide released from astrocytes and microglia in the optic nerve head region. Such cascades of events initiated by ischemia to the optic nerve head region ultimately cause ganglion cells to die at different rates.

Recent clinical findings with memantine should not mean that the idea of neuroprotection in glaucoma is abandoned.

Loss of vision in primary open‐angle glaucoma (glaucoma) is caused by retinal ganglion cells dying at a seemingly steady and variable rate in different patients. Present treatments for all glaucoma patients are inadequate and a goal to rectify this is to discover appropriate drugs or chemicals (neuroprotectants) that can be taken orally to slow down retinal ganglion cell death and have negligible side‐effects. It was therefore of great disappointment to learn earlier this year that the one clinical trial conducted to test the efficacy of memantine as a neuroprotectant for glaucoma was unsuccessful. In this article, I consider the mechanisms by which retinal ganglion cells may die in glaucoma and suggest that memantine may have benefited patients taking it but to a level that was difficult to detect with present methodologies. Ganglion cells are induced to die by different triggers in glaucoma, suggesting that neuroprotectants with multiple modes of actions are likely to reveal clearer results than was found for memantine. Therefore, the idea of neuroprotection in glaucoma must not be abandoned.