The role of aquaporin-4 in optic nerve head astrocytes in experimental glaucoma

Abstract

Purpose To study aquaporin channel expression in astrocytes of the mouse optic nerve (ON) and the response to IOP elevation in mice lacking aquaporin 4 (AQP4 null). Methods C57BL/6 (B6) and AQP4 null mice were exposed to bead-induced IOP elevation for 3 days (3D-IOP), 1 and 6 weeks. Mouse ocular tissue sections were immunolabeled against aquaporins 1(AQP1), 4(AQP4), and 9(AQP9). Ocular tissue was imaged to identify normal AQP distribution, ON changes, and axon loss after IOP elevation. Ultrastructure examination, cell proliferation, gene expression, and transport block were also analyzed. Results B6 mice had abundant AQP4 expression in Müller cells, astrocytes of retina and myelinated ON (MON), but minimal AQP4in prelaminar and unmyelinated ON (UON). MON of AQP4 nulls had smaller ON area, smaller axon diameter, higher axon density, and larger proportionate axon area than B6 (all p≤0.05). Bead-injection led to comparable 3D-IOP elevation (p = 0.42) and axonal transport blockade in both strains. In B6, AQP4 distribution was unchanged after 3D-IOP. At baseline, AQP1 and AQP9 were present in retina, but not in UON and this was unaffected after IOP elevation in both strains. In 3D-IOP mice, ON astrocytes and microglia proliferated, more in B6 than AQP4 null. After 6 week IOP elevation, axon loss occurred equally in the two mouse types (24.6%, AQP4 null vs. 23.3%, B6). Conclusion Lack of AQP4 was neither protective nor detrimental to the effects of IOP elevation. The minimal presence of AQP4 in UON may be a vital aspect of the regionally specific phenotype of astrocytes in the mouse optic nerve head.

Aquaporin 4 is not present in normal porcine and human lamina cribrosa

“…Our results reported here showing the lack of significant expression of AQP4 in the lamina cribrosa of human and porcine eyes is consistent with our previous findings with mise and rats and suggests the more universal finding, that these channels are likely not present in the equivalent of the lamina cribrosa in any mammalian eye. The minimal presence of AQP4 at the large animal lamina and the unmyelinated nerve of rodent eyes is highly evolutionarily conserved and therefore may be either advantageous for normal ONH function, and/or a protective influence for retinal ganglion cell axons from glaucoma damage [ 39 ]. There are several possible hypotheses for the potential benefit from this regional astrocytic characteristic.…”

“…The inability to expand in the closed ONH compartment, could prevent astrocytes could from enhancing the axonal transport obstruction in axons known to occur in glaucoma. We showed that the myelinated optic nerve in normal mice is larger in area than in AQP4 null mice, due to a reduced astrocytic volume in the nulls [ 39 ]. Previous research showed that increased IOP leads to greater fluid movement from the vitreous cavity through the ONH [ 41 ].…”

“…We used our published method for semi-quantifying fluorescence [ 39 ] using the pixel intensity value (PIV), calculated using FIJI (ImageJ, Bethesda MD). Longitudinal, cryopreserved porcine sections from 4 samples immunolabeled against AQP4 were imaged using the same acquisition settings.…”

“…While Immunolabeling of human secondary glaucoma eyes found reduced AQP9 in the retina, the presence or change in AQP4 in the ONH itself was not demonstrated [ 38 ]. We exposed C57BL/6 and AQP4 knock out mice to bead-induced intraocular pressure (IOP) elevation for 3 days, 1 and 6 weeks [ 39 ]. Wild type mice had abundant AQP4 expression in Müller cells, astrocytes of the retina and myelinated optic nerve, but minimal AQP4 in prelaminar and unmyelinated optic nerve by immunolabeling and gene expression, despite the presence of the DG complex.…”

Regional Gene Expression in the Retina, Optic Nerve Head, and Optic Nerve of Mice with Experimental Glaucoma and Optic Nerve Crush

“…The injection of microbeads into the anterior chamber produces IOP elevation known to cause RGC death that is maximal by 6 weeks [23] (Figure 4A, Figure S4, Table S1). Mice exposed to elevated IOP followed prior experience with bead injection [23, 24], having significant mean IOP elevation at three days, decreasing at two weeks and with minimal difference from baseline at six weeks (Figure S4, Table S1). Micro-dissected UON, MON, and retinal tissue was collected at three days (early, 3D), two weeks (middle, 2W), and six weeks (late, 6W) post-injection to characterize gene expression changes spanning the time course of this model (Figure 4A).…”