Sodium channel expression in optic nerve astrocytes chronically deprived of axonal contact

Abstract

Immunocytochemical and electrophysiological methods were used to examine the effect of retinal ablation on the expression of sodium channels within optic nerve astrocytes in situ and in vitro. Enucleation was performed at postanatal day 3 (P3), and electron microscopy of the enucleated optic nerves at P28–P40 revealed complete degeneration of retinal ganglion axons, resulting in optic nerves composed predominantly of astrocytes. In contrast to control (non‐enucleated) optic nerve astrocytes, which exhibited distinct sodium channel immunoreactivity following immunostaining with antibody 7493, the astrocytes in enucleated optic nerves did not display sodium channel immunoreactivity in situ. Cultures obtained from enucleated optic nerves consisted principally ( > 90%) of glial fibrillary acidic protein (GFAP)⁺/A2B5⁻ (“type‐1”) astrocytes, as determined by indirect immunofluorescence; GFAP⁺/A2B5⁺ (“type‐2”) astrocytes were not present, nor were GFAP⁻/A2B5⁺ (O‐2A) progenitor cells. Sodium channel immunoreactivity was not present in GFAP⁺/A2B5⁻ astrocytes obtained from enucleated optic nerves; in contrast, GFAP⁺/A2B5⁻ astrocytes from control optic nerves exhibited 7493 immunostaining for the first 4–6 days in culture. Sodium current expression, studied using whole‐cell patch‐clamp recording, was attenuated in cultured astrocytes derived from enucleated optic nerves. Whereas 39 of 50 type‐1 astrocytes cultured from intact optic nerves showed measurable sodium currents at 1–7 days in vitro, sodium currents were present in only 6 of 38 astrocytes cultured from enucleated optic nerves. Mean sodium current densities in astrocytes from the enucleated optic nerves (0.66 ± 0.3 pA/pF) were significantly smaller than in astrocytes from control optic nerves (7.15 ± 1.1 pA/pF). The h_χ‐curves of sodium currents were similar in A2B5⁻ astrocytes from enucleated and control rat optic nerves. These results suggest that there is neuronal modulation of sodium channel expression in type‐1 optic nerve astrocytes, and that, following chronic loss of axonal association in vivo, sodium channel expression is down‐regulated in this population of optic nerve astrocytes.