Electrical stimulation ameliorates light-induced photoreceptor degeneration in vitro via suppressing the proinflammatory effect of microglia and enhancing the neurotrophic potential of Müller cells

Abstract

Many types of electrical stimulation (ES) devices have been shown to promote the survival of degenerated neural cells, such as dopaminergic neurons in the medial forebrain bundle-transected rats, ischemic-injured cortical neurons and inner-and outer-nuclear-layer cells in degenerated retina. Using a rat photic injury model, our lab previously proved the neuroprotective effect of transcorneal electrical stimulation (TCES) on apoptotic photoreceptor cells. To delineate the mechanisms that might underlie this process, the effects of ES on light-damaged photoreceptor degeneration-induced microglia and Müller cell activation were investigated in the present in vitro study. Our data showed that ES (3 ms, 20 Hz, 300-1600 μA) increased survival among light-reared cone-derived cells (661W) cultured alongside microglia or Müller cells analyzed by LDH and TUNEL assays. The degree of rescue was found to depend on the different intensities of the ES current. The immunocytochemistry revealed that ES significantly decreased the numbers of activated microglia cells with ameboid shapes and increased the numbers of reactive gliotic Müller cells with larger soma when they were co-cultured with light-damaged 661W cells. Real-time RT-PCR and Western blotting indicated that ES which was applied to different co-cultures and 661W cell-conditioned media (661WCM)-treated glia cultures had a prominent inhibitive effect on the secretion of interleukin (IL)-1β and tumor necrosis factor (TNF)-α in microglia and a positive regulative effect on the production of brain-derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF) in Müller cells. The death rate of light-exposed 661W cells cultured with microglia was decreased significantly by the addition of neutralizing antibodies against IL-1β and TNF-α. On the other hand, the death rate of light-exposed 661W cells cultured with Müller cells was prominently increased when the co-culture was incubated in the presence of neutralizing antibody against BDNF while anti-CNTF neutralizing antibody did not induce additional exacerbation of the cell death among those 661W cells. These findings indicate the feasibility of using ES to create a nurturing environment for light-damaged photoreceptor cells. This environment is characterized by diminished microglial activation and fortified Müller cells reactive gliosis, which may have great potential in ameliorating photoreceptor damage. In this way, ES was here determined to be a novel, potent therapeutic option for delaying the progression of photoreceptor degeneration in patients suffering from retinitis pigmentosa (RP).