Abstract
Müller cells are mainly involved in controlling extracellular homeostasis in the retina, where intense neural activity alters ion concentrations and osmotic gradients, thus favoring cell swelling. This increase in cell volume is followed by a regulatory volume decrease response (RVD), which is known to be partially mediated by the activation of K+ and anion channels. However, the precise mechanisms underlying osmotic swelling and subsequent cell volume regulation in Müller cells have been evaluated by only a few studies. Although the activation of ion channels during the RVD response may alter transmembrane potential (Vm), no studies have actually addressed this issue in Müller cells. The aim of the present work is to evaluate RVD using a retinal Müller cell line (MIO-M1) under different extracellular ionic conditions, and to study a possible association between RVD and changes in Vm. Cell volume and Vm changes were evaluated using fluorescent probe techniques and a mathematical model. Results show that cell swelling and subsequent RVD were accompanied by Vm depolarization followed by repolarization. This response depended on the composition of extracellular media. Cells exposed to a hypoosmotic solution with reduced ionic strength underwent maximum RVD and had a larger repolarization. Both of these responses were reduced by K+ or Cl− channel blockers. In contrast, cells facing a hypoosmotic solution with the same ionic strength as the isoosmotic solution showed a lower RVD and a smaller repolarization and were not affected by blockers. Together, experimental and simulated data led us to propose that the efficiency of the RVD process in Müller glia depends not only on the activation of ion channels, but is also strongly modulated by concurrent changes in the membrane potential. The relationship between ionic fluxes, changes in ion permeabilities and ion concentrations –all leading to changes in Vm– define the success of RVD.
Highlights
Glial cells in the sensory retina (Muller cells) are mainly involved in controlling osmotic and ionic homeostasis [1,2]
Cells were grown as monolayers in the presence of Dulbecco’s Modified Eagle Medium (DMEM)/glutamax supplemented with 10% fetal calf serum (FCS), containing 5 mg/ml streptomycin and 5 U/ml penicillin at 37uC in a humidified atmosphere containing 5% CO2
The percentage of RVD at 10 minutes (% RVD10) is significantly higher with the HYPONaCl solution (Figure 2 insert). These results indicate that MIO-M1 cells respond to cell swelling by triggering RVD, the magnitude of this response depends on extracellular media composition
Summary
Glial cells in the sensory retina (Muller cells) are mainly involved in controlling osmotic and ionic homeostasis [1,2]. Retinal cells can be surrounded by a hypoosmotic environment, since light-evoked changes in the ionic composition of the extracellular fluid cause a decrease in osmolarity, favoring glial swelling [3]. It has been reported that Muller cells in situ show an effective control of cell volume, that prevents cell swelling, probably due to the presence of K+ channels Kir 4.1. The expression of these channels is altered in different pathologies such as retinal ischemia, ocular inflammation and diabetes, as well as in organ cultures [9,10,11,12]
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