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Two modes of stimulation by ammonia of taurine release from cultured rabbit Müller cells
L Faff1, A Reichenbach, J Albrecht
1Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
Abstract:
A previous study revealed that a 10-min ('acute') treatment of cultured Müller glia with ammonium ions (further referred to as 'ammonia') at 0.5-5 mM concentration stimulated the release of newly loaded taurine (Tau) by a cAMP-dependent, osmoresistant mechanism. Here we showed that a 24 h treatment of the cells with 1 mM ammonia increased both Tau release and intracellular cAMP content in a degree similar to acute treatment with 5 mM ammonia, and the effects were similarly resistant to an increase of medium tonicity by addition of 50 mM sucrose. A 65 min superfusion of the cells with a guanylate cyclase inhibitor [methylene blue (MB)], a protein kinase inhibitor (H7) and a calcium-free buffer containing 10 mM Mg2+ (OCa-10Mg) also increased Tau release and cAMP level in the cells. Acute treatment with 5 mM ammonia of cells pretreated for 24 h with 1 mM ammonia or for 65 min with MB, H7 or OCa-10Mg produced additional significant stimulation of Tau release, without further increasing the cAMP level in the cells. By contrast, a 10-min treatment with 65 mM KCl, which is a potent, cAMP-independent stimulus of Tau release in untreated Müller glia, produced no further enhancement of Tau release in ammonia-, MB-, H7 or OCa-10Mg-pretreated cells. The results indicate that acute treatment with ammonia, on top of treatments that evoke Tau release associated with an increase of cAMP, produces an extra Tau release that is cAMP-independent. Tau released by this extra ammonia treatment possibly originates from a different pool than Tau liberated by the pretreatments or 65 mM KCl.
Insights
Ammonia stimulates taurine release from Müller glia via a cAMP-dependent pathway. Further ammonia exposure triggers additional, cAMP-independent taurine release, suggesting a distinct cellular mechanism.
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- Previous studies indicated ammonia stimulates taurine release in Müller glia through a cAMP-dependent mechanism.
- Ammonia, at concentrations of 0.5-5 mM, triggers osmoresistant taurine release.
Purpose of the Study:
- To investigate the effects of prolonged ammonia treatment on taurine release and intracellular cAMP levels.
- To determine if ammonia-induced taurine release involves cAMP-independent pathways.
Main Methods:
- Cultured Müller glia were treated with varying concentrations and durations of ammonia.
- Inhibitors of guanylate cyclase (MB), protein kinase (H7), and calcium-free buffer (OCa-10Mg) were used.
- Cells were subsequently stimulated with ammonia or KCl to assess additional taurine release.
Main Results:
- Prolonged ammonia treatment (24h, 1mM) increased taurine release and cAMP levels, similar to acute high-dose ammonia (5mM).
- Inhibitors (MB, H7, OCa-10Mg) also increased taurine release and cAMP.
- Acute ammonia treatment after these pretreatments caused additional taurine release, independent of further cAMP increase.
- KCl, a cAMP-independent stimulus, did not enhance taurine release in pretreated cells.
Conclusions:
- Acute ammonia exposure induces a cAMP-independent taurine release in Müller glia, distinct from other stimuli.
- This additional taurine release may originate from a separate intracellular pool.
- The findings elucidate complex regulatory mechanisms of taurine release in glial cells.