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A. Feigenspan, T. Schubert, R. Weiler; Intracellular Calcium Is Regulated By Different Pathways in Horizontal Cells of the Mouse Retina . Invest. Ophthalmol. Vis. Sci. 2005;46(13):1121.
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Purpose: To study the regulation of intracellular calcium concentration in isolated horizontal cells of the mouse retina. Methods: Horizontal cells were enzymatically and mechanically dissociated from the retina and visually identified according to their unique morphology. Voltage–clamp and current–clamp recording in the whole–cell configuration were combined with calcium imaging techniques and performed on isolated horizontal cells. Results: All horizontal cells showed a non–inactivating inward current upon depolarization from a holding potential of –60 mV. This current activated at around –20 mV, reached a peak amplitude of –79.1 ± 2.9 pA at 10 mV (n=125) and reversed sign at around 50 mV. The current was insensitive to TTX (1 µM), and it was partially blocked by the L–type channel antagonist verapamil (100 µM). The N–type channel blocker ω–conotoxin GVIA (1 µM) induced an additional reduction of current amplitudes. We never observed transient calcium currents, suggesting expression of L– and N–type calcium channels, but not T–type channels. For imaging experiments, cells were incubated in Ringer solution containing 15 µM Fura–2 AM and 0.01% Pluronic F–127. The specific glutamate receptor agonists AMPA (100 µM) and kainate (100 µM) evoked calcium signals in the presence of extracellular cadmium (250 µM). Two agonists of the metabotropic glutamate receptor mGluR1, trans–ACPD (200 µM) and quisqualate (100 µM), had no effect on intracellular calcium in the presence of the ionotropic glutamate receptor antagonist CNQX (10 µM). However, the intracellular calcium concentration was increased by application of coffeine (10 mM), indicating release of calcium from internal stores via ryanodine receptors. Conclusions: These data show that several pathways cause an increase of intracellular calcium in horizontal cells of the mouse retina: (1) activation of L–type and N–type calcium channels by membrane depolarization, (2) binding of glutamate to ionotropic AMPA– and kainate–type receptors, (3) release of calcium from internal stores after activation of ryanodine receptors, but not by activation of metabotropic GluR1. The fine tuning of intracellular calcium concentration during excitatory synaptic activity may thus serve to regulate feedback mechanisms at the photoreceptor synapse.
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