Glial Ca2+ excitability plays a key role in reciprocal neuron-glia communication. In the retina, neuron-glia signaling is expected to be maximal in the dark, but the glial Ca2+ signal characteristics under such conditions have not been evaluated. To address this question, we used a bioluminescent imaging to monitor spontaneous Ca2+ changes under dark conditions, selectively in Müller cells, the principal retinal glial cells. By combining this imaging approach with network analysis, we demonstrate that activity in Müller cells is organized in networks of coactive cells, involving 2 to 16 cells located distantly and/or in clusters. We also report that spontaneous activity of small networks (2 to 6 Müller cells) repeat over time, sometimes in the same sequential order, revealing specific temporal dynamics. In addition, we show that networks of coactive glial cells are inhibited by TTX, indicating that ganglion and/or amacrine neuronal cells probably regulate Müller cell network properties. These results represent the first demonstration that spontaneous activity in adult Müller cells is patterned into correlated networks that display repeated sequences of coactivations over time. Furthermore, our bioluminescent technique provides a novel tool to study the dynamic characteristics of glial Ca2+ events in the retina under dark conditions, which should greatly facilitate future investigations of retinal dark-adaptive processes.