The authors developed a new class of genetically encoded calcium indicators, GCaMP6, which outperforms existing sensors in terms of sensitivity and speed. GCaMP6 was engineered through structure-based mutagenesis and neuron-based screening, and it demonstrated superior performance in cultured neurons and in vivo systems such as zebrafish, flies, and mice. In mouse visual cortex pyramidal neurons, GCaMP6 reliably detected single action potentials and orientation-tuned synaptic calcium transients in dendritic spines. Orientation tuning of these spines was stable over weeks, and the average tuning across spine populations predicted the tuning of their parent cells. In contrast, somata of GABAergic neurons showed little orientation tuning, but their dendrites contained highly tuned segments. GCaMP6 sensors provide new tools for studying the organization and dynamics of neural circuits over multiple spatial and temporal scales.The authors developed a new class of genetically encoded calcium indicators, GCaMP6, which outperforms existing sensors in terms of sensitivity and speed. GCaMP6 was engineered through structure-based mutagenesis and neuron-based screening, and it demonstrated superior performance in cultured neurons and in vivo systems such as zebrafish, flies, and mice. In mouse visual cortex pyramidal neurons, GCaMP6 reliably detected single action potentials and orientation-tuned synaptic calcium transients in dendritic spines. Orientation tuning of these spines was stable over weeks, and the average tuning across spine populations predicted the tuning of their parent cells. In contrast, somata of GABAergic neurons showed little orientation tuning, but their dendrites contained highly tuned segments. GCaMP6 sensors provide new tools for studying the organization and dynamics of neural circuits over multiple spatial and temporal scales.