Researchers developed ultra-sensitive fluorescent calcium sensors (GCaMP6) for imaging neuronal activity. These sensors outperformed existing ones in cultured neurons and in vivo in zebrafish, flies, and mice. GCaMP6 reliably detected single action potentials in neuronal somata and orientation-tuned synaptic calcium transients in individual dendritic spines. Orientation tuning of spines was stable over weeks, and averaged across spine populations predicted the tuning of their parent cell. GABAergic neurons showed little orientation tuning, but their dendrites had highly tuned segments. GCaMP6 provides new insights into neural circuit organization and dynamics across multiple spatial and temporal scales. The study used structure-based mutagenesis and neuron-based screening to develop GCaMP6, which has higher sensitivity and faster kinetics than previous versions. GCaMP6s were tested in cultured neurons and validated in in vivo systems. They showed improved sensitivity and faster kinetics compared to other GCaMP variants. GCaMP6s detected single action potentials with high reliability and resolved step-wise fluorescence increases in bursts of spikes. In the mouse visual cortex, GCaMP6 detected orientation-tuned synaptic calcium signals in dendritic spines. Spine responses were stable over weeks, and orientation tuning was consistent across spines. Spines often had distinct preferred orientations, and the orientation tuning of spines predicted the orientation tuning of their parent neurons. In GABAergic neurons, dendrites showed pronounced orientation-tuned domains, with multiple domains preferring different orientations. The orientation selectivity index (OSI) of dendritic segments was higher than for interneuron somata but lower than for pyramidal neurons. The large size of these domains may reflect spatially clustered input with shared orientation preference or a few strong inputs amplified by local postsynaptic mechanisms. The study concludes that GCaMP6 sensors have greatly improved properties and can be used for imaging large groups of neurons and synaptic compartments over multiple imaging sessions. These sensors are likely to find widespread applications in brain research and calcium signaling. Future efforts may focus on red fluorescent calcium indicators for deeper tissue imaging.Researchers developed ultra-sensitive fluorescent calcium sensors (GCaMP6) for imaging neuronal activity. These sensors outperformed existing ones in cultured neurons and in vivo in zebrafish, flies, and mice. GCaMP6 reliably detected single action potentials in neuronal somata and orientation-tuned synaptic calcium transients in individual dendritic spines. Orientation tuning of spines was stable over weeks, and averaged across spine populations predicted the tuning of their parent cell. GABAergic neurons showed little orientation tuning, but their dendrites had highly tuned segments. GCaMP6 provides new insights into neural circuit organization and dynamics across multiple spatial and temporal scales. The study used structure-based mutagenesis and neuron-based screening to develop GCaMP6, which has higher sensitivity and faster kinetics than previous versions. GCaMP6s were tested in cultured neurons and validated in in vivo systems. They showed improved sensitivity and faster kinetics compared to other GCaMP variants. GCaMP6s detected single action potentials with high reliability and resolved step-wise fluorescence increases in bursts of spikes. In the mouse visual cortex, GCaMP6 detected orientation-tuned synaptic calcium signals in dendritic spines. Spine responses were stable over weeks, and orientation tuning was consistent across spines. Spines often had distinct preferred orientations, and the orientation tuning of spines predicted the orientation tuning of their parent neurons. In GABAergic neurons, dendrites showed pronounced orientation-tuned domains, with multiple domains preferring different orientations. The orientation selectivity index (OSI) of dendritic segments was higher than for interneuron somata but lower than for pyramidal neurons. The large size of these domains may reflect spatially clustered input with shared orientation preference or a few strong inputs amplified by local postsynaptic mechanisms. The study concludes that GCaMP6 sensors have greatly improved properties and can be used for imaging large groups of neurons and synaptic compartments over multiple imaging sessions. These sensors are likely to find widespread applications in brain research and calcium signaling. Future efforts may focus on red fluorescent calcium indicators for deeper tissue imaging.