<p><em> In vivo</em> circuit and cellular level functional imaging is a critical tool for understanding the brain in action. High resolution imaging of mouse cortical neurons with two-photon microscopy has provided unique insights into cortical structure, function and plasticity. However, these studies are limited to head fixed animals, greatly reducing the behavioral complexity available for study. In this paper, we describe a procedure for performing chronic fluorescence microscopy with cellular-resolution across multiple cortical layers in freely behaving mice.

<p><span>Natural and drug rewards increase the motivational valence of stimuli in the environment that, through Pavlovian learning mechanisms, become conditioned stimuli that directly motivate behavior in the absence of the original unconditioned stimulus. While the hippocampus has received extensive attention for its role in learning and memory processes, less is known regarding its role in drug-reward associations.

<p><span>Learning to vocalize depends on the ability to adaptively modify the temporal and spectral features of vocal elements. Neurons that convey motor-related signals to the auditory system are theorized to facilitate vocal learning, but the identity and function of such neurons remain unknown. Here we identify a previously unknown neuron type in the songbird brain that transmits vocal motor signals to the auditory cortex. Genetically ablating these neurons in juveniles disrupted their ability to imitate features of an adult tutor's song.

<p><span>Memory-guided behavior requires maintenance of task-relevant information without sensory input, but the underlying circuit mechanism remains unclear. Calcium imaging in mice performing a delayed Go or No-Go task revealed robust delay activity in dorsomedial prefrontal cortex, with different pyramidal neurons signaling Go and No-Go action plans. Inhibiting pyramidal neurons by optogenetically activating somatostatin- or parvalbumin-positive interneurons, even transiently during the delay, impaired task performance, primarily by increasing inappropriate Go responses.

<p>Episodic memories initially require rapid synaptic plasticity within the hippocampus for their formation and are gradually consolidated in neocortical networks for permanent storage. However, the engrams and circuits that support neocortical memory consolidation have thus far been unknown.We found that neocortical prefrontal memory engram cells, which are critical for remote contextual fear memory, were rapidly generated during initial learning through inputs from both the hippocampal–entorhinal cortex network and the basolateral amygdala.

<p><span>The continuous addition of new dentate granule cells, exquisitely regulated by brain activity, renders the hippocampus plastic. However, how neural circuits encode experiences to impact the addition of adult-born neurons remains unknown. Here, we used endoscopic Ca</span>²⁺<span> imaging to track the real-time activity of individual dentate granule cells in freely-behaving mice.

<p><span>The brain’s ability to associate different stimuli is vital for long-term memory, but how neural ensembles encode associative memories is unknown. Here we studied how cell ensembles in the basal and lateral amygdala encode associations between conditioned and unconditioned stimuli (CS and US, respectively). Using a miniature fluorescence microscope, we tracked the Ca</span>²⁺<span>dynamics of ensembles of amygdalar neurons during fear learning and extinction over 6 days in behaving mice.

<p><span>Genetically encoded calcium indicators (GECIs) are mainly represented by two- or one-fluorophore-based sensors. One type of two-fluorophore-based sensor, carrying </span>Opsanus<span> troponin C (TnC) as the Ca</span>²⁺<span>-binding moiety, has two binding sites for calcium ions, providing a linear response to calcium ions. One-fluorophore-based sensors have four Ca</span>²⁺<span>-binding sites but are better suited for </span>in vivo<span> experiments.

<p><span>Memories about sensory experiences are tightly linked to the context in which they were formed. Memory contextualization is fundamental for the selection of appropriate behavioral reactions needed for survival, yet the underlying neuronal circuits are poorly understood. By combining </span><em>trans</em><span>-synaptic viral tracing and optogenetic manipulation, we found that the ventral hippocampus (vHC) and the amygdala, two key brain structures encoding context and emotional experiences, interact via multiple parallel pathways.

<p><span>Altered dopamine D2 receptor (D2R) binding in the striatum has been associated with abnormal motivation in neuropsychiatric disorders, including schizophrenia. Here, we tested whether motivational deficits observed in mice with upregulated D2Rs (D2R-OE</span>_dev<span> mice) are reversed by decreasing function of the striatopallidal “no-go” pathway.