A positively tuned voltage indicator for extended electrical recordings in the brain.

Genetically encoded voltage indicators (GEVIs) enable optical recording of electrical signals in the brain, providing subthreshold sensitivity and temporal resolution not possible with calcium indicators. However, one- and two-photon voltage imaging over prolonged periods with the same GEVI has not yet been demonstrated. Here, we report engineering of ASAP family GEVIs to enhance photostability by inversion of the fluorescence-voltage relationship. Two of the resulting GEVIs, ASAP4b and ASAP4e, respond to 100-mV depolarizations with ≥180% fluorescence increases, compared with the 50% fluorescence decrease of the parental ASAP3. With standard microscopy equipment, ASAP4e enables single-trial detection of spikes in mice over the course of minutes. Unlike GEVIs previously used for one-photon voltage recordings, ASAP4b and ASAP4e also perform well under two-photon illumination. By imaging voltage and calcium simultaneously, we show that ASAP4b and ASAP4e can identify place cells and detect voltage spikes with better temporal resolution than commonly used calcium indicators. Thus, ASAP4b and ASAP4e extend the capabilities of voltage imaging to standard one- and two-photon microscopes while improving the duration of voltage recordings.

Progressive long-term spatial memory loss following repeat concussive and subconcussive brain injury in mice, associated with dorsal hippocampal neuron loss, microglial phenotype shift, and vascular abnormalities.

There is considerable concern about the long-term deleterious effects of repeat head trauma on cognition, but little is known about underlying mechanisms and pathology. To examine this, we delivered four air blasts to the left side of the mouse cranium, a week apart, with an intensity that causes deficits when delivered singly and considered "concussive," or an intensity that does not yield significant deficits when delivered singly and considered "subconcussive." Neither repeat concussive nor subconcussive blast produced spatial memory deficits at 4 months, but both yielded deficits at 14 months, and dorsal hippocampal neuron loss. Hierarchical cluster analysis of dorsal hippocampal microglia across the three groups based on morphology and expression of MHCII, CX3CR1, CD68 and IBA1 revealed five distinct phenotypes. Types 1A and 1B microglia were more common in sham mice, linked to better neuron survival and memory, and appeared mildly activated. By contrast, 2B and 2C microglia were more common in repeat concussive and subconcussive mice, linked to poorer neuron survival and memory, and characterized by low expression levels and attenuated processes, suggesting they were de-activated and dysfunctional. In addition, endothelial cells in repeat concussive mice exhibited reduced CD31 and eNOS expression, which was correlated with the prevalence of type 2B and 2C microglia. Our findings suggest that both repeat concussive and subconcussive head injury engender progressive pathogenic processes, possibly through sustained effects on microglia that over time lead to increased prevalence of dysfunctional microglia, adversely affecting neurons and blood vessels, and thereby driving neurodegeneration and memory decline.

Non-spatial hippocampal behavioral timescale synaptic plasticity during working memory is gated by entorhinal inputs.

Behavioral timescale synaptic plasticity (BTSP) is a form of synaptic potentiation where the occurrence of a single large plateau potential in CA1 hippocampal neurons leads to the formation of reliable place fields during spatial learning tasks. We asked whether BTSP could also be a plasticity mechanism for generation of non-spatial responses in the hippocampus and what roles the medial and lateral entorhinal cortex (MEC and LEC) play in driving non-spatial BTSP. By performing simultaneous calcium imaging of dorsal CA1 neurons and chemogenetic inhibition of LEC or MEC while mice performed an olfactory working memory task' we discovered BTSP-like events which formed stable odor-specific fields. Critically' the success rate of calcium events generating a significant odor-field increased with event amplitude' and large events exhibited asymmetrical formation with the newly formed odor-fields preceding the timepoint of their induction event. We found that MEC and LEC play distinct roles in modulating BTSP: MEC inhibition reduced the frequency of large calcium events' while LEC inhibition reduced the success rate of odor-field generation. Using two-photon calcium imaging of LEC and MEC temporammonic axons projecting to CA1 we found that LEC projections to CA1 were strongly odor selective even early in task learning' while MEC projection odor-selectivity increased with task learning but remained weaker than LEC. Finally' we found that LEC and MEC inhibition both slowed representational drift of odor representations in CA1 across 48 hours. Altogether' odor-specific information from LEC and strong odor-timed activity from MEC are crucial for driving BTSP in CA1 which is a synaptic plasticity mechanism for generation of both spatial and non-spatial responses in the hippocampus that may play a role in explaining representational drift and one-shot learning of non-spatial information.

Differential Emergence and Stability of Sensory and Temporal Representations in Context-Specific Hippocampal Sequences.

Hippocampal spiking sequences encode external stimuli and spatiotemporal intervals, linking sequential experiences in memory, but the dynamics controlling the emergence and stability of such diverse representations remain unclear. Using two-photon calcium imaging in CA1 while mice performed an olfactory working-memory task, we recorded stimulus-specific sequences of "odor-cells" encoding olfactory stimuli followed by "time-cells" encoding time points in the ensuing delay. Odor-cells were reliably activated and retained stable fields during changes in trial structure and across days. Time-cells exhibited sparse and dynamic fields that remapped in both cases. During task training, but not in untrained task exposure, time-cell ensembles increased in size, whereas odor-cell numbers remained stable. Over days, sequences drifted to new populations with cell activity progressively converging to a field and then diverging from it. Therefore, CA1 employs distinct regimes to encode external cues versus their variable temporal relationships, which may be necessary to construct maps of sequential experiences.