CSF1R inhibitor PLX3397 depletes microglia in Mongolian gerbil Meriones unguiculatus, but not in syrian hamster Mesocricetus auratus.

Microglia are the residential immune cells in the central nervous system. Their roles as innate immune cells and regulators of synaptic remodeling are critical to the development and the maintenance of the brain. Numerous studies have depleted microglia to elucidate their involvement in healthy and pathological conditions. PLX3397, a blocker of colony stimulating factor 1 receptor (CSF1R), is widely used to deplete mouse microglia due to its non-invasiveness and convenience. Recently, other small rodents, including Syrian hamsters (Mesocricetus auratus) and Mongolian gerbils (Meriones unguiculatus), have been recognized as valuable animal models for studying brain functions and diseases. However, whether microglia depletion via PLX3397 is feasible in these species remains unclear. Here, we administered PLX3397 orally via food pellets to hamsters and gerbils. PLX3397 successfully depleted gerbil microglia but had no effect on microglial density in hamsters. Comparative analysis of the CSF1R amino acid sequence in different species hints that amino acid substitutions in the juxtamembrane domain may potentially contribute to the inefficacy of PLX3397 in hamsters.

Circadian control of sleep-related neuronal activity in lizards.

Although diurnal animals displaying monophasic sleep patterns exhibit periodic cycles of alternating slow-wave sleep (SWS) and rapid eye movement sleep (REMS), the regulatory mechanisms underlying these regular sleep cycles remain unclear. Here, we report that in the Australian dragon Pogona vitticeps exposed to constant darkness (DD), sleep behavior and sleep-related neuronal activity emerged over a 24-h cycle. However, the regularity of the REMS/SWS alternation was disrupted under these conditions. Notably, when the lizards were then exposed to 12 h of light after DD, the regularity of the sleep stages was restored. These results suggest that sleep-related neuronal activity in lizards is regulated by circadian rhythms and that the regularity of REMS and SWS cycling is influenced by daytime light exposure.

Methyl vinyl ketone impairs spatial memory and activates hippocampal glial cells in mice.

Memory is a fundamental brain function that can be affected by a variety of external factors including environmental pollutants. One of these pollutants is methyl vinyl ketone (MVK), a hazardous substance found in cigarettes, industrial wastes, and car exhaust. Humans can be exposed to MVK under many circumstances; however, it is unclear whether MVK affects higher-order brain functions such as memory. Here, we examined the memory performances of mice receiving systemic MVK administration. We found that 1 mg/kg of MVK impaired spatial memory. We also showed that 1 mg/kg MVK activated glial cells and altered glial functions in several subregions of the hippocampus, a brain region involved in learning and memory. These results suggest that MVK induces memory deficits and activates glial cells in hippocampal subregions.

Sleep sharp wave ripple and its functions in memory and synaptic plasticity.

Memory is one of the fundamental cognitive functions of brain. The formation and consolidation of memory depend on the hippocampus and sleep. Sharp wave ripple (SWR) is an electrophysiological event which is most frequently observed in the hippocampus during sleep. It represents a highly synchronized neuronal activity pattern which modulates numerous brain regions including the neocortex, subcortical areas, and the hippocampus itself. In this review, we discuss how SWRs link experiences to memories and what happens in the hippocampus and other brain regions during sleep by focusing on synaptic plasticity.

Ablation of microglia does not alter circadian rhythm of locomotor activity.

Microglia, as macrophages in the brain, are responsible for immune responses and synaptic remodeling. Although the function of microglia is regulated by circadian rhythms, it is still unclear whether microglia are involved in the generation and light entrainment of circadian rhythms of behavior. Here, we report that microglial depletion does not alter behavioral circadian rhythms. We depleted ~ 95% of microglia in the mouse brain by PLX3397, a CSF1R inhibitor, and analyzed the effect on the spontaneous behaviors of mice. We found that neither the free-running period under constant darkness nor light entrainment under jet-lag circumstances were influenced by the ablation of microglia. Our results demonstrate that the circadian rhythms of locomotor activity, an important output of the circadian clock in the brain, are likely a phenomenon not produced by microglia.

Methyl vinyl ketone disrupts neuronal survival and axonal morphogenesis.

Methyl vinyl ketone (MVK) is an environmental hazardous substrate which is mainly present in cigarette smoke, industrial waste, and exhaust gas. Despite many chances to be exposed to MVK, the cellular toxicity of MVK is largely unknown. Neurons are the main component of the brain, which is one the most vital organs to human beings. Nevertheless, the influence of MVK to neurons has not been investigated. Here, we determined whether MVK treatment negatively affects neuronal survival and axonal morphogenesis using primary hippocampal neuronal cultures. We treated hippocampal neurons with 0.1 µM to 3.0 µM MVK and observed a concentration-dependent increase of neuronal death rate. We also demonstrated that the treatment with a low concentration of MVK 0.1 µM or 0.3 µM inhibited axonal branching specifically without affecting axon outgrowth. Our results suggest that MVK is highly toxic to neurons.

A claustrum in reptiles and its role in slow-wave sleep.

The mammalian claustrum, owing to its widespread connectivity with other forebrain structures, has been hypothesized to mediate functions that range from decision-making to consciousness1. Here we report that a homologue of the claustrum, identified by single-cell transcriptomics and viral tracing of connectivity, also exists in a reptile-the Australian bearded dragon Pogona vitticeps. In Pogona, the claustrum underlies the generation of sharp waves during slow-wave sleep. The sharp waves, together with superimposed high-frequency ripples2, propagate to the entire neighbouring pallial dorsal ventricular ridge (DVR). Unilateral or bilateral lesions of the claustrum suppress the production of sharp-wave ripples during slow-wave sleep in a unilateral or bilateral manner, respectively, but do not affect the regular and rapidly alternating sleep rhythm that is characteristic of sleep in this species3. The claustrum is thus not involved in the generation of the sleep rhythm itself. Tract tracing revealed that the reptilian claustrum projects widely to a variety of forebrain areas, including the cortex, and that it receives converging inputs from, among others, areas of the mid- and hindbrain that are known to be involved in wake-sleep control in mammals4-6. Periodically modulating the concentration of serotonin in the claustrum, for example, caused a matching modulation of sharp-wave production there and in the neighbouring DVR. Using transcriptomic approaches, we also identified a claustrum in the turtle Trachemys scripta, a distant reptilian relative of lizards. The claustrum is therefore an ancient structure that was probably already present in the brain of the common vertebrate ancestor of reptiles and mammals. It may have an important role in the control of brain states owing to the ascending input it receives from the mid- and hindbrain, its widespread projections to the forebrain and its role in sharp-wave generation during slow-wave sleep.

Involvement of l-lactate in hippocampal dysfunction of type I diabetes.

Hippocampal neurons play a crucial role in memory formation. Accumulating evidence raises the possibility that hippocampal sharp-wave ripples (SW-Rs) are involved in memory consolidation. Here, we examined in an animal model of diabetes and found the amplitude of SW-Rs in diabetic mice were smaller than control group and were rescued by acute application of l-lactate, a major neural energy source. The cognitive impairment in diabetic mice was alleviated by intracerebroventricular l-lactate treatment. Our results suggested that l-lactate is important for hippocampal dysfunction in diabetes.

Central Histamine Boosts Perirhinal Cortex Activity and Restores Forgotten Object Memories.

BACKGROUND: A method that promotes the retrieval of lost long-term memories has not been well established. Histamine in the central nervous system is implicated in learning and memory, and treatment with antihistamines impairs learning and memory. Because histamine H3 receptor inverse agonists upregulate histamine release, the inverse agonists may enhance learning and memory. However, whether the inverse agonists promote the retrieval of forgotten long-term memory has not yet been determined. METHODS: Here, we employed multidisciplinary methods, including mouse behavior, calcium imaging, and chemogenetic manipulation, to examine whether and how the histamine H3 receptor inverse agonists, thioperamide and betahistine, promote the retrieval of a forgotten long-term object memory in mice. In addition, we conducted a randomized double-blind, placebo-controlled crossover trial in healthy adult participants to investigate whether betahistine treatment promotes memory retrieval in humans. RESULTS: The treatment of H3 receptor inverse agonists induced the recall of forgotten memories even 1 week and 1 month after training in mice. The memory recovery was mediated by the disinhibition of histamine release in the perirhinal cortex, which activated the histamine H2 receptor. Histamine depolarized perirhinal cortex neurons, enhanced their spontaneous activity, and facilitated the reactivation of behaviorally activated neuronal ensembles. A human clinical trial revealed that treatment of H3 receptor inverse agonists is specifically more effective for items that are more difficult to remember and subjects with poorer performance. CONCLUSIONS: These results highlight a novel interaction between the central histamine signaling and memory engrams.

Hippocampal ripples down-regulate synapses.

The specific effects of sleep on synaptic plasticity remain unclear. We report that mouse hippocampal sharp-wave ripple oscillations serve as intrinsic events that trigger long-lasting synaptic depression. Silencing of sharp-wave ripples during slow-wave states prevented the spontaneous down-regulation of net synaptic weights and impaired the learning of new memories. The synaptic down-regulation was dependent on the N-methyl-d-aspartate receptor and selective for a specific input pathway. Thus, our findings are consistent with the role of slow-wave states in refining memory engrams by reducing recent memory-irrelevant neuronal activity and suggest a previously unrecognized function for sharp-wave ripples.

Sphingosine 1­phosphate induced by hypoxia increases the expression of PAI­1 in HepG2 cells via HIF­1α.

Our group has recently reported that in the immortal human HepG2 liver cell line, sphingosine 1­phosphate (S1P) increases transcription of plasminogen activator inhibitor type­1 (PAI­1), the major physiological inhibitor of fibrinolysis, within 4 h. The present study aimed to elucidate the molecular mechanisms underlying this effect. PAI­1 expression was measured by reverse transcription­quantitative polymerase chain reaction and immunoblotting. It was demonstrated that S1P increased PAI­1 promoter activity but did not increase the activity of promoters lacking the hypoxia responsive element (HRE) 2. In addition, S1P transiently increased the concentration of hypoxia inducible factor (HIF)­1α, a transcription factor capable of binding to HRE. When HIF­1α was knocked down, the induction of transcription of PAI­1 by S1P was no longer observed. Sphingosine kinase (SPHK) activity is increased by hypoxia. It was demonstrated that increases in the concentration of the HIF­1α protein induced by hypoxia were prevented by treatment with SPHK inhibitor or S1P receptor antagonists. Thus, modification of the induction of HIF­1α by S1P, leading to increased transcription of PAI­1, may be an attractive therapeutic target for thrombosis and consequent inhibition of fibrinolysis associated with hypoxia.

Accurate detection of low signal-to-noise ratio neuronal calcium transient waves using a matched filter.

BACKGROUND: Calcium imaging has become a fundamental modality for studying neuronal circuit dynamics both in vitro and in vivo. However, identifying calcium events (CEs) from spectral data remains laborious and difficult, especially since the signal-to-noise ratio (SNR) often falls below 2. Existing automated signal detection methods are generally applied at high SNRs, leaving a large need for an automated algorithm that can accurately extract CEs from fluorescence intensity data of SNR 2 and below. NEW METHOD: In this work we develop a Matched filter for Multi-unit Calcium Event (MMiCE) detection to extract CEs from fluorescence intensity traces of simulated and experimentally recorded neuronal calcium imaging data. RESULTS: MMiCE reached perfect performance on simulated data with SNR ≥ 2 and a true positive (TP) rate of 98.27% (± 1.38% with a 95% confidence interval), and a false positive(FP) rate of 6.59% (± 2.56%) on simulated data with SNR 0.2. On real data, verified by patch-clamp recording, MMiCE performed with a TP rate of 100.00% (± 0.00) and a FP rate of 2.04% (± 4.10). COMPARISON WITH EXISTING METHOD(S): This high level of performance exceeds existing methods at SNRs as low as 0.2, which are well below those used in previous studies (SNR ≃ 5-10). CONCLUSION: Overall, the MMiCE detector performed exceptionally well on both simulated data, and experimentally recorded neuronal calcium imaging data. The MMiCE detector is accurate, reliable, well suited for wide-spread use, and freely available at sites.uci.edu/aggies or from the corresponding author.

Visual cortical prosthesis with a geomagnetic compass restores spatial navigation in blind rats.

Allocentric sense is one of the major components that underlie spatial navigation. In blind patients, the difficulty in spatial exploration is attributed, at least partly, to the deficit of absolute direction perception. In support of this notion, we announce that blind adult rats can perform spatial tasks normally when externally provided with real-time feedback of their head directions. Head-mountable microstimulators coupled with a digital geomagnetic compass were bilaterally implanted in the primary visual cortex of adult rats whose eyelids had been sutured. These "blind" rats were trained to seek food pellets in a T-shaped maze or a more complicated maze. Within tens of trials, they learned to manage the geomagnetic information source to solve the mazes. Their performance levels and navigation strategies were similar to those of normal sighted, intact rats. Thus, blind rats can recognize self-location through extrinsically provided stereotactic cues.

Ex vivo cultured neuronal networks emit in vivo-like spontaneous activity.

Spontaneous neuronal activity is present in virtually all brain regions, but neither its function nor spatiotemporal patterns are fully understood. Ex vivo organotypic slice cultures may offer an opportunity to investigate some aspects of spontaneous activity, because they self-restore their networks that collapsed during slicing procedures. In hippocampal networks, we compared the levels and patterns of in vivo spontaneous activity to those in acute and cultured slices. We found that the firing rates and excitatory synaptic activity in the in vivo hippocampus are more similar to those in slice cultures compared to acute slices. The soft confidence-weighted algorithm, a machine learning technique without human bias, also revealed that hippocampal slice cultures resemble the in vivo hippocampus in terms of the overall tendency of the parameters of spontaneous activity.

Dopamine receptor activation reorganizes neuronal ensembles during hippocampal sharp waves in vitro.

Hippocampal sharp wave (SW)/ripple complexes are thought to contribute to memory consolidation. Previous studies suggest that behavioral rewards facilitate SW occurrence in vivo. However, little is known about the precise mechanism underlying this enhancement. Here, we examined the effect of dopaminergic neuromodulation on spontaneously occurring SWs in acute hippocampal slices. Local field potentials were recorded from the CA1 region. A brief (1 min) treatment with dopamine led to a persistent increase in the event frequency and the magnitude of SWs. This effect lasted at least for our recording period of 45 min and did not occur in the presence of a dopamine D1/D5 receptor antagonist. Functional multineuron calcium imaging revealed that dopamine-induced SW augmentation was associated with an enriched repertoire of the firing patterns in SW events, whereas the overall tendency of individual neurons to participate in SWs and the mean number of cells participating in a single SW were maintained. Therefore, dopaminergic activation is likely to reorganize cell assemblies during SWs.

Unbalanced excitability underlies offline reactivation of behaviorally activated neurons.

Hippocampal sharp waves (SWs)/ripples represent the reactivation of neurons involved in recently acquired memory and are crucial for memory consolidation. By labeling active cells with fluorescent protein under the control of an immediate-early gene promoter, we found that neurons that had been activated while mice explored a novel environment were preferentially reactivated during spontaneous SWs in hippocampal slices in vitro. During SWs, the reactivated neurons received strong excitatory synaptic inputs as opposed to a globally tuned network balance between excitation and inhibition.

Subicular activation preceding hippocampal ripples in vitro.

Sharp wave-ripple complexes (SW-Rs), a transient form of high-frequency field oscillations observed in the hippocampus, are thought to mediate memory consolidation. They are initiated mainly in hippocampal CA3 area and propagate to the entorhinal cortex through the subiculum; however, little is known about how SW-Rs are initiated and propagate. Here, we used functional multineuronal calcium imaging to monitor SW-R-relevant neuronal activity from the subiculum at single-cell resolution. An unexpected finding was that a subset of subicular neurons was activated immediately before hippocampal SW-Rs. The SW-R-preceding activity was not abolished by surgical lesion of the CA1-to-subiculum projection, and thus, it probably arose from entorhinal inputs. Therefore, SW-Rs are likely to be triggered by entorhinal-to-CA3/CA1 inputs. Moreover, the subiculum is not merely a passive intermediate region that SW-Rs pass through, but rather, it seems to contribute to an active modification of neural information related to SW-Rs.

Layer III neurons control synchronized waves in the immature cerebral cortex.

Correlated spiking activity prevails in immature cortical networks and is believed to contribute to neuronal circuit maturation; however, its spatiotemporal organization is not fully understood. Using wide-field calcium imaging from acute whole-brain slices of rat pups on postnatal days 1-6, we found that correlated spikes were initiated in the anterior part of the lateral entorhinal cortex and propagated anteriorly to the frontal cortex and posteriorly to the medial entorhinal cortex, forming traveling waves that engaged almost the entire cortex. The waves were blocked by ionotropic glutamatergic receptor antagonists but not by GABAergic receptor antagonists. During wave events, glutamatergic and GABAergic synaptic inputs were balanced and induced UP state-like depolarization. Magnified monitoring with cellular resolution revealed that the layer III neurons were first activated when the waves were initiated. Consistent with this finding, layer III contained a larger number of neurons that were autonomously active, even under a blockade of synaptic transmission. During wave propagation, the layer III neurons constituted a leading front of the wave. The waves did not enter the parasubiculum; however, in some cases, they were reflected at the parasubicular border and propagated back in the opposite direction. During this reflection process, the layer III neurons in the medial entorhinal cortex maintained persistent activity. Thus, our data emphasize the role of layer III in early network behaviors and provide insight into the circuit mechanisms through which cerebral cortical networks maturate.

Muscarinic receptor activation disrupts hippocampal sharp wave-ripples.

Cholinergic muscarinic innervations to the hippocampus play a role in learning and memory. Here we report that pharmacological activation of muscarinic receptors eliminates sharp wave-ripple events in the mouse hippocampal CA1 region in vivo and in vitro. This effect was associated with a decorrelation of excitatory synaptic inputs and a net increase in inhibitory conductances in pyramidal neurons. Multineuron calcium imaging revealed that muscarinic activation altered the spatiotemporal pattern of network activities. Thus, cholinergic input is likely to contribute to a neuromodulatory switch of hippocampal network states, as proposed in the "two-stage" model of learning processes.