Anterior cingulate cortex projections to the dorsal medial striatum underlie insomnia associated with chronic pain.

Chronic pain often leads to the development of sleep disturbances. However, the precise neural circuit mechanisms responsible for sleep disorders in chronic pain have remained largely unknown. Here, we present compelling evidence that hyperactivity of pyramidal neurons (PNs) in the anterior cingulate cortex (ACC) drives insomnia in a mouse model of nerve-injury-induced chronic pain. After nerve injury, ACC PNs displayed spontaneous hyperactivity selectively in periods of insomnia. We then show that ACC PNs were both necessary for developing chronic-pain-induced insomnia and sufficient to mimic sleep loss in naive mice. Importantly, combining optogenetics and electrophysiological recordings, we found that the ACC projection to the dorsal medial striatum (DMS) underlies chronic-pain-induced insomnia through enhanced activity and plasticity of ACC-DMS dopamine D1R neuron synapses. Our findings shed light on the pivotal role of ACC PNs in developing chronic-pain-induced sleep disorders.

Facilitation of Behavioral and Cortical Emergence from Isoflurane Anesthesia by GABAergic Neurons in Basal Forebrain.

General anesthesia shares many similarities with natural sleep in behavior and electroencephalogram (EEG) patterns. The latest evidence suggests that general anesthesia and sleep-wake behavior may share overlapping neural substrates. The GABAergic neurons in the basal forebrain (BF) have recently been demonstrated to play a key role in controlling wakefulness. It was hypothesized that BF GABAergic neurons may participate in the regulation of general anesthesia. Here, using in vivo fiber photometry, we found that the activity of BF GABAergic neurons was generally inhibited during isoflurane anesthesia, having obviously decreased during the induction of anesthesia and being gradually restored during the emergence from anesthesia, in Vgat-Cre mice of both sexes. Activation of BF GABAergic neurons with chemogenetic and optogenetic approaches decreased sensitivity to isoflurane, delayed induction, and accelerated emergence from isoflurane anesthesia. Optogenetic activation of BF GABAergic neurons decreased EEG δ power and the burst suppression ratio (BSR) during 0.8% and 1.4% isoflurane anesthesia, respectively. Similar to the effects of activating BF GABAergic cell bodies, photostimulation of BF GABAergic terminals in the thalamic reticular nucleus (TRN) also strongly promoted cortical activation and behavioral emergence from isoflurane anesthesia. Collectively, these results showed that the GABAergic BF is a key neural substrate for general anesthesia regulation that facilitates behavioral and cortical emergence from general anesthesia via the GABAergic BF-TRN pathway. Our findings may provide a new target for attenuating the depth of anesthesia and accelerating emergence from general anesthesia.SIGNIFICANCE STATEMENT The basal forebrain (BF) is a key brain region controlling sleep-wake behavior. Activation of GABAergic neurons in the BF potently promotes behavioral arousal and cortical activity. Recently, many sleep-wake-related brain structures have been reported to participate in the regulation of general anesthesia. However, it is still unclear what role BF GABAergic neurons play in general anesthesia. In this study, we aim to reveal the role of BF GABAergic neurons in behavioral and cortical emergence from isoflurane anesthesia and elucidate the underlying neural pathways. Understanding the specific role of BF GABAergic neurons in isoflurane anesthesia would improve our understanding of the mechanisms of general anesthesia and may provide a new strategy for accelerating emergence from general anesthesia.

A glutamatergic basal forebrain to midbrain circuit mediates wakefulness and defensive behavior.

Defensive behavior, a group of responses that evolved due to threatening stimuli, is crucial for animal survival in the natural environment. For defensive measures to be timely and successful, a high arousal state and immediate sleep-to-wakefulness transition are required. Recently, the glutamatergic basal forebrain (BF) has been implicated in sleep-wake regulation; however, the associated physiological functions and underlying neural circuits remain unknown. Here, using in vivo fiber photometry, we found that BF glutamatergic neuron is activated by various threatening stimuli, including predator odor, looming threat, sound, and tail suspension. Optogenetic activation of BF glutamatergic neurons induced a series of context-dependent defensive behaviors in mice, including escape, fleeing, avoidance, and hiding. Similar to the effects of activated BF glutamatergic cell body, photoactivation of BF glutamatergic terminals in the ventral tegmental area (VTA) strongly drove defensive behaviors in mice. Using synchronous electroencephalogram (EEG)/electromyogram (EMG) recording, we showed that photoactivation of the glutamatergic BF-VTA pathway produced an immediate transition from sleep to wakefulness and significantly increased wakefulness. Collectively, our results clearly demonstrated that the glutamatergic BF is a key neural substrate involved in wakefulness and defensive behaviors, and encodes these behaviors through glutamatergic BF-VTA pathway. Overexcitation of the glutamatergic BF-VTA pathway may be implicated in clinical psychiatric diseases characterized by exaggerated defensive responses, such as autism spectrum disorders.

Parabrachial nucleus astrocytes regulate wakefulness and isoflurane anesthesia in mice.

Background: The parabrachial nucleus (PBN) is an important structure regulating the sleep-wake behavior and general anesthesia. Astrocytes in the central nervous system modulate neuronal activity and consequential behavior. However, the specific role of the parabrachial nucleus astrocytes in regulating the sleep-wake behavior and general anesthesia remains unclear. Methods: We used chemogenetic approach to activate or inhibit the activity of PBN astrocytes by injecting AAV-GFAabc1d-hM3Dq-eGFP or AAV-GFAabc1d-hM4Di-eGFP into the PBN. We investigated the effects of intraperitoneal injection of CNO or vehicle on the amount of wakefulness, NREM sleep and REM sleep in sleep-wake behavior, and on the time of loss of righting reflex, time of recovery of righting reflex, sensitivity to isoflurane, electroencephalogram (EEG) power spectrum and burst suppression ratio (BSR) in isoflurane anesthesia. Results: The activation of PBN astrocytes increased wakefulness amount for 4 h, while the inhibition of PBN astrocytes decreased total amount of wakefulness during the 3-hour post-injection period. Chemogenetic activation of PBN astrocytes decreased isoflurane sensitivity and shortened the emergence time from isoflurane-induced general anesthesia. Cortical EEG recordings revealed that PBN astrocyte activation decreased the EEG delta power and BSR during isoflurane anesthesia. Chemogenetic Inhibition of PBN astrocytes increased the EEG delta power and BSR during isoflurane anesthesia. Conclusion: PBN astrocytes are a key neural substrate regulating wakefulness and emergence from isoflurane anesthesia.

miR­93­5p regulates the occurrence and development of esophageal carcinoma epithelial cells by targeting TGFßR2.

Emerging studies have indicated that the dysregulation of microRNAs (miRNAs or miRs) plays a vital role in the development and metastasis of tumors. However, the role of miR­93­5p in esophageal carcinoma (EC) has not been extensively reported. The present study thus focused on the role of miR­93­5p and its downstream target in the occurrence and development of EC. Firstly, miRNA expression profiles associated with EC were accessed from the TCGA_ESCA dataset and analyzed. Subsequently, the expression patterns of miR­93­5p and TGFßR2 were characterized in the human esophageal cell line, Het­1A, and the human EC cell lines, TE­1, Eca­109 and EC9706, by RT­qPCR and western blot analysis. WST­1 assay, flow cytometry, Transwell assay, wound healing assay and bioinformatics analysis were used to explore their functions in EC cells. Finally, a dual­luciferase reporter assay was employed to determine the targeted association between miR­93­5p and TGFßR2. The results revealed that the expression of miR­93­5p was markedly higher in EC cell lines compared with that in the normal cell line. The overexpression of miR­93­5p facilitated cell proliferation, migration and invasion, and inhibited cell apoptosis. Additionally, TGFßR2 was identified as a functional target of miR­93­5p in EC cells, as judged by a series of in vitro experiments. Furthermore, it was found that the simultaneous overexpression of miR­93­5p and TGFßR2 almost had no effect on the biological behaviors of EC cells. On the whole, the present study demonstrates that miR­93­5p promotes the proliferation, migration and invasion, and inhibits the apoptosis of EC cells by targeting TGFßR2.

Basal forebrain GABAergic neurons promote arousal and predatory hunting.

Predatory hunting is an important approach for animals to obtain valuable nutrition and energy, which critically depends on heightened arousal. Yet the neural substrates underlying predatory hunting remain largely undefined. Here, we report that basal forebrain (BF) GABAergic neurons play an important role in regulating predatory hunting. Our results showed that BF GABAergic neurons were activated during the prey (cricket)-hunting and food feeding in mice. Optogenetic activation of BF GABAergic neurons evoked immediate predatory-like actions to both artificial and natural preys, significantly reducing the attack latency while increasing the attack probability and the number of killed natural prey (crickets). Similar to the effect of activating the soma of BF GABAergic neurons, photoactivation of their terminals in the ventral tegmental area (VTA) also strongly promotes predatory hunting. Moreover, photoactivation of GABAergic BF - VTA pathway significantly increases the intake of various food in mice. By synchronous recording of electroencephalogram and electromyogram, we showed that photoactivation of GABAergic BF - VTA pathway induces instant arousal and maintains long-term wakefulness. In summary, our results clearly demonstrated that the GABAergic BF is a key neural substrate for predatory hunting, and promotes this behavior through GABAergic BF - VTA pathway.