Search Results (27)

Accumulating studies have shown that astrocytes are essential for regulating neurons at both synaptic and circuit levels. The main mechanism of brain astrocytic intracellular Ca²⁺ activity is through the release of Ca²⁺ via the inositol 1,4,5-trisphosphate receptor type 2 (IP3R2) from the endoplasmic reticulum (ER). Studies using IP3R2 knockout mouse models (Itpr2^−/−) have shown that eliminating IP3R2 leads to a significant reduction in astrocytic Ca²⁺ activity However, there is ongoing controversy regarding the effect of this IP3R2-dependent reduction in astrocytic Ca²⁺ transients on neuronal activity. In our study, we employed dual-color two-photon Ca²⁺ imaging to study astrocytes and neurons simultaneously in vibrissa somatosensory cortex (vS1) in awake-behaving wild-type and Itpr2^−/− mice. We systematically characterized and compared both recorded astrocytic and neuronal Ca²⁺ activities in wild-type and Itpr2^−/− mice during various animal behaviors, particularly during the transition period from stillness to locomotion. We report that vS1 astrocytic Ca²⁺ elevation in both wild-type and Itpr2^−/− mice was significantly modulated by free whisking and locomotion. However, vS1 neurons were only significantly modulated by locomotion in wild-type mice, but not in Itpr2^−/− mice. Our study suggests a non-synaptic modulatory mechanism on functions of astrocytic IP3R2-dependent Ca²⁺ transients to local neurons. Full article

Brain function declines because of aging and several metabolites change their concentration. However, this decrease may be a consequence or a driver of aging. It has been described that taurine levels decrease with age and that taurine supplementation increases health span in mice and monkeys, finding taurine as a driver of aging. The frontal cortex is one of the most key areas studied to know the normal processes of cerebral aging, due to its relevant role in cognitive processes, emotion, and motivation. In the present work, we analyzed by intracerebral microdialysis in vivo in the prefrontal cortex of young (3 months) and old (24 months) awake rats, the basal- and K⁺-evoked release of taurine, and its precursors methionine and serine. The taurine/serine/methionine (TSM) ratio was also calculated as an index of transmethylation reactions. No changes were found in the basal levels of taurine, serine, or methionine between young and aged animals. On the contrary, a significant decrease in the K⁺-evoked release of serine and taurine appeared in aged rats when compared with young animals. No changes were seen in methionine. TSM ratio also decreased with age in both basal- and K⁺-stimulated conditions. Therefore, taurine and its related precursor serine decrease with age in the frontal cortex of aged animals under K⁺-stimulated but not basal conditions, which supports the importance of the decline of evoked taurine in its functions at the brain level, also supporting the idea proposed by other authors of a pharmacological and/or nutritional intervention to its restoration. A deficit of precursors for transmethylation reactions in the brain with age is also considered. Full article

The preoptic area of the hypothalamus is critical for regulation of brain–body interaction, including circuits that control vital signs such as body temperature and heart rate. The preoptic area contains approximately 70 molecularly distinct cell types. The Gabre gene is expressed in a subset of preoptic area cell types. It encodes the GABA receptor ε-subunit, which is thought to confer resistance to anesthetics at the molecular level, but the function of Gabre cells in the brain remains largely unknown. We generated and have extensively characterized a Gabre-cre knock-in mouse line and used chemogenetic tools to interrogate the function of Gabre cells in the preoptic area. Comparison with macaque GABRE expression revealed the conserved character of Gabre cells in the preoptic area. In awake mice, we found that chemogenetic activation of Gabre neurons in the preoptic area reduced body temperature, whereas chemogenetic inhibition had no effect. Furthermore, chemogenetic inhibition of Gabre neurons in the preoptic area decreased the heart rate, whereas chemogenetic activation had no effect under isoflurane anesthesia. These findings suggest an important role of preoptic Gabre neurons in maintaining vital signs such as body temperature and heart rate during wakefulness and under anesthesia. Full article

When exposed to X-rays, scintillators emit visible luminescence. X-ray-mediated optogenetics employs scintillators for remotely activating light-sensitive proteins in biological tissue through X-ray irradiation. This approach offers advantages over traditional optogenetics, allowing for deeper tissue penetration and wireless control. Here, we assessed the short-term safety and efficacy of candidate scintillator materials for neuronal control. Our analyses revealed that lead-free halide scintillators, such as Cs₃Cu₂I₅, exhibited significant cytotoxicity within 24 h and induced neuroinflammatory effects when injected into the mouse brain. In contrast, cerium-doped gadolinium aluminum gallium garnet (Ce:GAGG) nanoparticles showed no detectable cytotoxicity within the same period, and injection into the mouse brain did not lead to observable neuroinflammation over four weeks. Electrophysiological recordings in the cerebral cortex of awake mice showed that X-ray-induced radioluminescence from Ce:GAGG nanoparticles reliably activated 45% of the neuronal population surrounding the implanted particles, a significantly higher activation rate than europium-doped GAGG (Eu:GAGG) microparticles, which activated only 10% of neurons. Furthermore, we established the cell-type specificity of this technique by using Ce:GAGG nanoparticles to selectively stimulate midbrain dopamine neurons. This technique was applied to freely behaving mice, allowing for wireless modulation of place preference behavior mediated by midbrain dopamine neurons. These findings highlight the unique suitability of Ce:GAGG nanoparticles for X-ray-mediated optogenetics. The deep tissue penetration, short-term safety, wireless neuronal control, and cell-type specificity of this system offer exciting possibilities for diverse neuroscience applications and therapeutic interventions. Full article

The dorsal raphe nucleus (DRN) has gained attention owing to its involvement in various physiological functions, such as sleep–awake, feeding, and emotion, with its analgesic role being particularly significant. It is described as the “pain inhibitory nucleus” in the brain. The DRN has diverse projections from hypothalamus, midbrain, and pons. In turn, the DRN is a major source of projections to diverse cortex, limbic forebrain thalamus, and the midbrain and contains highly heterogeneous neuronal subtypes. The activation of DRN neurons in mice prevents the establishment of neuropathic, chronic pain symptoms. Chemogenetic or optogenetic inhibition neurons in the DRN are sufficient to establish pain phenotypes, including long-lasting tactile allodynia, that scale with the extent of stimulation, thereby promoting nociplastic pain. Recent progress has been made in identifying the neural circuits and cellular mechanisms in the DRN that are responsible for sensory modulation. However, there is still a lack of comprehensive review addressing the specific neuron types in the DRN involved in pain modulation. This review summarizes the function of specific cell types within DRN in the pain regulation, and aims to improve understanding of the mechanisms underlying pain regulation in the DRN, ultimately offering insights for further exploration. Full article

We have recently shown that the volatile anesthetics isoflurane and sevoflurane acutely enhance the brain uptake of the hydrophilic markers sucrose and mannitol about two-fold from an awake condition, while the combined injection of the anesthetic agents ketamine and xylazine has no effect. The present study investigated two small-molecule hydrophilic drugs with potential neurotoxicity, the antibiotic agents ceftazidime and gentamicin. Transport studies using an in vitro blood–brain barrier (BBB) model, a monolayer of induced pluripotent stem cell-derived human brain microvascular endothelial cells seeded on Transwells, and LC-MS/MS analysis demonstrated the low permeability of both drugs in the range of sucrose, with permeability coefficients of 6.62 × 10⁻⁷ ± 2.34 × 10⁻⁷ cm/s for ceftazidime and 7.38 × 10⁻⁷ ± 2.29 × 10⁻⁷ cm/s for gentamicin. In vivo brain uptake studies of ceftazidime or gentamicin after IV doses of 25 mg/kg were performed in groups of 5–6 mice anesthetized at typical doses for surgical procedures with either isoflurane (1.5–2% v/v) or ketamine/xylazine (100:10 mg/kg I.P.). The brain uptake clearance, K_in, for ceftazidime increased from 0.033 ± 0.003 μL min⁻¹ g⁻¹ in the ketamine/xylazine group to 0.057 ± 0.006 μL min⁻¹ g⁻¹ in the isoflurane group (p = 0.0001), and from 0.052 ± 0.016 μL min⁻¹ g⁻¹ to 0.101 ± 0.034 μL min⁻¹ g⁻¹ (p = 0.0005) for gentamicin. We did not test the dose dependency of the uptake, because neither ceftazidime nor gentamicin are known substrates of any active uptake or efflux transporters at the BBB. In conclusion, the present study extends our previous findings with permeability markers and suggests that inhalational anesthetic isoflurane increases the BBB permeability of hydrophilic small-molecule endobiotics or xenobiotics when compared to the injection of ketamine/xylazine. This may be of clinical relevance in the case of potential neurotoxic substances. Full article

Mice are increasingly used as models of human-acquired neurological or neurodevelopmental conditions, such as autism, schizophrenia, and Alzheimer’s disease. All these conditions involve central auditory processing disorders, which have been little investigated despite their potential for providing interesting insights into the mechanisms behind such disorders. Alterations of the auditory steady-state response to 40 Hz click trains are associated with an imbalance between neuronal excitation and inhibition, a mechanism thought to be common to many neurological disorders. Here, we demonstrate the value of presenting click trains at various rates to mice with chronically implanted pins above the inferior colliculus and the auditory cortex for obtaining easy, reliable, and long-lasting access to subcortical and cortical complex auditory processing in awake mice. Using this protocol on a mutant mouse model of autism with a defect of the Shank3 gene, we show that the neural response is impaired at high click rates (above 60 Hz) and that this impairment is visible subcortically—two results that cannot be obtained with classical protocols for cortical EEG recordings in response to stimulation at 40 Hz. These results demonstrate the value and necessity of a more complete investigation of central auditory processing disorders in mouse models of neurological or neurodevelopmental disorders. Full article

(1) From mouse to man, shaking behavior (head twitches and/or wet dog shakes) is a reliable readout of psychedelic drug action. Shaking behavior like psychedelia is thought to be mediated by serotonin 2A receptors on cortical pyramidal cells. The involvement of pyramidal cells in psychedelic-induced shaking behavior remains hypothetical, though, as experimental in vivo evidence is limited. (2) Here, we use cell type-specific voltage imaging in awake mice to address this issue. We intersectionally express the genetically encoded voltage indicator VSFP Butterfly 1.2 in layer 2/3 pyramidal neurons. We simultaneously capture cortical hemodynamics and cell type-specific voltage activity while mice display psychedelic shaking behavior. (3) Shaking behavior is preceded by high-frequency oscillations and overlaps with low-frequency oscillations in the motor cortex. Oscillations spectrally mirror the rhythmics of shaking behavior and reflect layer 2/3 pyramidal cell activity complemented by hemodynamics. (4) Our results reveal a clear cortical fingerprint of serotonin-2A-receptor-mediated shaking behavior and open a promising methodological avenue relating a cross-mammalian psychedelic effect to cell-type specific brain dynamics. Full article

Activation of intravesical protease activated receptors-4 (PAR4) results in bladder pain through the release of urothelial macrophage migration inhibitory factor (MIF) and high mobility group box-1 (HMGB1). We aimed to identify HMGB1 downstream signaling events at the bladder that mediate HMGB1-induced bladder pain in MIF-deficient mice to exclude any MIF-related effects. We studied whether oxidative stress and ERK activation are involved by examining bladder tissue in mice treated with intravesical disulfide HMGB1 for 1 h and analyzed with Western blot and immunohistochemistry. HMGB1 intravesical treatment increased urothelium 4HNE and phospho-ERK1/2 staining, suggesting that HMGB1 increased urothelial oxidative stress and ERK activation. Furthermore, we examined the functional roles of these events. We evaluated lower abdominal mechanical thresholds (an index of bladder pain) before and 24 h after intravesical PAR4 or disulfide HMGB1. Intravesical pre-treatments (10 min prior) included: N-acetylcysteine amide (NACA, reactive oxygen species scavenger) and FR180204 (FR, selective ERK1/2 inhibitor). Awake micturition parameters (voided volume; frequency) were assessed at 24 h after treatment. Bladders were collected for histology at the end of the experiment. Pre-treatment with NACA or FR significantly prevented HMGB1-induced bladder pain. No significant effects were noted on micturition volume, frequency, inflammation, or edema. Thus, HMGB1 activates downstream urothelial oxidative stress production and ERK1/2 activation to mediate bladder pain. Further dissection of HMGB1 downstream signaling pathway may lead to novel potential therapeutic strategies to treat bladder pain. Full article

JWH-018 is the most known compound among synthetic cannabinoids (SCs) used for their psychoactive effects. SCs-based products are responsible for several intoxications in humans. Cardiac toxicity is among the main side effects observed in emergency departments: SCs intake induces harmful effects such as hypertension, tachycardia, chest pain, arrhythmias, myocardial infarction, breathing impairment, and dyspnea. This study aims to investigate how cardio-respiratory and vascular JWH-018 (6 mg/kg) responses can be modulated by antidotes already in clinical use. The tested antidotes are amiodarone (5 mg/kg), atropine (5 mg/kg), nifedipine (1 mg/kg), and propranolol (2 mg/kg). The detection of heart rate, breath rate, arterial oxygen saturation (SpO2), and pulse distention are provided by a non-invasive apparatus (Mouse Ox Plus) in awake and freely moving CD-1 male mice. Tachyarrhythmia events are also evaluated. Results show that while all tested antidotes reduce tachycardia and tachyarrhythmic events and improve breathing functions, only atropine completely reverts the heart rate and pulse distension. These data may suggest that cardiorespiratory mechanisms of JWH-018-induced tachyarrhythmia involve sympathetic, cholinergic, and ion channel modulation. Current findings also provide valuable impetus to identify potential antidotal intervention to support physicians in the treatment of intoxicated patients in emergency clinical settings. Full article

Unwanted proteins and metabolic waste in cerebral spinal fluid are cleared from the brain by meningeal and nasal lymphatics and the perineural sheath of cranial nerves; however, the distribution and clearance of cerebral spinal fluid (CSF) along the subarachnoid space of the entire spinal cord is not fully understood. Cryo-fluorescence tomography (CFT) was used to follow the movement of tracers from the ventricular system of the brain down through the meningeal lining of the spinal cord and out to the spinal lymphatic nodes. Isoflurane-anesthetized mice were infused into the lateral cerebroventricle with 5.0 µL of quantum dots [QdotR 605 ITKTM amino (PEG)] over two mins. Mice were allowed to recover (ca 2–3 min) and remained awake and ambulatory for 5, 15, 30, 60, and 120 min after which they were euthanized, and the entire intact body was frozen at −80^°. The entire mouse was sectioned, and white light and fluorescent images were captured after each slice to produce high resolution three-dimensional volumes. Tracer appeared throughout the ventricular system and central canal of the spinal cord and the entire subarachnoid space of the CNS. A signal could be visualized in the nasal cavity, deep cervical lymph nodes, thoracic lymph nodes, and more superficial submandibular lymph nodes as early as 15 min post infusion. A fluorescent signal could be visualized along the dorsal root ganglia and down the proximal extension of the spinal nerves of the thoracic and lumbar segments at 30 min. There was a significant accumulation of tracer in the lumbar and sacral lymph nodes between 15–60 min. The dense fluorescent signal in the thoracic vertebrae noted at 5- and 15-min post infusion was significantly reduced by 30 min. Indeed, all signals in the spinal cord were ostensibly absent by 120 min, except for trace amounts in the coccyx. The brain still had some residual signal at 120 min. These data show that Qdots with a hydrodynamic diameter of 16–20 nm rapidly clear from the brain of awake mice. These data also clearly demonstrate the rapid distribution and efflux of traces along a major length of the vertebral column and the potential contribution of the spinal cord in the clearance of brain waste. Full article

We examined bladder function following spinal cord injury (SCI) by repeated urodynamic investigation (UDI), including external urethral sphincter (EUS) electromyography (EMG) in awake restrained mice and correlated micturition parameters to gene expression and morphological changes in the bladder. A partial bladder outlet obstruction (pBOO) model was used for comparison to elucidate both the common and specific features of obstructive and neurogenic lower urinary tract dysfunction (LUTD). Thirty female C57Bl/6J mice in each group received an implanted bladder catheter with additional electrodes placed next to the EUS in the SCI group. UDI assessments were performed weekly for 7 weeks (pBOO group) or 8 weeks (SCI group), after which bladders were harvested for histological and transcriptome analysis. SCI mice developed detrusor sphincter dyssynergia (DSD) one week after injury with high-pressure oscillations and a significantly increased maximal bladder pressure P_max and were unable to void spontaneously during the whole observation period. They showed an increased bladder-to-bodyweight ratio, bladder fibrosis, and transcriptome changes indicative of extracellular matrix remodeling and alterations of neuronal signaling and muscle contraction. In contrast, pBOO led to a significantly increased P_max after one week, which normalized at later time points. Increased bladder-to-bodyweight ratio and pronounced gene expression changes involving immune and inflammatory pathways were observed 7 weeks after pBOO. Comparative transcriptome analysis of SCI and pBOO bladders revealed the activation of Wnt and TGF-beta signaling in both the neurogenic and obstructive LUTD and highlighted FGF2 as a major upregulated transcription factor during organ remodeling. We conclude that SCI-induced DSD in mice leads to profound changes in neuronal signaling and muscle contractility, leading to bladder fibrosis. In a similar time frame, significant bladder remodeling following pBOO allowed for functional compensation, preserving normal micturition parameters. Full article

Several new psychoactive substances (NPS) are responsible for intoxication involving the cardiovascular and respiratory systems. Among NPS, synthetic cannabinoids (SCs) provoked side effects in humans characterized by tachycardia, arrhythmias, hypertension, breathing difficulty, apnoea, myocardial infarction, and cardiac arrest. Therefore, the present study investigated the cardio-respiratory (MouseOx Plus; EMKA electrocardiogram (ECG) and plethysmography TUNNEL systems) and vascular (BP-2000 systems) effects induced by 1-naphthalenyl (1-pentyl-1H-indol-3-yl)-methanone (JWH-018; 0.3–3–6 mg/kg) and Δ⁹-tetrahydrocannabinol (Δ⁹-THC; 0.3–3–6 mg/kg), administered in awake CD-1 male mice. The results showed that higher doses of JWH-018 (3–6 mg/kg) induced deep and long-lasting bradycardia, alternated with bradyarrhythmia, spaced out by sudden episodes of tachyarrhythmias (6 mg/kg), and characterized by ECG electrical parameters changes, sustained bradypnea, and systolic and transient diastolic hypertension. Otherwise, Δ⁹-THC provoked delayed bradycardia (minor intensity tachyarrhythmias episodes) and bradypnea, also causing a transient and mild hypertensive effect at the tested dose range. These effects were prevented by both treatment with selective CB₁ (AM 251, 6 mg/kg) and CB₂ (AM 630, 6 mg/kg) receptor antagonists and with the mixture of the antagonists AM 251 and AM 630, even if in a different manner. Cardio-respiratory and vascular symptoms could be induced by peripheral and central CB₁ and CB₂ receptors stimulation, which could lead to both sympathetic and parasympathetic systems activation. These findings may represent a starting point for necessary future studies aimed at exploring the proper antidotal therapy to be used in SCs-intoxicated patient management. Full article

Tremor is the most common movement disorder. Several drugs reduce tremor severity, but no cures are available. Propranolol, a β-adrenergic receptor blocker, is the leading treatment for tremor. However, the in vivo circuit mechanisms by which propranolol decreases tremor remain unclear. Here, we test whether propranolol modulates activity in the cerebellum, a key node in the tremor network. We investigated the effects of propranolol in healthy control mice and Car8^wdl/wdl mice, which exhibit pathophysiological tremor and ataxia due to cerebellar dysfunction. Propranolol reduced physiological tremor in control mice and reduced pathophysiological tremor in Car8^wdl/wdl mice to control levels. Open field and footprinting assays showed that propranolol did not correct ataxia in Car8^wdl/wdl mice. In vivo recordings in awake mice revealed that propranolol modulates the spiking activity of control and Car8^wdl/wdl Purkinje cells. Recordings in cerebellar nuclei neurons, the targets of Purkinje cells, also revealed altered activity in propranolol-treated control and Car8^wdl/wdl mice. Next, we tested whether propranolol reduces tremor through β₁ and β₂ adrenergic receptors. Propranolol did not change tremor amplitude or cerebellar nuclei activity in β₁ and β₂ null mice or Car8^wdl/wdl mice lacking β₁ and β₂ receptor function. These data show that propranolol can modulate cerebellar circuit activity through β-adrenergic receptors and may contribute to tremor therapeutics. Full article

The analysis of blood samples from mice treated with the PDE4 inhibitor Roflumilast revealed an unexpected reduction in serum potassium levels, while sodium and chloride levels were unaffected. Treatment with several structurally distinct PAN-PDE4 inhibitors, including Roflumilast, Rolipram, RS25344, and YM976 dose-dependently reduced serum potassium levels, indicating the effect is a class-characteristic property. PDE4 inhibition also induces hypothermia and hypokinesia in mice. However, while general anesthesia abrogates these effects of PDE4 inhibitors, potassium levels decrease to similar extents in both awake as well as in fully anesthetized mice. This suggests that the hypokalemic effects of PDE4 inhibitors occur independently of hypothermia and hypokinesia. PDE4 inhibition reduces serum potassium within 15 min of treatment, consistent with a rapid transcellular shift of potassium. Catecholamines promote the uptake of potassium into the cell via increased cAMP signaling. PDE4 appears to modulate these adrenoceptor-mediated effects, as PDE4 inhibition has no additional effects on serum potassium in the presence of saturating doses of the β-adrenoceptor agonist Isoprenaline or the α₂-blocker Yohimbine, and is partially blocked by pre-treatment with the β-blocker Propranolol. Together, these data suggest that PDE4 inhibitors reduce serum potassium levels by modulating the adrenergic regulation of cellular potassium uptake. Full article