Muscarinic Receptors and Alzheimer's Disease: New Perspectives and Mechanisms.

Alzheimer's disease (AD) is one of the most prevalent neurodegenerative diseases on a global scale. Historically, this pathology has been linked to cholinergic transmission, and despite the scarcity of effective therapies, numerous alternative processes and targets have been proposed as potential avenues for comprehending this complex illness. Nevertheless, the fundamental pathophysiological mechanisms underpinning AD remain largely enigmatic, with a growing body of evidence advocating for the significance of muscarinic receptors in modulating the brain's capacity to adapt and generate new memories. This review summarizes the current state of the art in the field of muscarinic receptors' involvement in AD. A specific key factor was the relationship between comorbidity and the emergence of new mechanisms.

Falls in hospitalized older adults and the use of fall risk-increasing drugs and anticholinergic medications in Colombia: a case‒control study.

Introduction: In-hospital falls are multicausal in older hospitalized patients. Drugs with anticholinergic load and psychotropic effects can increase the risk of falling. Objective: This study aimed to determine the associations between fall risk-increasing drugs (FRIDs) and the anticholinergic risk score (ARS) with falls in hospitalized older hospitalized patients. Methods: This was a case‒control study of patients ≥65 years of age of either sex treated in four clinics in Colombia between 2018 and 2020. Each patient who suffered a fall during hospitalization was matched with four hospitalized patients who did not. Sociodemographic, clinical, and pharmacologic variables and the use of the ARS and FRIDs were evaluated. The risk associated with FRIDs was estimated using conditional logistic regression. Results: There were 250 patients and 1,000 controls (ratio of 1:4), with a mean age of 77.4 ± 7.4 years and a predominance of men (n = 800, 64.0%). The majority of falls occurred during hospitalization (n = 192 patients, 76.8%). Polypharmacy, calcium channel blockers, antiepileptics, antipsychotics, sodium-glucose cotransporter type 2 inhibitors, and nonsteroidal anti-inflammatory drugs were associated with falls during hospitalization. With an ARS score of 3, the probability of falling during the hospital stay increased (aOR: 2.34; 95% CI: 1.64-3.32). Conclusion: There is an association between suffering a fall and the use of drugs with anticholinergic load or FRIDs in hospitalized adults more than 65 years of age in Colombia.

In vivo and in silico elucidation of possible potential and mechanisms involved in the analgesic action of ethanolic extract of Lavandula Stoechas.

OBJECTIVES: Our research focused on plant's ethanolic extract Lavandula stoechas flower part to investigate the potential analgesic effects and possible pathways involvements. METHODS: Four experimental tests were performed on Swiss albino mice with five animals in each group at different doses (50, 100, and 200mg/kg); formalin test, tail-flick test, acetic acid-induced writhing, and hot-plate test. The opioidergic, noradrenergic, cholinergic, and K channel blockers in the analgesic actions were also carried out for the potential route involvement. KEY FINDING: The percentage inhibition for abdominal writhing's and formalin activity showed a dose-dependent manner for early and late phases reducing abdominal writhing's and time period of licking, respectively. Tail immersion and hot-plate test demonstrated a substantial and dose-dependent increase in the latency time and time period of paw liking and jumping response respectively. GC-MS showed the abundantly present compounds were octadecatrienoic acid (34.35%), n-hexadecanoic acid (12.98%). In silico analyses have revealed three compounds that had good interactions with 6y3c receptor proteins, demonstrating strong binding affinities and satisfying docking parameters. CONCLUSIONS: Overall, these studies showed that ethanolic extract of L. stoechas is an important medicinal plant, with both central and peripheral antinociceptive and analgesic activities supporting its traditional use for therapeutic purposes.

Effect of sex on chronic stress induced alterations in hindbrain catecholaminergic system and autonomic dysfunction resulting in gastrointestinal dysmotility.

It has been reported that the clinical symptoms of functional dyspepsia (FD) exacerbate upon stress while the gender-related factors have been incompletely understood. This study aims to investigate the role of sex in chronic heterotypic stress (CHS)-induced autonomic and gastric motor dysfunction. For CHS, the rats were exposed to the combination of different stressors for 7 consecutive days. Subsequently, electrocardiography was recorded in anesthetized rats to evaluate heart rate variability (HRV) for the determination of autonomic outflow and sympathovagal balance. Solid gastric emptying (GE) was measured in control and CHS-loaded male and female rats. The immunoreactivities of catecholaminergic cell marker tyrosine hydroxylase (TH), choline acetyltransferase (ChAT), corticotropin releasing factor (CRF), and estrogen receptor (ER-α/ß) were evaluated in medullary and pontine brainstem sections by immunohistochemistry. Compared with the controls, CHS significantly delayed GE in males but not in females. There was no significant sex-related difference in parasympathetic indicator HF under either control or CHS conditions. Sympathetic indicator LF was significantly higher in control females compared to the males. The higher sympathetic output in females was found to be attenuated upon CHS; in contrast, the elevated sympathetic output was detected in CHS-loaded males. No sex- or stress-related effect was observed on ChAT immunoreactivity in the dorsal motor nucleus of N.vagus (DMV). In males, greater number of TH-ir cells was observed in the caudal locus coeruleus (LC), while they were more densely detected in the rostral LC of females. Regardless of sex, CHS elevated immunoreactivity of TH throughout the LC. Under basal conditions, greater number of TH-ir cells was detected in the rostral ventrolateral medulla (RVLM) of females. In contrast, CHS remarkably increased the number of TH-ir cells in the RVLM of males which was found to be decreased in females. There was no sex-related alteration in TH immunoreactivity in the nucleus tractus solitarius (NTS) of control rats, while CHS affected both sexes in a similar manner. Compared with females, CRF immunoreactivity was prominently observed in control males, while both of which were stimulated by CHS. ER-α/ß was found to be co-expressed with TH in the NTS and LC which exhibit no alteration related to either sex or stress status. These results indicate a sexual dimorphism in the catecholaminergic and the CRF system in brainstem which might be involved in the CHS-induced autonomic and visceral dysfunction occurred in males.

Contribution of transient receptor potential vanilloid 1 (TRPV1) channel to cholinergic contraction of rat bladder smooth muscle.

Transient receptor potential vanilloid 1 (TRPV1) is a calcium-permeable ion channel that is gated by the pungent constituent of red chili pepper, capsaicin, and by related chemicals from the group of vanilloids, in addition to noxious heat. It is expressed mostly in sensory neurons to act as a detector of painful stimuli produced by pungent chemicals and high temperatures. Although TRPV1 is also found outside the sensory nervous system, its expression and function in the bladder detrusor smooth muscle (DSM) remain controversial. Here, by using Ca2+ imaging and patch clamp on isolated rat DSM cells, in addition to tensiometry on multicellular DSM strips, we show that TRPV1 is expressed functionally in only a fraction of DSM cells, in which it acts as an endoplasmic reticulum Ca2+-release channel responsible for the capsaicin-activated [Ca2+]i rise. Carbachol-stimulated contractions of multicellular DSM strips contain a TRPV1-dependent component, which is negligible in the circular DSM but reaches ≤50% in the longitudinal DSM. Activation of TRPV1 in rat DSM during muscarinic cholinergic stimulation is ensured by phospholipase A2-catalysed derivation of arachidonic acid and its conversion by lipoxygenases to eicosanoids, which act as endogenous TRPV1 agonists. Immunofluorescence detection of TRPV1 protein in bladder sections and isolated DSM cells confirmed both its preferential expression in the longitudinal DSM sublayer and its targeting to the endoplasmic reticulum. We conclude that TRPV1 is an essential contributor to the cholinergic contraction of bladder longitudinal DSM, which might be important for producing spatial and/or temporal anisotropy of bladder wall deformation in different regions during parasympathetic stimulation. KEY POINTS: The transient receptor potential vanilloid 1 (TRPV1) heat/capsaicin receptor/channel is localized in the endoplasmic reticulum membrane of detrusor smooth muscle (DSM) cells of the rat bladder, operating as a calcium-release channel. Isolated DSM cells are separated into two nearly equal groups, within which the cells either show or do not show TRPV1-dependent [Ca2+]i rise. Carbachol-stimulated, muscarinic ACh receptor-mediated contractions of multicellular DSM strips contain a TRPV1-dependent component. This component is negligible in the circular DSM but reaches ≤50% in longitudinal DSM. Activation of TRPV1 in rat DSM during cholinergic stimulation involves phospholipase A2-catalysed derivation of arachidonic acid and its conversion by lipoxygenases to eicosanoids, which act as endogenous TRPV1 agonists.

Chronic inflammation with microglia senescence at basal forebrain: impact on cholinergic deficit in Alzheimer's brain haemodynamics.

Cholinergic innervation in the brain is involved in modulating neurovascular function including cerebral blood flow haemodynamics in response to neuronal activity. Cholinergic deficit is associated with pathophysiology in Alzheimer's disease, albeit the aetiology remains to be clarified. In the current study, neocortex cerebral blood flow response to acetylcholine was evaluated by Laser-Doppler Flowmetry (LDF) in 3xTgAD Alzheimer's disease model) and wild-type mice of two age groups. The peak of cerebral blood flow to acetylcholine (i.v.) from baseline levels (% ΔrCBF) was higher in young 3xTgAD versus in wild-type mice (48.35; 95% CI:27.03-69.67 versus 22.70; CI:15.5-29.91, P < 0.05); this was reversed in old 3xTgAD mice (21.44; CI:2.52-40.35 versus 23.25; CI:23.25-39). Choline acetyltransferase protein was reduced in neocortex, while cerebrovascular reactivity to acetylcholine was preserved in young 3×TgAD mice. This suggests endogenous acetylcholine deficit and possible cholinergic denervation from selected cholinergic nuclei within the basal forebrain. The early deposition of tauopathy moieties (mutant hTau and pTau181) and its coincidence in cholinergic cell clusters (occasionaly), were observed at the basal forebrain of 3xTgAD mice including substantia innominate, nucleus Basalis of Meynert and nucleus of horizontal limb diagonal band of Broca. A prominent feature was microglia interacting tauopathy and demonstrated a variety of morphology changes particularly when located in proximity to tauopathy. The microglia ramified phenotype was reduced as evaluated by the ramification index and Fractal analysis. Increased microglia senescence, identified as SASP (senescence-associated secretory phenotype), was colocalization with p16Ink4ɑ, a marker of irreversible cell-cycle arrest in old 3xTgAD versus wild-type mice (P = 0.001). The p16Ink4ɑ was also observed in neuronal cells bearing tauopathy within the basal forebrain of 3xTgAD mice. TNF-ɑ, the pro-inflammatory cytokine elevated persistently in microglia (Pearson's correlation coefficient = 0.62) and the loss of cholinergic cells in vulnerable basal forebrain environment, was indicated by image analysis in 3xTgAD mice, which linked to the cholinergic deficits in neocortex rCBF haemodynamics. Our study revealed the early change of CBF haemodynamics to acetylcholine in 3xTgAD model. As a major effector of brain innate immune activation, microglia SASP with age-related disease progression is indicative of immune cell senescence, which contributes to chronic inflammation and cholinergic deficits at the basal forebrain. Targeting neuroinflammation and senescence may mitigate cholinergic pathophysiology in Alzheimer's disease.

Sepsis-induced inflammatory demyelination in medullary visceral zone and cholinergic anti-inflammatory pathway: Insights from a Rat's model study.

Background: Our previous studies have demonstrated that the activated Cholinergic Anti-inflammatory Pathway (CAP) effectively suppresses systemic inflammation and immunity in early sepsis. Some parameters of Heart Rate Variability (HRV) could be used to reflect the regulatory activity of CAP. However, in the early stages of severe sepsis of some patients, the inflammatory storm can still result in multiple organs dysfunction and even death, suggesting they lose CAP's modulation ability. Since CAP is part of the vagus nerve and is directly innervated by the Medullary Visceral Zone (MVZ), we can reasonably concluded that pathological changes induced by MVZ's neuroinflammation should be responsible for CAP's dysfunction in modulating systemic inflammation in early sepsis. Methods: We conducted two independent septic experiments, the sepsis model rats were prepared by cecum ligation and puncture (CLP) method. In the first experiment, A total of 64 adult male Sprague-Dawley rats were included. Under the condition of sepsis and CAP's pharmacological activation or blockade, we investigated the MVZ's pathological changes, the functional state of key neurons including catecholaminergic and cholinergic neurons, key genes' expression such as Oligodendrocyte Transcription Factor 2 (Olig-2) mRNA, glial fibrillary acidic protein (GFAP) mRNA, and matrix metalloprotein (MMP) -9 mRNA, and CAP's activities reflected by HRV. The second experiment involved in 56 rats, through central anti-inflammation by feeding with 10 mg/ml minocycline sucrose solution as the only water source, or right vagus transection excepting for central anti-inflammation as a mean of the CAP's functional cancel, we confirmed that the neuroinflammation in MVZ affected systemic inflammation through CAP in sepsis. Results: In the first experiment, cholinergic and catecholaminergic neurons showed significant apoptosis with reduced expressions of TH, but the expression of CHAT remained relatively unaffected in MVZ in sepsis. HRV parameters representing the tone of the vagus nerve, such as SDNN, RMSSD, HF, SD1, and SD2, did not show significant differences among the three Septic Groups, although they all decreased significantly compared to the Control Group. The expressions of GFAP mRNA and MMP-9 mRNA were up-regulated, while the expression of Olig-2 mRNA was down-regulated in the Septic Groups. Intervention of CAP had a significant effect on cholinergic and catecholaminergic neurons' apoptosis, as well as the expressions of TH/CHAT and these key genes, but had little effect on HRV in sepsis. In the second experiment, the levels of TNF-α, IL-6, in serum and MVZ were significantly increased in sepsis. Central anti-inflammatory treatment reversed these changes. However, right vagotomy abolished the central anti-inflammatory effect. Conclusions: Our study uncovered that MVZ's neuroinflammation may play a crucial role in the uncontrolled systemic inflammation through inflammatory demyelination in MVZ, which disrupts CAP's modulation on the systemic inflammation in early sepsis.

The role of the Ventral Nucleus of the Trapezoid Body in the auditory prepulse inhibition of the acoustic startle reflex.

Cholinergic signaling is essential to mediate the auditory prepulse inhibition (PPI), an operational measure of sensorimotor gating, that refers to the reduction of the acoustic startle reflex (ASR) when a low-intensity, non-startling acoustic stimulus (the prepulse) is presented just before the onset of the acoustic startle stimulus. The cochlear root neurons (CRNs) are the first cells of the ASR circuit to receive cholinergic inputs from non-olivocochlear neurons of the ventral nucleus of the trapezoid body (VNTB) and subsequently decrease their neuronal activity in response to auditory prepulses. Yet, the contribution of the VNTB-CRNs pathway to the mediation of PPI has not been fully elucidated. In this study, we used the immunotoxin anti-choline acetyltransferase (ChAT)-saporin as well as electrolytic lesions of the medial olivocochlear bundle to selectively eliminate cholinergic VNTB neurons, and then assessed the ASR and PPI paradigms. Retrograde track-tracing experiments were conducted to precisely determine the site of lesioning VNTB neurons projecting to the CRNs. Additionally, the effects of VNTB lesions and the integrity of the auditory pathway were evaluated via auditory brain responses tests, ChAT- and FOS-immunohistochemistry. Consequently, we established three experimental groups: 1) intact control rats (non-lesioned), 2) rats with bilateral lesions of the olivocochlear bundle (OCB-lesioned), and 3) rats with bilateral immunolesions affecting both the olivocochlear bundle and the VNTB (OCB/VNTB-lesioned). All experimental groups underwent ASR and PPI tests at several interstimulus intervals before the lesion and 7, 14, and 21 days after it. Our results show that the ASR amplitude remained unaffected both before and after the lesion across all experimental groups, suggesting that the VNTB does not contribute to the ASR. The%PPI increased across the time points of evaluation in the control and OCB-lesioned groups but not in the OCB/VNTB-lesioned group. At the ISI of 50 ms, the OCB-lesioned group exhibited a significant increase in%PPI (p < 0.01), which did not occur in the OCB/VNTB-lesioned group. Therefore, the ablation of cholinergic non-olivocochlear neurons in the OCB/VNTB-lesioned group suggests that these neurons contribute to the mediation of auditory PPI at the 50 ms ISI through their cholinergic projections to CRNs. Our study strongly reinforces the notion that auditory PPI encompasses a complex mechanism of top-down cholinergic modulation, effectively attenuating the ASR across different interstimulus intervals within multiple pathways.

Cholinergic nucleus degeneration and its association with gait impairment in Parkinson's disease.

BACKGROUND: The contribution of cholinergic degeneration to gait disturbance in Parkinson's disease (PD) is increasingly recognized, yet its relationship with dopaminergic-resistant gait parameters has been poorly investigated. We investigated the association between comprehensive gait parameters and cholinergic nucleus degeneration in PD. METHODS: This cross-sectional study enrolled 84 PD patients and 69 controls. All subjects underwent brain structural magnetic resonance imaging to assess the gray matter density (GMD) and volume (GMV) of the cholinergic nuclei (Ch123/Ch4). Gait parameters under single-task (ST) and dual-task (DT) walking tests were acquired using sensor wearables in PD group. We compared cholinergic nucleus morphology and gait performance between groups and examined their association. RESULTS: PD patients exhibited significantly decreased GMD and GMV of the left Ch4 compared to controls after reaching HY stage > 2. Significant correlations were observed between multiple gait parameters and bilateral Ch123/Ch4. After multiple testing correction, the Ch123/Ch4 degeneration was significantly associated with shorter stride length, lower gait velocity, longer stance phase, smaller ankle toe-off and heel-strike angles under both ST and DT condition. For PD patients with HY stage 1-2, there were no significant degeneration of Ch123/4, and only right side Ch123/Ch4 were corrected with the gait parameters. However, as the disease progressed to HY stage > 2, bilateral Ch123/Ch4 nuclei showed correlations with gait performance, with more extensive significant correlations were observed in the right side. CONCLUSIONS: Our study demonstrated the progressive association between cholinergic nuclei degeneration and gait impairment across different stages of PD, and highlighting the potential lateralization of the cholinergic nuclei's impact on gait impairment. These findings offer insights for the design and implementation of future clinical trials investigating cholinergic treatments as a promising approach to address gait impairments in PD.

Increased spontaneous activity and progressive suppression of adult neurogenesis in the hippocampus of rat offspring after maternal exposure to imidacloprid.

Imidacloprid (IMI) is a widely used neonicotinoid insecticide that poses risks for developmental neurotoxicity in mammals. The present study investigated the effects of maternal exposure to IMI on behaviors and adult neurogenesis in the hippocampal dentate gyrus (DG) of rat offspring. Dams were exposed to IMI via diet (83, 250, or 750 ppm in diet) from gestational day 6 until day 21 post-delivery on weaning, and offspring were maintained until adulthood on postnatal day 77. In the neurogenic niche, 750-ppm IMI decreased numbers of late-stage neural progenitor cells (NPCs) and post-mitotic immature granule cells by suppressing NPC proliferation and ERK1/2-FOS-mediated synaptic plasticity of granule cells on weaning. Suppressed reelin signaling might be responsible for the observed reductions of neurogenesis and synaptic plasticity. In adulthood, IMI at ≥ 250 ppm decreased neural stem cells by suppressing their proliferation and increasing apoptosis, and mature granule cells were reduced due to suppressed NPC differentiation. Behavioral tests revealed increased spontaneous activity in adulthood at 750 ppm. IMI decreased hippocampal acetylcholinesterase activity and Chrnb2 transcript levels in the DG on weaning and in adulthood. IMI increased numbers of astrocytes and M1-type microglia in the DG hilus, and upregulated neuroinflammation and oxidative stress-related genes on weaning. In adulthood, IMI increased malondialdehyde level and number of M1-type microglia, and downregulated neuroinflammation and oxidative stress-related genes. These results suggest that IMI persistently affected cholinergic signaling, induced neuroinflammation and oxidative stress during exposure, and increased sensitivity to oxidative stress after exposure in the hippocampus, causing hyperactivity and progressive suppression of neurogenesis in adulthood. The no-observed-adverse-effect level of IMI for offspring behaviors and hippocampal neurogenesis was determined to be 83 ppm (5.5-14.1 mg/kg body weight/day).

Effects of hot bath therapy on cholinergic urticaria with hypohidrosis or anhidrosis.

Cholinergic urticaria with hypohidrosis or anhidrosis (CUHA) can impair quality of life due to itching, tingling, and reduced sweating. Current treatment options for CUHA include antihistamines, pulsed steroids, and sweat-promoting therapies such as exercise or hot baths. However, the efficacy of these therapies, particularly hot bath therapy, has yet to be established. We evaluated the efficacy of hot bath therapy in patients with CUHA. We enrolled eight patients who underwent hot bath therapy between January 2010 and August 2022. Patients had a half-body bath in a bathtub filled with hot water (40-43°C) for 30-60 minutes daily for 3-7 days. After treatment, pain improved in three (42.9%) patients, urticaria improved in four (50%) patients, and anhidrosis improved in five (62.5%) patients without any severe adverse events. Because hot bath therapy is easily performed, it should be considered a treatment option for patients with CUHA.

New insights into chronic inducible urticaria.

PURPOSE OF REVIEW: Chronic inducible urticaria (CIndU) is a group of long-persisting and challenging to manage diseases, characterized by recurrent wheals and angioedema induced by definite triggers. In this review, we address recent findings on CIndU pathogenesis, diagnosis as well as its treatment, and we discuss novel potential targets that may lead to the development of more effective therapies for CIndU patients. RECENT ADVANCES: Meaningful advances in the understanding of its pathogenesis have been reported in the last decades. Novel CIndU-specific patient-reported outcome measures enable a closer and better evaluation of patients. CIndU is a hard-to-treat disease that highly impairs quality of life (QoL) of affected patients. Provocation tests allow to diagnose CIndU subtypes. The only licensed and recommended treatment for CIndU are second generation non-sedating H1-antihistamines, which lack efficacy in many cases. Omalizumab off-label use has been assessed in all types of CIndU with overall good outcomes. Promising emerging therapies currently assessed in chronic spontaneous urticaria are paving the path for novel treatments for CIndU.

[Immunological mechanism of Ermiao Powder improving inflammation in rats with collagen-induced arthritis through α7nAChR-JAK2/STAT3 pathway].

This study investigated the immunological mechanisms of Ermiao powder in the treatment of rheumatoid arthritis rats through the alpha 7 nicotinic acetylcholine receptor(α7nAChR)-Janus kinases 2(JAK2)/signal transducer and activator of transcription 3(STAT3) signaling pathway. A total of 56 female Wistar rats were randomly divided into the normal group(HG, n=8), collagen-induced arthritis(CIA) model group(CM, n=8), vagotomy group(VA, n=8), sham group(SH, n=8), Ermiao Powder treatment model group(EM, n=8), Ermiao Powder treatment for vagotomy group(EV, n=8) and Ermiao Powder treatment for sham group(ES, n=8). Following the establishment of CIA models in all groups except the HG group, the rats underwent unilateral vagotomy and sham operation(only the vagus nerve was separated). Drug treatment was started 7 days after surgery and continued for 35 days. The body weight and joints of rats were recorded, the pathological changes of the spleen of rats were observed, the contents of interleukin-6(IL-6), interleukin-1ß(IL-1ß) and tumor necrosis factor-α(TNF-α) in serum were detected by enzyme-linked immunosorbent assay(ELISA), and the mRNA and protein expression of α7nAChR-JAK2/STAT3 pathway core genes in spleen were detected by qRT-PCR, Western blot and immunohistochemistry. RESULTS:: showed that CM group(compared with HG group) and VA group(compared with CM group and SH group) had significantly decreased body weight(P<0.05, P<0.01), increased arthritis score(P<0.05, P<0.01), swollen ankle joints with deformity, and increased and enlarged lymph nodes in the spleen. There were also notable increases in the serum levels of IL-6, IL-1ß and TNF-α(P<0.05, P<0.01), and in the mRNA expressions of JAK2 and STAT3 in the spleen(P<0.05, P<0.01). The protein levels of phosphorylated JAK2(p-JAK2)/JAK2 and phospho-STAT3(p-STAT3)/STAT3 were significantly increased(P<0.05, P<0.01), and the number of JAK2, p-JAK2, STAT3 and p-STAT3 cells increased(P<0.05, P<0.01). EM group(compared with CM group) and ES group(compared with SH group) exhibited significantly increased body weight(P<0.01), decreased arthritis scores(P<0.05, P<0.01), reduced swelling of ankle joint, and decreased number and volume of lymph nodes in the spleen. Furthermore, serum levels of IL-6, IL-1ß, and TNF-α decreased(P<0.05, P<0.01), the mRNA expression of JAK2 and STAT3 in spleen decreased(P<0.05, P<0.01), the protein levels of p-JAK2/JAK2 and p-STAT3/STAT3 decreased(P<0.05, P<0.01), and the number of JAK2, p-JAK2, STAT3 and p-STAT3 cells decreased(P<0.05, P<0.01), whereas the mRNA and protein expressions of α7nAChR were significantly increased(P<0.01). Compared with the VA group, there was no significant differences in weight gain and arthritis scores in the EV group. The number of lymph nodes in the spleen was not significantly reduced and the volume was still large, suggesting the inflammation was not significantly improved. The serum levels of IL-6, IL-1ß and TNF-α were not significantly different, and there were no significant differences in α7nAChR, JAK2, and STAT3 mRNA expression in the spleen. The protein expression levels of p-JAK2/JAK2 and α7nAChR in spleen were lower(P<0.05, P<0.01), while p-STAT3/STAT3 protein expression was not significantly different. Besides, the two groups had no significant difference in the number of JAK2, p-JAK2, STAT3, and p-STAT3 cells. The results suggested that unilateral vagotomy promoted the increase of phosphorylated JAK2 and STAT3 expressions and exacerbated inflammation. In contrast, Ermiao Powder alleviated the inflammation in rheumatoid arthritis rats by activating the α7nAChR-mediated JAK2/STAT3 pathway through the vagus nerve, suggesting that the α7nAchR-JAK2/STAT3 pathway may be a potential target for the treatment of rheumatoid arthritis.

Vitamin D attenuates monosodium glutamate-induced behavioural anomalies, metabolic dysregulation, cholinergic impairment, oxidative stress, and astrogliosis in rats.

BACKGROUND: Monosodium glutamate (MSG) is a commonly used flavor enhancer that has raised concerns due to its potential adverse effects on various organs. This study explored the neuroprotective potential of Vitamin D, a beneficial micronutrient, in mitigating MSG-induced neurotoxicity. MATERIALS AND METHODS: Adult male Wistar rats were categorized into five groups: control (2 ml/kg PBS orally for 30 days), MSG (40 mg/kg orally for 30 days), VIT-D (oral cholecalciferol; 500 IU/kg for 30 days), MSG+VIT-D (MSG for 30 days followed by VIT-D for another 30 days), and VIT-D/MSG (concurrent VIT-D and MSG for 30 days). The rats underwent neurobehavioral, histochemical, and biochemical analyses following the treatments. RESULTS: MSG treatment caused a decline in both long and short-term memory, along with reduced exploratory and anxiogenic behavior, mitigated by vitamin D treatment. MSG exposure also induced impaired behavior, dyslipidemia, oxidative stress, lipid peroxidation, altered cholinergic transmission, and increased chromatolysis and neuroinflammation in the frontal cortex, hippocampus, and cerebellum. CONCLUSIONS: VIT-D demonstrated a mitigating effect on MSG-induced adverse outcomes, highlighting its potential to attenuate neurodegenerative cascades. This investigation contributes to understanding MSG-associated neurotoxicity and suggests vitamin D as a valuable and potential intervention for neuroprotection.

Sex differences in the distribution and density of regulatory interneurons in the striatum.

Introduction: Dysfunction of the cortico-basal circuitry - including its primary input nucleus, the striatum - contributes to neuropsychiatric disorders, such as autism and Tourette Syndrome (TS). These conditions show marked sex differences, occurring more often in males than in females. Regulatory interneurons, such as cholinergic interneurons (CINs) and parvalbumin-expressing GABAergic fast spiking interneurons (FSIs), are implicated in human neuropsychiatric disorders such as TS, and ablation of these interneurons produces relevant behavioral pathology in male mice, but not in females. Here we investigate sex differences in the density and distribution of striatal interneurons. Methods: We use stereological quantification of CINs, FSIs, and somatostatin-expressing (SOM) GABAergic interneurons in the dorsal striatum (caudate-putamen) and the ventral striatum (nucleus accumbens) in male and female mice. Results: Males have a higher density of CINs than females, especially in the dorsal striatum; females have equal distribution between dorsal and ventral striatum. FSIs showed similar distributions, with a greater dorsal-ventral density gradient in males than in females. SOM interneurons were denser in the ventral than in the dorsal striatum, with no sex differences. Discussion: These sex differences in the density and distribution of FSIs and CINs may contribute to sex differences in basal ganglia function, particularly in the context of psychopathology.

Specific Mode Electroacupuncture Stimulation Mediates the Delivery of NGF Across the Hippocampus Blood-Brain Barrier Through p65-VEGFA-TJs to Improve the Cognitive Function of MCAO/R Convalescent Rats.

Cognitive impairment frequently presents as a prevalent consequence following stroke, imposing significant burdens on patients, families, and society. The objective of this study was to assess the effectiveness and underlying mechanism of nerve growth factor (NGF) in treating post-stroke cognitive dysfunction in rats with cerebral ischemia-reperfusion injury (MCAO/R) through delivery into the brain using specific mode electroacupuncture stimulation (SMES). From the 28th day after modeling, the rats were treated with NGF mediated by SMES, and the cognitive function of the rats was observed after treatment. Learning and memory ability were evaluated using behavioral tests. The impact of SMES on blood-brain barrier (BBB) permeability, the underlying mechanism of cognitive enhancement in rats with MCAO/R, including transmission electron microscopy, enzyme-linked immunosorbent assay, immunohistochemistry, immunofluorescence, and TUNEL staining. We reported that SMES demonstrates a safe and efficient ability to open the BBB during the cerebral ischemia repair phase, facilitating the delivery of NGF to the brain by the p65-VEGFA-TJs pathway.

Differentiation of SH-SY5Y neuroblastoma cells using retinoic acid and BDNF: a model for neuronal and synaptic differentiation in neurodegeneration.

There has been much interest in the use of cell culture models of neurones, to avoid the animal welfare and cost issues of using primary and human-induced pluripotent stem cell (hiPSC)-derived neurones respectively. The human neuroblastoma cell line, SH-SY5Y, is extensively used in laboratories as they can be readily expanded, are of low cost and can be differentiated into neuronal-like cells. However, much debate remains as to their phenotype once differentiated, and their ability to recapitulate the physiology of bona fide neurones. Here, we characterise a differentiation protocol using retinoic acid and BDNF, which results in extensive neurite outgrowth/branching within 10 days, and expression of key neuronal and synaptic markers. We propose that these differentiated SH-SY5Y cells may be a useful substitute for primary or hiPSC-derived neurones for cell biology studies, in order to reduce costs and animal usage. We further propose that this characterised differentiation timecourse could be used as an in vitro model for neuronal differentiation, for proof-of principle studies on neurogenesis, e.g. relating to neurodegenerative diseases. Finally, we demonstrate profound changes in Tau phosphorylation during differentiation of these cells, suggesting that they should not be used for neurodegeneration studies in their undifferentiated state.

Cilostazol Ameliorates Motor Dysfunction and Schwann Cell Impairment in Streptozotocin-Induced Diabetic Rats.

This study investigated the effects of cilostazol on motor dysfunction, spinal motor neuron abnormalities, and schwannopathy in rats with diabetes. Diabetes mellitus (DM) was induced in rats via femoral intravenous streptozotocin (STZ) injection (60 mg/kg). After successful DM induction, cilostazol was administered on day 15 via oral gavage (100 mg/kg/day) for 6 weeks until sacrifice. Behavioral assays, including motor function, were performed weekly. The sciatic nerve, L5 spinal cord, and spinal ventral root were collected to evaluate the expression of the glial fibrillary acidic protein (GFAP), myelin protein zero (P0), and choline acetyltransferase (ChAT) by immunofluorescence and Western blotting. DM rats displayed decreased running speeds, running distances, and toe spread but increased foot pressure. In addition, loss of non-myelinating Schwann cells and myelin sheaths was observed in the sciatic nerve and L5 spinal ventral root. Reduced numbers of motor neurons were also found in the L5 spinal ventral horn. Cilostazol administration significantly potentiated running speed and distance; increased hind paw toe spread; and decreased foot pressure. In the sciatic nerve and L5 spinal ventral root, cilostazol treatment significantly improved non-myelinated Schwann cells and increased myelin mass. ChAT expression in motor neurons in the spinal ventral horn was improved, but not significantly. Cilostazol administration may protect sensorimotor function in diabetic rats.

Functionally linked amygdala and prefrontal cortical regions are innervated by both single and double projecting cholinergic neurons.

Cholinergic cells have been proposed to innervate simultaneously those cortical areas that are mutually interconnected with each other. To test this hypothesis, we investigated the cholinergic innervation of functionally linked amygdala and prefrontal cortical regions. First, using tracing experiments, we determined that cholinergic cells located in distinct basal forebrain (BF) areas projected to the different nuclei of the basolateral amygdala (BLA). Specifically, cholinergic cells in the ventral pallidum/substantia innominata (VP/SI) innervated the basal nucleus (BA), while the horizontal limb of the diagonal band of Broca (HDB) projected to its basomedial nucleus (BMA). In addition, cholinergic neurons in these two BF areas gave rise to overlapping innervation in the medial prefrontal cortex (mPFC), yet their axons segregated in the dorsal and ventral regions of the PFC. Using retrograde-anterograde viral tracing, we demonstrated that a portion of mPFC-projecting cholinergic neurons also innervated the BLA, especially the BA. By injecting retrograde tracers into the mPFC and BA, we found that 28% of retrogradely labeled cholinergic cells were double labeled, which typically located in the VP/SI. In addition, we found that vesicular glutamate transporter type 3 (VGLUT3)-expressing neurons within the VP/SI were also cholinergic and projected to the mPFC and BA, implicating that a part of the cholinergic afferents may release glutamate. In contrast, we uncovered that GABA is unlikely to be a co-transmitter molecule in HDB and VP/SI cholinergic neurons in adult mice. The dual innervation strategy, i.e., the existence of cholinergic cell populations with single as well as simultaneous projections to the BLA and mPFC, provides the possibility for both synchronous and independent control of the operation in these cortical areas, a structural arrangement that may maximize computational support for functionally linked regions. The presence of VGLUT3 in a portion of cholinergic afferents suggests more complex functional effects of cholinergic system in cortical structures.

Neuroinflammation in the medullary visceral zone exert a powerful impaction on the systemic inflammation in sepsis through cholinergic anti-inflammatory pathway.

To investigate whether sepsis-induced neuroinflammation of medulla visceral zone (MVZ) predominates the systemic inflammation through cholinergic anti-inflammatory pathway (CAP), and to explore the effect of central anti-inflammation on systemic inflammation. 112 adult Sprague-Dawley male rats were randomly divided into sepsis experimental group (n = 56) and neuroinflammation experimental group (n = 56). The two experimental groups were individually randomly divided into control group (n = 8), model group (n = 16), central anti-inflammatory group (n = 16) and vagus transection group (n = 16). Rats in two control groups were administered with saline at the dose of 6 mL/kg intraperitoneally or with 25 µL artificial cerebrospinal fluid injected into forth ventricle once a day for 3 days. Rats in two model groups were administered with Lipopolysaccharide (LPS) at the dose of 6 mg/kg intraperitoneally or with 25 µg/25 µL LPS injected into forth ventricle once a day for 3 days. Rats in two central anti-inflammatory groups were fed with 10 mg/mL minocycline sucrose solution as the only water source for 4 days prior to be treated as the model groups of their own, and feeding style was continued until the end of the experiment. Rats in the two vagus transection groups were undergone right vagotomy and 7 days of adaptive feeding prior to be treated as the same as those in the central anti-inflammatory group of their own. The Murine Sepsis Score (MSS), mortality rate and heat rate variability (HRV) were recorded during the last 3 days of intervention. Then the rats were sacrificed and blood samples were collected for ELISA analysis to detect the serum level of inflammatory cytokines such as TNF-α, IL-6, and IL-10. The expression of TNF-α and IL-6 in medulla oblongata were analyzed by Western blot. The correlation and regression analysis among the expression levels of cytokines in medulla oblongata, HRV indexes and serum inflammatory cytokines were performed. The mortality rate and MSS of the sepsis model group and the MVZ's neuroinflammation model group were significantly higher than those of their own control group, and the central anti-inflammation reduced the mortality rate and MSS scores of the two model groups, while the right vagotomy abolished the effect of central anti-inflammatory. In the sepsis model group and the MVZ's neuroinflammation model group, the levels of TNF-α, IL-6, and other cytokines in serum and MVZ were significantly increased, and HRV indexes (SDNN, RMSSD, LF, HF, LF/HF) were significantly decreased (P = 0.000). Central anti-inflammatory treatment reversed the above changes. However, right vagotomy abolished the central anti-inflammatory effect. Correlation and regression analysis showed that there was a significant linear correlation among the expression of inflammatory factors in MVZ, the indexes of HRV and the levels of serum cytokines. Our study shows that sepsis-induced MVZ's neuroinflammation exert a powerful influence on the systemic inflammation through CAP in sepsis. Central anti-inflammation effectively improves systemic inflammation through inhibiting MVZ's neuroinflammation in sepsis. The time domain and frequency domain indexes of HRV can reflect the regulatory effect of CAP and the degree of inflammation of MVZ, which may be potentially used to monitor the condition and treatment effectiveness of sepsis patients.