Endogenous α7 nAChR Agonist SLURP1 Facilitates Escherichia coli K1 Crossing the Blood-Brain Barrier.

Alpha 7 nicotinic acetylcholine receptor (α7 nAChR) is critical for the pathogenesis of Escherichia coli (E. coli) K1 meningitis, a severe central nervous system infection of the neonates. However, little is known about how E. coli K1 manipulates α7 nAChR signaling. Here, through employing immortalized cell lines, animal models, and human transcriptional analysis, we showed that E. coli K1 infection triggers releasing of secreted Ly6/Plaur domain containing 1 (SLURP1), an endogenous α7 nAChR ligand. Exogenous supplement of SLURP1, combined with SLURP1 knockdown or overexpression cell lines, showed that SLURP1 is required for E. coli K1 invasion and neutrophils migrating across the blood-brain barrier (BBB). Furthermore, we found that SLURP1 is required for E. coli K1-induced α7 nAChR activation. Finally, the promoting effects of SLURP1 on the pathogenesis of E. coli K1 meningitis was significantly abolished in the α7 nAChR knockout mice. These results reveal that E. coli K1 exploits SLURP1 to activate α7 nAChR and facilitate its pathogenesis, and blocking SLURP1-α7 nAChR interaction might represent a novel therapeutic strategy for E. coli K1 meningitis.

Aß1-16 controls synaptic vesicle pools at excitatory synapses via cholinergic modulation of synapsin phosphorylation.

Amyloid beta (Aß) is linked to the pathology of Alzheimer's disease (AD). At physiological concentrations, Aß was proposed to enhance neuroplasticity and memory formation by increasing the neurotransmitter release from presynapse. However, the exact mechanisms underlying this presynaptic effect as well as specific contribution of endogenously occurring Aß isoforms remain unclear. Here, we demonstrate that Aß1-42 and Aß1-16, but not Aß17-42, increased size of the recycling pool of synaptic vesicles (SV). This presynaptic effect was driven by enhancement of endogenous cholinergic signalling via α7 nicotinic acetylcholine receptors, which led to activation of calcineurin, dephosphorylation of synapsin 1 and consequently resulted in reorganization of functional pools of SV increasing their availability for sustained neurotransmission. Our results identify synapsin 1 as a molecular target of Aß and reveal an effect of physiological concentrations of Aß on cholinergic modulation of glutamatergic neurotransmission. These findings provide new mechanistic insights in cholinergic dysfunction observed in AD.

Mesenchymal Stem Cells or Interleukin-6 Improve Episodic Memory of Mice Lacking α7 Nicotinic Acetylcholine Receptors.

Nicotinic acetylcholine receptors of α7 subtype (α7 nAChRs) are involved in regulating cognition, inflammation and cell survival. Neuroinflammation is accompanied by the decrease of α7 nAChRs in the brain and impairment of memory. We show here that α7-/- mice possess pro-inflammatory phenotype and demonstrate worse episodic memory compared to wild-type mice. Previously we reported that mesenchymal stem cells (MSCs) restored episodic memory of lipopolysaccharide-treated wild-type mice. The aim of this study was to examine if MSCs or their soluble factors improve memory of α7-/- mice. The α7-specific signal (ELISA) and α7+ cells (IHC) were found in the brain of α7-/- mice on days 7 and 14 after intravenous injection of α7+ MSCs from either human umbilical cord (hMSCs) or mouse placenta (mMSCs). The intravenously injected MSCs or intraperitoneally injected hMSCs-conditioned medium transiently improved episodic memory of α7-/- mice and decreased cytochrome c release from their brain mitochondria under the effect of Ca2+. Either MSCs or conditioned medium stimulated an IL-6 increase in the brain, which coincided with the improvement of episodic memory. Injections of recombinant IL-6 also improved episodic memory of α7-/- mice accompanied by the up-regulation of α3, α4, ß2 and ß4 nAChR subunits in the brain. It is concluded that MSCs, injected intravenously, penetrate the brain of α7-/- mice and persist there for at least 2 weeks. They improve episodic memory of mice and make their mitochondria more resistant to apoptogenic influence. One of the soluble factors responsible for the memory improvement is IL-6.

Nicotinic acetylcholine receptor subunit α7-knockout mice exhibit degraded auditory temporal processing.

The CHRNA7 gene that encodes the α7-subunit of the nicotinic acetylcholine receptor (α7-nAChR) has been associated with some autism spectrum disorders and other neurodevelopmental conditions characterized, in part, by auditory and language impairment. These conditions may include auditory processing disorders that represent impaired timing of neural activity, often accompanied by problems understanding speech. Here, we measure timing properties of sound-evoked activity via the auditory brainstem response (ABR) of α7-nAChR knockout mice of both sexes and wild-type colony controls. We find a significant timing delay in evoked ABR signals that represents midbrain activity in knockouts. We also examine spike-timing properties of neurons in the inferior colliculus, a midbrain nucleus that exhibits high levels of α7-nAChR during development. We find delays of evoked responses along with degraded spiking precision in knockout animals. We find similar timing deficits in responses of neurons in the superior paraolivary nucleus and ventral nucleus of the lateral lemniscus, which are brainstem nuclei thought to shape temporal precision in the midbrain. In addition, we find that other measures of temporal acuity including forward masking and gap detection are impaired for knockout animals. We conclude that altered temporal processing at the level of the brainstem in α7-nAChR-deficient mice may contribute to degraded spike timing in the midbrain, which may underlie the observed timing delay in the ABR signals. Our findings are consistent with a role for the α7-nAChR in types of neurodevelopmental and auditory processing disorders and we identify potential neural targets for intervention.NEW & NOTEWORTHY Disrupted signaling via the α7-nicotinic acetylcholine receptor (α7-nAChR) is associated with neurodevelopmental disorders that include impaired auditory processing. The underlying causes of dysfunction are not known but a common feature is abnormal timing of neural activity. We examined temporal processing of α7-nAChR knockout mice and wild-type controls. We found degraded spike timing of neurons in knockout animals, which manifests at the level of the auditory brainstem and midbrain.

α7nAChR Deletion Aggravates Myocardial Infarction and Enhances Systemic Inflammatory Reaction via mTOR-Signaling-Related Autophagy.

Alpha7 nicotinic acetylcholine receptor (α7nAChR) has been previously reported to play an alleviative role in myocardial infarction (MI). In this study, we investigated its specific mechanism. α7nAChR-/- mice and its control (α7nAChR+/+) were used for the study of α7nAChR. Left anterior descending coronary artery occlusion was conducted for the creation of mice MI model and lipopolysaccharide (LPS) was used as inflammatory stressor in murine peritoneal macrophages. Triphenyltetrazolium chloride (TTC) staining and echocardiography was used for the detection of infarct size and cardiac function, respectively. Western blot was conducted for the testing of autophagy-related proteins and enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (RT-PCR) was used for the testing of proinflammatory cytokines. Rapamycin was used for the induction of autophagy through inhibiting mammalian target of rapamycin (mTOR)-related signaling. We found that knocking out α7nAChR enhanced the cardiac infarct size and damaged cardiac function in MI. α7nAChR deficiency increased the levels of several proinflammatory cytokines in serum and spleen from MI mice as well as murine macrophages under inflammatory stress. α7nAChR deletion decreased the level of autophagy in spleen from MI mice and macrophages under inflammatory stress. Rapamycin alleviated the cardiac function and systemic inflammatory reaction in MI mice as well as inflammatory reaction in macrophages under inflammatory stress, which was attenuated by knocking out α7nAChR. Our current study investigated the mechanism of α7nAChR-mediated cardio-protective and anti-inflammatory effect related to mTOR-related autophagy, which might provide a novel insight in the treatment of MI.

Proteomic Investigation of Murine Neuronal α7-Nicotinic Acetylcholine Receptor Interacting Proteins.

The α7-nicotinic acetylcholine receptor (α7-nAChR) is a ligand-gated ion channel that is expressed widely in vertebrates and is the principal high-affinity α-bungarotoxin (α-bgtx) binding protein in the mammalian CNS. α7-nAChRs associate with proteins that can modulate its properties. The α7-nAChR interactome is the summation of proteins interacting or associating with α7-nAChRs in a protein complex. To identify an α7-nAChR interactome in neural tissue, we isolated α-bgtx-affinity protein complexes from wild-type and α7-nAChR knockout (α7 KO) mouse whole brain tissue homogenates using α-bgtx-affinity beads. Affinity precipitated proteins were trypsinized and analyzed with an Orbitrap Fusion mass spectrometer. Proteins isolated with the α7-nAChR specific ligand, α-bgtx, were determined to be α7-nAChR associated proteins. The α7-nAChR subunit and 120 additional proteins were identified. Additionally, 369 proteins were identified as binding to α-bgtx in the absence of α7-nAChR expression, thereby identifying nonspecific proteins for α7-nAChR investigations using α-bgtx enrichment. These results expand on our previous investigations of α7-nAChR interacting proteins using α-bgtx-affinity bead isolation by controlling for differences between α7-nAChR and α-bgtx-specific proteins, developing an improved protein isolation methodology, and incorporating the latest technology in mass spectrometry. The α7-nAChR interactome identified in this study includes proteins associated with the expression, localization, function, or modulation of α7-nAChRs, and it provides a foundation for future studies to elucidate how these interactions contribute to human disease.

Effects of neuromuscular presynaptic muscarinic M1 receptor blockade on rocuronium-induced neuromuscular blockade in immobilized tibialis anterior muscles.

This in vivo study tested the hypothesis that the modulation of acetylcholine (ACh) release by the M1 muscarinic receptor (mAChR) in the neuromuscular junction of disused muscles may affect the tensions of the muscles during the neuromuscular monitoring of a rocuronium-induced neuromuscular block and compared the results with those obtained from normal muscles. A total of 20 C57BL/6 (wild-type) and 10 α7 knock out (α7KO) mice were used in this experiment. As a pre-experimental procedure, knee and ankle joints of right hind limbs were fixed by needle pinning at the 90° flexed position. After 2 weeks, the main experiment was performed. Both tendons of the tibialis anterior (TA) muscles were obtained, and the muscle tensions were recorded while the dose-responses of rocuronium were measured three times in the same mouse by the serial administration of pirenzepine (0, 0.001 and 0.01 µg/g). Weight losses were observed after 2 weeks of immobilization in both groups, and a decrease in the mass of TA muscles at the immobilized side was observed compared to those of the contralateral nonimmobilized side. Tension depression of the TA muscles at immobilized side of the α7KO group was faster than those of the wild-type group, but these differences decreased after the administration of pirenzepine. The tension depressions were similar regardless of the pirenzepine doses at the same side in the group. Tension depression may become more rapid in the α7 AChR-expressed disused muscles by the decreased release of ACh release upon neuronal firing by the blockade of facilitatory M1 mAChR.

Deficiency of α7 nicotinic acetylcholine receptor attenuates bleomycin-induced lung fibrosis in mice.

α7 nicotinic acetylcholine receptor (α7 nAChR, coded by Chrna7) is indispensible in dampening proinflammatory responses. However, whether α7 nAChR would play a role in regulating bleomycin (BLM)-induced lung fibrosis is less investigated. Here, we intratracheally challenged wildtype and Chrna7-/- mice with BLM to elicit lung fibrosis. Taken advantage of this model, we measured body weight loss, lung fibrogenic genes (Acta2, Col1a1, Fsp1, and Fstl1), histology, Masson's trichrome staining, hydroxyproline levels, and expression of α-SMA at protein levels in the BLM-challenged lung for evaluating severity of lung fibrosis. We also pretreated human fibroblasts (MRC5 cell line) and isolated mouse lung fibroblasts with GTS-21 (an α7 nAChR agonist) to study its effects on TGF-ß-stimulated profibrotic profiles. We found that lung Chrna7 expression and CD4+CHAT+ (Choline acetyltransferase, an enzyme for local acetylcholine synthesis) cells were 12-fold and 4.5-fold respectively elevated in the early stage of lung fibrosis. Deletion of Chrna7 prevented body weight loss and reduced lung fibrogenic genes (Acta2, Col1a1, Fsp1, and Fstl1) and Arg 1 (coding arginase 1). Deletion of Chrna7 attenuated lung arginase 1+Ly6C+ cells, Masson's trichrome staining, hydroxyproline levels, and expression of α-SMA at protein levels in BLM-challenged mice. Mechanistically, activation of α7 nAChR in human fibroblasts increased TGF-ß-induced phosphorylation of Smad2/3 and transcription of fibrogenic genes (Acta2, Col1a1). In isolated mouse lung fibroblasts, activation of α7 nAChR also enhanced TGF-ß induced-transcription of fibrogenic genes; however, deletion of Chrna7 diminished these effects. Taken together, deficiency of α7 nAChR could suppress the development of BLM-induced lung fibrosis. Thus, α7 nAChR might be a novel therapeutic target for treating lung fibrosis.

The nicotinic acetylcholine receptor α7 subunit is an essential negative regulator of bone mass.

The nicotinic receptor α7nAchR reportedly regulates vagal nerve targets in brain and cardiac tissue. Here we show that nAchR7-/- mice exhibit increased bone mass due to decreased osteoclast formation, accompanied by elevated osteoprotegerin/RANKL ratios in serum. Vagotomy in wild-type mice also significantly increased the serum osteoprotegerin/RANKL ratio, and elevated bone mass seen in nAchR7-/- mice was reversed in α7nAchR/osteoprotegerin-doubly-deficient mice. α7nAchR loss significantly increased TNFα expression in Mac1-positive macrophages, and TNFα increased the osteoprotegerin/RANKL ratio in osteoblasts. Targeting TNFα in nAchR7-/- mice normalized both serum osteoprotegerin/RANKL ratios and bone mass. Administration of nicotine, an α7nAchR ligand, to wild-type mice increased serum RANKL levels. Thus, vagal nerve stimulation of macrophages via α7nAchR regulates bone mass by modulating osteoclast formation.

Nicotine withdrawal-induced inattention is absent in alpha7 nAChR knockout mice.

RATIONALE: Smoking is the leading cause of preventable death in the USA, but quit attempts result in withdrawal-induced cognitive dysfunction and predicts relapse. Greater understanding of the neural mechanism(s) underlying these cognitive deficits is required to develop targeted treatments to aid quit attempts. OBJECTIVES: We examined nicotine withdrawal-induced inattention in mice lacking the α7 nicotinic acetylcholine receptor (nAChR) using the five-choice continuous performance test (5C-CPT). METHODS: Mice were trained in the 5C-CPT prior to osmotic minipump implantation containing saline or nicotine. Experiment 1 used 40 mg kg-1 day-1 nicotine treatment and tested C57BL/6 mice 4, 28, and 52 h after pump removal. Experiment 2 used 14 and 40 mg kg-1 day-1 nicotine treatment in α7 nAChR knockout (KO) and wildtype (WT) littermates tested 4 h after pump removal. Subsets of WT mice were killed before and after pump removal to assess changes in receptor expression associated with nicotine administration and withdrawal. RESULTS: Nicotine withdrawal impaired attention in the 5C-CPT, driven by response inhibition and target detection deficits. The overall attentional deficit was absent in α7 nAChR KO mice despite response disinhibition in these mice. Synaptosomal glutamate mGluR5 and dopamine D4 receptor expression were reduced during chronic nicotine but increased during withdrawal, potentially contributing to cognitive deficits. CONCLUSIONS: The α7 nAChR may underlie nicotine withdrawal-induced deficits in target detection but is not required for response disinhibition deficits. Alterations to the glutamatergic and dopaminergic pathways may also contribute to withdrawal-induced attentional deficits, providing novel targets to alleviate the cognitive symptoms of withdrawal during quit attempts.

Hepatic vagus nerve regulates Kupffer cell activation via α7 nicotinic acetylcholine receptor in nonalcoholic steatohepatitis.

BACKGROUND: Nonalcoholic fatty liver disease ranges from simple steatosis to nonalcoholic steatohepatitis (NASH). Kupffer cells play a central role in promoting hepatic inflammation, which leads to the development of NASH. We investigated the anti-inflammatory effect of hepatic vagus-mediated stimulation of the α7 nicotinic acetylcholine receptor (α7nAChR) on Kupffer cells in NASH pathogenesis. METHODS: Wild-type (WT) mice undergoing hepatic vagotomy (HV) were fed a methionine- and choline-deficient (MCD) diet for 1 week. α7nAChR knockout (α7KO) chimeric mice were generated by transplanting α7KO bone marrow cells into irradiated and Kupffer cell-deleted WT recipients. Kupffer cells were isolated from WT mice and treated with α7nAChR agonist under stimulation by lipopolysaccharide and/or palmitic acid. RESULTS: HV aggravated MCD diet-induced NASH in both steatosis and inflammation. The hepatic inflammatory response, including the upregulation of tumor necrosis factor alpha (TNFα), interleukin (IL)-12, and monocyte chemoattractant protein 1 (MCP-1), was accelerated in HV mice, accompanied by the downregulation of PPARα pathway genes. Kupffer cells were highly activated via the phosphorylation and nuclear translocation of nuclear factor-kappa B (NF-κB) in MCD diet-fed HV mice. The α7nAchR agonist suppressed the inflammatory response of primary Kupffer cells induced by lipopolysaccharide and palmitic acid by attenuating the NF-κB cascade. α7KO chimeric mice fed an MCD diet for 1 week developed advanced NASH with highly activated Kupffer cells. The hepatic expression of TNFα, IL-12, and MCP-1 was upregulated in α7KO chimeric mice, accompanied by abnormal lipid metabolism. CONCLUSIONS: Hepatic vagus activity regulates the inflammatory response of Kupffer cells via α7nAChR in NASH development.

Chrna7 deficient mice manifest no consistent neuropsychiatric and behavioral phenotypes.

The alpha7 nicotinic acetylcholine receptor, encoded by the CHRNA7 gene, has been implicated in various psychiatric and behavioral disorders, including schizophrenia, bipolar disorder, epilepsy, autism, Alzheimer's disease, and Parkinson's disease, and is considered a potential target for therapeutic intervention. 15q13.3 microdeletion syndrome is a rare genetic disorder, caused by submicroscopic deletions on chromosome 15q. CHRNA7 is the only gene in this locus that has been deleted entirely in cases involving the smallest microdeletions. Affected individuals manifest variable neurological and behavioral phenotypes, which commonly include developmental delay/intellectual disability, epilepsy, and autism spectrum disorder. Subsets of patients have short attention spans, aggressive behaviors, mood disorders, or schizophrenia. Previous behavioral studies suggested that Chrna7 deficient mice had attention deficits, but were normal in baseline behavioral responses, learning, memory, and sensorimotor gating. Given a growing interest in CHRNA7-related diseases and a better appreciation of its associated human phenotypes, an in-depth behavioral characterization of the Chrna7 deficient mouse model appeared prudent. This study was designed to investigate whether Chrna7 deficient mice manifest phenotypes related to those seen in human individuals, using an array of 12 behavioral assessments and electroencephalogram (EEG) recordings on freely-moving mice. Examined phenotypes included social interaction, compulsive behaviors, aggression, hyperactivity, anxiety, depression, and somatosensory gating. Our data suggests that mouse behavior and EEG recordings are not sensitive to decreased Chrna7 copy number.

Involvement of arterial baroreflex and nicotinic acetylcholine receptor α7 subunit pathway in the protection of metformin against stroke in stroke-prone spontaneously hypertensive rats.

Stroke is a leading cause of mortality and disability worldwide. There is growing evidence that metformin (Met) has potent neuroprotective effects; however, its mechanisms remain unclear. We examined the role of the arterial baroreflex and cholinergic-α7 nicotinic acetylcholine receptor (α7nAChR) anti-inflammory pathway in the beneficial effects of Met against stroke. Stroke-prone spontaneously hypertensive rats (SHRSP) were used to observe stroke development indicated by lifespan of SHRSP and the ischemic injury induced by permanent middle cerebral artery occlusion (MCAO). Sinoaortic denervation was used to inactivate the arterial baroreflex. MCAO were also performed in α7nAChR knockout (KO) mice. Briefly, Met increased the life span of SHRSP and reduced the infarct area induced by MCAO. Met also improved the function of arterial baroreflex. The beneficial effects of Met on stroke were markedly attenuated by blunting the arterial baroreflex. Met up-regulated the expression of vesicular acetylcholine transporter (VAChT) and α7nAChR, down-regulated the level of pro-inflammtory cytokines in serum and peri-infarct of ischemic brain. Arterial baroreflex dysfunction decreased the expression of VAchT and α7nAChR, showed upward tendency in the level of pro-inflammtory cytokines. Most importantly, arterial baroreflex dysfunction nearly abolished such effect of Met on cholinergic signaling. In addition, the α7nAChR KO mice also had significantly worse ischemic damage induced by MCAO, and neuroprotection of Met disappeared in α7nAChR KO mice. In conclusion, Met improved the arterial baroreflex function, and then enhancing cholinergic anti-inflammatory pathway in an α7nAChR-dependent manner, thereby effectively prevent ischemic induced brain injury and delayed stroke onset in SHRSP.

Nicotine Accelerates Atherosclerosis in Apolipoprotein E-Deficient Mice by Activating α7 Nicotinic Acetylcholine Receptor on Mast Cells.

OBJECTIVE: Cigarette smoking is an independent risk factor for atherosclerosis. Nicotine, the addictive component of cigarettes, induces mast cell (MC) release and contributes to atherogenesis. The purpose of this study was to determine whether nicotine accelerates atherosclerosis through MC-mediated mechanisms and whether MC stabilizer prevents this pathological process. APPROACH AND RESULTS: Nicotine administration increased the size of atherosclerotic lesions in apolipoprotein E-deficient (Apoe-/-) mice fed a fat-enriched diet. This was accompanied by enhanced intraplaque macrophage content and lipid deposition but reduced collagen and smooth muscle cell contents. MC deficiency in Apoe-/- mice (Apoe-/-KitW-sh/W-sh) diminished nicotine-induced atherosclerosis. Nicotine activated bone marrow-derived MCs in vitro, which was inhibited by a MC stabilizer disodium cromoglycate or a nonselective nicotinic acetylcholine receptor blocker mecamylamine. Further investigation revealed that α7 nicotinic acetylcholine receptor was a target for nicotine activation in MCs. Nicotine did not change atherosclerotic lesion size of Apoe-/-KitW-sh/W-sh mice reconstituted with MCs from Apoe-/-α7nAChR-/- animals. CONCLUSIONS: Activation of α7 nicotinic acetylcholine receptor on MCs is a mechanism by which nicotine enhances atherosclerosis.

α7 Nicotinic Acetylcholine Receptor Relieves Angiotensin II-Induced Senescence in Vascular Smooth Muscle Cells by Raising Nicotinamide Adenine Dinucleotide-Dependent SIRT1 Activity.

OBJECTIVE: α7 nicotinic acetylcholine receptor (α7nAChR) is a subtype of nAChR and has been reported to be involved in hypertension end-organ damage. In this study, we tested the role of α7nAChR in angiotensin II (Ang II)-induced senescence of vascular smooth muscle cells (VSMCs). APPROACH AND RESULTS: Expression of α7nAChR was not influenced by Ang II. Ang II induced remarkable senescent phenotypes in rodent and human VSMCs, including increased senescence-associated ß-galactosidase activity, phosphorylation of H2A.X(Ser139), phosphorylation of Chk1(Ser317), reduced replication, and downregulation of proliferating cell nuclear antigen. Activation of α7nAChR with a selective agonist PNU-282987 blocked Ang II-induced senescence in cultured VSMCs. Moreover, PNU-282987 treatment attenuated the Ang II infusion-induced VSMC senescence in wild-type but not in α7nAChR(-/-) mice. PNU-282987 reduced the Ang II-enhanced reactive oxygen species, lipid peroxidation, and the expression of NADPH oxidase 1, NADPH oxidase 4, and p22(phox) in cultured VSMCs isolated from wild-type but not in α7nAChR(-/-) mice. Furthermore, PNU-282987 diminished Ang II-induced prosenescence signaling pathways, including p53, acetyl-p53, p21, and p16(INK4a). Finally, although α7nAChR activation by PNU-282987 did not affect the Ang II-induced downregulation of sirtuin 1 (SIRT1), it significantly increased intracellular NAD(+) levels, and thereby enhanced SIRT1 activity in an AMP-dependent protein kinase-independent manner. Depletion of SIRT1 by knockdown or SIRT1 inhibitor EX527 abrogated the antisenescence effect of α7nAChR against Ang II. CONCLUSIONS: Our results demonstrate that activation of α7nAChR alleviates Ang II-induced VSMC senescence through promoting NAD(+)-SIRT1 pathway, suggesting that α7nAChR may be a potential therapeutic target for the treatment of Ang II-associated vascular aging disorders.

Central Insulin Action Activates Kupffer Cells by Suppressing Hepatic Vagal Activation via the Nicotinic Alpha 7 Acetylcholine Receptor.

Central insulin action activates hepatic IL-6/STAT3 signaling, which suppresses the gene expression of hepatic gluconeogenic enzymes. The vagus nerve plays an important role in this centrally mediated hepatic response; however, the precise mechanism underlying this brain-liver interaction is unclear. Here, we present our findings that the vagus nerve suppresses hepatic IL-6/STAT3 signaling via α7-nicotinic acetylcholine receptors (α7-nAchR) on Kupffer cells, and that central insulin action activates hepatic IL-6/STAT3 signaling by suppressing vagal activity. Indeed, central insulin-mediated hepatic IL-6/STAT3 activation and gluconeogenic gene suppression were impeded in mice with hepatic vagotomy, pharmacological cholinergic blockade, or α7-nAchR deficiency. In high-fat diet-induced obese and insulin-resistant mice, control of the vagus nerve by central insulin action was disturbed, inducing a persistent increase of inflammatory cytokines. These findings suggest that dysregulation of the α7-nAchR-mediated control of Kupffer cells by central insulin action may affect the pathogenesis of chronic hepatic inflammation in obesity.

α7 nicotinic ACh receptor-deficient mice exhibit sustained attention impairments that are reversed by ß2 nicotinic ACh receptor activation.

BACKGROUND AND PURPOSE: Disruptions of executive function, including attentional deficits, are a hallmark of a number of diseases. ACh in the prefrontal cortex regulates attentive behaviour; however, the role of α7 nicotinic ACh receptor (α7nAChR) in attention is contentious. EXPERIMENTAL APPROACH: In order to probe attention, we trained both wild-type and α7nAChR knockout mice on a touch screen-based five-choice serial reaction time task (5-CSRT). Following training procedures, we then tested sustained attention using a probe trial experiment. To further differentiate the role of specific nicotinic receptors in attention, we then tested the effects of both α7nAChR and ß2nAChR agonists on the performance of both wild-type and knockout mice on the 5-CSRT task. KEY RESULTS: At low doses, α7nAChR agonists improved attentional performance of wild-type mice, while high doses had deleterious effects on attention. α7nAChR knockout mice displayed deficits in sustained attention that were not ameliorated by α7nAChR agonists. However, these deficits were completely reversed by the administration of a ß2nAChR agonist. Furthermore, administration of a ß2nAChR agonist in α7nAChR knockout mice elicited similar biochemical response in the prefrontal cortex as the administration of α7nAChR agonists in wild-type mice. CONCLUSIONS AND IMPLICATIONS: Our experiments reveal an intricate relationship between distinct nicotinic receptors to regulate attentional performance and provide the basis for targeting ß2nAChRs pharmacologically to decrease attentional deficits due to a dysfunction in α7nAChRs.

α7 and ß2 Nicotinic Acetylcholine Receptor Subunits Form Heteromeric Receptor Complexes that Are Expressed in the Human Cortex and Display Distinct Pharmacological Properties.

The existence of α7ß2 nicotinic acetylcholine receptors (nAChRs) has recently been demonstrated in both the rodent and human brain. Since α7-containing nAChRs are promising drug targets for schizophrenia and Alzheimer's disease, it is critical to determine whether α7ß2 nAChRs are present in the human brain, in which brain areas, and whether they differ functionally from α7 nAChR homomers. We used α-bungarotoxin to affinity purify α7-containing nAChRs from surgically excised human temporal cortex, and found that α7 subunits co-purify with ß2 subunits, indicating the presence of α7ß2 nAChRs in the human brain. We validated these results by demonstrating co-purification of ß2 from wild-type, but not α7 or ß2 knock-out mice. The pharmacology and kinetics of human α7ß2 nAChRs differed significantly from that of α7 homomers in response to nAChR agonists when expressed in Xenopus oocytes and HEK293 cells. Notably, α7ß2 heteromers expressed in HEK293 cells display markedly slower rise and decay phases. These results demonstrate that α7 subunits in the human brain form heteromeric complexes with ß2 subunits, and that human α7ß2 nAChR heteromers respond to nAChR agonists with a unique pharmacology and kinetic profile. α7ß2 nAChRs thus represent an alternative mechanism for the reported clinical efficacy of α7 nAChR ligands.

Impaired synaptic plasticity in the visual cortex of mice lacking α7-nicotinic receptor subunit.

The primary visual cortex (V1) is the first step in visual information processing and its function may be modulated by acetylcholine through nicotinic receptors (nAChRs). Since our previous work demonstrated that visual acuity and cortical spatial resolution limit were significantly reduced in α7 knock-out (KO) mice in the absence of retinal alterations, we decided to characterize the contribution of homomeric α7 nicotinic receptors (α7nAChRs) to visual information processing at the cortical level. We evaluated long-term forms of synaptic plasticity in occipital slices containing V1 from α7 KO mice and in wild-type (WT) slices perfused with nAChRs selective blocking agents. In α7 KO mice slices, electrophysiological recordings demonstrated the absence of long-term potentiation (LTP) and long-term depression (LTD) in layer II/III after the stimulation of different intracortical pathways (layer IV or II/III). Furthermore, the acute and selective blockade of α7nAChRs in slices from WT mice with either α-bungarotoxin or methyllycaconitine did not alter the expression of LTP and LTD. Conversely, the perfusion with the unspecific nAChRs antagonist mecamylamine impaired LTP and LTD. Our results suggest the presence of impaired synaptic plasticity in the V1 of α7 KO mice and indicate a different contribution of nAChRs to visual cortex function.

Hematopoietic α7 nicotinic acetylcholine receptor deficiency increases inflammation and platelet activation status, but does not aggravate atherosclerosis.

BACKGROUND: The autonomic nervous system attenuates inflammation through activation of the α7 nicotinic acetylcholine receptor (α7nAChR), a pathway termed the cholinergic anti-inflammatory reflex. Interestingly, α7nAChR is expressed on immune cells and platelets, both of which play a crucial role in the development of atherosclerosis. OBJECTIVE: To investigate the role of hematopoietic α7nAChR in inflammation and platelet function in atherosclerotic ldlr(-/-) mice and to identify its consequences for atherosclerotic lesion development. METHODS: Bone marrow from α7nAChR(-/-) mice or wild-type littermates was transplanted into irradiated ldlr(-/-) mice. After a recovery period of 8 weeks, the mice were fed an atherogenic Western-type diet for 7 weeks. RESULTS: Hematopoietic α7nAChR deficiency clearly increased the number of leukocytes in the peritoneum (2.6-fold, P < 0.001), blood (2.9-fold; P < 0.01), mesenteric lymph nodes (2.0-fold; P < 0.001) and spleen (2.2-fold; P < 0.01), indicative of an increased inflammatory status. Additionally, expression of inflammatory mediators was increased in peritoneal leukocytes (TNFα, 1.6-fold, P < 0.01; CRP, 1.8-fold, P < 0.01) as well as in the spleen (TNFα, 1.6-fold, P < 0.01). The lack of α7nAChR on platelets from these mice increased the expression of active integrin αIIb ß3 upon stimulation by ADP (1.9-fold, P < 0.01), indicating increased activation status, while incubation of human platelets with an α7nAChR agonist decreased aggregation (-35%, P < 0.05). Despite the large effects of hematopoietic α7nAChR deficiency on inflammatory status and platelet function, it did not affect atherosclerosis development or composition of lesions. CONCLUSIONS: Hematopoietic α7nAChR is important for attenuation of inflammatory responses and maintaining normal platelet reactivity, but loss of hematopoietic α7nAChR does not aggravate development of atherosclerosis.