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ObjectivesNever-smoking women in Xuanwei (XW), China, have some of the highest lung cancer rates in the country. This has been attributed to the combustion of smoky coal used for indoor cooking and heating. The aim of this study was to evaluate the spectrum of cause-specific mortality in this unique population, including among those who use smokeless coal, considered ‘cleaner’ coal in XW, as this has not been well-characterised.DesignCohort study.SettingXW, a rural region of China where residents routinely burn coal for indoor cooking and heating.ParticipantsAge-adjusted, cause-specific mortality rates between 1976 and 2011 were calculated and compared among lifetime smoky and smokeless coal users in a cohort of 42 420 men and women from XW. Mortality rates for XW women were compared with those for a cohort of predominately never-smoking women in Shanghai.ResultsMortality in smoky coal users was driven by cancer (41%), with lung cancer accounting for 88% of cancer deaths. In contrast, cardiovascular disease (CVD) accounted for 32% of deaths among smokeless coal users, with 7% of deaths from cancer. Total cancer mortality was four times higher among smoky coal users relative to smokeless coal users, particularly for lung cancer (standardised rate ratio (SRR)=17.6). Smokeless coal users had higher mortality rates of CVD (SRR=2.9) and pneumonia (SRR=2.5) compared with smoky coal users. These patterns were similar in men and women, even though XW women rarely smoked cigarettes. Women in XW, regardless of coal type used, had over a threefold higher rate of overall mortality, and most cause-specific outcomes were elevated compared with women in Shanghai.ConclusionsCause-specific mortality burden differs in XW based on the lifetime use of different coal types. These observations provide evidence that eliminating all coal use for indoor cooking and heating is an important next step in improving public health particularly in developing countries.

As a cis‐acting non‐depolarizing neuromuscular blocker through a nicotinic acetylcholine receptor (nAChR), cisatracurium (CAC) is widely used in anaesthesia and intensive care units. nAChR may be present on Leydig cells to mediate the action of CAC. Here, by Western blotting, immunohistochemistry and immunofluorescence, we identified that CHRNA4 (a subunit of nAChR) exists only on rat adult Leydig cells. We studied the effect of CAC on the synthesis of testosterone in rat adult Leydig cells and mouse MLTC‐1 tumour cells. Rat Leydig cells and MLTC‐1 cells were treated with CAC (5, 10 and 50 μmol/L) or nAChR agonists (50 μmol/L nicotine or 50 μmol/L lobeline) for 12 hours, respectively. We found that CAC significantly increased testosterone output in rat Leydig cells and mouse MLTC‐1 cells at 5 μmol/L and higher concentrations. However, nicotine and lobeline inhibited testosterone synthesis. CAC increased intracellular cAMP levels, and nicotine and lobeline reversed this change in rat Leydig cells. CAC may increase testosterone synthesis in rat Leydig cells and mouse MLTC‐1 cells by up‐regulating the expression of Lhcgr and Star. Up‐regulation of Scarb1 and Hsd3b1 expression by CAC was also observed in rat Leydig cells. In addition to cAMP signal transduction, CAC can induce ERK1/2 phosphorylation in rat Leydig cells. In conclusion, CAC binds to nAChR on Leydig cells, and activates cAMP and ERK1/2 phosphorylation, thereby up‐regulating the expression of key genes and proteins in the steroidogenic cascade, resulting in increased testosterone synthesis in Leydig cells.

To observe the therapeutic effect of moxa fume in the treatment of chronic rhinosinusitis（CRS） and the effect of acupuncture plus smokeless moxibustion or smoky moxibustion on the expression of thymic stromal lymphopoietin (TSLP) and pituitary adenylate cyclase activating polypeptide (PACAP) proteins in the sinus mucosal tissue in CRS mice.Sixty male C57BL/6J mice were randomly divided into 6 groups, namely normal control, sham operation, CRS model, medication, acupuncture plus smokeless moxibustion (Acu+smokeless Moxi) and acupuncture plus smoky moxibustion (Acu+smoky Moxi) groups, with 20 mice in each group. The CRS model was established by inserting a piece of polyporous sponge filled with streptococcus pneumoniae into the maxillary sinus after operation. The mice in the sham operation group received skin incision after opening the maxillary sinus. Mice of the medication group received gavage of clarithromycin 0.103 g·kgAt the end of the treatment, mice of the model group still had symptoms of nasal obstruction and runny nose, but those of the 3 treatment groups were obviously relieved in the nasal symptoms. H.E. staining showed an obvious chronic inflammatory reaction in the sinus mucosa, uneven distribution of the mucosal epithelium and necrotic and exfoliated epithelial cells, hyperplasia of fibrous tissue in the submucosa, etc. in the model group, which were relatively milder in the medication, Acu+smokeless Moxi and Acu+smoky Moxi groups, while no obvious inflammation was found in the normal group and sham operation group. In comparison with the normal group, no significant changes were found in the expression levels of PACAP and TSLP in the sham operation group (Acupuncture combined with smoky moxibustion can down-regulate the expression of TSLP protein in the nasal sinus mucosa in CRS mice, which maybe contribute to its effect in reducing the inflammatory reaction and nasal symptoms.

Methamphetamine is a potent psychostimulant with high abuse rates. Currently, there is no Food and Drug Administration-approved pharmacotherapy for methamphetamine addiction. Ideally, a pharmacotherapy should selectively decrease methamphetamine self-administration without affecting responding for other reinforcers. One way to test this is with the use of a multiple schedule of reinforcement, in which drug and food are available in alternating components within a session. The present study evaluated GZ-793A, a vesicular monoamine transporter-2 inhibitor, and varenicline, a partial agonist at α4β2 and full agonist at α7 nicotinic acetylcholine receptors, for their ability to decrease methamphetamine and food self-administration using a multiple schedule of reinforcement. Male Sprague-Dawley rats self-administered methamphetamine (0.03 mg/kg/intravenous infusion) and food pellets under a multiple schedule of reinforcement. GZ-793A or varenicline was administered before multiple schedule sessions. GZ-793A (5 and 20 mg/kg) significantly decreased methamphetamine intake compared with saline and did not alter food-maintained responding. In contrast, varenicline decreased methamphetamine intake less specifically across time. The results suggest that vesicular monoamine transporter-2 inhibition may be a viable pharmacological target for the treatment of methamphetamine-use disorders.

Juxtapulmonary receptors (J) lying in the lung parenchyma are stimulated naturally by any condition that produces interstitial oedema, transient increases in interstitial volume and pressure or raised pulmonary capillary pressure. There is no information available about the level of their stimulation in patients with idiopathic pulmonary hypertension (IPH) who have high levels of pulmonary artery systolic pressures. The aim of the present study therefore was to find the level of these receptors activity in these patients at their prevailing pulmonary artery systolic pressures. This was done by the established method of determining the dose of i.v. lobeline that gives rise to threshold levels of sensations in the upper chest areas and accelerates respiration. In IPH patients it was found to be as high as 31.6 ± 5.6 μg/kg i.e., twice as much as that known for healthy individuals which is 15 μg/kg. This shows an enhanced stimulation of J receptors in IPH patients. Expectedly when pulmonary artery systolic pressure falls with pulmonary bed vasodilator medication given to IPH patients, a reduction in the natural stimulus of J receptors would also occur leading to a fall in their activity and hence that of the quantum of their reflexes of respiratory acceleration and inhibition of exercise. This finding provides the first insight of a neural mechanism that could be influenced to produce its effects when pulmonary artery systolic pressure falls by pulmonary vasodilator medication.

Evidence indicates that the brain nicotinic acetylcholine receptor (nAChRs) ligand lobeline reduces depression-like behaviors, ethanol drinking, and nicotine withdrawal-induced depression-like behaviors. The purpose of the present study was to determine the effects of lobeline on ethanol abstinence-induced depression-like behavior and associated neuroadaptive changes in mice. Adult C57BL/6J male mice were allowed to drink 10% ethanol for 4 weeks using a two-bottle choice procedure. Mice were tested after 24 h and 14 days of ethanol abstinence in a forced swim test (FST), a measure for depression-like behavior. Acute lobeline treatment (1 mg/kg) significantly reduced immobility time compared to controls after 24 h and 14 days of abstinence. In addition, abstinence from chronic ethanol exposure reduced serotonin levels in the hippocampus, which was reversed by acute lobeline treatment. Repeated lobeline treatment (1 mg/kg, once daily) for 14 days during ethanol abstinence also significantly reduced FST immobility in mice exposed to ethanol. Chronic ethanol exposure significantly reduced the number of 5-bromo 2′-deoxyuridine (BrdU)-positive cells in the dentate gyrus of the hippocampus, indicating decreased hippocampal cell proliferation. Abstinence from chronic ethanol exposure also decreased brain-derived neurotrophic factor (BDNF) in the dentate gyrus and CA3 region of the hippocampus. In contrast, repeated lobeline treatment significantly increased both BrdU- and BDNF-positive cells. Taken together, our results indicate that lobeline produced antidepressant-like effects, likely by targeting brain β2-containing nAChRs, serotonergic neurotransmission, and/or hippocampal cell proliferation. Therefore, lobeline may have therapeutic utility to treat alcohol abstinence-induced depression.

Despite increased methamphetamine use worldwide, pharmacotherapies are not available to treat methamphetamine use disorder. The vesicular monoamine transporter-2 (VMAT2) is an important pharmacological target for discovery of treatments for methamphetamine use disorder. VMAT2 inhibition by the natural product, lobeline, reduced methamphetamine-evoked dopamine release, methamphetamine-induced hyperlocomotion, and methamphetamine self-administration in rats. Compared to lobeline, lobelane exhibited improved affinity and selectivity for VMAT2 over nicotinic acetylcholine receptors. Lobelane inhibited neurochemical and behavioral effects of methamphetamine, but tolerance developed to its behavioral efficacy in reducing methamphetamine self-administration, preventing further development. The lobelane analog, R-N-(1,2-dihydroxypropyl)-2,6-cis-di-(4-methoxyphenethyl)piperidine hydrochloride (GZ-793A), potently and selectively inhibited VMAT2 function and reduced neurochemical and behavioral effects of methamphetamine. However, GZ-793A exhibited potential to induce ventricular arrhythmias interacting with human-ether-a-go-go (hERG) channels. Herein, a new lead, R-3-(4-methoxyphenyl)-N-(1-phenylpropan-2-yl)propan-1-amine (GZ-11610), from the novel scaffold (N-alkyl(1-methyl-2-phenylethyl)amine) was evaluated as a VMAT2 inhibitor and potential therapeutic for methamphetamine use disorder. GZ-11610 was 290-fold selective for VMAT2 over dopamine transporters, suggesting that it may lack abuse liability. GZ-11610 was 640- to 3500-fold selective for VMAT2 over serotonin transporters and nicotinic acetylcholine receptors. GZ-11610 exhibited > 1000-fold selectivity for VMAT2 over hERG, representing a robust improvement relative to our previous VMAT2 inhibitors. GZ-11610 (3-30 mg/kg, s.c. or 56-300 mg/kg, oral) reduced methamphetamine-induced hyperactivity in methamphetamine-sensitized rats. Thus, GZ-11610 is a potent and selective inhibitor of VMAT2, may have low abuse liability and low cardiotoxicity, and after oral administration is effective and specific in inhibiting the locomotor stimulant effects of methamphetamine, suggesting further investigation as a potential therapeutic for methamphetamine use disorder.

Evidence suggests that lobeline, a nicotinic acetylcholine receptor ligand, has antidepressant-like properties in mice. The present study investigated the possible additive or synergistic effects of lobeline in combination with commonly used antidepressants, such as reboxetine, fluoxetine, or bupropion, using the tail suspension test (TST) and the forced swim test (FST) in C57BL/6J mice. Reboxetine (5 or 10 mg/kg, i.p.), fluoxetine (5 or 10 mg/kg, i.p.), or bupropion (2 or 4 mg/kg, i.p.) were administered 30 min before TST or FST. A fixed dose of lobeline (1 mg/kg, i.p.) was injected 15 min prior to tests. Co-administration of lobeline and reboxetine, fluoxetine, or bupropion significantly reduced immobility time in the TST and FST in comparison to the antidepressants used alone. The results suggest that lobeline enhanced the effects of reboxetine, fluoxetine, or bupropion in mice. Therefore, lobeline or similar nicotinic receptor ligand may have therapeutic potential as an adjunct for the treatment of major depression.

Aim Lobeline is a natural alkaloid derived from Lobelia inflata that has been investigated as a clinical candidate for the treatment of alcoholism. In a pre-clinical trial, lobeline decreased the preference for and consumption of ethanol, due to the modulation of the nicotinic acetylcholine receptor. However, the interaction between lobeline and ethanol is poorly known and thus there are safety concerns. The present study was conducted to evaluate the mutagenic and genotoxic effects of lobeline and assess its modulation of ethanol-induced toxicological effects. Main methods CF-1 male mice were divided into five groups. Groups received an intraperitoneal injection of saline solution, lobeline (5 or 10 mg/kg), ethanol (2.5 g/kg), or lobeline plus ethanol, once a day for three consecutive days. Genotoxicity was evaluated in peripheral blood using the alkaline comet assay. The mutagenicity was evaluated using both Salmonella /microsome assay in TA1535, TA97a, TA98, TA100, and TA102 Salmonella typhimurium strains and the micronucleus test in bone marrow. Possible liver and kidney injuries were evaluated using biochemical analysis. Key findings Lobeline did not show genotoxic or mutagenic effects and did not increase the ethanol-induced genotoxic effects in blood. Lobeline also protected blood cells against oxidative damage induced by hydrogen peroxide. Biochemical parameters were not altered, indicating no liver or kidney injuries or alterations in lipid and carbohydrate metabolisms. Significance These findings suggest that lobeline does not induce gene or chromosomal mutations, and that this lack of genetic toxicity is maintained in the presence of ethanol, providing further evidence of the safety of this drug to treat alcohol dependence.

Evidence suggests that neuronal nicotinic acetylcholine receptor (nAChR) ligand lobeline has antidepressant-like properties. The present study investigated the effects of lobeline on nicotine withdrawal-induced depression-like behavior. Adult C57BL/6J mice were exposed to nicotine (200 μg/ml) in drinking solution for 3 weeks. During withdrawal, depression-like behavior was measured by the forced swim test (FST). We also determined norepinephrine (NE) levels in the prefrontal cortex (PFC) and hippocampus during nicotine withdrawal. Furthermore, we determined the effects of repeated treatment with lobeline or a selective α4β2 nAChR ligand 3-(pyridine-3-yl)-cytisine on brain-derived neurotrophic factor (BDNF) and phosphorylated cAMP-responsive element binding (p-CREB) protein expression in the hippocampus. Withdrawal from chronic nicotine increased immobility time in the FST, a measure for depression-like behavior. Pretreatment with lobeline significantly decreased immobility time during nicotine withdrawal. In addition, pretreatment with lobeline attenuated nicotine withdrawal-induced increased NE levels in the PFC and hippocampus. Further, repeated treatment with lobeline or 3-(pyridine-3-yl)-cytisine decreased immobility time in the FST and reduced withdrawal-induced increased BDNF and p-CREB expression in the hippocampus. Taken together, our results indicate that lobeline attenuated nicotine withdrawal-induced depression-like behavior likely by targeting brain nAChRs, noradrenergic neurotransmission, and/or hippocampal BDNF. Thus, lobeline may have some potential to prevent smoking relapse by counteracting nicotine withdrawal-induced depression in humans.

Lobeline is a natural alkaloid with high affinity for nicotinic acetylcholine receptors, and it is a promising candidate for addiction treatment in human beings. This work evaluated the toxicological profile of lobeline with different behavioural models and investigated its effect on DNA damage (comet assay and micronucleus test) in mice. Acute administration of lobeline (5 or 10 mg/kg; i.p.) did not impair the parameters measured in the habituation and inhibitory avoidance test, suggesting that it has no effect on memory acquisition in these tasks. Lobeline did not affect the number or the latency to the first head-dip in the hole board test, indicating that it was not anxiolytic/anxiogenic in this model. No genotoxic effects were observed in blood, liver and brain tissues collected 24 hr after the single injection of lobeline (both doses). There was no increase in micronucleus frequency in mice treated with lobeline, indicating the absence of toxicity in bone marrow of the animals. Therefore, the acute treatment with high doses of lobeline did not impair the behavioural parameters measured in this work. Additionally, the drug was not able to produce DNA damage.

Despite the high prevalence of use of methamphetamine (METH), there is no FDA-approved pharmacological treatment available currently for METH addiction. The vesicular monoamine transporter (VMAT2) has been proposed as a novel target to treat METH abuse. GZ-793A, a lobelane analog and selective VMAT2 inhibitor, has been shown previously to decrease METH self-administration specifically when administered via the subcutaneous route in rats. Since oral administration is the preferred clinical route, the present experiments determined if oral administration of GZ-793A would decrease specifically METH self-administration. Experiments 1 and 2 assessed the dose-effect functions of oral administration of GZ-793A (30–240 mg/kg) on intravenous METH self-administration and food-maintained responding, respectively. Experiments 3 and 4 assessed the time-course (20–180 min pretreatment) of oral administration of GZ-793A on METH self-administration and food-maintained responding, respectively. Oral administration of GZ-793A dose-dependently decreased METH self-administration, with the highest dose (240 mg/kg) producing an 85% decrease compared to control baseline. The decrease in METH self-administration produced by GZ-793A (120 mg/kg) lasted at least 180 min. In contrast, GZ-793A failed to alter food-maintained responding at any of the doses or pretreatment intervals tested. The oral effectiveness and the specificity of GZ-793A to decrease methamphetamine self-administration support the feasibility of developing VMAT2 inhibitors as treatments for METH abuse.

Objective: In preclinical studies, lobeline inhibited hyperactivity induced by nicotine and amphetamine, and improved performance and learning in studies utilizing radial-arm maze and spatial-discrimination water maze. This laboratory proof-of-concept study investigated lobeline as a treatment for ADHD symptoms in adults (31.11 ± 7.08 years). Method: Using cognitive tasks and self-report measures, the effects of lobeline (0, 7.5, 15, or 30 mg, s.l.) and methylphenidate (0, 15, or 30 mg, p.o.) were assessed in nine volunteers with ADHD. Results: Evidence suggested that lobeline could modestly improve working memory in adults with ADHD, but no significant improvement in attention was observed. Lobeline administration was associated with mild adverse side effects (nausea). Conclusion: Further investigation of lobeline on working memory may be warranted.

Lobeline and lobelane inhibit the behavioral and neurochemical effects of methamphetamine via an interaction with the vesicular monoamine transporter-2 (VMAT2). However, lobeline has high affinity for nicotinic receptors, and tolerance develops to the behavioral effects of lobelane. A water-soluble analog of lobelane, R-N-(1,2–dihydroxypropyl)-2,6-cis-di-(4-methoxyphenethyl)piperidine hydrochloride (GZ-793A), also interacts selectively with VMAT2 to inhibit the effects of methamphetamine, but does not produce behavioral tolerance. The current study further evaluated the mechanism underlying the GZ-793A-mediated inhibition of the neurochemical effects of methamphetamine. In contrast to lobeline, GZ-793A does not interact with the agonist recognition site on α4β2* and α7* nicotinic receptors. GZ-793A (0.3–100 μM) inhibited methamphetamine (5 μM)-evoked fractional dopamine release from rat striatal slices, and did not evoke dopamine release in the absence of methamphetamine. Furthermore, GZ-793A (1–100 μM) inhibited neither nicotine (30 μM)-evoked nor electrical field-stimulation-evoked (100 Hz/1min) fractional dopamine release. Unfortunately, GZ-793A inhibited [(3)H]dofetilide binding to human-ether-a-go-go related gene channels expressed on human embryonic kidney cells, and further, prolonged action potentials in rabbit cardiac Purkinje fibers, suggesting the potential for GZ-793A to induce ventricular arrhythmias. Thus, GZ-793A selectively inhibits the neurochemical effects of methamphetamine and lacks nicotinic receptor interactions; however, development as a pharmacotherapy for methamphetamine use disorders will not be pursued due to its potential cardiac liabilities.

In screening a library of plant extracts from ~1000 species native to the Southeastern United States, Lobelia cardinalis was identified as containing nicotinic acetylcholine receptor (nicAchR) binding activity which was relatively non-selective for the α4β2- and α7-nicAchR subtypes. This nicAchR binding profile is atypical for plant-derived nicAchR ligands, the majority of which are highly selective for α4β2-nicAchRs. Its potential therapeutic relevance is noteworthy since agonism of α4β2- and α7-nicAchRs is associated with anti-inflammatory and neuroprotective properties. Bioassay-guided fractionation of L. cardinalis extracts led to the identification of lobinaline, a complex binitrogenous alkaloid, as the main source of the unique nicAchR binding profile. Purified lobinaline was a potent free radical scavenger, displayed similar binding affinity at α4β2- and α7-nicAchRs, exhibited agonist activity at nicAchRs in SH-SY5Y cells, and inhibited [(3)H]-dopamine (DA) uptake in rat striatal synaptosomes. Lobinaline significantly increased fractional [(3)H] release from superfused rat striatal slices preloaded with [(3)H]-DA, an effect that was inhibited by the non-selective nicAchR antagonist mecamylamine. In vivo electrochemical studies in urethane-anesthetized rats demonstrated that lobinaline locally applied in the striatum significantly prolonged clearance of exogenous DA by the dopamine transporter (DAT). In contrast, lobeline, the most thoroughly investigated Lobelia alkaloid, is an α4β2-nicAchR antagonist, a poor free radical scavenger, and is a less potent DAT inhibitor. These previously unreported multifunctional effects of lobinaline make it of interest as a lead to develop therapeutics for neuropathological disorders that involve free radical generation, cholinergic, and dopaminergic neurotransmission. These include neurodegenerative conditions, such as Parkinson's disease, and drug abuse.

Alcohol and nicotine are commonly abused drugs in humans and evidence suggests that neuronal nicotinic acetylcholine receptors (nAChRs) in the midbrain dopamine system are common targets for the neurobehavioral interactions between alcohol (ethanol) and nicotine. The present study examined the efficacy of nAChR ligands with different pharmacological profiles such as cytisine, lobeline and dihydro-β-erythroidine (DHβE) to modulate chronic nicotine-induced increase in ethanol intake by C57BL/6J mice, using a two-bottle choice procedure. After establishment of baseline ethanol preference (10%, v/v), animals received daily subcutaneous injections of saline, nicotine (0.4 mg/kg) or different doses of cytisine, lobeline or DHβE 15 min prior to nicotine, for 10 days. Ethanol and water were presented immediately after the last (saline or nicotine) injection and fluid levels were monitored for post 1 h and 2 h treatment. Compared to control, nicotine injection significantly increased mean ethanol intake over 10 days, at both post 1 h and 2 h. Pretreatment with cytisine (0.5, 1.5 or 3.0 mg/kg) or lobeline (4.0 or 10.0 mg/kg) significantly reduced nicotine-induced increase in ethanol intake post 1 h and 2 h, without affecting water consumption. DHβE (0.5 or 2.0 mg/kg) failed to suppress nicotine-induced ethanol intake across 2 h post injection. These results indicate that nAChR-mediated signaling is critical in regulating nicotine-induced ethanol drinking behaviors.

To investigate the anticonvulsant activity of the lobeline isolated from the Lobelia nicotianaefolia in chemoconvulsant-induced seizures and its biochemical mechanism by investigating relationship between seizure activities and altered gamma amino butyric acid (GABA) in brain of mice in Pentylenetetrazol (PTZ) seizure models.The anticonvulsant activity of the isolated lobeline (5, 10, 20 and 30 mg/kg, i.p.) was investigated in PTZ and strychnine induced seizures in mice and the effect of isolated lobeline on brain GABA level in seizures induced by PTZ. Diazepam was used as reference anticonvulsant drugs for comparison.Isolated lobeline (10, 20 and 30 mg/kg, i.p.) significantly delayed and antagonized (P0.050-0.001) the onset of PTZ-induced seizures. It also antagonized strychnine induced seizures. The mortality was also prevented in the test group of animals. In biochemical evaluation, isolated lobeline (5, 10 and 20 mg/kg, i.p.) significantly increased the brain GABA level. And at dose of 30 mg/kg GABA level showed slight decrease in PTZ model.In our findings, isolated lobeline (20mg/kg) exhibited potent anticonvulsant activity against PTZ induced seizures. Also a biochemical evaluation suggested significant increase in barain GABA level at 20 mg/kg i.p. of isolated lobeline. Hence, we may propose that lobeline reduces epileptic seizures by enhancing the GABA release supporting the GABAergic mechanism.

Background: Genetic alteration of muscle cell glycosylation in muscular dystrophy models has ameliorated disease. Results: A high throughput screen identified a small molecule, lobeline, which altered muscle cell glycosylation and improved laminin binding. Conclusion: Lobeline increased abundance of sarcolemmal glycoproteins and increased laminin binding in an N-glycan-dependent manner. Significance: A novel approach revealed an unexpected role for N-glycans in muscle cell function., Duchenne muscular dystrophy is an X-linked disorder characterized by loss of dystrophin, a cytoskeletal protein that connects the actin cytoskeleton in skeletal muscle cells to extracellular matrix. Dystrophin binds to the cytoplasmic domain of the transmembrane glycoprotein β-dystroglycan (β-DG), which associates with cell surface α-dystroglycan (α-DG) that binds laminin in the extracellular matrix. β-DG can also associate with utrophin, and this differential association correlates with specific glycosylation changes on α-DG. Genetic modification of α-DG glycosylation can promote utrophin binding and rescue dystrophic phenotypes in mouse dystrophy models. We used high throughput screening with the plant lectin Wisteria floribunda agglutinin (WFA) to identify compounds that altered muscle cell surface glycosylation, with the goal of finding compounds that increase abundance of α-DG and associated sarcolemmal glycoproteins, increase utrophin usage, and increase laminin binding. We identified one compound, lobeline, from the Prestwick library of Food and Drug Administration-approved compounds that fulfilled these criteria, increasing WFA binding to C2C12 cells and to primary muscle cells from wild type and mdx mice. WFA binding and enhancement by lobeline required complex N-glycans but not O-mannose glycans that bind laminin. However, inhibiting complex N-glycan processing reduced laminin binding to muscle cell glycoproteins, although O-mannosylation was intact. Glycan analysis demonstrated a general increase in N-glycans on lobeline-treated cells rather than specific alterations in cell surface glycosylation, consistent with increased abundance of multiple sarcolemmal glycoproteins. This demonstrates the feasibility of high throughput screening with plant lectins to identify compounds that alter muscle cell glycosylation and identifies a novel role for N-glycans in regulating muscle cell function.

Current therapies for attention deficit hyperactivity disorder (ADHD) have varying efficacy in individuals with fetal alcohol spectrum disorders (FASD), suggesting that alternative therapeutics are needed. Developmental exposure to ethanol produces changes in dopamine (DA) systems, and DA has also been implicated in ADHD pathology. In the current study, lobeline, which interacts with proteins in dopaminergic presynaptic terminals, was evaluated for its ability to attenuate neonatal ethanol-induced locomotor hyperactivity and alterations in dopamine transporter (DAT) function in striatum and prefrontal cortex (PFC). From postnatal days (PND) 1–7, male and female rat pups were intubated twice daily with either 3 g/kg ethanol or milk, or were not intubated (non-intubated control) as a model for “third trimester” ethanol exposure. On PND 21 and 22, pups received acute lobeline (0, 0.3, 1, or 3 mg/kg), and locomotor activity was assessed. On PND 23–25, pups again received an acute injection of lobeline (1 or 3 mg/kg), and DAT kinetic parameters ( Km and V max ) were determined. Results demonstrated that neonatal ethanol produced locomotor hyperactivity on PND 21 that was reversed by lobeline (1 and 3 mg/kg). Although striatal DAT function was not altered by neonatal ethanol or acute lobeline, neonatal ethanol exposure increased the V max for DAT in the PFC, suggesting an increase in DAT function in PFC. Lobeline ameliorated this effect on PFC V max at the same doses that decreased hyperactivity. Methylphenidate, the gold standard therapeutic for ADHD, was also evaluated for comparison with lobeline. Methylphenidate decreased DAT V max and Km in PFC from ethanol-treated pups. Thus, lobeline and methylphenidate differentially altered DAT function following neonatal ethanol exposure. Collectively, these findings provide support that lobeline may be a useful pharmacotherapy for some of the deficits associated with neonatal ethanol exposure.

Previous research suggests that the vesicular monoamine transporter-2 (VMAT2) is a novel target for the treatment of methamphetamine (METH) abuse.The effects GZ-793A, a novel, selective, and potent lobelane analog, on the rewarding effects of METH, cocaine, and palatable food in rats were determined.GZ-793A (3-30 mg/kg, s.c.) was administered 20 min prior to each session in which the groups of rats pressed a lever for infusions of METH (0.03 mg/kg/infusion), cocaine (0.3 mg/kg/infusion), or food pellets. Tolerance to repeated GZ-793A (15 mg/kg, s.c. for 7 days) on METH self-administration and food-maintained responding was determined. The ability of increasing doses of METH (0.001-0.56 mg/kg, i.v.) to surmount inhibition produced by GZ-793A (15 mg/kg, s.c.) was determined. Self-administration of GZ-793A (0.01-0.3 mg/kg/infusion, i.v.) was tested as a substitute for METH infusion. GZ-793A (15 mg/kg, s.c.) was administered 20 min prior to METH or saline conditioning in a place preference test.GZ-793A specifically decreased METH self-administration, without the development of tolerance. Increasing the unit dose of METH did not surmount the inhibition produced by GZ-793A on METH self-administration. GZ-793A did not serve as a substitute for self-administered METH. GZ-793A blocked METH-induced conditioned place preference (CPP) and did not induce CPP alone.These results indicate that VMAT2 is a viable target for pharmacological inhibition of METH reward and that GZ-793A represents a new lead in the discovery of a treatment for METH abuse.

Lobeline, a nicotinic receptor antagonist and neurotransmitter transporter inhibitor, is a candidate pharmacotherapy for methamphetamine abuse. meso-Transdiene (MTD), a lobeline analog, lacks nicotinic receptor affinity, retains affinity for vesicular monoamine transporter 2 (VMAT2), and, surprisingly, has enhanced affinity for dopamine (DA) and serotonin transporters [DA transporter (DAT) and serotonin transporter (SERT), respectively]. In the current study, MTD was evaluated for its ability to decrease methamphetamine self-administration in rats relative to food-maintained responding. MTD specifically decreased methamphetamine self-administration, extending our previous work. Classical structure-activity relationships revealed that more conformationally restricted MTD analogs enhanced VMAT2 selectivity and drug likeness, whereas affinity at the dihydrotetrabenazine binding and DA uptake sites on VMAT2 was not altered. Generally, MTD analogs exhibited 50- to 1000-fold lower affinity for DAT and were equipotent or had 10-fold higher affinity for SERT, compared with MTD. Representative analogs from the series potently and competitively inhibited [(3)H]DA uptake at VMAT2. (3Z,5Z)-3,5-bis(2,4-dichlorobenzylidene)-1-methylpiperidine (UKMH-106), the 3Z,5Z-2,4-dichlorophenyl MTD analog, had improved selectivity for VMAT2 over DAT and importantly inhibited methamphetamine-evoked DA release from striatal slices. In contrast, (3Z,5E)-3,5-bis(2,4-dichlorobenzylidene)-1-methylpiperidine (UKMH-105), the 3Z,5E-geometrical isomer, inhibited DA uptake at VMAT2, but did not inhibit methamphetamine-evoked DA release. Taken together, these results suggest that these geometrical isomers interact at alternate sites on VMAT2, which are associated with distinct pharmacophores. Thus, structural modification of the MTD molecule resulted in analogs exhibiting improved drug likeness and improved selectivity for VMAT2, as well as the ability to decrease methamphetamine-evoked DA release, supporting the further evaluation of these analogs as treatments for methamphetamine abuse.

Using a physiological model of acutely increasing venous return into the lungs, i.e. by applying and then releasing lower body negative pressure (LBNP) to mimic the natural stimulus of juxtapulmonary capillary (J) or pulmonary C fibre receptors, produced an immediate and significant reduction in the amplitude of the Hoffman (H) reflex by 81±4% (P=0.001) in a majority of subjects 70% (n=5). Accompanying this was a notable change in the respiratory pattern with tidal volume (V(T)) increasing in all subjects from (mean) 0.462±.038 to 0.777±.061l/min (P=0.001) and the respiratory rate (F(R)) in 40% from 14±1 to 24±0.8 breaths/min. A feeling of pressure in throat, upper chest was reported by all and a shortness of breath-by 70% of the subjects. These were similar in nature to the respiratory sensations felt with threshold doses of intravenous lobeline, a well-established chemical stimulant of J receptors. All effects lasted for 15-20s and within a minute the parameters resumed their earlier control values. In animals, respiratory augmentation and locomotion inhibition are well-established reflexes of J receptors - this simultaneous though transitory reduction in H reflex amplitude reflecting change in the excitability of the motoneurone pool and appearance of respiratory effects, is the first demonstration in human subjects of the two reflexes appearing in response to a sudden increase in pulmonary blood flow that mimics the natural stimulus of these receptors.

Both lobeline and lobelane attenuate methamphetamine self-administration in rats by decreasing methamphetamine-induced dopamine release via interaction with vesicular monoamine transporter-2 (VMAT2). A novel derivative of nor-lobelane, cis-2,5-di-(2-phenethyl)-pyrrolidine hydrochloride (UKCP-110), and its trans-isomers, (2R,5R)-trans-di-(2-phenethyl)-pyrrolidine hydrochloride (UKCP-111) and (2S,5S)-trans-di-(2-phenethyl)-pyrrolidine hydrochloride (UKCP-112), were evaluated for inhibition of [3H]dihydrotetrabenazine binding and [3H]dopamine uptake by using a rat synaptic vesicle preparation to assess VMAT2 interaction. Compounds were evaluated for inhibition of [3H]nicotine and [3H]methyllycaconitine binding to assess interaction with the major nicotinic receptor subtypes. In addition, compounds were evaluated for inhibition of methamphetamine-evoked endogenous dopamine release by using striatal slices. The most promising compound, UKCP-110, was evaluated for its ability to decrease methamphetamine self-administration and methamphetamine discriminative stimulus cues and for its effect on food-maintained operant responding. UKCP-110, UKCP-111, and UKCP-112 inhibited [3H]dihydrotetrabenazine binding (Ki = 2.66 ± 0.37, 1.05 ± 0.10, and 3.80 ± 0.31 μM, respectively) and had high potency inhibiting [3H]dopamine uptake (Ki = 0.028 ± 0.001, 0.046 ± 0.008, 0.043 ± 0.004 μM, respectively), but lacked affinity at nicotinic receptors. Although the trans-isomers did not alter methamphetamine-evoked dopamine release, UKCP-110 inhibited (IC50 = 1.8 ± 0.2 μM; Imax = 67.18 ± 6.11 μM) methamphetamine-evoked dopamine release. At high concentrations, UKCP-110 also increased extracellular dihydroxyphenylacetic acid. It is noteworthy that UKCP-110 decreased the number of methamphetamine self-infusions, while having no effect on food-reinforced behavior or the methamphetamine stimulus cue. Thus, UKCP-110 represents a new lead in the development of novel pharmacotherapies for the treatment of methamphetamine abuse.

Vesicular monoamine transporter-2 (VMAT2) is a viable target for development of pharmacotherapies for psychostimulant abuse. Lobeline (1) is a potent antagonist at alpha4beta2 * nicotinic acetylcholine receptors, has moderate affinity (K(i)=5.46microM) for VMAT2, and is being investigated currently as a clinical candidate for treatment of psychostimulant abuse. A series of carboxylic acid and sulfonic acid ester analogs 2-20 of lobeline were synthesized and evaluated for interaction with alpha4beta2 * and alpha7 * neuronal nicotinic acetylcholine receptors (nAChRs), the dopamine transporter (DAT), serotonin transporter (SERT) and VMAT2. Both carboxylic acid and sulfonic acid esters had low affinity at alpha7 * nAChRs. Similar to lobeline (K(i)=4nM), sulfonic acid esters had high affinity at alpha4beta2 * (K(i)=5-17nM). Aromatic carboxylic acid ester analogs of lobeline (2-4) were 100-1000-fold less potent than lobeline at alpha4beta2 * nAChRs, whereas aliphatic carboxylic acid ester analogs were 10-100-fold less potent than lobeline at alpha4beta2 *. Two representative lobeline esters, the 10-O-benzoate (2) and the 10-O-benzenesulfonate (10) were evaluated in the (36)Rb(+) efflux assay using rat thalamic synaptosomes, and were shown to be antagonists with IC(50) values of 0.85microM and 1.60microM, respectively. Both carboxylic and sulfonic acid esters exhibited a range of potencies (equipotent to 13-45-fold greater potency compared to lobeline) for inhibiting DAT and SERT, respectively, and like lobeline, had moderate affinity (K(i)=1.98-10.8microM) for VMAT2. One of the more interesting analogs, p-methoxybenzoic acid ester 4, had low affinity at alpha4beta2 * nAChRs (K(i)=19.3microM) and was equipotent with lobeline, at VMAT2 (K(i)=2.98microM), exhibiting a 6.5-fold selectivity for VMAT2 over alpha4beta2 nAChRs. Thus, esterification of the lobeline molecule may be a useful structural modification for the development of lobeline analogs with improved selectivity at VMAT2.

Behavioural studies have yielded results that show lobeline has the ability to attenuate d-methamphetamine self-administration. Further in vivo and in vitro studies have demonstrated a blockade of μ-opioid receptors with lobeline. The present investigation examined the ability of lobeline to attenuate heroin intravenous (i.v.) self-administration when administered prior to testing. Male Sprague—Dawley rats were surgically implanted with jugular catheters and trained to lever press for i.v. heroin infusions (18 μg/kg) under a fixed ratio-2 schedule wherein two active lever presses resulted in heroin delivery. Rats then were tested for heroin self-administration after pretreatment with subcutaneous lobeline injections (0.3, 1.0, or 3.0 mg/kg, 15 min prior to testing sessions). At doses of 1.0 and 3.0 mg/kg, lobeline attenuated self-administration of heroin. The results suggest a potential for lobeline to be used in pharmacotherapy for opioid abuse.

Neuronal nicotinic acetylcholine receptors (nAChRs) are implicated in the reinforcing effects of many drugs of abuse, including ethanol. The present study examined the efficacy of cytisine, a nAChR partial agonist, and lobeline, a putative nAChR antagonist, on the maintenance of ethanol drinking by HAD-2 rats. Adult male HAD-2 rats were given access to ethanol (15 and 30%, with ad libitum access to water and food) 22 h/day for 12 weeks, beginning at 60 days of age, after which cytisine (0.0, 0.5, and 1.5 mg/kg) was tested for 3 consecutive days. The rats were given an 18-day washout period and were then tested with lobeline (0.0, 1.0, and 5.0 mg/kg) for 3 consecutive days. Ethanol intake was measured at 1, 4, and 22 h postinjection. Rats were injected intraperitoneally just before lights out (1200 h). There was a significant main effect of cytisine treatment on the second test day, with the 1.5 mg/kg dose significantly reducing ethanol intake at the 1- and 4-h time-points, relative to saline, and the 0.5 mg/kg dose inducing a significant reduction at the 4-h time-point. Conversely, lobeline treatment resulted in significant main effects of treatment for all three time-points within each test day, with the 5.0 mg/kg dose significantly reducing ethanol intake, relative to saline, at each time-point within each test day. These findings provide further evidence that activity at the nAChR influences ethanol intake and is a promising target for pharmacotherapy development for the treatment of alcohol dependence and relapse.

Respiratory sensations of eight patients with mitral stenosis in response to i.v. lobeline and 6-min walk before percutaneous mitral valvulotomy (PMV) were 'being short of breath', pressure in chest, tracheo-bronchial irritation, a desire to cough, persistent dry cough, chest pain and were qualitatively similar amongst 75% (P=0.005) of the patients. A week after PMV lobeline evoked similar sensations but the threshold dose decreased from 32.4+/-3.8 to 24.1+/-3.2 microg/kg (P=0.001) and pulmonary artery wedge pressure (PAwP), signifying reduction in pulmonary congestion, from 23.1+/-1.4 to 14.3+/-3.4 mm Hg (P0.001). Distance walked in 6 min increased from 217+/-58 to 319+/-51.6m; and mitral valve area from 0.63+/-0.01 to 1.43+/-0.26 cm(2) (P0.001). A fall in lobeline-sensation threshold dose indicated reduction in pulmonary congestion and stimulus to juxtapulmonary/J (or pulmonary C fibre) receptors which suggests that they had contributed to the respiratory and viscerosomatic symptoms seen before PMV.

Lobeline is currently being evaluated in clinical trials as a methamphetamine abuse treatment. Lobeline interacts with nicotinic receptor subtypes, dopamine transporters (DATs), and vesicular monoamine transporters (VMAT2s). Methamphetamine inhibits VMAT2 and promotes dopamine (DA) release from synaptic vesicles, resulting ultimately in increased extracellular DA. The present study generated structure-activity relationships by defunctionalizing the lobeline molecule and determining effects on [3H]dihydrotetrabenazine binding, inhibition of [3H]DA uptake into striatal synaptic vesicles and synaptosomes, the mechanism of VMAT2 inhibition, and inhibition of methamphetamine-evoked DA release. Compared with lobeline, the analogs exhibited greater potency inhibiting DA transporter (DAT) function. Saturated analogs, lobelane and nor-lobelane, exhibited high potency (Ki = 45 nM) inhibiting vesicular [3H]DA uptake, and lobelane competitively inhibited VMAT2 function. Lobeline and lobelane exhibited 67- and 35-fold greater potency, respectively, in inhibiting VMAT2 function compared to DAT function. Lobelane potently decreased (IC50 = 0.65 μM; Imax = 73%) methamphetamine-evoked DA overflow, and with a greater maximal effect compared with lobeline (IC50 = 0.42 μM, Imax = 56.1%). These results provide support for VMAT2 as a target for inhibition of methamphetamine effects. Both trans-isomers and demethylated analogs of lobelane had reduced or unaltered potency inhibiting VMAT2 function and lower maximal inhibition of methamphetamine-evoked DA release compared with lobelane. Thus, defunctionalization, cis-stereochemistry of the side chains, and presence of the piperidino N-methyl are structural features that afford greatest inhibition of methamphetamine-evoked DA release and enhancement of selectivity for VMAT2. The current results reveal that lobelane, a selective VMAT2 inhibitor, inhibits methamphetamine-evoked DA release and is a promising lead for the development of a pharmacotherapeutic for methamphetamine abuse.

Lobeline is a partial nicotinic agonist and is currently being investigated as a therapeutic drug for several addictive disorders particularly for smoking cessation. The present study evaluated the effects of repeated (continuous and recurring) administration of lobeline on alcohol consumption (10% alcohol vs. water) and alcohol preference using a 2-bottle choice test procedure. Male C57BL/6J mice were individually housed and acclimatized to 10% alcohol. Immediately following the last day of alcohol acclimatization and attainment of consistent drinking pattern, mice (n = 5/group) received subcutaneous injections of lobeline (3, 5, or 10 mg/kg) or saline. Groups received either repeated–recurring (3 injections, given every other day) or repeated–continuous (daily injections for 5 days) subcutaneous injections of lobeline. Fluid consumption (alcohol and water) was recorded daily. Results showed that lobeline significantly reduced alcohol consumption and alcohol preference during the repeated (recurring and continuous) administration phases, while total fluid consumption remained unchanged. These results provide support that nicotinic receptor based drugs may be useful as potential treatments for alcoholism.

The alkaloid lobeline inhibits the function of vesicular monoamine and dopamine transporters and diminishes the behavioral and neurochemical effects of nicotine and amphetamines. In the present study, we examined the interaction of systemic administration of lobeline on breakpoint scores on a progressive ratio (PR) schedule of intracranial self-stimulation (ICSS) of the medial forebrain bundle (MFB). Rats were run in two 30 min sessions, separated by a 10 min timeout period. At the end of the first session, each rat was injected with either 0, 0.5, 1.0 or 2.0 mg/kg (i.p.) lobeline. Positive controls known to suppress and to augment ICSS responding included the adrenergic antagonist prazosin (0, 0.5 and 2.0 mg/kg, i.p.) and the psychostimulant cocaine (0, 1.25, and 5.0 mg/kg, i.p.). Analyses of changes in average PR breakpoint scores between the 2 sessions revealed that lobeline significantly suppressed PR scores at doses of 0.5, 1.0 and 2.0 mg/kg, as did 0.5 mg/kg and 2.0 mg/kg prazosin. These changes are unlikely to reflect motoric effects of these drugs inasmuch as neither lobeline nor prazosin alter locomotion at these doses. In contrast, PR breakpoint scores were significantly increased at 5.0 mg/kg cocaine, a dose that is sufficient to elevate locomotion in the rat. These results are consistent with the view that lobeline modulates brain reinforcement processes.

Lobelane, a minor alkaloid of Lobelia inflata and a synthetic, des-oxy analog of lobeline, has good affinity for the vesicular monoamine transporter and the dopamine transporter. The current study examined the ability of lobelane to specifically decrease methamphetamine self-administration. Rats were trained on a fixed ratio 5 schedule of reinforcement to self-administer methamphetamine (0.05 mg/kg/infusion, i.v.) or to respond for sucrose pellets. Upon reaching stable responding, rats were pretreated with lobelane (0.1, 1, 3, 5.6, or 10 mg/kg, s.c.) or saline, 15 min prior to the operant session. To assess the effect of repeated lobelane on methamphetamine self-administration, rats were pretreated with lobelane (5.6 or 10 mg/kg, s.c.) for 7 sessions. Behavioral specificity was further investigated by assessing the effects of lobelane (0.1, 1, 3, 5, or 10 mg/kg, s.c.) or saline on locomotor activity. Within the dose range tested, lobelane dose-dependently decreased methamphetamine self-administration, while having no effect on sucrose-maintained responding. Locomotor activity was decreased following only the highest dose of lobelane (10 mg/kg). Across repeated pretreatments, tolerance developed to the effect of lobelane on methamphetamine self-administration, demonstrating that the ability of lobelane to specifically decrease methamphetamine self-administration is a transient effect. Thus, taken together, the results show that although lobelane interacts with the pharmacological targets believed to be responsible for its ability to decrease methamphetamine self-administration, removal of the oxygen functionalities from the lobeline molecule may have afforded a compound with an altered pharmacokinetic and/or pharmacodynamic profile.

Acute lung injury (ALI) is a severe, life-threatening medical condition whose pathogenesis is linked to neutrophil infiltration of the lung. Activation and recruitment of neutrophils to the lung is mostly attributed to the production of chemokines NO, IL-6, for instance. This study aims to investigate lobeline ability in reducing NO production, and nitric oxide synthase (iNOs) expression. Lobeline was tested by inhibiting phosphorylation of mitogen-activated protein kinases (MAPKs), NF-κB and IκBα in LPS-stimulated RAW 264.7 cells. When RAW 264.7 macrophages were given lobeline with LPS, a significant concentration-dependent inhibition of NO production was detected. In vivo tests, mice were either treated with normal saline, 10mg/kg dexmethasone or 5, 10, 20mg/kg lobeline intraperitoneally, and after an hour, the administration of 5mg/kg of LPS was given intratracheally. External performance, cytokines, MAPK pathways and antioxidative enzymes (AOEs) were also carried out to evaluate the effects of these drugs. This is the first investigation in which lobeline was found to effectively inhibit acute lung edema, which may provide a potential target for treating ALI. Lobeline may utilize MAPKs pathways as well as AOEs activity to attenuate LPS-induced nonspecific pulmonary inflammation.

Lobeline has high affinity for nicotinic receptors and alters presynaptic dopamine storage and release in brain. Moreover, lobeline decreases the reinforcing and locomotor-activating properties of methamphetamine, suggesting that lobeline may be a pharmacotherapy for psychostimulant abuse. This study determined if lobeline alters cocaine-induced hyperactivity and if lobeline alters the induction and/or expression of sensitization to cocaine. On Days 1-12, male rats were administered lobeline (0.3 or 1.0 mg/kg) or saline, placed in an automated activity monitor for 20 min, administered cocaine (10, 20 or 30 mg/kg) or saline and returned to the monitor for 60 min. On Day 13, the effect of lobeline on the induction and expression of sensitization to cocaine was determined. Lobeline did not alter the effect of cocaine after acute injection. However, 1.0 mg/kg lobeline attenuated cocaine (10 and 20 mg/kg)-induced hyperactivity after repeated administration and prevented the development of sensitization to these cocaine doses. Interestingly, 0.3 mg/kg lobeline augmented cocaine (10 mg/kg)-induced hyperactivity after repeated administration. Lobeline did not alter the effect of 30 mg/kg cocaine. The present results indicate a complex interaction of lobeline with cocaine and support other research indicating a role for nicotinic receptors in the development of sensitization to psychostimulants.

There is evidence that the cholinergic nicotinic system is involved in the modulation of anxiety. Anxiolytic and anxiogenic effects of nicotine agonists have been reported in mice. Bupropion is an antidepressant drug which may alleviate some symptoms of nicotine withdrawal, although its effects on anxiety are not clear. It has been suggested that the interaction between bupropion and nicotinic mechanisms could be complex. The aim of the present study was to investigate acute effects of co-administration of bupropion and nicotinic agonists on the elevated plus-maze test in NMRI mice. Effects of nicotine, lobeline, and cytisine (0.35 and 0.175 mg/kg), administered alone or combined with bupropion (20 mg/kg) were tested in the plus-maze. Results indicated that nicotine (0.35 mg/kg) decreased number and percentage of entries and time spent in open arms, and increased percentage of protected stretched attend posture. Bupropion (20 mg/kg) plus lobeline (0.175 mg/kg) increased percentage of time spent in open arms, without altering total or closed arm entries. Our findings suggest that the highest dose of nicotine induces anxiogenic effects, which are counteracted when co-administered with bupropion. The combination of bupropion with a low dose of lobeline seems to have an anxiolytic profile in the conventional parameters of the plus-maze, although ethological measures do not support it clearly.

Factors related to parental smoking and parenting practice have a big effect on adolescent smoking. More in-depth information about these relationships can be used to inform interventions. This study investigated Maori, Pacific Islander, New Zealand European and Asian parent attitudes and practices in relation to smoking uptake in children.Parents of children aged 8-15 years, recruited through the researchers' community networks and the media, participated in focus groups or interviews run by ethnically matched facilitators. Areas addressed included parents' beliefs about children smoking, their actions, and their suggestions for tobacco control activities. The data were analysed thematically.Parents believed that parental smoking, peer pressure and smoking role models in the media influenced smoking uptake in children. They said they would be disappointed if their children started smoking, but their confidence in influencing them varied. Many talked to their children about the health consequences of smoking, including their own negative perceptions of smoking and smokers. Most had smoke-free homes. The parents who smoked tried to avoid smoking around children. There was a good deal of commonality across the different ethnic groups. Important differences related to the provision of interventions.Parents believed in the value of a smoke-free lifestyle and wanted to protect their children from smoking. Important strategies to prevent smoking in children may include supporting parents to quit, informing them that discouraging children of any age from smoking can be effective, and providing culturally appropriate education and resources to facilitate parent/child communication about smoking.

In order to examine, whether the lobeline-induced cough is a true reflex or a voluntary effort to get rid of its irritating sensations in the upper respiratory tract, we systematically studied the cough response to lobeline, of subjects who were unable to make conscious discriminations i.e. were either comatose ( n = 4) or anaesthetized ( n = 5). 8 μg/kg lobeline injected into the right atrium of one and 29 μg/kg intravenously (i.v.) into another evenly and spontaneously breathing comatose subject produced a cough after 4 s and 12 s, respectively. Cough was repeatable and showed a dose response relationship i.e., its latency decreasing and its duration/intensity increasing with the dose. In a third subject, capable only of weak spontaneous respiration, a relatively high dose injected into the right atrium (44 μg/kg) generated a pronounced cough-like respiratory movement superimposed on the artificial ventilation and also during the apnoea after disconnecting the pump. No respiratory response was evoked in a fourth subject who had no evidence of brainstem reflexes. In five normals, cough was elicited with a mean dose of 35 ± 5 μg/kg i.v. (latency 14 ± 2 s; duration 10 ± 3 s). After thiopental anaesthesia, injecting 41 ± 7 μg/kg produced a cough within 13 ± 2 s that lasted for 12 ± 2 s. It may be noted that neither the later dose nor the latency or duration of cough that it produced were significantly different from the pre anaesthesia values ( P > 0.05). These two sets of results show unequivocally that the lobeline-induced cough is evoked reflexely; its magnitude in the conscious state could vary by subjective influences. We discuss the likelihood of its origin from juxtapulmonary capillary receptors.

Rationale. Previous results demonstrated that pretreatment with lobeline attenuates d-methamphetamine self-administration in rats. Objective. The present experiments determined if lobeline serves as a reinforcer, if it decreases d-methamphetamine-induced reinstatement of d-methamphetamine self-administration, and if it activates the mesolimbic and nigrostriatal dopamine (DA) pathways in Sprague-Dawley male rats. Methods. The ability of intravenous (IV) lobeline (0.015–0.15 mg/kg per infusion) to engender responding and the ability of lobeline (0.015 and 0.05 mg/kg per infusion) to substitute for d-methamphetamine was determined using the self-administration paradigm. Experiments were also performed to determine if lobeline (1.0 and 3.0 mg/kg) reinstates responding for d-methamphetamine or alters the ability of d-methamphetamine (1.0 mg/kg per infusion) to reinstate responding following extinction. The effect of lobeline (3.0 mg/kg) or d-methamphetamine (1.0 and 3.0 mg/kg) on DA and dihydroxyphenylacetic acid (DOPAC) levels in the nucleus accumbens and striatum were also determined. Results. Lobeline was not self-administered and did not substitute for d-methamphetamine. Also, lobeline did not reinstate responding for d-methamphetamine following extinction nor did it alter d-methamphetamine-induced reinstatement. Furthermore, lobeline did not alter DA or DOPAC levels in the either the nucleus accumbens or striatum. Conclusions. Taken together, the present results indicate that lobeline decreases d-methamphetamine self-administration by decreasing reward, not by acting as a substitute reinforcer.

The present study investigated the effects of two herbal components (BACCOFF™ and DIPSTOP™) of a commercially available smokeless tobacco treatment program for reducing subjective withdrawal symptoms during deprivation. One component, BACCOFF™, is a non-nicotinic chew. The second component, DIPSTOP™, is a liquid containing the alkaloid lobeline, which to some extent mimics peripheral nicotinic effects. All participants (N = 22 males) were placed in four conditions: BACCOFF™ + DIPSTOP™, BACCOFF™ + placebo control, DIPSTOP™ only, and placebo control only. The conditions involved 48 h of deprivation, and subjects were exposed to one condition per week for 4 weeks. Withdrawal measures were taken at baseline, 24 h, and 48 h of deprivation. Individuals were randomly assigned, and conditions were counterbalanced. Results showed that BACCOFF™, as compared with DIPSTOP™, significantly reduced withdrawal symptoms but not craving. These data suggest that behavioral/sensory substitutes’ influence on withdrawal might be routed through the product’s ability to approximate the preferred moist snuff.

We have reported that brain nicotinic acetylcholine receptor (nAChR) ligand lobeline has antidepressant-like effects in mice. The present study examined the effects of lobeline on chronic unpredictable stress (CUS)-induced depression-like behavior, deficits in brain-derived neurotrophic factor (BDNF) expression and cell proliferation in the hippocampus. Adult C57BL/6J mice were exposed to CUS for 6 weeks. Lobeline (1 or 4 mg/kg, s.c.) or saline was administered once daily during the last 14 days of CUS. CUS-exposed mice showed increased immobility time in the FST compared to control. Pretreatment with lobeline (1 mg/kg) significantly reduced immobility time in the CUS-exposed mice. Twenty-four hour following lobeline or saline treatment, BDNF expression or cell proliferation was measured in the hippocampus using Western blotting and bromodeoxyuridine immunohistochemistry, respectively. Lobeline (1 mg/kg) treatment prevented CUS-induced reduction in BDNF expression and cell proliferation in the hippocampus. Overall, our findings suggest that antidepressant-like effects of lobeline could involve nAChR mediated signaling, BDNF expression, and/or hippocampal cell proliferation.

1. The sensations evoked by the injection of lobeline into the right antecubital vein were studied in 8 subjects after bilateral lung transplantation and 10 control subjects. In control subjects, two distinct sensations were experienced. There was an early noxious sensation (onset approximately 10 s) followed by a late sensation of breathlessness (onset approximately 26 s) associated with involuntary hyperventilation. The early sensation was accompanied by respiratory and cardiovascular changes. 2. In contrast to control subjects, the early respiratory events and the noxious sensations evoked by injections of lobeline (18-60 microg kg(-1)) did not occur in subjects with recent bilateral lung transplantation. This suggests that the early respiratory sensations are mediated by the discharge of receptors in the lungs. 3. The late hyperventilation and the accompanying sensation of breathlessness occurred in both transplant and control subjects and are therefore likely to be mediated by receptors elsewhere in the body, presumably systemic arterial chemoreceptors stimulated by lobeline. 4. In control subjects, but not transplant subjects, there was a consistent decrease in mean arterial pressure associated with the lobeline injection. This suggests that pulmonary afferents mediate the hypotension. 5. For transplant subjects studied more than a year after transplantation, there was some evidence that the noxious respiratory sensations evoked by lobeline had returned. This suggests that some functional reinnervation of pulmonary afferents may occur.

We investigated the physiological role of native low- and high-affinity nicotinic acetylcholine receptors (nAChRs) in regulating dopamine (DA) release from striatal DA terminals. To evaluate the functional interactions of the two receptor subtypes, nicotine (which interacts with both high- and low-affinity nAChRs) and lobeline (which selectively interacts with high-affinity nAChRs) were perfused through a microdialysis probe implanted into the striatum of freely moving rats. The DA content of successive dialysates was quantified by HPLC with an electrochemical detector. A short-lasting (1-min) perfusion of nicotine or lobeline dose-dependently increased the DA content of striatal dialysates. A second application of the same dose of nicotine resulted in an attenuated DA increase, compared with the increase elicited by the first application; however, the DA increase elicited by a second application of lobeline was similar to that of the first lobeline application. The nicotine-induced response was not attenuated when it followed a lobeline perfusion; in contrast, if the nicotine perfusion preceded that of lobeline, the lobeline-induced response was attenuated. In the presence of mecamylamine (a noncompetitive nAChR antagonist), the increase in DA content of striatal dialysate samples induced by either nicotine or lobeline was attenuated. However, in the presence of methyllycaconitine (a preferential antagonist for low-affinity α 7 homomeric nAChRs), the nicotine response was attenuated but that of lobeline was unaffected. These results suggest that the functional inactivation of striatal nAChRs requires the simultaneous activation of both low- and high-affinity nAChRs. Since lobeline is devoid of reinforcing properties, one might infer that the reinforcing properties of nicotine require the simultaneous activation of high- and low-affinity brain nAChRs.

The nicotinic compounds (-)-cytisine, (-)-lobeline, (+/-)-epibatidine, (S)-3methyl-5-(1-methyl-2-pyrrolidinyl)isoaxzole (ABT-418), (-)-nicotine, and cocaine were compared in an acute self-administration model using drug-naive mice that could self-administer intravenous infusions contingent on nose poking (fixed ratio 1 with no time out). Although the nose pokes of yoked control mice were unaffected by unit dose, inverted U-shaped unit dose response curves were seen with cocaine (up to 0.26 mg/kg/infusion), nicotine (up to 0.175 mg/kg/infusion), cytisine (up to 0.125 mg/kg/infusion), and lobeline (up to 1.25 mg/kg/infusion) in mice receiving infusions contingent upon nose poke responses. Epibatidine (up to 1.25 microg/kg/infusion) and ABT-418 (up to 0.125 mg/kg/infusion) failed to exhibit inverted U-shaped unit dose response curves. The present studies demonstrate that cytisine and lobeline, but not ABT-418 or epibatidine, were self-administered by drug-naive mice in a manner similar to cocaine and nicotine. These findings are discussed in terms of potency and selectivity at the alpha4beta2 nicotinic acetylcholine receptor subunit combination.

Background In Eisenmenger syndrome (ES), oral phosphodiesterase type-5 inhibitors, which are preferential pulmonary vasodilators, reduce the elevated pulmonary artery pressure and pulmonary vascular resistance index by increasing cyclic guanosine monophosphate (cGMP). However, no information is available as to how pulmonary vasodilatation alleviates the accompanying dyspnoea and improves patient's exercising ability. Objectives As the natural stimulus of juxtapulmonary capillary (J) receptors is an increase in interstitial pressure, the aim was to estimate their threshold level stimulation chemically by intravenous lobeline, before and after 6weeks of sildenafil therapy in treatment-naive ES patients. Methods Nine Eisenmenger syndrome patients [mean age=26 (SD=1.6) years] underwent 6MWT and an exercise test before and 6weeks after oral sildenafil (20mg 3× D). Their respiratory responses to threshold doses of intravenous lobeline were determined at both these stages. Results After 6weeks of sildenafil therapy, the 6MWD [from 453.3 (SD=50.9) m to 516.6 (SD=48.9) m; P =0.001] and the duration of exercise with the modified Bruce protocol from 7min 53s (SD=0.04) to 10min 44s (SD=0.88) ( P =0.001) improved significantly. However, the improvement in oxygen saturation was not noteworthy. The lobeline dose required to produce threshold level of respiratory effects was higher in ES patients [37.5 (SD=3.4) μg/kg] and with sildenafil therapy it fell significantly [20.6 (SD=1.8) μg/kg; P =0.001]. Conclusions J receptor threshold doses were elevated in ES patients and fell significantly with sildenafil therapy that was associated with improved exercise tolerance, implying thereby a role of J receptors in producing dyspnea in ES patients.

In normal subjects, if an acoustic startle stimulus is immediately preceded by a small brief change in background noise intensity, the magnitude of the subsequent startle response is decreased. This prepulse inhibition (PPI) of an acoustic startle response has been shown to be associated with sensorimotor gating. PPI is disrupted in schizophrenic patients and has been linked to attentional disorders characteristic of this disease. We tested the effects of (−)-nicotine, (0.19, 0.62, and 1.9 μmol/kg IP) (equivalent to 0.03, 0.1, and 0.3 mg/kg base) and the nicotinic cholinergic receptor (nAChR) channel blocker, mecamylamine (5.0 and 50 μmol/kg IP) (equivalent to 1.0 and 10.0 mg/kg) on PPI of the acoustic startle response in the rat. Nicotine increased the PPI at the lowest prepulse signal levels but not at the stronger levels. Mecamylamine was without effect at 5.0 μmol/kg, but the 50μmol/kg dose decreased the inhibition at both weak and strong prepulse (PP) levels. Mecamylamine (5.0 μmol/kg) pretreatment did not block the (−)-nicotine-induced increase in PPI. Lobeline (0.19, 0.62, 1.9, and 6.2 μmol/kg IP) (equivalent to 0.071, 0.23, 0.71, and 2.3 mg/kg) was without effect. These results are consistent with a mecamylamine-insensitive effect of nicotine to improve gating in normal rats. The nAChR subtype involved in producing nicotine's increase of PPI needs further investigation.

Rats were trained to discriminate 1.9 μmol/kg (−)-nicotine (0.3 mg/kg) from saline in a standard two-bar operant conditioning paradigm with food reinforcement. The effect of neuronal nicotinic acetylcholine receptor (nAChR) agonists and antagonists were verified, and the participation of dopaminergic receptors subtypes in the expression of the (−)-nicotine cue was investigated with cis-flupentixol (D1-D2 antagonist), haloperidol (D2 antagonist) and clozapine (D4 antagonist). The stereoselectivity of the behavioral response was indicated by the 10-fold less sensitivity to (+)-nicotine in (−)-nicotine-trained rats. (±)-Anabasine and (−)-cytisine exhibited partial agonist profiles at the 1.9 μmol/kg dose while (−)-lobeline was devoid of any effect in doses up to 19 μmol/kg. (−)-Lobeline did not show antagonist properties in this paradigm. The nicotinic channel blockers mecamylamine, chlorisondamine and hexamethonium were inactive on their own but mecamylamine and chlorisondamine were able to block the effect of (−)-nicotine. Clozapine attenuated the (−)-nicotine cue while cis-flupentixol and haloperidol were ineffective. Similar doses of cis-flupentixol significantly blocked the locomotr stimulant effect of (−)-nicotine in rats indicating that blockade of dopaminergic receptors was achieved at the doses used in the drug discrimination studies. These data suggest that the discriminative stimulus properties of (−)-nicotine are mediated through neuronal nAChRs and involves the activation of dopaminergic receptors of the D4 subtype.

Juxta-pulmonary capillary (J or pulmonary C fiber) receptors are stimulated by an increase in pulmonary blood flow and give rise to respiratory acceleration and related sensations and inhibit exercise. However, the reverse, i.e., the effect of reducing pulmonary blood flow on their reflexes, is as yet not known. This was investigated by carrying out a placebo-controlled study on the acute effects of a single dose (0.4 mg) of sublingual glyceryl nitrate (GTN), known to shift blood from the central to the peripheral circulation, on the respiratory parameters of exercising healthy subjects and on their responses to i.v. lobeline. In 10 subjects, GTN use delayed the first appearance of respiratory sensations from 9.08 ± 0.9 min to 11 min ( P = 0.002), reduced the increase in minute ventilation by the end of 10 min of exercise ( P = 0.003) and increased its duration by 1–4 s and doubled it in the remaining one subject. In a majority of 8 of them, the effect of GTN on i.v. lobeline-induced respiratory reflexes and sensations was a significant increase in the dose required ( P = 0.006) for producing threshold effects and in the latency of their appearance ( P = 0.003). The latter feature points to a reduction in blood flow in the lung parenchyma where these receptors are located and to which they are sensitive. As this would have led to a reduced stimulation of these receptors, it would account for the delayed appearance of respiratory symptoms, a reduction in ventilatory increase and prolongation of exercise duration. We demonstrated a mechanism of reducing the stimulus level of J receptors by reducing pulmonary blood flow by means of pharmacological sequestration with GTN use, which then led to a reduction in the magnitude of respiratory and viscerosomatic reflexes, while noting at the same time that changes in blood flow in the pulmonary bed do not directly influence limb muscles, tendons and joints which also determine exercise output.

(R)-3-[2,6-cis-Di(4-methoxyphenethyl)piperidin-1-yl]propane-1,2-diol (GZ-793A) inhibits methamphetamine-evoked dopamine release from striatal slices and methamphetamine self-administration in rats. GZ-793A potently and selectively inhibits dopamine uptake at the vesicular monoamine transporter-2 (VMAT2). This study determined GZ-793A's ability to evoke [³H]dopamine release and inhibit methamphetamine-evoked [³H]dopamine release from isolated striatal synaptic vesicles. Results show GZ-793A concentration-dependent [³H]dopamine release; nonlinear regression revealed a two-site model of interaction with VMAT2 (High- and Low-EC₅₀ = 15.5 nM and 29.3 μM, respectively). Tetrabenazine and reserpine completely inhibited GZ-793A-evoked [³H]dopamine release, however, only at the High-affinity site. Low concentrations of GZ-793A that interact with the extravesicular dopamine uptake site and the High-affinity intravesicular DA release site also inhibited methamphetamine-evoked [³H]dopamine release from synaptic vesicles. A rightward shift in the methamphetamine concentration-response was evident with increasing concentrations of GZ-793A, and the Schild regression slope was 0.49 ± 0.08, consistent with surmountable allosteric inhibition. These results support a hypothetical model of GZ-793A interaction at more than one site on the VMAT2 protein, which explains its potent inhibition of dopamine uptake, dopamine release via a High-affinity tetrabenazine- and reserpine-sensitive site, dopamine release via a Low-affinity tetrabenazine- and reserpine-insensitive site, and a low-affinity interaction with the dihydrotetrabenazine binding site on VMAT2. GZ-793A inhibition of the effects of methamphetamine supports its potential as a therapeutic agent for the treatment of methamphetamine abuse.

Exposure to early life stress has been suggested to increase an individual’s vulnerability to methamphetamine (MA) dependence. Although there is no cure for drug dependence, the opioid and vesicular monoamine transporter 2 (VMAT2) systems may be useful targets for treatment insofar as they play pivotal roles in the neurochemistry of addiction. Here we investigated the effects of naltrexone (opioid antagonist) and lobeline (VMAT2 inhibitor) on MA-induced place preference in adolescent rodents subjected to early life trauma (maternal separation, MS) and controls, as well as the effects on dopamine and serotonin levels in the striatum. We found: (1) maternal separation attenuated methamphetamine-induced place preference; (2) lobeline and naltrexone treatment had differential effects on serotonin and dopamine concentrations in the striatum, naltrexone increased serotonin levels in the maternally separated animals. The hypothesized effect of early adversity increasing MA-induced place preference may not be apparent in adolescence. However the data are consistent with the hypothesis that early life stress influences neurochemical pathways that predispose an individual to drug dependence.

Lobelane, a chemically defunctionalized saturated analog of lobeline, has increased selectivity for the vesicular monoamine transporter 2 (VMAT2) compared with the parent compound. Lobelane inhibits methamphetamine-evoked dopamine (DA) release and decreases methamphetamine self-administration. Unfortunately, tolerance develops to the ability of lobelane to decrease these behavioral effects of methamphetamine. Lobelane has low water solubility, which is problematic for drug development. The aim of the current study was to determine the pharmacological effect of replacement of the N-methyl moiety with a chiral N-1,2-dihydroxypropyl (N-1,2-diol) moiety, which enhances water solubility, altering the configuration of the N-1,2-diol moiety and incorporating phenyl ring substituents into the analogs. To determine VMAT2 selectivity, structure-activity relationships also were generated for inhibition of DA and serotonin transporters. Analogs with the highest potency for inhibiting DA uptake at VMAT2 and at least 10-fold selectivity were evaluated further for ability to inhibit methamphetamine-evoked DA release from superfused striatal slices. (R)-3-[2,6-cis-di(4-methoxyphenethyl)piperidin-1-yl]propane-1,2-diol (GZ-793A), the (R)-4-methoxyphenyl-N-1,2-diol analog, and (R)-3-[2,6-cis-di(1-naphthylethyl)piperidin-1-yl]propane-1,2-diol (GZ-794A), the (R)-1-naphthyl-N-1,2-diol analog, exhibited the highest potency (K(i) ∼30 nM) inhibiting VMAT2, and both analogs inhibited methamphetamine-evoked endogenous DA release (IC(50) = 10.6 and 0.4 μM, respectively). Thus, the pharmacophore for VMAT2 inhibition accommodates the N-1,2-diol moiety, which improves drug-likeness and enhances the potential for the development of these clinical candidates as treatments for methamphetamine abuse.