Additive Protective Effects of Donepezil and Nicotine Against Salsolinol-Induced Cytotoxicity in SH-SY5Y Cells

Although the etiology of Parkinson’s disease (PD) remains elusive, a number of toxins including elevated salsolinol, an endogenous metabolite of dopamine may contribute to its pathology. It was reported recently that nicotine may have protective effects against salsolinol-induced toxicity in human neuroblastoma derived SH-SY5Y cells and that these effects of nicotine are mediated by nicotinic receptors. Donepezil (Aricept) is a reversible non-competitive acetylcholinesterase inhibitor that is approved for use in mild to moderate Alzheimer’s disease. The increase in acetylcholine concentrations is believed to be the major contributory factor in donepezil’s therapeutic efficacy. However, cholinesterase inhibitors may also directly interact with nicotinic receptors and possess neuroprotective properties. In this study, we sought to determine whether donepezil may have protective effects against salsolinol-induced toxicity in SH-SY5Y cells and whether the combination of donepezil and nicotine may result in additive protection. Moreover, it was of interest to elucidate the role of nicotinic receptors as well as cell cycle and apoptosis in mechanism of action of these compounds. SH-SY5Y cells were exposed to 0.6 mM salsolinol with and without various drug pretreatments for 48 h. Nicotine (50 μM) resulted in approximately 54% protection and donepezil (5 μM) resulted in approximately 40% protection, and the combination of the two resulted in an additive (approximately 93%) protection against salsolinol-induced toxicity. Salsolinol caused an arrest of the cells in G₁-phase of cell cycle and an increase in apoptotic indices that were blocked by the combination of donepezil and nicotine. Mecamylamine, a non-selective nicotinic receptor antagonist completely blocked the effects of nicotine and partially attenuated the effects of donepezil. A combination of atropine, a muscarinic receptor antagonist and mecamylamine completely blocked the effects of donepezil, indicating involvement of both nicotinic and muscarinic receptors in donepezil’s actions. The findings suggest a therapeutic potential for the combination of donepezil and nicotine in PD.     

Neuroprotective effects of nicotine against salsolinol-induced cytotoxicity: Implications for parkinson’s disease

Parkinson's disease is associated with degeneration of dopaminergic cell bodies in the substantia nigra. It has been suggested that salsolinol, an endogenous metabolite of dopamine, may be involved in this process. An inverse relationship between Parkinson's disease and smoking (nicotine intake) has been observed in epidemiological studies. Moreover, neuroprotective effects of nicotine in various experimental models have been observed. In this study we sought to determine whether salsolinol-induced cytotoxicity in SH-SY5Y human neuroblastoma cells, a cloned cell line which expresses dopaminergic activity, could also be prevented by nicotine pretreatment, and if so, which nicotinic receptors may mediate the actions of nicotine. Exposure of SH-SY5Y cells to 0.8 mM salsolinol for 24 hours resulted in approximately 80% cell death as determined by 3,[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Pretreatment of cells with 0.1 mM nicotine resulted in inhibition of salsolinol-induced cytotoxicity. The effects of nicotine were blocked by mecamylamine, a non-selective nicotinic antagonist as well as conotoxins with selective antagonism against alpha3-containing nicotinic receptor subunits. The effects of nicotine were not affected by dihydro-beta-erythroidine or methyllycaconitine, selective antagonists against alpha4-beta2 or alpha7 nicotinic receptors, respectively. It is suggested that selective nicotinic agonists may be of therapeutic potential in at least a subpopulation of Parkinsonian patients.     

Salsolinol, a dopamine-derived tetrahydroisoquinoline, induces cell death by causing oxidative stress in dopaminergic SH-SY5Y cells, and the said effect is attenuated by metallothionein

The endogenous neurotoxin, 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (salsolinol), has been considered a potential neurotoxin in the etiology of Parkinson's disease (PD). Salsolinol and N-methyl(R)-salsolinol were identified in the brains and cerebrospinal fluid (CSF) of PD patients. Oxidative stress is known to be one of the major contributing factors in the cascade that may finally leads to the cell death in PD. The present study was undertaken to understand the role of salsolinol in oxidative-mediated neuronal toxicity in dopaminergic SH-SY5Y cells, and the neuroprotective effects of metallothionein (MT) against salsolinol toxicity in MT overexpressing (MT(trans)) fetal mesencephalic cells. Salsolinol increased the production of reactive oxygen species (ROS) and significantly decreased glutathione (GSH) levels and cell viability in SH-SY5Y cells. Salsolinol also decreased intracellular ATP levels and induced nuclear condensation in these cells. Salsolinol-induced depletion in cell viability was completely prevented by N-acetylcysteine in SH-SY5Y cells, and also prevented by MT in MT(trans) fetal mesencephalic cells compared to control(wt) cells. The extent of nuclear condensation and caspase activation was also less in MT(trans) cells than control(wt) cells. These results suggest that salsolinol causes oxidative stress by decreasing the levels of GSH and by increasing ROS production, and these events may lead to the death of dopaminergic cell. Furthermore, MT overexpression may protect dopaminergic neurons against salsolinol-induced neurotoxicity, most probably by the inhibition of oxidative stress and apoptotic pathways including caspase-3 activation.     

Toxic Effects of Low Alcohol and Nicotine Combinations in SH-SY5Y Cells are Apoptotically Mediated

It is well established that combination of heavy drinking and smoking has severe health consequences. However, at relatively low concentrations, both alcohol and nicotine may have beneficial effects including neuroprotection. Thus, protective effects of low alcohol concentration against beta-amyloid-induced toxicity in organotypic hippocampal slices and protective effects of nicotine against salsolinol-induced toxicity in human-derived neuroblastoma cells (SH-SY5Y) have been reported. In this study, we sought to determine whether alcohol might also be protective against salsolinol-induced toxicity in SH-SY5Y cells and whether the combination of low doses of alcohol and nicotine might have an additive or synergistic effect. Pre-exposure of SH-SY5Y cells to either ethanol (1 or 10 mM) or nicotine (20 or 50 μM) significantly attenuated salsolinol-induced toxicity. However, contrary to the expectation the combination of low doses of alcohol and nicotine not only did not provide any synergistic or additive protective effect, but exacerbated salsolinol-induced toxicity. Indeed, simple combination of low alcohol and nicotine resulted in significant toxicity in SH-SY5Y cells. This toxicity, reflected in a reduction in cell viability was associated with an increase in apoptosis as determined by caspase-3 measurement. These in vitro results suggest that combination of even low concentrations of alcohol and nicotine may activate apoptotic mechanisms that can lead to cell toxicity and detrimental consequences.     

Protective Effects of Curcumin Against Rotenone and Salsolinol-Induced Toxicity: Implications for Parkinson’s Disease

Parkinson’s disease (PD) is a debilitating neurodegenerative disorder that results from the loss of or damage to dopaminergic cells in the substantia nigra. Exposure to either the pesticide rotenone or the endogenous neurotoxin salsolinol has been shown to mimic this dopaminergic cell loss. In this study, we first sought to determine whether combination of rotenone and salsolinol would result in an additive or synergistic toxicity. For this purpose we utilized SH-SY5Y cells, a human neuroblastoma cell line that is commonly used to model dopaminergic neurodegeneration. We then tested whether curcumin, a natural plant compound with known health benefits including potential neuroprotective properties, could also protect against rotenone and/or salsolinol-induced toxicity. Moreover, since apoptotic mechanism has been implicated in toxicity of these compounds the anti-apoptotic effect of curcumin was also evaluated. Our results indicate a synergistic toxicity of low concentrations of rotenone (1 and 5 µM) and salsolinol (25 and 50 µM) that was associated with apoptosis as determined by cell flow cytometry. There was also an increase in caspase-3 levels. Pretreatment with curcumin (1–10 µM) dose-dependently attenuated rotenone and/or salsolinol-induced toxicity and the associated apoptosis. These results suggest that exposure to a combination of rotenone and salsolinol may contribute to the pathology of PD, and that curcumin has a therapeutic potential in this disease.     

Protective Effects of Nicotine Against Aminochrome-Induced Toxicity in Substantia Nigra Derived Cells: Implications for Parkinson’s Disease

Parkinson's disease is a debilitating progressive neurodegenerative disorder that results from the loss of or damage to dopaminergic cells containing neuromelanin in the substantia nigra (SN). The underlying neurodegenerative mechanism(s), however, remain elusive. Aminochrome, the precursor of neuromelanin is an endogenous substance capable of inducing selective neurotoxicity to dopaminergic neurons in SN. Nicotine, on the other hand, may offer protective effects against dopaminergic cell damage induced by various neurotoxins including MPTP and salsolinol. In this study, we sought to determine whether nicotine may also protect against aminochrome-induced toxicity in SN derived RCSN-3 cells. Exposure of RCSN-3 cells to a combination of aminochrome (50?μM) and dicoumarol (50?μM) for 48?h induced approximately 70?% cell death. Pretreatment with nicotine, dose-dependently blocked this toxicity. The effects of nicotine in turn were dose-dependently blocked by mecamylamine, a non-selective nicotinic receptor antagonist. These results suggest involvement of nicotinic receptors in protective effects of nicotine against aminochrome-induced toxicity and provide further evidence for possible therapeutic effects of nicotine or nicotinic agonists in Parkinson's disease.     

Existence and characterization of Salsolinol synthase in neuronal cells and rat brain

Salsolinol (1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline), the endogenous dopamine-derived catechol isoquinolines whose structure is similar with MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine), may be a possible candidate of dopaminergic neurotoxins to elicit Parkinson’s disease (PD). Catechol isoquinolines can selectively target dopaminergic neurons, leading to dopaminergic neuronal death. However, the formation and nosogenesis of these toxins remains unclear. Salsolinol synthase is a novel enzyme which condensate dopamine and acetaldehyde to salsolinol. It is the first key enzyme in the metabolic pathway of catechol isoquinolines which directly affects salsolinol and its derivative metabolism in vivo. It is also one kind of Pictet-Spenglerase, which has been little studied and need more characterization. PC12 cells and rat brains were performed to illustrate the existence of salsolinol synthase in our study. The results indicate that salsolinol synthase is a low molecular weight protein, showing enhanced activity with increase in dopamine concentration. It is suggested that salsolinol synthase is sensitive to strong acid and stable to high-temperature. In this research, existence of salsolinol synthase was confirmed in vivo, and also provided some new evidences to elucidate the endogenous catechol isoquinoline neurotoxin substances involved in the pathogenesis of PD.     

PACAP Protects Against Salsolinol-Induced Toxicity in Dopaminergic SH-SY5Y Cells: Implication for Parkinson’s Disease

Pituitary adenylate cyclase-activating polypeptide (PACAP) is an endogenous 38 amino acid containing neuropeptide with various cytoprotective functions including neuroprotection. Administration of PACAP has been shown to reduce damage induced by ischemia, trauma, or exogenous toxic substances. Moreover, mice deficient in PACAP are more vulnerable to damaging insults. In this study, we sought to determine whether PACAP may also be protective against salsolinol-induced toxicity in SH-SY5Y cells and, if so, elucidate its mechanism(s) of action. Salsolinol (SALS) is an endogenous dopamine metabolite with selective toxicity to nigral dopaminergic neurons, which are directly implicated in Parkinson’s disease (PD). SH-SY5Y cells, derived from human neuroblastoma cells, express high levels of dopaminergic activity and are used extensively as a model to study these neurons. Exposure of SH-SY5Y cells to 400 μM SALS for 24 h resulted in approximately 50 % cell death that was mediated by apoptosis as determined by cell flow cytometry and increases in caspase-3 levels. Cellular toxicity was also associated with reductions in brain-derived neurotrophic factor and phosphorylated cyclic AMP response element-binding protein. Pretreatment with PACAP dose-dependently attenuated SALS-induced toxicity and the associated apoptosis and the chemical changes. PACAP receptor antagonist PACAP6-38, in turn, dose-dependently blocked the effects of PACAP. Neither PACAP nor PACAP antagonist had any effect of its own on cellular viability. These results suggest the protective effects of PACAP in a cellular model of PD. Hence, PACAP or its agonists could be of therapeutic benefit in PD.     

Pharmacological consequences of nicotinergic plus serotonergic manipulations

The present study investigates the effects of concurrent manipulations of nicotinic cholinergic receptors (nicotinic cholinergic agonist: nicotine 0.03, 0.1, 0.3 mg/kg, nicotinic cholinergic antagonist: mecamylamine 7.5 mg/kg) and serotonin neurons (p-chlorophenylalanine (PCPA), 400/kg mg on each of 3 days) on spatial navigation (water maze, WM) and passive avoidance (PA) performance. Nicotine did not affect PA performance but at the highest dose slightly impaired WM performance. PCPA did not affect WM navigation or PA performance in saline or nicotine-treated rats. Nicotine restored WM and PA performance defect in mecamylamine pretreated rats. PCPA aggravated the WM defect and decreased the WM performance-improving effect of nicotine in mecamylamine pretreated rats. PCPA did not aggravate the PA performance defect of mecamylamine but completely blocked the PA performance-improving effect of nicotine in mecamylamine pretreated rats. These results suggest that serotonergic and nicotinergic cholinergic systems jointly modulate performance in WM and PA tests.     

Influence of various drugs on the voluntary intake of nicotine by rats

The effects of the dopaminergic agonists (L-dopa, pergolide) and antagonists (haloperidol, clozapin) and a cholinergic agonist (tacrine) and antagonist (mecamylamine) on the voluntary intake of nicotine were investigated with the 2-bottle paradigm with the test drugs being dissolved directly in the drinking fluid of the animals. This method was found to be a reliable procedure to quickly screen compounds with specific sites of action in the brain for their effects on the voluntary intake of nicotine or perhaps other substances of abuse as well. L-dopa, pergolide and haloperidol did not affect the intake of nicotine, whereas tacrine increased it slightly and clozapine and mecamylamine markedly. These results indicate that blockade of nicotinic and dopaminergic D4 receptors partially reduce the desired effect of nicotine by forcing the animals to consume more of this substance.     

Nicotine modulates expression of amyloid precursor protein and amyloid precursor-like protein 2 in mouse brain and in SH-SY5Y neuroblastoma cells

Epidemiological studies indicate that tobacco smoking can be protective against neurodegenerative diseases such as Alzheimer's disease (AD) and Parkinson's disease (PD). The objective of the present study was to examine the changes in gene expression induced by chronic oral nicotine administration (100 mug/ml in 2% saccharin for 14 days), with special emphasis on amyloid precursor protein (APP) and its homologue, amyloid precursor-like protein 2 (APLP2), in different brain regions of C57BL/6 mice using a pathway-focused microarray. Our results revealed that nicotine stimulated mRNA expression of APP in the amygdala (64%; P = 0.003) and hippocampus (32%; P = 0.034) and of APLP2 in the amygdala (39%; P = 0.002). These results were verified by quantitative real-time RT-PCR except that expression of APLP2 was also significantly upregulated by nicotine in the hippocampus. In addition, in vitro nicotine treatment of SH-SY5Y neuroblastoma cells resulted in a significant increase in expression of APP protein, soluble APP, and APLP2, whereas co-treatment with mecamylamine (an antagonist of nicotinic acetylcholine receptors) attenuated the stimulating effect of nicotine on APP and APLP2 expression. These findings suggest that nicotine treatment facilitates the increase in the expression of mRNA and protein of the APP and APLP2 genes in rat brain and SH-SY5Y neuroblastoma cells.     

Cardiovascular responses to microinjections of nicotine into the caudal ventrolateral medulla of the rat

This study focuses on the role of nicotinic receptors located in the caudal ventrolateral medullary depressor area (CVLM) in regulating/modulating cardiovascular function. Blood pressure and heart rate were monitored by standard techniques in urethane-anesthetized, artificially ventilated, adult male Wistar rats. Multi-barreled glass-micropipettes (tip size 20-40 microm) were used to make microinjections (100 nl) into the CVLM. Concentrations of nicotine ranging from 250 micromto 10 mM were microinjected unilaterally into the CVLM. The maximum depressor and bradycardic responses were elicited by a 1 mM concentration of nicotine. Sequential microinjections of mecamylamine (1 mM), an antagonist for nicotinic receptors containing alpha3beta4 subunits, then alpha-bungarotoxin (1 microm), an antagonist for nicotinic receptors containing alpha-7 subunits, were made into the CVLM. Microinjecting a combination of a nicotinic receptor blocker and toxin resulted in the complete blockade of the cardiovascular responses induced by nicotine (1 mM, 100 nl). These results indicate that: (1) nicotinic receptors are present in the CVLM; (2) activation of these receptors results in depressor and bradycardic responses; (3) for a complete blockade of nicotine-induced cardiovascular responses, it is necessary to use a combination of mecamylamine and alpha-bungarotoxin; (4) since mecamylamine and alpha-bungarotoxin are known to block nicotinic receptors containing alpha3beta4 and alpha-7 subunits, respectively, two different subtypes of nicotinic receptors (one which contains a combination of alpha3beta4 subunits, and one which contains alpha-7 subunits) must be present in the CVLM; and (5) it is not clear whether these two subtypes of nicotinic receptor are located on the same or different populations of CVLM-neurons.     

Tacrine interacts with different sites on nicotinic receptor subtypes in SH-SY5Y neuroblastoma and M10 cells

The effect of chronic treatment with the cholinesterase inhibitor tacrine on nicotinic receptor subtypes was investigated in human SH-SY5Y neuroblastoma cells and in a fibroblast cell line (M10 cells) stably transfected with alpha4beta2 nicotinic receptors. Tacrine significantly increased the number of nicotinic receptors in SH-SY5Y cells, in a concentration dependent manner (10(-9) to 10(-4) M), when using [3H]epibatidine as labelled ligand. Chronic tacrine treatment of M10 cells significantly increased and decreased the number of alpha4beta2 nicotinic receptors in a concentration dependent manner (10(-9) to 5 x 10(-6) M and 2 x 10(-5) to 10(-4) M, respectively). The tacrine induced increase of nicotinic receptors in SH-SY5Y cells, was not blocked in the presence of the nicotinic antagonists tubocurarine or mecamylamine. A further increase in the number of nicotinic receptors was, however, observed in the presence of mecamylamine. This study demonstrates that the effect of tacrine on the number of nicotinic receptor subtypes is different in human SH-SY5Y neuroblastoma and M10 cells. The up-regulation of different nicotinic receptor subtypes obtained with tacrine might be achieved through interaction via different binding sites on the receptor, i.e. the acetylcholine binding site as well as an allosteric site.     

Influence of Central Nicotinic Receptors on Arachidonylcyclopropylamide (ACPA)-Induced Antinociception in Mice

The nicotinic cholinergic receptors have been reported to be involved in several actions of cannabinoids (e.g., bradycardia, hypothermia). However, the influence of central cholinergic system on cannabinoids antinociceptive effect has not been reported. This study investigated the possible part played by nicotinic cholinergic modulator drugs on the antinociceptive effect of central administration of arachidonylcyclopropylamide (ACPA) in mice. The antinociceptive effects of intracerebroventricular (i.c.v.) administration of ACPA using the formalin test have been studied in mice. The effects of nicotine or mecamylamine (a nicotinic cholinergic antagonist) on ACPA analgesia are also studied. i.c.v. administration of ACPA (0.004-1 μg/mice) induced antinociceptive effect in mice. i.c.v. administration of nicotine (0.1 or 0.5 μg/mice) or mecamylamine (2 μg/mice) potentiated or antagonized ACPA antinociceptive effects, respectively. It is concluded that ACPA-induced analgesia is influenced by central nicotinic cholinergic activity.     

Nicotine inhibits ethanol-induced toxicity in cultured cerebral cortical cells

The high incidence of smoking among alcoholics may be partially due to nicotine's ability to counteract some of the adverse effects of ethanol on motor coordination and/or cognitive functions. Neuroprotective effects of nicotine on ethanol-induced toxicity in cerebellar granular cells have been observed. In this study, we sought to determine whether similar protection is observed in neocortical cells and if so, what specific nicotinic receptor subtypes may be mediating the actions of nicotine. Primary cultures of neocortical cells were prepared from 20-day embryos obtained from time-pregnant Sprague-Dawley rats. Cells were cultured for 10 days and were then exposed for 3 days to various concentrations of ethanol with and without pretreatment with nicotine and nicotinic antagonists. Cellular toxicity was evaluated by measuring the lactate dehydrogenase level. Administration of ethanol (10-100 mM) resulted in a dose-dependent toxicity. Pretreatment with nicotine 5-20 microM resulted in a dose-dependent protection against ethanol-induced toxicity. The effects of nicotine were blocked by pretreatment with nicotinic antagonists such as mecamylamine (1-20 microM), dihydro-beta-erythroidine (DHBE) 50 nM-1.0 microM and methyllycaconitine (MLA) 5 nM-1 microM in a dose-dependent manner. Compared to previous studies, higher ethanol concentrations were required to induce toxicity in neocortical vs cerebellar granule cells. Moreover, the effects of nicotine in the neocortical cells were blocked by lower concentrations of MLA, but higher concentrations of DHBE compared to cerebellar cells. Collectively, the results suggest differential sensitivity of various neuronal populations to the toxic effect of ethanol. Furthermore, protective effects of nicotine against alcohol in various regions appear to be mediated by different nicotinic receptor subtypes. The neuroprotective effect of nicotine against ethanol-induced toxicity may be a contributing factor to the high incidence of smoking among alcoholics.     

Nicotine reduces A beta in the brain and cerebral vessels of APPsw mice

Ten days treatment with nicotine reduced insoluble amyloid A beta 1-40 and Alpha beta 1-42 peptides by 80% in the cortex of 9-month-old APPsw mice, which is more than that observed in 14.5-month-old mice following nicotine treatment for 5.5 months. A reduction in A beta associated with cerebral vessels was observed in addition to that deposited as parenchymal plaques after 5.5 months treatment. The diminution in A beta peptides observed was not accompanied by changes in brain alpha, beta or gamma secretase-like activities, NGF or BDNF protein expression measured in brain homogenates. A significant increase in sAPP was observed after nicotine treatment of SH-SY5Yneuroblastoma cells that could be blocked by the nicotinic antagonist mecamylamine. Attenuation of elevated [(125)I]-alpha bungarotoxin binding (alpha 7) in APPsw mice was observed after 5.5 months nicotine treatment. Both these observations suggest that the reduction in insoluble A beta by nicotine might be in part mediated via the alpha 7 nicotinic receptor. Further studies are required to identify potential mechanisms of the nicotine's amyloid-reducing effect.     

Influence of central nicotinic receptors on arachidonylcyclopropylamide (ACPA)-induced antinociception in mice

The nicotinic cholinergic receptors have been reported to be involved in several actions of cannabinoids (e.g., bradycardia, hypothermia). However, the influence of central cholinergic system on cannabinoids antinociceptive effect has not been reported. This study investigated the possible part played by nicotinic cholinergic modulator drugs on the antinociceptive effect of central administration of arachidonylcyclopropylamide (ACPA) in mice. The antinociceptive effects of intracerebroventricular (i.c.v.) administration of ACPA using the formalin test have been studied in mice. The effects of nicotine or mecamylamine (a nicotinic cholinergic antagonist) on ACPA analgesia are also studied. i.c.v. administration of ACPA (0.004-1 microg/mice) induced antinociceptive effect in mice. i.c.v. administration of nicotine (0.1 or 0.5 microg/mice) or mecamylamine (2 microg/mice) potentiated or antagonized ACPA antinociceptive effects, respectively. It is concluded that ACPA-induced analgesia is influenced by central nicotinic cholinergic activity.     

Protective effects of nicotine on ethanol-induced toxicity in cultured cerebellar granule cells

Alcoholism is associated with a higher incidence of smoking. In addition to the stimulatory effects of both ethanol and nicotine on the mesolimbic reward pathway, nicotine's ability to counteract some of the adverse effects of ethanol (e.g. ataxia) may be a powerful incentive for alcohol consumers to increase their tobacco (nicotine) intake. The cerebellum is believed to play an important role in ethanol-induced ataxia. In this study, we sought to test the hypothesis that nicotine would protect against toxic effects of ethanol in primary cultures of cerebellar granule cells. Moreover, it was postulated that the effects of nicotine would be mediated through nicotinic receptors. Primary cultures of cerebellar granule cells were prepared from 20-day embryos obtained from timed-pregnant Sprague Dawley rats. Cells were cultured for 10 days and were then exposed for 3 days to various concentrations of ethanol with and without pretreatment with nicotine and nicotinic antagonists. Cellular toxicity was evaluated by measuring the lactate dehydrogenase level. Administration of ethanol (10–100 mM) resulted in a dose-dependent toxicity. Pretreatment with nicotine 1–20 μM resulted in a dose-dependent protection against ethanol-induced toxicity. The effects of nicotine were blocked by pretreatment with nicotinic antagonists such as mecamylamine 1–20 μM, dihydro-β-erythroidine 1.0 nM–1.0 μM and methyllycaconitine 5 nM-5 μM in a dose-dependent manner. Thus, ethanol-induced cytotoxicity in primary cultures of cerebellar granule cells is blocked by pretreatment with nicotine. The effects of nicotine, in turn, may be blocked by nicotinic antagonists, implicating both high and low affinity nicotinic receptors in mediating the actions of nicotine. The exact mechanism of ethanol-induced toxicity and/or neuroprotection through activation of nicotinic receptors in this paradigm remains to be elucidated. The neuroprotective effect of nicotine against ethanol-induced toxicity in cerebellar neurons may be a contributing factor to the high incidence of smoking among alcoholics.     

Stimulation of nicotinic receptors in the lateral septal nucleus increases anxiety

The present study investigated the role of nicotinic receptors in the lateral septum in the modulation of anxiety. The effects of direct injections of nicotine into the lateral septum were first investigated in two tests of anxiety, social interaction and elevated plus-maze tests. Intra-septal injection of nicotine (1 and 4 microgram) induced consistent anxiogenic effects in both tests. The reversal of nicotinic effects with mecamylamine was then studied in the social interaction test. Intra-septal injection of mecamylamine at a low dose (15 ng) induced an anxiolytic effect, suggesting the presence of intrinsic cholinergic tone increasing anxiety. At higher doses (30-50 ng), mecamylamine was without effect in the social interaction test, but blocked the anxiogenic effects of nicotine (4 microgram). These findings provide further evidence for the role of the lateral septum in the modulation of anxiety and suggest that cholinergic projections to this brain area facilitate anxiety through nicotinic receptors.     

Nicotine Induces Sensitization of Turning Behavior in 6-Hydroxydopamine Lesioned Rats

Nicotinic drugs have been proposed as putative drugs to treat Parkinson’s disease (PD). In this study, we investigated whether nicotine can sensitize parkinsonian animals to the effect of dopaminergic drugs. Testing this hypothesis is important because nicotine has been shown to present neuroprotective and acute symptomatic effects on PD, but few studies have addressed the question of whether it may induce long-lasting effects on dopamine neurotransmission. We tested this hypothesis in the 6-hydroxydopamine (6-OHDA) rat model of PD. A pretreatment of these rats with 0.1–1.0 mg/kg nicotine induced a dose-dependent sensitization of the turning behavior when the animals were challenged with the dopamine receptor agonist apomorphine 24 h later. In agreement with previous studies, while apomorphine induced contraversive turns, nicotine, as well as amphetamine, induced ipsiversive turns in the 6-OHDA rats. This result suggests that, like amphetamine, nicotine induces turning behavior by promoting release of dopamine in the non-lesioned striatum of the rats. However, it is unlikely that the release of dopamine may also explain the nicotine-induced sensitization of turning behavior. First, the dopamine amount that could be released in the lesioned hemi-striatum by the nicotine pretreatment was minimum—less than 3%, as detected by HPLC–EC. Second, a pretreatment with amphetamine did not induce this behavioral sensitization. A pretreatment with apomorphine-induced sensitization, but it was minimal when compared to that induced by nicotine. Therefore, it is unlikely that the sensitization of the turning behavior induced by nicotine was consequent of the release of dopamine. However, the expression of such sensitization seems to depend on the activation of dopaminergic receptors, since it was seen when the nicotine-sensitized animals were challenged with apomorphine, but not with a second nicotine challenge. These findings are relevant for PD drug therapy since they suggest that the doses of dopaminergic drugs used to treat PD could be reduced if a nicotinic drug were co-administered.