Role of prediction error and the cholinergic system on memory reconsolidation processes in mice.

The ability to make predictions based on stored information is a general coding strategy. A prediction error (PE) is a mismatch between expected and current events. Our memories, like ourselves, are subject to change. Thus, an acquired memory can become active and update its content or strength by a labilization-reconsolidation process. Within the reconsolidation framework, PE drives the updating of consolidated memories. In the past our lab has made key progresses showing that a blockade in the central cholinergic system during reconsolidation can cause memory impairment, while reinforcement of cholinergic activity enhances it. In the present work we determined that PE is a necessary condition for memory to reconsolidate in an inhibitory avoidance task using both male and female mice. Depending on the intensity of the unconditioned stimulus (US) used during training, a negative (higher US intensity) or positive (lower US intensity/no US) PE on a retrieval session modified the behavioral response on a subsequent testing session. Furthermore, we demonstrated that the cholinergic system modulates memory reconsolidation only when PE is detected. In this scenario administration of oxotremorine, scopolamine or nicotine after memory reactivation either enhanced or impaired memory reconsolidation in a sex-specific manner.

Effects of diazinon on the lymphocytic cholinergic system of Nile tilapia fish (Oreochromis niloticus).

Fish rearing under intensive farming conditions can be easily disturbed by pesticides, substances that have immunotoxic properties and may predispose to infections. Organophosphorus pesticides (OPs) are widely used in agricultural activities; however, the mechanism of immunotoxicity of these substances is unclear. The aim of this study was to evaluate the effect of diazinon pesticides (OPs) on the cholinergic system of immune cells as a possible target of OP immunotoxicity. We evaluated ACh levels and cholinergic (nicotinic and muscarinic) receptor concentration. Additionally, AChE activity was evaluated in mononuclear cells of Nile tilapia (Oreochromis niloticus), a freshwater fish mostly cultivated in tropical regions around the world. The obtained results indicate that acute exposure to diazinon induces an increase in ACh concentration and a decrease in nAChR and mAChR concentrations and AChE activity in fish immune cells, This suggests that the non-neuronal lymphocytic cholinergic system may be the main target in the mechanism of OP immunotoxicity. This study contributes to the understanding of the mechanisms of immunotoxicity of pollutants and may help to take actions for animal health improvement.

Pattern of Maxillofacial Fractures in Severe Multiple Trauma Patients: A 7-year Prospective Study

The incidence of facial trauma is high. This study has the primary objective of documenting and cataloging maxillofacial fractures in polytrauma patients. From a total of 1229 multiple trauma cases treated at the Emergency Room of the Santo Antonio Hospital - Oporto Hospital Center, Portugal, between August 2001 and December 2007, 251 patients had facial wounds and 209 had maxillofacial fractures. Aged ranged form 13 to 86 years. The applied selective method was based on the presence of facial wound with Abbreviated Injury Scale ≥1. Men had a higher incidence of maxillofacial fractures among multiple trauma patients (86.6%) and road traffic accidents were the primary cause of injuries (69.38%). Nasoorbitoethmoid complex was the most affected region (67.46%) followed by the maxilla (57.42%). The pattern and presentation of maxillofacial fractures had been studied in many parts of the world with varying results. Severe multiple trauma patients had different patterns of maxillofacial injuries. The number of maxillofacial trauma is on the rise worldwide as well as the incidence of associated sequelae. Maxillofacial fractures on multiple trauma patients were more frequent among males and in road traffic crashes. Knowing such data is elementary. The society should have a key role in the awareness of individuals and in prevention of road traffic accidents.

É alta a incidência de traumas na face. Este estudo teve por objetivo documentar e catalogar as fraturas maxilofaciais em pacientes com politraumatismos. De um total de 1229 casos de politraumatizados tratados na Sala de Emergência do Hospital de Santo António - Centro Hospitalar do Porto, Portugal, entre Agosto de 2001 e Dezembro de 2007, 251 pacientes tiveram ferimentos na face e 209 apresentaram fraturas maxilofaciais. As idades variaram de 13 a 86 anos. O método de seleção baseou-se na presença de ferimentos na face com Abreviated Injury Scale ≥1. Os homens apresentaram maior incidência de fraturas maxilofaciais (86,6%) entre os pacientes com múltiplos traumatismos na face e os acidentes de trânsito foram a causa principal dos traumatismos (69,38%). A região mais afetada foi o complexo naso-órbito-etmoidal (67,46%), seguido pela maxila (57,42%). O padrão e a apresentação das fraturas maxilofaciais tem sido estudado em muitas regiões do mundo com resultados variados. Pacientes com politraumatizados graves apresentaram padrões diferentes de traumatismos maxilofaciais. O número de traumatismos maxilofaciais tem aumentado à escala mundial, assim como a incidência das sequelas associadas. Entre os pacientes com traumatismos múltiplos, a maioria pertencia ao sexo masculino, assim como a causa mais frequente foram os acidentes automobilísticos. É elementar o conhecimento destes dados. A sociedade tem um papel primordial nos cuidados individuais e na prevenção dos acidentes de trânsito.

Mechanisms of the antinociceptive action of (-) epicatechin obtained from the hydroalcoholic fraction of Combretum leprosum Mart & Eic in rodents.

BACKGROUND: The mechanisms of the antinociceptive activity of (-) epicatechin (EPI), a compound isolated from the hydroalcoholic fraction of Combreum leprosum Mart & Eicher. METHODS: were assessed in the model of chemical nociception induced by glutamate (20 µmol/paw). To evaluate the mechanisms involved, the animals , male Swiss mice (25-30 g), received EPI (50 mg/kg p.o.) after pretreatment with naloxone (2 mg/kg s.c. opioid antagonist), glibenclamide (2 mg/kg s.c. antagonist K + channels sensitive to ATP), ketanserin (0.3 mg/kg s.c. antagonist of receptor 5-HT(2A)), yoimbine (0.15 mg/kg s.c. α2 adrenergic receptor antagonist), pindolol (1 mg/kg s.c. 5-HT1(a)/1(b) receptor antagonist), atropine (0.1 mg/kg s.c. muscarinic antagonist) and caffeine (3 mg/kg s.c. adenosine receptor antagonist), ondansetron (0.5 mg/kg s.c. for 5-HT(3) receptor) and L-arginine (600 mg/kg i.p.). RESULTS: The antinociceptive effect of EPI was reversed by pretreatment with naloxone and glibenclamide, ketanserin, yoimbine, atropine and pindolol, which demonstrates the involvement of opioid receptors and potassium channels sensitive to ATP, the serotoninergic (receptor 5HT(1A) and 5HT(2A)), adrenergic (receptor alpha 2) and cholinergic (muscarinic receptor) systems in the activities that were observed. The effects of EPI, however, were not reversed by pretreatment with caffeine, L-arginine or ondansetron, which shows that there is no involvement of 5HT(3) receptors or the purinergic and nitrergic systems in the antinociceptive effect of EPI. In the Open Field and Rotarod test, EPI had no significant effect, which shows that there was no central nervous system depressant or muscle relaxant effect on the results. CONCLUSIONS: This study demonstrates that the antinociceptive activity of EPI in the glutamate model involves the participation of the opioid system, serotonin, adrenergic and cholinergic.

Cognitive enhancers: focus on modulatory signaling influencing memory consolidation.

Biological research has unraveled many of the molecular and cellular mechanisms involved in the formation of long-lasting memory, providing new opportunities for the development of cognitive-enhancing drugs. Studies of drug enhancement of cognition have benefited from the use of pharmacological treatments given after learning, allowing the investigation of mechanisms regulating the consolidation phase of memory. Modulatory systems influencing consolidation processes include stress hormones and several neurotransmitter and neuropeptide systems. Here, we review some of the findings on memory enhancement by drug administration in animal models, and discuss their implications for the development of cognitive enhancers.

Alterations in cholinergic sensitivity of respiratory neurons induced by pre-natal nicotine: a mechanism for respiratory dysfunction in neonatal mice.

Nicotine may link cigarette smoking during pregnancy with sudden infant death syndrome (SIDS). Pre-natal nicotine leads to diminished ventilatory responses to hypercarbia and reduced central chemoreception in mice at post-natal days 0-3. We studied how pre-natal nicotine exposure changes the cholinergic contribution to central respiratory chemoreception in neonatal isolated brainstem-spinal cord and slice preparations. Osmotic minipumps, implanted subcutaneously into 5-7 days pregnant mice, delivered saline or nicotine ditartrate 60 mg kg(-1) d(-1) for up to 28 days. In control preparations, acidification of the superfusion medium from pH 7.4 to 7.3 increased the frequency and reduced the amplitude of fictive respiration. In nicotine-exposed neonatal mice, the reduction in amplitude induced by acidification was reduced. In control preparations, atropine suppressed respiratory responses to acidification, while hexamethonium did not. By contrast, in nicotine-exposed preparations, hexamethonium blocked chemosensory responses but atropine did not. Our results indicate that pre-natal nicotine exposure switches cholinergic mechanisms of central chemosensory responses from muscarinic receptors to nicotinic receptors. Modification of the cholinergic contribution to central chemoreception may produce respiratory dysfunctions, as suggested by receptor-binding studies in victims of SIDS.

Filicene obtained from Adiantum cuneatum interacts with the cholinergic, dopaminergic, glutamatergic, GABAergic, and tachykinergic systems to exert antinociceptive effect in mice.

In the present study, we describe the antinociceptive effect of filicene, a triterpene isolated from Adiantum cuneatum (Adiantaceae) leaves, in several models of pain in mice. When evaluated against acetic acid-induced abdominal constrictions, filicene (10, 30 and 60 mg/kg, i.p.) produced dose-related inhibition of the number of constrictions, being several times more potent [ID(50)=9.17 (6.27-13.18) mg/kg] than acetaminophen [ID(50)=18.8 (15.7-22.6) mg/kg], diclofenac [ID(50)=12.1(9.40-15.6) mg/kg] and acetylsalicylic acid [ID(50)=24.0(13.1-43.8) mg/kg] in the same doses as those used for the standard drugs. Filicene also produced dose-related inhibition of the pain caused by capsaicin and glutamate, with mean ID(50) values of 11.7 (8.51-16.0) mg/kg and <10 mg/kg, respectively. Its antinociceptive action was significantly reversed by atropine, haloperidol, GABA(A) and GABA(B) antagonists (bicuculline and phaclofen, respectively), but was not affected by L-arginine-nitric oxide, serotonin, adrenergic and the opioid systems. Together, these results indicate that the mechanisms involved in its action are not completely understood, but seem to involve interaction with the cholinergic, dopaminergic, glutamatergic, GABAergic and tachykinergic systems.

Ryanodine is a positive modulator of acetylcholine receptor gating in cochlear hair cells.

The efferent synaptic specialization of hair cells includes a near-membrane synaptic cistern, whose presence suggests a role for internal calcium stores in cholinergic inhibition. Calcium release channels from internal stores include 'ryanodine receptors', whose participation is usually demonstrated by sensitivity to the eponymous plant alkaloid, ryanodine. However, use of this and other store-active compounds on hair cells could be confounded by the unusual pharmacology of the alpha9alpha10-containing hair cell nicotinic cholinergic receptor (nAChR), which has been shown to be antagonized by a broad spectrum of compounds. Surprisingly, we found that ryanodine, rather than antagonizing, is a positive modulator of the alpha9alpha10 nAChR expressed in Xenopus oocytes, the first such compound to be found. The effect of ryanodine was to increase the apparent affinity and efficacy for acetylcholine (ACh). Correspondingly, ACh-evoked currents through the isolated cholinergic receptors of inner hair cells in excised mouse cochleas were approximately doubled by 200 microM ryanodine, a concentration that inhibits gating of the ryanodine receptor itself. This unusual positive modulation was not unique to the mammalian receptor. The response to ACh of chicken 'short' hair cells likewise was enhanced in the presence of 100 microM ryanodine. This facilitatory effect on current through the AChR could enhance brief ( approximately 1 s) activation of associated calcium-dependent K(+) (SK) channels in both chicken short hair cells and rat outer hair cells. This novel effect of ryanodine provides new opportunities for the design of compounds that potentiate alpha9alpha10-mediated responses and for potential inner ear therapeutics based on this interaction.

Carotid body chemosensory activity and ventilatory chemoreflexes in cats persist after combined cholinergic-purinergic block.

Acetylcholine (ACh) and ATP have been proposed as excitatory co-transmitters operating at synapses between glomus cells and sensory nerve endings of the carotid body (CB). To test such hypothesis, we performed experiments on cats under pentobarbitone anesthesia and breathing spontaneously. Cholinergic and purinergic agonists and antagonists were given into one common carotid artery. Chemoreflex ventilatory changes initiated from the ipsilateral CB or chemosensory activity from the ipsilateral carotid nerve were recorded. Agonists ACh, nicotine, epibatidine, ATP, betagamma-methylene-ATP and gammaS-ATP induced transient chemoreflex enhancements of ventilation or increased chemosensory activity. When given in combination, mecamylamine and suramin suppressed both nicotine- and ATP-induced ventilatory chemoreflexes or chemosensory responses. However, neither chemoreflex hyperventilation induced by brief hypoxic exposures or steady-state hypoxic levels, nor chemosensory excitation elicited by these maneuvers were eliminated. Asphyxia-induced chemosensory excitation was not reduced by combined blockade of ACh and ATP receptors. Furthermore, ventilatory or chemosensory depression evoked by 100% O2 tests was unmodified, thus evidencing that basal chemosensory drive in normoxia was not suppressed by combined cholinergic-purinergic blockade. Therefore, although ACh and ATP may participate in chemoexcitation of the CB, their involvement fails to explain the origin of chemosensory discharges from synaptic transmission between glomus cells and chemosensory nerve endings of the CB.

Effects of combined cholinergic-purinergic block upon cat carotid body chemoreceptors in vitro.

Since acetylcholine (ACh) and ATP have been proposed as excitatory co-transmitters at synapses between glomus cells and sensory nerve endings of the carotid body (CB), we tested such hypothesis by studying the effects of combined cholinergic-purinergic block on the chemosensory activity recorded from cat's carotid bodies perfused and/or superfused in vitro. The preparations were bathed with Tyrode's solution, either normoxic (PO2=98.5+/-13.5 Torr) or hypoxic (PO2=31.8+/-5.2 Torr), and the frequency of chemosensory impulses (fchi) was recorded from the carotid (sinus) nerve. Dose-response curves for fchi increases evoked by intra-stream boluses of acetylcholine, nicotine and ATP were studied. A combination of mecamylamine 2 microM and suramin 50 microM, applied through the perfusate or superfusate, suppressed nicotine- and ATP-induced increases in fchi, but the basal chemosensory activity in normoxia and the chemosensory excitation elicited by hypoxic superfusion were preserved, although variably reduced in most preparations. Thus, in spite of the excitatory effects provoked by applying ACh and ATP to the perfused/superfused CB in vitro, a co-release of these substances cannot account entirely for the chemosensory excitation induced by hypoxic stimulation of the CB.

Modulation of working, short- and long-term memory by nicotinic receptors in the basolateral amygdala in rats.

Male Wistar rats were exposed to one-trial step-down inhibitory avoidance training using a 0.5 mA footshock. Through bilaterally implanted indwelling cannulae, they received bilateral 0.5 microL infusions of saline, mecamylamine (1.0 or 10.0 microg/side), or nicotine (0.6 or 3.0 microg/side) into the basolateral complex of the amygdaloid nucleus (BLA). Infusions were either 10 min before training (Experiment 1) or 4 min after training (Experiment 2). In Experiment 1, the animals were tested three times: first for working memory (WM) 5 s after training, then for short-term memory (STM) 90 min later, and finally for long-term memory (LTM) 24 h later. Mecamylamine depressed and nicotine enhanced WM, STM, and LTM. In Experiment 2, the treatments were given after WM was presumably over. Again, mecamylamine inhibited and nicotine enhanced STM and LTM. The results indicate that nAChRs in BLA participate in the regulation of WM formation and STM and LTM acquisition and consolidation.

Cholinergic-opioidergic interaction in the central amygdala induces antinociception in the guinea pig.

Several studies have demonstrated the involvement of the central nucleus of the amygdala (CEA) in the modulation of defensive behavior and in antinociceptive regulation. In a previous study, we demonstrated the existence of a cholinergic-opioidergic interaction in the CEA, modulating the defensive response of tonic immobility in guinea pigs. In the present study, we investigated a similar interaction in the CEA, but now involved in the regulation of the nociceptive response. Microinjection of carbachol (2.7 nmol) and morphine (2.2 nmol) into the CEA promoted antinociception up to 45 min after microinjection in guinea pigs as determined by a decrease in the vocalization index in the vocalization test. This test consists of the application of a peripheral noxious stimulus (electric shock into the subcutaneous region of the thigh) that provokes the emission of a vocalization response by the animal. Furthermore, the present results demonstrated that the antinociceptive effect of carbachol (2.7 nmol; N = 10) was blocked by previous administration of atropine (0.7 nmol; N = 7) or naloxone (1.3 nmol; N = 7) into the same site. In addition, the decrease in the vocalization index induced by the microinjection of morphine (2.2 nmol; N = 9) into the CEA was prevented by pretreatment with naloxone (1.3 nmol; N = 11). All sites of injection were confirmed by histology. These results indicate the involvement of the cholinergic and opioidergic systems of the CEA in the modulation of antinociception in guinea pigs. In addition, the present study suggests that cholinergic transmission may activate the release of endorphins/enkephalins from interneurons of the CEA, resulting in antinociception.

Central muscarinic receptors signal pilocarpine-induced salivation.

Although cholinergic agonists such as pilocarpine injected peripherally can act directly on salivary glands to induce salivation, it is possible that their action in the brain may contribute to salivation. To investigate if the action in the brain is important to salivation, we injected pilocarpine intraperitoneally after blockade of central cholinergic receptors with atropine methyl bromide (atropine-mb). In male Holtzman rats with stainless steel cannulas implanted into the lateral ventricle and anesthetized with ketamine, atropine-mb (8 and 16 nmol) intracerebroventricularly reduced the salivation induced by pilocarpine (4 micro mol/kg) intraperitoneally (133 + 42 and 108 + 22 mg/7 min, respectively, vs. saline, 463 + 26 mg/7 min), but did not modify peripheral cardiovascular responses to intravenous acetylcholine. Similar doses of atropine-mb intraperitoneally also reduced pilocarpine-induced salivation. Therefore, systemically injected pilocarpine also enters the brain and acts on central muscarinic receptors, activating autonomic efferent fibers to induce salivation.

Cholinergic responses of seminal vesicles isolated from rats exposed perinatally to hydrocortisone.

This study was performed in order to investigate the cholinomimetic response of seminal vesicles isolated from rats treated with hydrocortisone acetate during perinatal life. At the adult phase, the body weight and the wet weight of the seminal vesicle of these animals were unchanged. However, these male rats exhibited a significant reduction in plasma testosterone concentration. A significant increase in the sensitivity of the seminal vesicle to acetylcholine was also observed. Despite this, there was a significant reduction in the maximum contractile response of the organ to this transmitter. These results indicate that exposure to hydrocortisone during the critical period of brain sexual differentiation has a long-term effect on testosterone production of male rats. In addition, physiological levels of cortisone in perinatal life are also essential to support the contractile response pattern of the seminal vesicle to acetylcholine in adult life, probably crucial to the reproductive process.

Opioid receptors mediate a postsynaptic facilitation and a presynaptic inhibition at the afferent synapse of axolotl vestibular hair cells.

This study was designed to determine the effects of opiate drugs on the electrical activity of afferent neurons and on the ionic currents of hair cells from semicircular canals. Experiments were done on larval axolotls (Ambystoma tigrinum). The multiunit spike activity of afferent neurons was recorded in the isolated inner ear under both resting conditions and mechanical stimulation. Ionic currents were recorded using voltage clamp of hair cells isolated from the semicircular canal. In the isolated inner-ear preparation, microperfusion of either non-specific opioid receptor antagonist naloxone (10 nM to 1 mM), mu receptor agonist [D-Ala(2), N-Me-Phe(4),Gly(5)-ol]-enkephalin (1 pM to 10 microM), or kappa receptor antagonist nor-binaltorphimine (10 nM to 100 microM) elicited a dose-dependent long-lasting (>5 min) increase of the electrical discharge of afferent neurons. The mu receptor agonist funaltrexamine (1 nM to 100 microM) and the kappa receptor agonist U-50488 (1 nM to 10 microM) diminished the basal spike discharge of vestibular afferents. The delta receptor agonist D-Pen(2)-D-Pen(5)-enkephalin (1 nM to 10 mM) and the antagonist naltrindole (1 nM to 10 mM) were without a significant effect. The only drug that displayed a significant action on hair-cell ionic currents was trans-(+/-)-3,4-dichloro-N-methyl-N-(2-[1-pyrrolidinyl]-cyclohexyl) benzeneacetamide methanesulfonate (U-50488) that reduced the Ca(2+) current in a dose-dependent fashion. On its own, mu receptor agonist [D-Ala(2), N-Me-Phe(4),Gly(5)-ol]-enkephalin (0.01 and 10 microM) significantly potentiated the response of afferent neurons to the excitatory amino acid agonist (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (0.1 microM), while synaptic transmission was blocked by the use of high-Mg(2+), low-Ca(2+) solutions. Our data indicate that the activity of vestibular afferent neurons may be regulated in a complex fashion by opioid receptors: mu opioid receptors mediating an excitatory, postsynaptic modulatory input to afferent neurons, and kappa receptors mediating an inhibitory, presynaptic input to hair cells.

Chronic ethanol intake and ageing effects on cortical and basal forebrain cholinergic parameters: morphometric and biochemical studies.

Abstract Twenty-eight Wistar rats treated orally with 20% ethanol solution, were divided into two groups: adult group (n = 19) and aged group (n = 9) consisting of animals aged 4 and 12 months, respectively, at the beginning of the treatment. Neurons from the basal nucleus region were counted and the percentage of choline acetyltransferase-immunoreactive cholinergic neurons was determined in adjacent sections. Acetylcholine release and choline-acetyltransferase activity in the cerebral cortex were assessed in the same animals. Nutritional parameters of the ethanol treated animals were monitored and found to be normal. Chronic exposure to ethanol did not result in global neuronal loss or loss of cholinergic neurons in the basal nucleus region. However, a greater expression of ChAT-immunoreactivity in the basal nucleus region and a tendency toward increased ChAT activity in the cerebral cortex of the control and treated aged animals, compared respectively to adult ones, were observed. These findings suggest adaptive changes of the aged rats in response to the possible cholinergic hypofunction, manifested as a decreased release of acetylcholine under stimulated conditions.

Stressful and behavioral conditions that affect reversible cardiac arrest in the Nile tilapia, Oreochromis niloticus (Teleostei).

The objective of the present investigation was to study the reversible cardiac arrest (RCA) to visual stimuli in the unrestrained Nile tilapia (Oreochromis niloticus) as well as the modulation of this response and its behavioral component (arousal/orientation or startle response) by external and internal factors that interfere with alertness and emotionality. The study was preceded by the determination of the autonomic receptors that contribute to the establishment of the heart rate (HR) and the RCA. Systemic injection of atropine and propranolol showed that a double cardiac autonomic control is present in the tilapia. Basal HR was 79.8+/-1.8 beats min(-1) and HR assessed after double autonomic blockade was 74.1+/-3.3 beats min(-1). The mean interbeat interval was 0.79+/-0.40 s during baseline recording and the magnitude of RCA induced by a moving shadow (2.67+/-0.22 s) was higher than that induced by light (1.53+/-1.11 s). RCA is peripherally mediated by muscarinic receptors for it is abolished by atropine but not by propranolol. Stressful conditions like handling the animal outside the water or a nociceptive stimulus (subcutaneous 2% or 3% formalin injection) reduced the cardiac interbeat interval. A subanesthetic dose of barbiturate (5 mg kg(-1)) inhibited RCA induced by a moving shadow stimulus and the startle response, suggesting that an ideal degree of vigilance is necessary for its occurrence. Benzodiazepine injections (1.0 and 2.0 mg kg(-1)) abolished the reduction in magnitude of RCA induced by handling stress and facilitated the startle response, seen in the dry-cold season, in a dose-dependent manner. These data suggest that drugs that act on alertness and on emotionality modulate the magnitude of cardiac interbeat intervals and the corresponding behavioral response.

Participation of the cholinergic system in the excessive grooming behavior induced by neuropeptide (N) glutamic acid (E) isoleucine (I) amide (NEI).

The neuropeptide (N) glutamic acid (E) isoleucine (I) amide (NEI) injected into the ventral tegmental area (VTA) or intraventricularly (icv) induces excessive grooming behavior (EGB) and motor activity (MA). Here, we studied whether the cholinergic system is involved in the NEI-induced behavior. The present results demonstrate that atropine, a general muscarinic antagonist, injected icv previous to NEI, suppresses the behavior provoked by icv injections of the peptide, whereas the prior icv injection of dyhidro-beta-erythroidine, a general nicotinic antagonist, did not affect the EGB and MA induced by the peptide. From the experimental evidence, it is suggested that NEI may act specifically on a cholinergic afferent to dopaminergic cells. Also, the results appear to indicate that a neural target, different from the dopamine system, may be activated by the peptide to elicit behavioral changes, such as EGB.

The noncompetitive inhibitor quinacrine modifies the desensitization kinetics of muscle acetylcholine receptors.

Quinacrine has been shown to act as a noncompetitive inhibitor of the nicotinic acetylcholine receptor (nAChR). However, its mechanism of action is still a matter of controversy. We analyzed in detail the action of quinacrine at both the single-channel and macroscopic current levels. The main effect of quinacrine is a profound concentration-dependent decrease in both the frequency of opening events and the duration of clusters elicited by high acetylcholine concentrations. Quinacrine also significantly increases (40-fold at 30 microM) the decay rate of macroscopic currents elicited by rapid perfusion of acetylcholine to outside-out patches. This decay is still well-described by a single exponential. Quinacrine has very little effect on the peak amplitude of the response, suggesting that it acts mainly on open channels. The recovery from desensitization after removal of acetylcholine is delayed in the presence of quinacrine. Results from both single-channel and macroscopic current recordings indicate that quinacrine increases the rate of nAChR desensitization and stabilizes the desensitized state. Interestingly, in equilibrium agonist-binding assays, quinacrine does not promote the typical high-affinity desensitized state. Thus, quinacrine seems to induce an intermediate state exhibiting the permeability but not the agonist binding properties of desensitization.

Tobacco cembranoids block behavioral sensitization to nicotine and inhibit neuronal acetylcholine receptor function.

Cembranoids are cyclic diterpenoids found in tobacco and in marine invertebrates. The present study established that tobacco cembranoids inhibit behavioral sensitization to nicotine in rats and block several types of nicotine acetylcholine receptors (AChRs). 1) At the behavioral level, rat locomotor activity induced by nicotine was significantly increased after seven daily nicotine injections. This sensitization to nicotine was blocked by mecamylamine (1 mg/kg) and by the cembranoids eunicin, eupalmerin acetate (EUAC), and (4R)-2,7,11-cembratriene-4-6-diol (4R), each at 6 mg/kg. None of these compounds modified locomotor activity of nonsensitized rats. 2) In cells expressing human AChRs, cembranoids blocked carbamoylcholine-induced (86)Rb(+) flux with IC(50) in the low micromolar range. The cell lines used were the SH-EP1-halpha4beta2 cell line heterologously expressing human alpha4beta2-AChR, the SH-SY5Y neuroblastoma line naturally expressing human ganglionic alpha3beta4-AChR, and the TE671/RD cell line naturally expressing embryonic muscle alpha1beta1gammadelta-AChR. The tobacco cembranoids tested were 4R and its diastereoisomer 4S, and marine cembranoids tested were EUAC and 12,13-bisepieupalmerin. 3) At the molecular level, tobacco (4R and 4S) and marine (EUAC) cembranoids blocked binding of the noncompetitive inhibitor [(3)H]tenocyclidine to AChR from Torpedo californica electric organ. IC(50) values were in the submicromolar to low-micromolar range, with 4R displaying an order of magnitude higher potency than its diastereoisomer, 4S.