Pica--a model of nausea? Species differences in response to cisplatin

Rats lack the emetic reflex but exhibit pica in response to stimuli that induce emesis in species with an emetic reflex, hence it has been proposed that pica may be analogous to emesis in species lacking the reflex. In the present study, we investigated whether pica was present in Suncus murinus (with an emetic reflex) as well as in rats and mice (without emetic reflex) to provide a further insight to the validity of pica as a model for nausea/vomiting. Cisplatin (6 mg/kg, i.p.) induced pica in rats, indicated by a significant increase in kaolin consumption at 24 h (but not 48 h) post-treatment whereas we failed to demonstrate this effect in mice (inbred or outbred strain, 6 or 20 mg/kg i.p.) and whilst cisplatin (20 mg/kg, i.p.) induced emesis in Suncus, kaolin intake was not significantly affected. Furthermore, cisplatin significantly increased the weight of gastric contents at 48 h post-injection in rats and mice indicating delayed gastric emptying whereas this effect was not present in Suncus. These results show that Suncus and two strains of mice, unlike rats, do not develop pica in response to cisplatin which suggests that the consumption of kaolin induced by cisplatin may not be associated with whether or not an emetic reflex is present. The differences in ingestive behaviour and gastric response between species with and without an emetic reflex in response to cisplatin treatment as well as the difference between mice and rats, is discussed in relation to the selection of models for the study of nausea and vomiting.     

Conditioned food aversion in Suncus murinus (house musk shrew) - a new model for the study of nausea in a species with an emetic reflex

The lack of a small animal model with an emetic reflex in which the relationship between conditioned food aversion and emesis could be investigated prompted a study of the insectivore, Suncus murinus (the house musk shrew). A novel food (either tuna or chicken cat food) was paired (C+) with a single exposure to either nicotine (4 mg/kg sc), motion (1 Hz, 4 cm, 10 min) or lithium chloride (100 mg/kg ip) or was paired (C-) with either saline or sham exposure to motion. Nicotine and motion both induced emesis (retching/vomiting) but lithium chloride did not. All three treatments produced a conditioned food aversion after a single pairing with consumption of C+ food. When given a choice between the two foods, S. murinus given lithium chloride, motion exposure and nicotine consumed, respectively, only 25%, 23% and 1% of their total intake from the C+ food. This study shows that a conditioned food aversion can be readily induced in S. murinus and that the induction of emesis can be uncoupled from food aversion. S. murinus provides a promising new model in which the relationship between emesis, nausea and conditioned food aversion can be investigated.     

Inhibited drinking and pica in rats following 2-deoxy-D-glucose

A sequence of studies sought to determine if 2-deoxy-D-glucose (2DG) produces malaise as a side effect. Drug treatment at dosages at least as low as 250 mg/kg triggered an immediate inhibition of fluid intake. In nondeprived animals, 750 mg/kg but not lower dosages of 2DG evoked a delayed but statistically significant level of pica as measured by kaolin consumption. Twenty-four hour food deprivation failed to produce pica, and previous claims that kaolin intake is a specific index of gastrointestinal distress therefore received additional support. Thirsty rats injected with 750 mg/kg 2DG displayed a delayed pattern of pica largely similar to that previously observed with undeprived subjects; although, under optimal circumstances, significant increases in kaolin consumption may be evident as soon as one hour post-injection. In general, these data indicated that malaise may be an important influence of 2DG; but dosage and time course parameters associated with post-drug reductions in drinking suggested that a separable consummatory inhibition effect may also occur.     

Intraperitoneally injected cholecystokinin-octapeptide activates pica in rats.

Cholecystokinin-octapeptide (CCK-8) was injected intraperitoneally into rats to see if it could cause them to eat kaolin (clay)--a pica behavior which has been shown to indicate gastric distress. In the first study, a single large dose of CCK-8 (20 micrograms/kg) failed to produce pica. In the second study, 4 smaller doses of CCK-8 (8 micrograms/kg), 30 min apart, produced significant ingestion of kaolin compared to the baseline condition of vehicle injections. The pica was comparable to that observed in another group of rats given a toxic dose of LiCl (127 mg/kg, IP). It is concluded that intraperitoneal injections of CCK-8 can induce a state of gastric distress in the rat.     

Appropriateness of kaolin consumption as an index of motion sickness in the rat

The appropriateness of kaolin consumption, one form of pica, as an index of motion sickness in the rat was examined. Unlike other motion sickness indices, the use of kaolin consumption results in a bitonic function across daily rotation sessions. This bitonic function is not predicted from any theory of motion sickness (viz., the Sensory Rearrangement Theory), rather an inverse relationship should exist between the severity of motion sickness and repeated exposure to the effective motion (i.e., habituation). The results of Experiments 1 and 2 support the continued use of kaolin consumption as an index of motion sickness in the rat. A response interference process is proposed to account for the first portion of the bitonic kaolin consumption function with grooming possibly representing a higher probability behavior than kaolin consumption. Experiment 3 examined and confirmed that kaolin consumption indexes the process of rehabituation to an effective motion. This extends the number of principles that are characteristic of motion sickness exhibited by species capable of emesis and supports the continued use of kaolin consumption as an index of motion sickness and general gastrointestinal malaise in the rat.     

Poison induced pica in rats.

Two experiments investigating the effects of poisoning on the consumption of non-nutritive substances are reported. In the first experiment, rats were poisoned with lithium chloride or Red Squill and offered a choice between food and soil. In the second experiment, rats were poisoned with cyclophosphamide and offered a choice between food and kaolin. Following treatment, poisoned rats in both experiments increased their consumption of the non-nutritive substances. Additionally, rats poisoned with logarithmic doses consumed amounts of the non-nutritive substances proportionate to the amount of poison administered. It was suggested that increased pica is an illness-response behavior of the rat, analogous to vomiting in other species, which can be used as an easily quantifiable behavioral assay of noxious drug effects.     

Pica: a species relevant behavioral assay of motion sickness in the rat

Motion sickness was produced in rats by placing them in an apparatus that rotated about two different axes simultaneously. In three separate experiments, rats subject to rotational stimulation engaged in pica, the consumption of non-nutritive substances. The use of pica as a behavioral assay of motion sickness also demonstrated the principal characteristics of motion sickness shown by species in which emesis is used as the criterion of illness, including variability, summation, habituation, and recovery. The restuls indicate that pica is a naturally occurring response of the rat to illness, that might profitably be used to investigate the etiology of motion sickness and to screen for the emetic properties of drugs. It is suggested that the criterion of emesis insisted upon by most students of motion sickness be discarded in favor of species relevant criteria.     

Anti-emetic activity of ghrelin in ferrets exposed to the cytotoxic anti-cancer agent cisplatin

Emesis may be modulated via multiple mechanisms. The actions of ghrelin suggest an ability to couple an induction of hunger with preparation of the stomach for ingestion of food. Such a process might reduce any tendency to vomit, so an anti-emetic activity of ghrelin was investigated in the ferret cisplatin-induced emesis model. In controls, intra-peritoneal cisplatin (10 mg/kg) induced 41.4+/-8.4 episodes of emesis comprising 310.4+/-55.3 retches and 28.8+/-6.9 vomits during the 6h observation; the latency to onset of the first emetic episode was 108.9+/-4.8 min. Intra-peritoneal ghrelin (1mg/kg, split as a 30 min pre- and 30 min-post dose) did not induce a change in behaviour or modify cisplatin-induced emesis (p>0.05). Intracerebroventricular (i.c.v.) administration (third ventricle) was achieved via a pre-implanted cannula. At the first emetic episode following cisplatin, ghrelin or vehicle (20 microl saline) was administered i.c.v. During the 30 min following the initial episode of emesis, control animals exhibited 18.0+/-2.6 emetic episodes comprising 160.3+/-24.1 retches and 13.8+/-2.7 vomits. Ghrelin 10 microg i.c.v. reduced the number of retches by 61.5% (p<0.05) and at a dose of 30 microg i.c.v. ghrelin reduced the number of episodes, individual retches and vomits by 74.4 (p<0.05), 80.4 (p<0.01), and 72.5% (p<0.05), respectively. At subsequent time periods there were no differences between ghrelin- or saline-treated animals (p>0.05). An ability of ghrelin to reduce emesis is consistent with a role in modulating gastro-intestinal functions and identifies a novel approach to the treatment of emesis.     

Antinociceptive,Anti-Inflammatory and Antipyretic Activities of the Leaf Methanol Extract of Ruta Graveolens L. (Rutaceae) in Mice and Rats

Background

Ruta graveolens has been used to treat toothache, earache, rheumatism and fever with little scientific evidence corroborating these uses.

Materials and Methods

The leaf methanol extract of Ruta graveolens was evaluated for antinociceptive activity using the acetic acid writhing and hot-plate tests in mice, also anti-inflammatory and antipyretic activities using the carrageenan-induced oedema and E. coli-induced pyrexia tests in rats, respectively.

Results

R. graveolens (100 mg/kg, i.p.), significantly reduced the number of acetic acid-induced writhes by 54 %. R. graveolens (400 mg/kg, i.p.), significantly delayed the reaction time in mice to thermal stimulation 15, 30, 45, and 60 min after treatment. Combined treatment of the lowest and sub-effective doses of the leaf methanol extract (25 mg/kg, i.p.), and indomethacin (10 mg/kg, i.p.) significantly reduced the number of acetic acid-induced writhes in mice. The leaf methanol extract of R. graveolens (50 – 400 mg/kg, i.p.), significantly reduced carrageenan-induced oedema over the 4 h period of testing. Combined treatment of the lowest doses of R. graveolens (25 mg/kg, i.p.) and indomethacin (2 mg/kg, i.p.) produced a significant reduction in carrageenan-induced oedema over the 4 h period of testing. R. graveolens (100 -400 mg/kg, i.p.) significantly reduced E. coli-induced pyrexia over the 5 h period of testing. Given together, the lowest dose of R. graveolens (25 mg/kg, i.p.) and pentoxifylline (10 mg/kg, i.p.) produced a significant reduction in pyrexia induced by E. coli (50 µg/kg, i.m.) over the 5 h period of measurement. The LD₅₀ value obtained for R. graveolens was greater than 4000mg/kg (p.o), suggesting that the plant species may be safe in or nontoxic to mice.

Conclusion

The data obtained indicate that R. graveolens has antinociceptive, anti-inflammatory and antipyretic activities, justifying the use of the plant species by traditional medicine practitioners in the management and treatment of pain, inflammation and fever.     

The compartment model for chronically implanted voltammetric electrodes in the rat brain

Diazepam (0.4-0.8 mg/kg i.p.) reduced the spontaneous activity in the electromyogram (EMG) recorded from the gastrocnemius-soleus muscle of spastic mutant Han--Wistar rats in a dose-dependent manner. The depressant effect of diazepam (0.8 mg/kg) on the EMG activity was antagonized by Ro 15-1788 (5 mg/kg i.p.) and by picrotoxin (2 mg/kg i.p.), but not by bicuculline (2 mg/kg i.p.). These results suggest that diazepam depresses EMG activity of mutant rats by mechanisms related to the interaction of the drug with GABA-independent benzodiazepine receptors.     

Neuropharmacological profile of aqueous extract of Anaphe venata larva (Notondotidae) in rats

Consumption of Anaphe larva had been reported to cause seasonal ataxia and impaired consciousness. Therefore this study examined the neuropharmacological and mechanism(s) of action of aqueous extract of Anaphe venata in rats. Behavioural effects namely rearing, stretching, sniffing and ataxia were determined after the intraperitoneal administration of aqueous extract of Anaphe larva in rats. Animals were divided into groups and graded doses (100, 200 and 400 mg/kg, i.p.) of extract were administered. The control group was administered normal saline (vehicle). The effects of scopolamine (3 mg/kg, i.p.), flumazenil (2 mg/kg, i.p.), naloxone (2.5 mg/kg, i.p.), and thiamine (1 mg/kg, i.p.) on the observed behavioral changes were also examined. The effects of the extract administered intraperitoneally at a dose of 200 mg/kg on the amphetamine-induced stereotypy and locomotion were evaluated. Aqueous anaphe extract induced significant (p< 0.01) stretching and ataxia behavioural effects while it inhibited rearing behaviour when compared with the vehicle-treated group. However, it had no significant effect on sniffing behaviour. Scopolamine reversed all the effects of the extract on rearing, stretching and ataxia. Both Flumazenil and naloxone only reversed the effects of the extract on stretching and ataxia-induced behaviours significantly. However, thiamine potentiated both stretching and ataxia-induced behaviours. The extract inhibited the amphetamine-induced stereotype behaviour and locomotion. In conclusion, these results showed that these anaphe-induced behavioural effects are mediated via cholinergic, GABAergic, opioidergic and dopaminergic receptor systems with strong muscarinic-cholinergic receptors involvement in ataxia-induced behaviour. We therefore suggest that muscranic-cholinergic like drugs may be of benefit in the management of patients that present with clinical condition of seasonal ataxia.     

N-methyl-D-aspartate receptors mediate activation of the c-fos proto-oncogene in a model of brain injury

The proto-oncogene c-fos is rapidly and transiently induced in the CNS by a variety of stimuli. Brain injury, disruption of pia-arachnoid in a limited area, is one of the situations that leads to a dramatic increase in c-fos immunoreactivity. This increase is limited to the lesioned hemisphere. Injections of atropine (25 mg/kg, i.p.), naltrexone (5 mg/kg, i.p.), nifedipine (5 mg/kg, i.p.), and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (20 mg/kg, i.p.), prior to the injury, did not affect the activation of c-fos as assessed by immunohistochemistry in adult Sprague-Dawley rats perfused 2 h after the lesion. The non-competitive N-methyl-D-aspartate antagonists ketamine (100 mg/kg, i.p.) and MK-801 [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate] (1 and 3 mg/kg, i.p.) markedly reduced c-fos activation. Phencyclidine (10 mg/kg, i.p.) produced a slight reduction in damage-induced fos activation. This study suggests that c-fos activation in this particular model is N-methyl-D-aspartate receptor-mediated and supports the idea that the fos proto-oncogene might play a role in plasticity and/or neurotoxic changes following brain damage.     

Modulation of short-term social memory in rats by adenosine A1 and A(2A) receptors

The recognition of an unfamiliar juvenile rat by an adult rat has been shown to imply short-term memory processes. The present study was designed to examine the role of adenosine receptors in the short-term social memory of rats using the social recognition paradigm. Adenosine (5.0-10.0 mg/kg), the selective adenosine A1 receptor agonist 2-chloro-N6-cyclopentyladenosine (CCPA, 0.025-0.05 mg/kg) and the selective adenosine A(2A) receptor agonist N6-[2-(3,5-dimethoxyphenyl)-2-(2-methylphenyl)ethyl]adenosine (DPMA, 1.0-5.0 mg/kg), given by i.p. route 30 min before the test, disrupted the juvenile recognition ability of adult rats. This negative effect of adenosine (5.0 mg/kg, i.p.) on social memory was prevented by pretreatment with the non-selective adenosine receptor antagonist caffeine (10.0 mg/kg, i.p.), the adenosine A1 antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 1.0 mg/kg, i.p.) and the adenosine A(2A) antagonist 4-(2-[7-amino-2-{2-furyl}{1,2,4}triazolo-{2,3-a}{1,3,5}triazin-5-yl-amino]ethyl)phenol (ZM241385, 1.0 mg/kg, i.p.). Furthermore, acute administration of caffeine (10.0-30.0 mg/kg, i.p.), DPCPX (1.0-3.0 mg/kg, i.p.) or ZM241385 (0.5-1.0 mg/kg, i.p.) improved the short-term social memory in a specific manner. These results indicate that adenosine modulates the short-term social memory in rats by acting on both A1 and A(2A) receptors, with adenosine receptor agonists and antagonists, respectively, disrupting and enhancing the social memory.     

Cisplatin-induced long-term dynorphin A-immunoreactivity in cell somata of rat area postrema neurons

We evaluated long-term dynorphin A-immunoreactivity in the rat area postrema (AP) after the administration of cisplatin. First, rats were given 1, 5 and 10 mg/kg body weight cisplatin (i.p.) and their behavior was monitored for 72 h. We observed a delayed increase in pica 24-72 h after injection, compared to the 24 h before injection. We attributed this to the cisplatin injection. Pica was defined as an increase in the intake of non-nutritional matter such as kaolin. Administration of 1, 5 and 10 mg/kg cisplatin led to an increase in kaolin intake on day 1. Administration of 5 and 10 mg/kg of cisplatin led to decreased intake of laboratory chow (MF) on days 1–3, but 10 mg/kg cisplatin causes an excessive aggravation of their condition. Following this behavioral experiment, we immunohistochemically examined the induction of dynorphin A in the AP at 24, 48 and 72 h post-administration of 1 and 5 mg/kg cisplatin. Administration of 5 mg/kg cisplatin caused dynorphin A to accumulate gradually in the neurosoma of the AP neurons, and the numbers of positive AP neurosomata at 48 and 72 h post-administration were higher than following an equal dosage of 0.9% NaCl. These findings suggest that dynorphin A increases in the central nervous system for a long time following administration, and causes certain behavioral and clinical changes, including those related to appetite and nausea.     

Effects of N-methyl-d-aspartate (dizocilpine) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (LY215490) receptor antagonists on the voiding reflex induced by perineal stimulation in the neonatal rat

The present study was undertaken to examine the role of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate and N-methyl-D-aspartate glutamate receptors in the regulation of voiding reflexes induced by perineal stimulation in the neonatal rat. Four-, six- and 10-day-old awake rats were used in the experiments and perineal stimulation was applied using the tip of a 1-ml syringe to evoke voiding. Voided volume and residual volume were measured. In 10-day-old rats, LY215490 (3-10 mg/kg, i.p.), a competitive alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor antagonist, significantly inhibited reflex voiding, increasing the residual volume 34-53-fold. A 3 mg/kg dose decreased the urine release by 55%, whereas 10 mg/kg totally suppressed the voiding reflex induced by the perineal stimulation. LY215490 (10 mg/kg, i.p.) produced similar effects in four- and six-day-old rats. Dizocilpine (1-3 mg/kg, i.p.), a non-competitive N-methyl-D-aspartate receptor antagonist, also significantly decreased the urine release (62-82%) and increased residual volume (180-230-fold). Combined administration of LY215490 (1 mg/kg, i.p.) and dizocilpine (0.3 mg/kg, i.p.) to 10-day-old rats, in doses that individually had no effect on perineal stimulation-evoked voiding, depressed voided volume by 65%. These results indicate that, in neonatal rats, glutamatergic transmission in the spinal cord has an essential role in reflex micturition induced by perineal stimulation, and that facilitatory interactions between alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate and N-methyl-D-aspartate glutamatergic mechanisms are important for voiding, as noted previously in adult rats.     

<Emphasis Type="Italic">Scutellaria baicalensis</Emphasis> decreases ritonavir-induced nausea

Background  

Protease inhibitors, particularly ritonavir, causes significant gastrointestinal disturbances such as nausea, even at low doses. This ritonavir-induced nausea could be related to its oxidative stress in the gut. Alleviation of drug-induced nausea is important in effectively increasing patients' compliance and improving their quality of life. Conventional anti-emetic drugs can only partially abate the symptoms in these patients, and their cost has also been a concern. Rats respond to nausea-producing emetic stimuli by increasing consumption of non-nutritive substances like kaolin or clay, a phenomenon known as pica. In this study, we used this rat pica model to evaluate the effects of Scutellaria baicalensis, a commonly used oriental herbal medicine, on ritonavir-induced nausea.     

Chronic morphine treatment exaggerates the suppressive effects of sucrose and cocaine, but not lithium chloride, on saccharin intake in Sprague-Dawley rats

Three experiments examined the effect of chronic morphine treatment on cocaine-, sucrose-, and lithium chloride (LiCl)-induced suppression of saccharin intake in Sprague-Dawley rats. All rats were either water- or food-deprived and then implanted subcutaneously with 1 morphine (75 mg) or vehicle pellet for 5 days. They were then given brief access to 0.15% saccharin and soon thereafter injected with either cocaine (10 mg/kg s.c.), LiCl (0.009 M, 1.33 ml/100 g body weight i.p.), or saline, or, in Experiment 2, given a 2nd access period to either a preferred 1.0 M sucrose solution or the same 0.15% saccharin solution. There was 1 taste-drug or taste-taste pairing per day for a number of days. The results showed that a history of chronic morphine treatment exaggerated the suppressive effects of a rewarding sucrose solution and cocaine but not those of the aversive agent, LiCl. These data provide further support for the reward comparison hypothesis.     

Antiamnesic properties of some neuropeptides and the involvement of neurotransmitters in the rats.

The effects of certain neuropeptides on electroconvulsive (ECS) shock-induced amnesia were studied in rats. The ECS was applied immediately after the learning a one-trial passive avoidance paradigm. The peptides--rat atrial natriuretic peptide (rANP 1-28, ANP), porcine brain natriuretic peptide (pBNP 1-32, BNP), rat calcitonin gene-related peptide (CGRP) and bombesin--were injected into the lateral brain ventricle 30 min after the ECS. In order to study the possible role of neurotransmitters in mediating the action of the peptides on amnesia, the animals were treated immediately after the ECS with, doses which themselves did not modify either the learning itself or the amnesic action of the ECS. All the above peptides attenuated the ECS-induced amnesia, but the transmitters involved in these actions differed. The anticonvulsive action of ANP was prevented by haloperidol (10 micrograms/kg i. p.), propranolol (10 mg/kg i. p.) and atropine (2 mg/kg i. p.). Phenoxybenzamine (2.0 mg/kg i. p.), bicuculline (1 mg/kg i. p.), methysergide (5 mg/kg i. p.) and naloxone (0.3 mg/kg i. p.) had no effect. Besides alpha-adrenergic and cholinergic receptor blockers the beta-adrenergic blocker propranolol was also effective in preventing the antiamnesic action of the BNP. As concerns the action of bombesin, only haloperidol was effective. Alpha- and beta-adrenergic and cholinergic receptor blockers and opiates are involved in the antiamnesic properties of CGRP. The results showed that, despite the fact that the studied, peptides attenuated the ECS-induced amnesia, different transmitters are involved in their action.     

Protease inhibitor-induced nausea and vomiting is attenuated by a peripherally acting, opioid-receptor antagonist in a rat model

Background  

Protease inhibitors such as ritonavir can cause nausea and vomiting which is the most common reason for discontinuation. Rats react to nauseous and emetic stimuli by increasing their oral intake of non-nutritive substances like kaolin, known as pica behavior. In this study, we evaluated the effects of methylnaltrexone, a peripherally acting mu-opioid receptor antagonist that does not affect analgesia, on ritonavir-induced nausea and vomiting in a rat pica model.     

Peripheral administration of CDP-choline and its cholinergic metabolites increases serum insulin: muscarinic and nicotinic acetylcholine receptors are both involved in their actions

The present study was designed to test the effects of CDP-choline and its metabolites on serum insulin concentrations in rats and to investigate the involvements of cholinergic and adrenergic receptors in the effect. Intraperitoneal (i.p.) administration of CDP-choline (200-600 micromol/kg) increased serum insulin in a dose- and time-related manner. Equivalent doses (200-600 micromol/kg; i.p.) of phosphocholine or choline also increased serum insulin dose-dependently. Serum-free choline concentrations increased several-fold following i.p. administration of CDP-choline, phosphocholine or choline itself. In contrast, equivalent doses of cytidine monophosphate and cytidine failed to alter serum insulin concentrations. The increases in serum insulin induced by i.p. 600 micromol/kg of CDP-choline, phosphocholine or choline were abolished by pretreatment with the ganglionic nicotinic acetylcholine receptor antagonist hexamethonium (15 mg/kg; i.p.), or by the muscarinic receptor antagonist atropine methylnitrate (2 mg/kg; i.p.). Pretreatment with prazosin (0.5 mg/kg; i.p.), an alpha(1)-adrenoceptor antagonist, or yohimbine (5 mg/kg, i.p.), an alpha(2)-adrenoceptor antagonist, enhanced slightly the increases in serum insulin in response to 600 micromol/kg of CDP-choline, phosphocholine and choline. Serum insulin also increased following central administration of choline; the effect was blocked by intracerebroventricularly injected atropine, mecamylamine or hemicholinium-3 (HC-3). It is concluded that CDP-choline or its cholinergic metabolites phosphocholine and choline increases circulating insulin concentrations by increasing muscarinic and nicotinic cholinergic neurotransmission in the insulin secreting beta-cells.